METHODS AND COMPOSITIONS FOR DIAGNOSIS AND PROGNOSIS OF RENAL

INJURY AND RENAL FAILURE

[0001] The present application claims the benefit of U.S. Provisional Patent Application 62/455,503 filed February 6, 2017, which is hereby incorporated by reference in its entirety including all tables, figures and claims.

BACKGROUND OF THE INVENTION

[0002] The following discussion of the background of the invention is merely provided to aid the reader in understanding the invention and is not admitted to describe or constitute prior art to the present invention.

[0003] The kidney is responsible for water and solute excretion from the body. Its functions include maintenance of acid-base balance, regulation of electrolyte concentrations, control of blood volume, and regulation of blood pressure. As such, loss of kidney function through injury and/or disease results in substantial morbidity and mortality. A detailed discussion of renal injuries is provided in Harrison's Principles of Internal Medicine, 17 ^th Ed., McGraw Hill, New York, pages 1741-1830, which are hereby incorporated by reference in their entirety. Renal disease and/or injury may be acute or chronic. Acute and chronic kidney disease are described as follows (from Current Medical Diagnosis & Treatment 2008, 47 ^th Ed, McGraw Hill, New York, pages 785-815, which are hereby incorporated by reference in their entirety): "Acute renal failure is worsening of renal function over hours to days, resulting in the retention of nitrogenous wastes (such as urea nitrogen) and creatinine in the blood. Retention of these substances is called azotemia. Chronic renal failure (chronic kidney disease) results from an abnormal loss of renal function over months to years".

[0004] Acute renal failure (ARF, also known as acute kidney injury, or AKI) is an abrupt (typically detected within about 48 hours to 1 week) reduction in glomerular filtration. This loss of filtration capacity results in retention of nitrogenous (urea and creatinine) and non- nitrogenous waste products that are normally excreted by the kidney, a reduction in urine output, or both. It is reported that ARF complicates about 5% of hospital admissions, 4-15% of cardiopulmonary bypass surgeries, and up to 30% of intensive care admissions. ARF may be categorized as prerenal, intrinsic renal, or postrenal in causation. Intrinsic renal disease can be further divided into glomerular, tubular, interstitial, and vascular abnormalities. Major causes of ARF are described in the following table, which is adapted from the Merck Manual, 17 ed., Chapter 222, and which is hereby incorporated by reference in their entirety:

Type Risk Factors

Prerenal

ECF volume depletion Excessive diuresis, hemorrhage, GI losses, loss of

intravascular fluid into the extravascular space (due to ascites, peritonitis, pancreatitis, or burns), loss of skin and mucus membranes, renal salt- and water-wasting states

Low cardiac output Cardiomyopathy, MI, cardiac tamponade, pulmonary

embolism, pulmonary hypertension, positive-pressure mechanical ventilation

Low systemic vascular Septic shock, liver failure, antihypertensive drugs

resistance

Increased renal vascular NSAIDs, cyclosporines, tacrolimus, hypercalcemia, resistance anaphylaxis, anesthetics, renal artery obstruction, renal

vein thrombosis, sepsis, hepatorenal syndrome

Decreased efferent ACE inhibitors or angiotensin II receptor blockers arteriolar tone (leading to

decreased GFR from

reduced glomerular

transcapillary pressure,

especially in patients with

bilateral renal artery

stenosis)

Intrinsic Renal

Acute tubular injury Ischemia (prolonged or severe prerenal state): surgery,

hemorrhage, arterial or venous obstruction; Toxins:

NSAIDs, cyclosporines, tacrolimus, aminoglycosides, foscarnet, ethylene glycol, hemoglobin, myoglobin, ifosfamide, heavy metals, methotrexate, radiopaque contrast agents, streptozotocin

Acute glomerulonephritis ANCA-associated: Crescentic glomerulonephritis,

polyarteritis nodosa, Wegener's granulomatosis; Anti- GBM glomerulonephritis: Goodpasture's syndrome;

Immune-complex: Lupus glomerulonephritis,

postinfectious glomerulonephritis, cryoglobulinemic glomerulonephritis

Acute tubulointerstitial Drug reaction (eg, β-lactams, NSAIDs, sulfonamides, nephritis ciprofloxacin, thiazide diuretics, furosemide, phenytoin, allopurinol, pyelonephritis, papillary necrosis

Acute vascular Vasculitis, malignant hypertension, thrombotic

nephropathy microangiopathies, scleroderma, atheroembolism

Infiltrative diseases Lymphoma, sarcoidosis, leukemia

Postrenal

Tubular precipitation Uric acid (tumor lysis), sulfonamides, triamterene,

acyclovir, indinavir, methotrexate, ethylene glycol ingestion, myeloma protein, myoglobin Type Risk Factors

Ureteral obstruction Intrinsic: Calculi, clots, sloughed renal tissue, fungus

ball, edema, malignancy, congenital defects; Extrinsic:

Malignancy, retroperitoneal fibrosis, ureteral trauma

during surgery or high impact injury

Bladder obstruction Mechanical: Benign prostatic hyperplasia, prostate

cancer, bladder cancer, urethral strictures, phimosis,

paraphimosis, urethral valves, obstructed indwelling

urinary catheter; Neurogenic: Anticholinergic drugs, upper or lower motor neuron lesion

[0005] In the case of ischemic ARF, the course of the disease may be divided into four phases. During an initiation phase, which lasts hours to days, reduced perfusion of the kidney is evolving into injury. Glomerular ultrafiltration reduces, the flow of filtrate is reduced due to debris within the tubules, and back leakage of filtrate through injured epithelium occurs. Renal injury can be mediated during this phase by reperfusion of the kidney. Initiation is followed by an extension phase which is characterized by continued ischemic injury and inflammation and may involve endothelial damage and vascular congestion. During the maintenance phase, lasting from 1 to 2 weeks, renal cell injury occurs, and glomerular filtration and urine output reaches a minimum. A recovery phase can follow in which the renal epithelium is repaired and GFR gradually recovers. Despite this, the survival rate of subjects with ARF may be as low as about 60%.

[0006] Acute kidney injury caused by radiocontrast agents (also called contrast media) and other nephrotoxins such as cyclosporine, antibiotics including aminoglycosides and anticancer drugs such as cisplatin manifests over a period of days to about a week. Contrast induced nephropathy (CIN, which is AKI caused by radiocontrast agents) is thought to be caused by intrarenal vasoconstriction (leading to ischemic injury) and from the generation of reactive oxygen species that are directly toxic to renal tubular epithelial cells. CIN classically presents as an acute (onset within 24-48h) but reversible (peak 3-5 days, resolution within 1 week) rise in blood urea nitrogen and serum creatinine.

[0007] A commonly reported criteria for defining and detecting AKI is an abrupt (typically within about 2-7 days or within a period of hospitalization) elevation of serum creatinine. Although the use of serum creatinine elevation to define and detect AKI is well established, the magnitude of the serum creatinine elevation and the time over which it is measured to define AKI varies considerably among publications. Traditionally, relatively large increases in serum creatinine such as 100%, 200%, an increase of at least 100% to a value over 2 mg/dL and other definitions were used to define AKI. However, the recent trend has been towards using smaller serum creatinine rises to define AKI. The relationship between serum creatinine rise, AKI and the associated health risks are reviewed in Praught and Shlipak, Curr Opin Nephrol Hypertens 14:265-270, 2005 and Chertow et al, J Am Soc Nephrol 16: 3365-3370, 2005, which, with the references listed therein, are hereby incorporated by reference in their entirety. As described in these publications, acute worsening renal function (AKI) and increased risk of death and other detrimental outcomes are now known to be associated with very small increases in serum creatinine. These increases may be determined as a relative (percent) value or a nominal value. Relative increases in serum creatinine as small as 20% from the pre-injury value have been reported to indicate acutely worsening renal function (AKI) and increased health risk, but the more commonly reported value to define AKI and increased health risk is a relative increase of at least 25%. Nominal increases as small as 0.3 mg/dL, 0.2 mg/dL or even 0.1 mg/dL have been reported to indicate worsening renal function and increased risk of death. Various time periods for the serum creatinine to rise to these threshold values have been used to define AKI, for example, ranging from 2 days, 3 days, 7 days, or a variable period defined as the time the patient is in the hospital or intensive care unit. These studies indicate there is not a particular threshold serum creatinine rise (or time period for the rise) for worsening renal function or AKI, but rather a continuous increase in risk with increasing magnitude of serum creatinine rise.

[0008] One study (Lassnigg et all, J Am Soc Nephrol 15: 1597-1605, 2004, hereby incorporated by reference in its entirety) investigated both increases and decreases in serum creatinine. Patients with a mild fall in serum creatinine of -0.1 to -0.3 mg/dL following heart surgery had the lowest mortality rate. Patients with a larger fall in serum creatinine (more than or equal to -0.4 mg/dL) or any increase in serum creatinine had a larger mortality rate. These findings caused the authors to conclude that even very subtle changes in renal function (as detected by small creatinine changes within 48 hours of surgery) seriously effect patient's outcomes. In an effort to reach consensus on a unified classification system for using serum creatinine to define AKI in clinical trials and in clinical practice, Bellomo et ah, Crit Care. 8(4):R204-12, 2004, which is hereby incorporated by reference in its entirety, proposes the following classifications for stratifying AKI patients:

"Risk": serum creatinine increased 1.5 fold from baseline OR urine production of <0.5 ml/kg body weight/hr for 6 hours;

"Injury": serum creatinine increased 2.0 fold from baseline OR urine production <0.5 ml/kg/hr for 12 h; "Failure": serum creatinine increased 3.0 fold from baseline OR creatinine >355 μιηοΐ/ΐ (with a rise of >44) or urine output below 0.3 ml/kg/hr for 24 h or anuria for at least 12 hours;

And included two clinical outcomes:

"Loss": persistent need for renal replacement therapy for more than four weeks.

"ESRD": end stage renal disease— the need for dialysis for more than 3 months.

These criteria are called the RIFLE criteria, which provide a useful clinical tool to classify renal status. As discussed in Kellum, Crit. Care Med. 36: S 141-45, 2008 and Ricci et al., Kidney Int. 73, 538-546, 2008, each hereby incorporated by reference in its entirety, the RIFLE criteria provide a uniform definition of AKI which has been validated in numerous studies.

[0009] More recently, Mehta et al, Crit. Care 11:R31 (doi: 10.1186.cc5713), 2007, hereby incorporated by reference in its entirety, proposes the following similar classifications for stratifying AKI patients, which have been modified from RIFLE:

"Stage I": increase in serum creatinine of more than or equal to 0.3 mg/dL (> 26.4 μιηοΙ/L) or increase to more than or equal to 150% (1.5-fold) from baseline OR urine output less than 0.5 mL/kg per hour for more than 6 hours;

"Stage Π": increase in serum creatinine to more than 200% (> 2-fold) from baseline OR urine output less than 0.5 mL/kg per hour for more than 12 hours;

"Stage III": increase in serum creatinine to more than 300% (> 3-fold) from baseline OR serum creatinine > 354 μιηοΙ/L accompanied by an acute increase of at least 44 μιηοΙ/L OR urine output less than 0.3 mL/kg per hour for 24 hours or anuria for 12 hours.

[0010] The CIN Consensus Working Panel (McCollough et al, Rev Cardiovasc Med.

2006;7(4):177-197, hereby incorporated by reference in its entirety? uses a serum creatinine rise of 25% to define Contrast induced nephropathy (which is a type of

AKI) .Although various groups propose slightly different criteria for using serum

creatinine to detect AKI, the consensus is that small changes in serum creatinine, such as 0.3 mg/dL or 25%, are sufficient to detect AKI (worsening renal function) and that the magnitude of the serum creatinine change is an indicator of the severity of the AKI and mortality risk.

[0011] Although serial measurement of serum creatinine over a period of days is an accepted method of detecting and diagnosing AKI and is considered one of the most important tools to evaluate AKI patients, serum creatinine is generally regarded to have several limitations in the diagnosis, assessment and monitoring of AKI patients. The time period for serum creatinine to rise to values (e.g., a 0.3 mg/dL or 25% rise) considered diagnostic for AKI can be 48 hours or longer depending on the definition used. Since cellular injury in AKI can occur over a period of hours, serum creatinine elevations detected at 48 hours or longer can be a late indicator of injury, and relying on serum creatinine can thus delay diagnosis of AKI.

Furthermore, serum creatinine is not a good indicator of the exact kidney status and treatment needs during the most acute phases of AKI when kidney function is changing rapidly. Some patients with AKI will recover fully, some will need dialysis (either short term or long term) and some will have other detrimental outcomes including death, major adverse cardiac events and chronic kidney disease. Because serum creatinine is a marker of filtration rate, it does not differentiate between the causes of AKI (pre-renal, intrinsic renal, post-renal obstruction, atheroembolic, etc) or the category or location of injury in intrinsic renal disease (for example, tubular, glomerular or interstitial in origin). Urine output is similarly limited, Knowing these things can be of vital importance in managing and treating patients with AKI.

[0012] These limitations underscore the need for better methods to detect and assess AKI, particularly in the early and subclinical stages, but also in later stages when recovery and repair of the kidney can occur. Furthermore, there is a need to better identify patients who are at risk of having an AKI.

BRIEF SUMMARY OF THE INVENTION

[0013] It is an object of the invention to provide methods and compositions for evaluating renal function in a subject. As described herein, measurement of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24 (collectively referred to herein as "kidney injury markers, and individually as a "kidney injury marker") can be used for diagnosis, prognosis, risk stratification, staging, monitoring, categorizing and determination of further diagnosis and treatment regimens in subjects suffering or at risk of suffering from an injury to renal function, reduced renal function, and/or acute renal failure (also called acute kidney injury).

[0014] These kidney injury markers may be used, individually or in panels comprising a plurality of kidney injury markers, for risk stratification (that is, to identify subjects at risk for a future injury to renal function, for future progression to reduced renal function, for future progression to ARF, for future improvement in renal function, etc.); for diagnosis of existing disease (that is, to identify subjects who have suffered an injury to renal function, who have progressed to reduced renal function, who have progressed to ARF, etc.); for monitoring for deterioration or improvement of renal function; and for predicting a future medical outcome, such as improved or worsening renal function, a decreased or increased mortality risk, a decreased or increased risk that a subject will require renal replacement therapy (i.e., hemodialysis, peritoneal dialysis, hemofiltration, and/or renal transplantation, a decreased or increased risk that a subject will recover from an injury to renal function, a decreased or increased risk that a subject will recover from ARF, a decreased or increased risk that a subject will progress to end stage renal disease, a decreased or increased risk that a subject will progress to chronic renal failure, a decreased or increased risk that a subject will suffer rejection of a transplanted kidney, etc.

[0015] In a first aspect, the present invention relates to methods for evaluating renal status in a subject. These methods comprise performing an assay method that is configured to detect one or more kidney injury markers of the present invention in a body fluid sample obtained from the subject. The assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are then correlated to the renal status of the subject. This correlation to renal status may include correlating the assay result(s) to one or more of risk stratification, diagnosis, prognosis, staging, classifying and monitoring of the subject as described herein. Thus, the present invention utilizes one or more kidney injury markers of the present invention for the evaluation of renal injury.

[0016] In certain embodiments, the methods for evaluating renal status described herein are methods for risk stratification of the subject; that is, assigning a likelihood of one or more future changes in renal status to the subject. In these embodiments, the assay result(s) is/are correlated to one or more such future changes. The following are preferred risk stratification embodiments.

[0017] In preferred risk stratification embodiments, these methods comprise determining a subject' s risk for a future injury to renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of

Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a likelihood of such a future injury to renal function. For example, the measured concentration(s) may each be compared to a threshold value. For a "positive going" kidney injury marker, an increased likelihood of suffering a future injury to renal function is assigned to the subject when the measured concentration is above the threshold, relative to a likelihood assigned when the measured concentration is below the threshold. For a "negative going" kidney injury marker, an increased likelihood of suffering a future injury to renal function is assigned to the subject when the measured concentration is below the threshold, relative to a likelihood assigned when the measured concentration is above the threshold.

[0018] In other preferred risk stratification embodiments, these methods comprise determining a subject's risk for future reduced renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a likelihood of such reduced renal function. For example, the measured concentrations may each be compared to a threshold value. For a "positive going" kidney injury marker, an increased likelihood of suffering a future reduced renal function is assigned to the subject when the measured concentration is above the threshold, relative to a likelihood assigned when the measured concentration is below the threshold. For a "negative going" kidney injury marker, an increased likelihood of future reduced renal function is assigned to the subject when the measured concentration is below the threshold, relative to a likelihood assigned when the measured concentration is above the threshold.

[0019] In still other preferred risk stratification embodiments, these methods comprise determining a subject's likelihood for a future improvement in renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a likelihood of such a future improvement in renal function. For example, the measured concentration(s) may each be compared to a threshold value. For a "positive going" kidney injury marker, an increased likelihood of a future improvement in renal function is assigned to the subject when the measured concentration is below the threshold, relative to a likelihood assigned when the measured concentration is above the threshold. For a "negative going" kidney injury marker, an increased likelihood of a future improvement in renal function is assigned to the subject when the measured concentration is above the threshold, relative to a likelihood assigned when the measured concentration is below the threshold.

[0020] In yet other preferred risk stratification embodiments, these methods comprise determining a subject's risk for progression to ARF, and the result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a likelihood of such progression to ARF. For example, the measured

concentration(s) may each be compared to a threshold value. For a "positive going" kidney injury marker, an increased likelihood of progression to ARF is assigned to the subject when the measured concentration is above the threshold, relative to a likelihood assigned when the measured concentration is below the threshold. For a "negative going" kidney injury marker, an increased likelihood of progression to ARF is assigned to the subject when the measured concentration is below the threshold, relative to a likelihood assigned when the measured concentration is above the threshold.

[0021] And in other preferred risk stratification embodiments, these methods comprise determining a subject's outcome risk, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a likelihood of the occurrence of a clinical outcome related to a renal injury suffered by the subject. For example, the measured concentration(s) may each be compared to a threshold value. For a "positive going" kidney injury marker, an increased likelihood of one or more of: acute kidney injury, progression to a worsening stage of AKI, mortality, a requirement for renal replacement therapy, a requirement for withdrawal of renal toxins, end stage renal disease, heart failure, stroke, myocardial infarction, progression to chronic kidney disease, etc., is assigned to the subject when the measured concentration is above the threshold, relative to a likelihood assigned when the measured concentration is below the threshold. For a "negative going" kidney injury marker, an increased likelihood of one or more of: acute kidney injury, progression to a worsening stage of AKI, mortality, a requirement for renal replacement therapy, a requirement for withdrawal of renal toxins, end stage renal disease, heart failure, stroke, myocardial infarction, progression to chronic kidney disease, etc., is assigned to the subject when the measured concentration is below the threshold, relative to a likelihood assigned when the measured concentration is above the threshold.

[0022] In such risk stratification embodiments, preferably the likelihood or risk assigned is that an event of interest is more or less likely to occur within 180 days of the time at which the body fluid sample is obtained from the subject. In particularly preferred embodiments, the likelihood or risk assigned relates to an event of interest occurring within a shorter time period such as 18 months, 120 days, 90 days, 60 days, 45 days, 30 days, 21 days, 14 days, 7 days, 5 days, 96 hours, 72 hours, 48 hours, 36 hours, 24 hours, 12 hours, or less. A risk at 0 hours of the time at which the body fluid sample is obtained from the subject is equivalent to diagnosis of a current condition.

[0023] In preferred risk stratification embodiments, the subject is selected for risk stratification based on the pre-existence in the subject of one or more known risk factors for prerenal, intrinsic renal, or postrenal ARF. For example, a subject undergoing or having undergone major vascular surgery, coronary artery bypass, or other cardiac surgery; a subject having pre-existing congestive heart failure, preeclampsia, eclampsia, diabetes mellitus, hypertension, coronary artery disease, proteinuria, renal insufficiency, glomerular filtration below the normal range, cirrhosis, serum creatinine above the normal range, or sepsis; or a subject exposed to NSAIDs, cyclosporines, tacrolimus, aminoglycosides, foscarnet, ethylene glycol, hemoglobin, myoglobin, ifosfamide, heavy metals, methotrexate, radiopaque contrast agents, or streptozotocin are all preferred subjects for monitoring risks according to the methods described herein. This list is not meant to be limiting. By "pre-existence" in this context is meant that the risk factor exists at the time the body fluid sample is obtained from the subject. In particularly preferred embodiments, a subject is chosen for risk stratification based on an existing diagnosis of injury to renal function, reduced renal function, or ARF.

[0024] In other embodiments, the methods for evaluating renal status described herein are methods for diagnosing a renal injury in the subject; that is, assessing whether or not a subject has suffered from an injury to renal function, reduced renal function, or ARF. In these embodiments, the assay result(s) is/are correlated to the occurrence or nonoccurrence of a change in renal status. The following are preferred diagnostic embodiments. [0025] In preferred diagnostic embodiments, these methods comprise diagnosing the occurrence or nonoccurrence of an injury to renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of such an injury. For example, each of the measured concentration(s) may be compared to a threshold value. For a positive going marker, an increased likelihood of the occurrence of an injury to renal function is assigned to the subject when the measured concentration is above the threshold (relative to the likelihood assigned when the measured concentration is below the threshold); alternatively, when the measured concentration is below the threshold, an increased likelihood of the nonoccurrence of an injury to renal function may be assigned to the subject (relative to the likelihood assigned when the measured concentration is above the threshold). For a negative going marker, an increased likelihood of the occurrence of an injury to renal function is assigned to the subject when the measured concentration is below the threshold (relative to the likelihood assigned when the measured concentration is above the threshold); alternatively, when the measured concentration is above the threshold, an increased likelihood of the nonoccurrence of an injury to renal function may be assigned to the subject (relative to the likelihood assigned when the measured concentration is below the threshold).

[0026] In other preferred diagnostic embodiments, these methods comprise diagnosing the occurrence or nonoccurrence of reduced renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of

Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of an injury causing reduced renal function. For example, each of the measured concentration(s) may be compared to a threshold value. For a positive going marker, an increased likelihood of the occurrence of an injury causing reduced renal function is assigned to the subject when the measured concentration is above the threshold (relative to the likelihood assigned when the measured concentration is below the threshold); alternatively, when the measured concentration is below the threshold, an increased likelihood of the nonoccurrence of an injury causing reduced renal function may be assigned to the subject (relative to the likelihood assigned when the measured concentration is above the threshold). For a negative going marker, an increased likelihood of the occurrence of an injury causing reduced renal function is assigned to the subject when the measured concentration is below the threshold (relative to the likelihood assigned when the measured concentration is above the threshold); alternatively, when the measured concentration is above the threshold, an increased likelihood of the nonoccurrence of an injury causing reduced renal function may be assigned to the subject (relative to the likelihood assigned when the measured concentration is below the threshold).

[0027] In yet other preferred diagnostic embodiments, these methods comprise diagnosing the occurrence or nonoccurrence of ARF, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of an injury causing ARF. For example, each of the measured concentration(s) may be compared to a threshold value. For a positive going marker, an increased likelihood of the occurrence of ARF is assigned to the subject when the measured concentration is above the threshold (relative to the likelihood assigned when the measured concentration is below the threshold); alternatively, when the measured concentration is below the threshold, an increased likelihood of the nonoccurrence of ARF may be assigned to the subject (relative to the likelihood assigned when the measured concentration is above the threshold). For a negative going marker, an increased likelihood of the occurrence of ARF is assigned to the subject when the measured concentration is below the threshold (relative to the likelihood assigned when the measured concentration is above the threshold); alternatively, when the measured concentration is above the threshold, an increased likelihood of the nonoccurrence of ARF may be assigned to the subject (relative to the likelihood assigned when the measured concentration is below the threshold).

[0028] In still other preferred diagnostic embodiments, these methods comprise diagnosing a subject as being in need of renal replacement therapy, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of

Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a need for renal replacement therapy. For example, each of the measured concentration(s) may be compared to a threshold value. For a positive going marker, an increased likelihood of the occurrence of an injury creating a need for renal replacement therapy is assigned to the subject when the measured concentration is above the threshold (relative to the likelihood assigned when the measured concentration is below the threshold); alternatively, when the measured concentration is below the threshold, an increased likelihood of the nonoccurrence of an injury creating a need for renal replacement therapy may be assigned to the subject (relative to the likelihood assigned when the measured concentration is above the threshold). For a negative going marker, an increased likelihood of the occurrence of an injury creating a need for renal replacement therapy is assigned to the subject when the measured concentration is below the threshold (relative to the likelihood assigned when the measured concentration is above the threshold); alternatively, when the measured concentration is above the threshold, an increased likelihood of the nonoccurrence of an injury creating a need for renal replacement therapy may be assigned to the subject (relative to the likelihood assigned when the measured concentration is below the threshold).

[0029] In still other preferred diagnostic embodiments, these methods comprise diagnosing a subject as being in need of renal transplantation, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of

Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a need for renal transplantation. For example, each of the measured concentration(s) may be compared to a threshold value. For a positive going marker, an increased likelihood of the occurrence of an injury creating a need for renal transplantation is assigned to the subject when the measured concentration is above the threshold (relative to the likelihood assigned when the measured concentration is below the threshold); alternatively, when the measured concentration is below the threshold, an increased likelihood of the nonoccurrence of an injury creating a need for renal transplantation may be assigned to the subject (relative to the likelihood assigned when the measured concentration is above the threshold). For a negative going marker, an increased likelihood of the occurrence of an injury creating a need for renal transplantation is assigned to the subject when the measured concentration is below the threshold (relative to the likelihood assigned when the measured concentration is above the threshold); alternatively, when the measured concentration is above the threshold, an increased likelihood of the nonoccurrence of an injury creating a need for renal transplantation may be assigned to the subject (relative to the likelihood assigned when the measured concentration is below the threshold).

[0030] In still other embodiments, the methods for evaluating renal status described herein are methods for monitoring a renal injury in the subject; that is, assessing whether or not renal function is improving or worsening in a subject who has suffered from an injury to renal function, reduced renal function, or ARF. In these embodiments, the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of a change in renal status. The following are preferred monitoring embodiments.

[0031] In preferred monitoring embodiments, these methods comprise monitoring renal status in a subject suffering from an injury to renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of a change in renal status in the subject. For example, the measured concentration(s) may be compared to a threshold value. For a positive going marker, when the measured concentration is above the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is below the threshold, an improvement of renal function may be assigned to the subject. For a negative going marker, when the measured concentration is below the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is above the threshold, an improvement of renal function may be assigned to the subject.

[0032] In other preferred monitoring embodiments, these methods comprise monitoring renal status in a subject suffering from reduced renal function, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of a change in renal status in the subject. For example, the measured concentration(s) may be compared to a threshold value. For a positive going marker, when the measured concentration is above the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is below the threshold, an improvement of renal function may be assigned to the subject. For a negative going marker, when the measured concentration is below the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is above the threshold, an improvement of renal function may be assigned to the subject.

[0033] In yet other preferred monitoring embodiments, these methods comprise monitoring renal status in a subject suffering from acute renal failure, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of

Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of a change in renal status in the subject. For example, the measured concentration(s) may be compared to a threshold value. For a positive going marker, when the measured concentration is above the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is below the threshold, an improvement of renal function may be assigned to the subject. For a negative going marker, when the measured concentration is below the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is above the threshold, an improvement of renal function may be assigned to the subject.

[0034] In other additional preferred monitoring embodiments, these methods comprise monitoring renal status in a subject at risk of an injury to renal function due to the pre-existence of one or more known risk factors for prerenal, intrinsic renal, or postrenal ARF, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the occurrence or nonoccurrence of a change in renal status in the subject. For example, the measured concentration(s) may be compared to a threshold value. For a positive going marker, when the measured concentration is above the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is below the threshold, an improvement of renal function may be assigned to the subject. For a negative going marker, when the measured concentration is below the threshold, a worsening of renal function may be assigned to the subject; alternatively, when the measured concentration is above the threshold, an improvement of renal function may be assigned to the subject.

[0035] In yet other preferred monitoring embodiments, these methods comprise monitoring renal status in a subject having, or at risk of, an injury to renal function for future persistence of acute kidney injury. "Future persistence" as used herein refers to an existing acute renal injury that will continue for a period selected from the group consisting of 21 days, 14 days, 7 days, 5 days, 96 hours, 72 hours, 48 hours, 36 hours, 24 hours, and 12 hours. In certain embodiments the subject has an acute kidney injury at the time the sample is obtained. This is not meant to imply that the subject must have an acute kidney injury at the time the sample is obtained, but rather that the subject, upon onset of an acute kidney injury, suffers from an acute kidney injury that will persist. In various embodiments, the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the future persistence of the acute kidney injury in the subject. For example, the measured concentration(s) may be compared to a threshold value. For a positive going marker, when the measured concentration is above the threshold, a future persistence of acute kidney injury may be assigned to the subject; alternatively, when the measured concentration is below the threshold, afuture improvement of renal function may be assigned to the subject. For a negative going marker, when the measured concentration is below the threshold, a future persistence of acute kidney injury may be assigned to the subject; alternatively, when the measured concentration is above the threshold, a future improvement of renal function may be assigned to the subject.

[0036] In still other embodiments, the methods for evaluating renal status described herein are methods for classifying a renal injury in the subject; that is, determining whether a renal injury in a subject is prerenal, intrinsic renal, or postrenal; and/or further subdividing these classes into subclasses such as acute tubular injury, acute glomerulonephritis acute

tubulointerstitial nephritis, acute vascular nephropathy, or infiltrative disease; and/or assigning a likelihood that a subject will progress to a particular RIFLE stage. In these embodiments, the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2,

Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to a particular class and/or subclass. The following are preferred classification embodiments.

[0037] In preferred classification embodiments, these methods comprise determining whether a renal injury in a subject is prerenal, intrinsic renal, or postrenal; and/or further subdividing these classes into subclasses such as acute tubular injury, acute glomerulonephritis acute tubulointerstitial nephritis, acute vascular nephropathy, or infiltrative disease; and/or assigning a likelihood that a subject will progress to a particular RIFLE stage, and the assay result(s), for example a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2, Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24, is/are correlated to the injury classification for the subject. For example, the measured concentration may be compared to a threshold value, and when the measured concentration is above the threshold, a particular classification is assigned;

alternatively, when the measured concentration is below the threshold, a different classification may be assigned to the subject.

[0038] A variety of methods may be used by the skilled artisan to arrive at a desired threshold value for use in these methods. For example, the threshold value may be determined from a population of normal subjects by selecting a concentration representing the 75 ^th , 85 ^th , 90 ^th , 95 ^th , or 99 ^th percentile of a kidney injury marker measured in such normal subjects.

Alternatively, the threshold value may be determined from a "diseased" population of subjects, e.g., those suffering from an injury or having a predisposition for an injury (e.g., progression to ARF or some other clinical outcome such as death, dialysis, renal transplantation, etc.), by selecting a concentration representing the 75 ^th , 85 ^th , 90 ^th , 95 ^th , or 99 ^th percentile of a kidney injury marker measured in such subjects. In another alternative, the threshold value may be determined from a prior measurement of a kidney injury marker in the same subject; that is, a temporal change in the level of a kidney injury marker in the subject may be used to assign risk to the subject.

[0039] The foregoing discussion is not meant to imply, however, that the kidney injury markers of the present invention must be compared to corresponding individual thresholds. Methods for combining assay results can comprise the use of multivariate logistical regression, loglinear modeling, neural network analysis, n-of-m analysis, decision tree analysis, calculating ratios of markers, etc. This list is not meant to be limiting. In these methods, a composite result which is determined by combining individual markers may be treated as if it is itself a marker; that is, a threshold may be determined for the composite result as described herein for individual markers, and the composite result for an individual patient compared to this threshold.

[0040] The ability of a particular test to distinguish two populations can be established using ROC analysis. For example, ROC curves established from a "first" subpopulation which is predisposed to one or more future changes in renal status, and a "second" subpopulation which is not so predisposed can be used to calculate a ROC curve, and the area under the curve provides a measure of the quality of the test. Preferably, the tests described herein provide a ROC curve area greater than 0.5, preferably at least 0.6, more preferably 0.7, still more preferably at least 0.8, even more preferably at least 0.9, and most preferably at least 0.95.

[0041] In certain aspects, the measured concentration of one or more kidney injury markers, or a composite of such markers, may be treated as continuous variables. For example, any particular concentration can be converted into a corresponding probability of a future reduction in renal function for the subject, the occurrence of an injury, a classification, etc. In yet another alternative, a threshold that can provide an acceptable level of specificity and sensitivity in separating a population of subjects into "bins" such as a "first" subpopulation (e.g., which is predisposed to one or more future changes in renal status, the occurrence of an injury, a classification, etc.) and a "second" subpopulation which is not so predisposed. A threshold value is selected to separate this first and second population by one or more of the following measures of test accuracy: an odds ratio greater than 1, preferably at least about 2 or more or about 0.5 or less, more preferably at least about 3 or more or about 0.33 or less, still more preferably at least about 4 or more or about 0.25 or less, even more preferably at least about 5 or more or about 0.2 or less, and most preferably at least about 10 or more or about 0.1 or less; a specificity of greater than 0.5, preferably at least about 0.6, more preferably at least about 0.7, still more preferably at least about 0.8, even more preferably at least about 0.9 and most preferably at least about 0.95, with a corresponding sensitivity greater than 0.2, preferably greater than about 0.3, more preferably greater than about 0.4, still more preferably at least about 0.5, even more preferably about 0.6, yet more preferably greater than about 0.7, still more preferably greater than about 0.8, more preferably greater than about 0.9, and most preferably greater than about 0.95; a sensitivity of greater than 0.5, preferably at least about 0.6, more preferably at least about 0.7, still more preferably at least about 0.8, even more preferably at least about 0.9 and most preferably at least about 0.95, with a corresponding specificity greater than 0.2, preferably greater than about 0.3, more preferably greater than about 0.4, still more preferably at least about 0.5, even more preferably about 0.6, yet more preferably greater than about 0.7, still more preferably greater than about 0.8, more preferably greater than about 0.9, and most preferably greater than about 0.95; at least about 75% sensitivity, combined with at least about 75% specificity; a positive likelihood ratio (calculated as sensitivity/(l-specificity)) of greater than 1, at least about 2, more preferably at least about 3, still more preferably at least about 5, and most preferably at least about 10; or a negative likelihood ratio (calculated as (l-sensitivity)/specificity) of less than 1, less than or equal to about 0.5, more preferably less than or equal to about 0.3, and most preferably less than or equal to about 0.1.

The term "about" in the context of any of the above measurements refers to +/- 5% of a given measurement.

[0042] Multiple thresholds may also be used to assess renal status in a subject. For example, a "first" subpopulation which is predisposed to one or more future changes in renal status, the occurrence of an injury, a classification, etc., and a "second" subpopulation which is not so predisposed can be combined into a single group. This group is then subdivided into three or more equal parts (known as tertiles, quartiles, quin tiles, etc., depending on the number of subdivisions). An odds ratio is assigned to subjects based on which subdivision they fall into. If one considers a tertile, the lowest or highest tertile can be used as a reference for comparison of the other subdivisions. This reference subdivision is assigned an odds ratio of 1. The second tertile is assigned an odds ratio that is relative to that first tertile. That is, someone in the second tertile might be 3 times more likely to suffer one or more future changes in renal status in comparison to someone in the first tertile. The third tertile is also assigned an odds ratio that is relative to that first tertile.

[0043] In certain embodiments, the assay method is an immunoassay. Antibodies for use in such assays will specifically bind a full length kidney injury marker of interest, and may also bind one or more polypeptides that are "related" thereto, as that term is defined hereinafter. Numerous immunoassay formats are known to those of skill in the art. Preferred body fluid samples are selected from the group consisting of urine, blood, serum, saliva, tears, and plasma.

[0044] The foregoing method steps should not be interpreted to mean that the kidney injury marker assay result(s) is/are used in isolation in the methods described herein. Rather, additional variables or other clinical indicia may be included in the methods described herein. For example, a risk stratification, diagnostic, classification, monitoring, etc. method may combine the assay result(s) with one or more variables measured for the subject selected from the group consisting of demographic information (e.g., weight, sex, age, race), medical history (e.g., family history, type of surgery, pre-existing disease such as aneurism, congestive heart failure, preeclampsia, eclampsia, diabetes mellitus, hypertension, coronary artery disease, proteinuria, renal insufficiency, or sepsis, type of toxin exposure such as NSAIDs,

cyclosporines, tacrolimus, aminoglycosides, foscarnet, ethylene glycol, hemoglobin, myoglobin, ifosfamide, heavy metals, methotrexate, radiopaque contrast agents, or

streptozotocin), clinical variables (e.g., blood pressure, temperature, respiration rate), risk scores (APACHE score, PREDICT score, TIMI Risk Score for UA/NSTEMI, Framingham Risk Score), a glomerular filtration rate, an estimated glomerular filtration rate, a urine production rate, a serum or plasma creatinine concentration, a urine creatinine concentration, a fractional excretion of sodium, a urine sodium concentration, a urine creatinine to serum or plasma creatinine ratio, a urine specific gravity, a urine osmolality, a urine urea nitrogen to plasma urea nitrogen ratio, a plasma BUN to creatnine ratio, a renal failure index calculated as urine sodium / (urine creatinine / plasma creatinine), a serum or plasma neutrophil gelatinase (NGAL) concentration, a urine NGAL concentration, a serum or plasma cystatin C

concentration, a serum or plasma cardiac troponin concentration, a serum or plasma BNP concentration, a serum or plasma NTproBNP concentration, and a serum or plasma proBNP concentration. Other measures of renal function which may be combined with one or more kidney injury marker assay result(s) are described hereinafter and in Harrison's Principles of Internal Medicine, 17 ^th Ed., McGraw Hill, New York, pages 1741-1830, and Current Medical Diagnosis & Treatment 2008, 47 ^th Ed, McGraw Hill, New York, pages 785-815, each of which are hereby incorporated by reference in their entirety.

[0045] When more than one marker is measured, the individual markers may be measured in samples obtained at the same time, or may be determined from samples obtained at different (e.g., an earlier or later) times. The individual markers may also be measured on the same or different body fluid samples. For example, one kidney injury marker may be measured in a serum or plasma sample and another kidney injury marker may be measured in a urine sample. In addition, assignment of a likelihood may combine an individual kidney injury marker assay result with temporal changes in one or more additional variables.

[0046] In various related aspects, the present invention also relates to devices and kits for performing the methods described herein. Suitable kits comprise reagents sufficient for performing an assay for at least one of the described kidney injury markers, together with instructions for performing the described threshold comparisons.

[0047] In certain embodiments, reagents for performing such assays are provided in an assay device, and such assay devices may be included in such a kit. Preferred reagents can comprise one or more solid phase antibodies, the solid phase antibody comprising antibody that detects the intended biomarker target(s) bound to a solid support. In the case of sandwich immunoassays, such reagents can also include one or more detectably labeled antibodies, the detectably labeled antibody comprising antibody that detects the intended biomarker target(s) bound to a detectable label. Additional optional elements that may be provided as part of an assay device are described hereinafter.

[0048] Detectable labels may include molecules that are themselves detectable (e.g., fluorescent moieties, electrochemical labels, eel (electrochemical luminescence) labels, metal chelates, colloidal metal particles, etc.) as well as molecules that may be indirectly detected by production of a detectable reaction product (e.g., enzymes such as horseradish peroxidase, alkaline phosphatase, etc.) or through the use of a specific binding molecule which itself may be detectable (e.g., a labeled antibody that binds to the second antibody, biotin, digoxigenin, maltose, oligohistidine, 2,4-dintrobenzene, phenylarsenate, ssDNA, dsDNA, etc.).

[0049] Generation of a signal from the signal development element can be performed using various optical, acoustical, and electrochemical methods well known in the art. Examples of detection modes include fluorescence, radiochemical detection, reflectance, absorbance, amperometry, conductance, impedance, interferometry, ellipsometry, etc. In certain of these methods, the solid phase antibody is coupled to a transducer (e.g., a diffraction grating, electrochemical sensor, etc) for generation of a signal, while in others, a signal is generated by a transducer that is spatially separate from the solid phase antibody (e.g., a fluorometer that employs an excitation light source and an optical detector). This list is not meant to be limiting. Antibody-based biosensors may also be employed to determine the presence or amount of analytes that optionally eliminate the need for a labeled molecule.

DETAILED DESCRIPTION OF THE INVENTION

[0050] The present invention relates to methods and compositions for diagnosis, differential diagnosis, risk stratification, monitoring, classifying and determination of treatment regimens in subjects suffering or at risk of suffering from injury to renal function, reduced renal function and/or acute renal failure through measurement of one or more kidney injury markers. In various embodiments, a measured concentration of one or more markers selected from the group consisting of Metalloproteinase inhibitor 1, Metalloproteinase inhibitor 2,

Metalloproteinase inhibitor 4, C-C motif chemokine 15, C-C motif chemokine 18, C-C motif chemokine 23, and/or, C-C motif chemokine 24 or one or more markers related thereto, and optionally one or more additional kidney injury markers known in the art, are correlated to the renal status of the subject.

[0051] For purposes of this document, the following definitions apply:

As used herein, an "injury to renal function" is an abrupt (within 14 days, preferably within 7 days, more preferably within 72 hours, and still more preferably within 48 hours) measurable reduction in a measure of renal function. Such an injury may be identified, for example, by a decrease in glomerular filtration rate or estimated GFR, a reduction in urine output, an increase in serum creatinine, an increase in serum cy statin C, a requirement for renal replacement therapy, etc. "Improvement in Renal Function" is an abrupt (within 14 days, preferably within 7 days, more preferably within 72 hours, and still more preferably within 48 hours) measurable increase in a measure of renal function. Preferred methods for measuring and/or estimating GFR are described hereinafter.

As used herein, "reduced renal function" is an abrupt (within 14 days, preferably within 7 days, more preferably within 72 hours, and still more preferably within 48 hours) reduction in kidney function identified by an absolute increase in serum creatinine of greater than or equal to 0.1 mg/dL (> 8.8 μιηοΙ/L), a percentage increase in serum creatinine of greater than or equal to 20% (1.2-fold from baseline), or a reduction in urine output (documented oliguria of less than 0. 5 ml/kg per hour).

As used herein, "acute renal failure" or "ARF" is an abrupt (within 14 days, preferably within 7 days, more preferably within 72 hours, and still more preferably within 48 hours) reduction in kidney function identified by an absolute increase in serum creatinine of greater than or equal to 0.3 mg/dl (> 26.4 μιηοΐ/ΐ), a percentage increase in serum creatinine of greater than or equal to 50% (1. 5-fold from baseline), or a reduction in urine output (documented oliguria of less than 0.5 ml/kg per hour for at least 6 hours). This term is synonymous with "acute kidney injury" or "AKI."

[0052] In this regard, the skilled artisan will understand that the signals obtained from an immunoassay are a direct result of complexes formed between one or more antibodies and the target biomolecule (i.e., the analyte) and polypeptides containing the necessary epitope(s) to which the antibodies bind. While such assays may detect the full length biomarker and the assay result be expressed as a concentration of a biomarker of interest, the signal from the assay is actually a result of all such "immunoreactive" polypeptides present in the sample. Expression of biomarkers may also be determined by means other than immunoassays, including protein measurements (such as dot blots, western blots, chromatographic methods, mass spectrometry, etc.) and nucleic acid measurements (mRNA quatitation). This list is not meant to be limiting.

[0053] As used herein, the term "Metalloproteinase inhibitor 1" refers to one or more polypeptides present in a biological sample that are derived from the Metalloproteinase inhibitor 1 precursor (Swiss-Prot P01033 (SEQ ID NO: 1)).

10 20 30 40 50 60

MAPFEPLASG ILLLLWLIAP SRACTCVPPH PQTAFCNSDL VIRAKFVGTP EVNQTTLYQR

70 80 90 100 110 120

YEIKMTKMYK GFQALGDAAD IRFVYTPAME SVCGYFHRSH NRSEEFLIAG KLQDGLLHIT

130 140 150 160 170 180

TCSFVAPWNS LSLAQRRGFT KTYTVGCEEC TVFPCLSIPC KLQSGTHCLW TDQLLQGSEK

190 200

GFQSRHLACL PREPGLCTWQ SLRSQIA

[0054] The following domains have been identified in Metalloproteinase inhibitor 1 :

Residues Length Domain ID

1-23 23 signal sequence

24-207 184 Metalloproteinase inhibitor 1

[0055] As used herein, the term "Metalloproteinase inhibitor 2" refers to one or more polypeptides present in a biological sample that are derived from the Metalloproteinase inhibitor 2 precursor (human precursor: Swiss-Prot P16035 (SEQ ID NO: 2)).

10 20 30 40 50 60

MGAAARTLRL ALGLLLLATL LRPADACSCS PVHPQQAFCN ADWIRAKAV SEKEVDSGND

70 80 90 100 110 120

IYGNPIKRIQ YEIKQIKMFK GPEKDIEFIY TAPSSAVCGV SLDVGGKKEY LIAGKAEGDG

130 140 150 160 170 180

KMHITLCDFI VPWDTLSTTQ KKSLNHRYQM GCECKITRCP MIPCYISSPD ECLWMDWVTE

190 200 210 220

KNINGHQAKF FACIKRSDGS CAWYRGAAPP KQEFLDIEDP [0056] The following domains have been identified in Metalloproteinase inhibitor 2:

Residues Length Domain ID

1-26 26 Signal peptide

27-220 194 Metalloproteinase inhibitor 2

[0057] As used herein, the term "Metalloproteinase inhibitor 4" refers to one or polypeptides present in a biological sample that are derived from the Metalloproteinase inhibitor 4 precursor (human precursor: Swiss-Prot Q99727 (SEQ ID NO: 3)).

10 20 30 40 50 60

MPGSPRPAPS WVLLLRLLAL LRPPGLGEAC SCAPAHPQQH ICHSALVIRA KISSEKVVPA

70 80 90 100 110 120

SADPADTEKM LRYEIKQIKM FKGFEKVKDV QYIYTPFDSS LCGVKLEANS QKQYLLTGQV

130 140 150 160 170 180

LSDGKVFIHL CNYIEPWEDL SLVQRESLNH HYHLNCGCQI TTCYTVPCTI SAPNECLWTD

190 200 210 220

WLLERKLYGY QAQHYVCMKH VDGTCSWYRG HLPLRKEFVD IVQP

[0058] The following domains have been identified in Metalloproteinase inhibitor 4:

Residues Length Domain ID

1-29 29 Signal sequence

30-224 195 Metalloproteinase inhibitor 4

[0059] As used herein, the term "C-C motif chemokine 15" refers to one or more polypeptides present in a biological sample that are derived from the C-C motif chemokine 15 precursor (human precursor: Swiss-Prot Q16663 (SEQ ID NO: 4)).

10 20 30 40 50

MKVSVAALSC LMLVAVLGSQ AQFINDAETE LMMSKLPLEN PVVLNSFHFA

60 70 80 90 100

ADCCTSYISQ SIPCSLMKSY FETSSECSKP GVIFLTKKGR QVCAKPSGPG

110

VQDCMKKLKP YSI

[0060] The following domains have been identified in C-C motif chemokine 15: Residues Length Domain ID

1-21 21 Signal peptide

22-113 92 C-C motif chemokine 15

43-113 71 C-C motif chemokine 15 (22-92)

46-113 68 C-C motif chemokine 15 (25-92)

50-113 64 C-C motif chemokine 15 (29-92)

[0061] As used herein, the term "C-C motif chemokine 18" refers to one or more polypeptides present in a biological sample that are derived from the C-C motif chemokine 18 precursor (human precursor: Swiss-Prot P55774 (SEQ ID NO: 5)).

10 20 30 40 50 60

MKGLAAALLV LVCTMALCSC AQVGTNKELC CLVYTSWQIP QKFIVDYSET SPQCPKPGVI

70 80

LLTKRGRQIC ADPNKKWVQK YISDLKLNA

[0062] The following domains have been identified in C-C motif chemokine 18:

Residues Length Domain ID

1-20 20 Signal peptide

21-89 69 C-C motif chemokine 18

21-88 68 CCL 18 (1-68)

23-89 67 CCL 18 (3-69)

24-89 66 CCL 18 (4-69)

[0063] As used herein, the term "C-C motif chemokine 23" refers to one or more polypeptides present in a biological sample that are derived from the C-C motif chemokine 23 precursor (Swiss-Prot P55773 (SEQ ID NO: 6))

10 20 30 40 50

MKVSVAALSC LMLVTALGSQ ARVTKDAETE FMMSKLPLEN PVLLDRFHAT

60 70 80 90 100

SADCCISYTP RSIPCSLLES YFETNSECSK PGVIFLTKKG RRFCANPSDK

110 120

QVQVCVRMLK LDTRIKTRKN

[0064] The following domains have been identified in C-C motif chemokine 23: Residues Length Domain ID

1-21 21 signal peptide

22- 120 99 C-C motif chemokine 23

40-120 81 C-C motif chemokine 23 (19-99)

43-120 78 C-C motif chemokine 23 (22-99)

48-120 73 C-C motif chemokine 23 (27-99)

51-120 70 C-C motif chemokine 23 (30-99)

[0065] As used herein, the term "C-C motif chemokine 24" refers to one or more polypeptides present in a biological sample that are derived from the C-C motif chemokine 24 precursor (Swiss-Prot 000175 (SEQ ID NO: 7))

10 2 0 30 4 0 50

MAGLMT IVTS LLFLGVCAHH I I PTGSWIP SPCCMFFVSK RI PENRWSY

60 7 0 80 90 1 00

QLS SRS TCLK AGVIFTTKKG QQFCGDPKQE WVQRYMKNLD AKQKKASPRA

1 10

RAVAVKGPVQ RYP GNQTTC

[0066] The following domains have been identified in C-C motif chemokine 24:

Residues Length Domain ID

1-26 26 signal peptide

27- 119 93 C-C motif chemokine 24

[0067] As used herein, the term "relating a signal to the presence or amount" of an analyte reflects this understanding. Assay signals are typically related to the presence or amount of an analyte through the use of a standard curve calculated using known concentrations of the analyte of interest. As the term is used herein, an assay is "configured to detect" an analyte if an assay can generate a detectable signal indicative of the presence or amount of a physiologically relevant concentration of the analyte. Because an antibody epitope is on the order of 8 amino acids, an immunoassay configured to detect a marker of interest will also detect polypeptides related to the marker sequence, so long as those polypeptides contain the epitope(s) necessary to bind to the antibody or antibodies used in the assay. [0068] The term "related marker" as used herein with regard to a biomarker such as one of the kidney injury markers described herein refers to one or more fragments, variants, etc., of a particular marker or its biosynthetic parent that may be detected as a surrogate for the marker itself or as independent biomarkers. The term also refers to one or more polypeptides present in a biological sample that are derived from the biomarker precursor complexed to additional species, such as binding proteins, receptors, heparin, lipids, sugars, etc.

[0069] The term "positive going" marker as that term is used herein refer to a marker that is determined to be elevated in subjects suffering from a disease or condition, relative to subjects not suffering from that disease or condition. The term "negative going" marker as that term is used herein refer to a marker that is determined to be reduced in subjects suffering from a disease or condition, relative to subjects not suffering from that disease or condition.

[0070] The term "subject" as used herein refers to a human or non-human organism. Thus, the methods and compositions described herein are applicable to both human and veterinary disease. Further, while a subject is preferably a living organism, the invention described herein may be used in post-mortem analysis as well. Preferred subjects are humans, and most preferably "patients," which as used herein refers to living humans that are receiving medical care for a disease or condition. This includes persons with no defined illness who are being investigated for signs of pathology.

[0071] Preferably, an analyte is measured in a sample. Such a sample may be obtained from a subject, or may be obtained from biological materials intended to be provided to the subject. For example, a sample may be obtained from a kidney being evaluated for possible

transplantation into a subject, and an analyte measurement used to evaluate the kidney for preexisting damage. Preferred samples are body fluid samples.

[0072] The term "body fluid sample" as used herein refers to a sample of bodily fluid obtained for the purpose of diagnosis, prognosis, classification or evaluation of a subject of interest, such as a patient or transplant donor. In certain embodiments, such a sample may be obtained for the purpose of determining the outcome of an ongoing condition or the effect of a treatment regimen on a condition. Preferred body fluid samples include blood, serum, plasma, cerebrospinal fluid, urine, saliva, sputum, and pleural effusions. In addition, one of skill in the art would realize that certain body fluid samples would be more readily analyzed following a fractionation or purification procedure, for example, separation of whole blood into serum or plasma components. [0073] The term "diagnosis" as used herein refers to methods by which the skilled artisan can estimate and/or determine the probability ("a likelihood") of whether or not a patient is suffering from a given disease or condition. In the case of the present invention, "diagnosis" includes using the results of an assay, most preferably an immunoassay, for a kidney injury marker of the present invention, optionally together with other clinical characteristics, to arrive at a diagnosis (that is, the occurrence or nonoccurrence) of an acute renal injury or ARF for the subject from which a sample was obtained and assayed. That such a diagnosis is "determined" is not meant to imply that the diagnosis is 100% accurate. Many biomarkers are indicative of multiple conditions. The skilled clinician does not use biomarker results in an informational vacuum, but rather test results are used together with other clinical indicia to arrive at a diagnosis. Thus, a measured biomarker level on one side of a predetermined diagnostic threshold indicates a greater likelihood of the occurrence of disease in the subject relative to a measured level on the other side of the predetermined diagnostic threshold.

[0074] Similarly, a prognostic risk signals a probability ("a likelihood") that a given course or outcome will occur. A level or a change in level of a prognostic indicator, which in turn is associated with an increased probability of morbidity (e.g., worsening renal function, future ARF, or death) is referred to as being "indicative of an increased likelihood" of an adverse outcome in a patient.

[0075] Marker Assays

[0076] In general, immunoassays involve contacting a sample containing or suspected of containing a biomarker of interest with at least one antibody that specifically binds to the biomarker. A signal is then generated indicative of the presence or amount of complexes formed by the binding of polypeptides in the sample to the antibody. The signal is then related to the presence or amount of the biomarker in the sample. Numerous methods and devices are well known to the skilled artisan for the detection and analysis of biomarkers. See, e.g., U.S. Patents 6,143,576; 6,113,855; 6,019,944; 5,985,579; 5,947,124; 5,939,272; 5,922,615;

5,885,527; 5,851,776; 5,824,799; 5,679,526; 5,525,524; and 5,480,792, and The Immunoassay Handbook, David Wild, ed. Stockton Press, New York, 1994, each of which is hereby incorporated by reference in its entirety, including all tables, figures and claims.

[0077] The assay devices and methods known in the art can utilize labeled molecules in various sandwich, competitive, or non-competitive assay formats, to generate a signal that is related to the presence or amount of the biomarker of interest. Suitable assay formats also include chromatographic, mass spectrographic, and protein "blotting" methods. Additionally, certain methods and devices, such as biosensors and optical immunoassays, may be employed to determine the presence or amount of analytes without the need for a labeled molecule. See, e.g., U.S. Patents 5,631,171 ; and 5,955,377, each of which is hereby incorporated by reference in its entirety, including all tables, figures and claims. One skilled in the art also recognizes that robotic instrumentation including but not limited to Beckman ACCESS®, Abbott AXSYM®, Roche ELECSYS®, Dade Behring STRATUS® systems are among the immunoassay analyzers that are capable of performing immunoassays. But any suitable immunoassay may be utilized, for example, enzyme-linked immunoassays (ELISA), radioimmunoassays (RIAs), competitive binding assays, and the like.

[0078] Antibodies or other polypeptides may be immobilized onto a variety of solid supports for use in assays. Solid phases that may be used to immobilize specific binding members include include those developed and/or used as solid phases in solid phase binding assays. Examples of suitable solid phases include membrane filters, cellulose-based papers, beads (including polymeric, latex and paramagnetic particles), glass, silicon wafers, microparticles, nanoparticles, TentaGels, AgroGels, PEGA gels, SPOCC gels, and multiple- well plates. An assay strip could be prepared by coating the antibody or a plurality of antibodies in an array on solid support. This strip could then be dipped into the test sample and then processed quickly through washes and detection steps to generate a measurable signal, such as a colored spot. Antibodies or other polypeptides may be bound to specific zones of assay devices either by conjugating directly to an assay device surface, or by indirect binding. In an example of the later case, antibodies or other polypeptides may be immobilized on particles or other solid supports, and that solid support immobilized to the device surface.

[0079] Biological assays require methods for detection, and one of the most common methods for quantitation of results is to conjugate a detectable label to a protein or nucleic acid that has affinity for one of the components in the biological system being studied. Detectable labels may include molecules that are themselves detectable (e.g., fluorescent moieties, electrochemical labels, metal chelates, etc.) as well as molecules that may be indirectly detected by production of a detectable reaction product (e.g., enzymes such as horseradish peroxidase, alkaline phosphatase, etc.) or by a specific binding molecule which itself may be detectable (e.g., biotin, digoxigenin, maltose, oligohistidine, 2,4-dintrobenzene, phenylarsenate, ssDNA, dsDNA, etc.). [0080] Preparation of solid phases and detectable label conjugates often comprise the use of chemical cross-linkers. Cross-linking reagents contain at least two reactive groups, and are divided generally into homofunctional cross-linkers (containing identical reactive groups) and heterofunctional cross-linkers (containing non-identical reactive groups). Homobifunctional cross-linkers that couple through amines, sulfhydryls or react non- specifically are available from many commercial sources. Maleimides, alkyl and aryl halides, alpha-haloacyls and pyridyl disulfides are thiol reactive groups. Maleimides, alkyl and aryl halides, and alpha- haloacyls react with sulfhydryls to form thiol ether bonds, while pyridyl disulfides react with sulfhydryls to produce mixed disulfides. The pyridyl disulfide product is cleavable. Imidoesters are also very useful for protein-protein cross-links. A variety of heterobifunctional cross- linkers, each combining different attributes for successful conjugation, are commercially available.

[0081] In certain aspects, the present invention provides kits for the analysis of the described kidney injury markers. The kit comprises reagents for the analysis of at least one test sample which comprise at least one antibody that a kidney injury marker. The kit can also include devices and instructions for performing one or more of the diagnostic and/or prognostic correlations described herein. Preferred kits will comprise an antibody pair for performing a sandwich assay, or a labeled species for performing a competitive assay, for the analyte.

Preferably, an antibody pair comprises a first antibody conjugated to a solid phase and a second antibody conjugated to a detectable label, wherein each of the first and second antibodies that bind a kidney injury marker. Most preferably each of the antibodies are monoclonal antibodies. The instructions for use of the kit and performing the correlations can be in the form of labeling, which refers to any written or recorded material that is attached to, or otherwise accompanies a kit at any time during its manufacture, transport, sale or use. For example, the term labeling encompasses advertising leaflets and brochures, packaging materials, instructions, audio or video cassettes, computer discs, as well as writing imprinted directly on kits.

[0082] Antibodies

[0083] The term "antibody" as used herein refers to a peptide or polypeptide derived from, modeled after or substantially encoded by an immunoglobulin gene or immunoglobulin genes, or fragments thereof, capable of specifically binding an antigen or epitope. See, e.g.

Fundamental Immunology, 3rd Edition, W.E. Paul, ed., Raven Press, N.Y. (1993); Wilson (1994; J. Immunol. Methods 175:267-273; Yarmush (1992) J. Biochem. Biophys. Methods 25:85-97. The term antibody includes antigen-binding portions, i.e., "antigen binding sites," (e.g., fragments, subsequences, complementarity determining regions (CDRs)) that retain capacity to bind antigen, including (i) a Fab fragment, a monovalent fragment consisting of the VL, VH, CL and CHI domains; (ii) a F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; (iii) a Fd fragment consisting of the VH and CHI domains; (iv) a Fv fragment consisting of the VL and VH domains of a single arm of an antibody, (v) a dAb fragment (Ward et al., (1989) Nature 341 :544-546), which consists of a VH domain; and (vi) an isolated complementarity determining region (CDR). Single chain antibodies are also included by reference in the term "antibody."

[0084] Antibodies used in the immunoassays described herein preferably specifically bind to a kidney injury marker of the present invention. The term "specifically binds" is not intended to indicate that an antibody binds exclusively to its intended target since, as noted above, an antibody binds to any polypeptide displaying the epitope(s) to which the antibody binds.

Rather, an antibody "specifically binds" if its affinity for its intended target is about 5-fold greater when compared to its affinity for a non-target molecule which does not display the appropriate epitope(s). Preferably the affinity of the antibody will be at least about 5 fold, preferably 10 fold, more preferably 25-fold, even more preferably 50-fold, and most preferably 100-fold or more, greater for a target molecule than its affinity for a non-target molecule. In preferred embodiments, Preferred antibodies bind with affinities of at least about 10 ⁷ M ^"1 , and preferably between about 10 ⁸ M ^"1 to about 10 ⁹ NT ¹ , about 10 ⁹ M ^"1 to about 10 ¹⁰ NT ¹ , or about 10 ¹⁰ M ¹ to about 10 ¹² M ¹ .

[0085] Affinity is calculated as Kd = k ₀ ff/k _on (k ₀ ff is the dissociation rate constant, K _on is the association rate constant and Kd is the equilibrium constant). Affinity can be determined at equilibrium by measuring the fraction bound (r) of labeled ligand at various concentrations (c). The data are graphed using the Scatchard equation: r/c = K(n-r): where r = moles of bound ligand/mole of receptor at equilibrium; c = free ligand concentration at equilibrium; K = equilibrium association constant; and n = number of ligand binding sites per receptor molecule. By graphical analysis, r/c is plotted on the Y-axis versus r on the X-axis, thus producing a Scatchard plot. Antibody affinity measurement by Scatchard analysis is well known in the art. See, e.g., van Erp et al., J. Immunoassay 12: 425-43, 1991 ; Nelson and Griswold, Comput. Methods Programs Biomed. 27: 65-8, 1988.

[0086] The term "epitope" refers to an antigenic determinant capable of specific binding to an antibody. Epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains and usually have specific three dimensional structural characteristics, as well as specific charge characteristics. Conformational and

nonconformational epitopes are distinguished in that the binding to the former but not the latter is lost in the presence of denaturing solvents.

[0087] Numerous publications discuss the use of phage display technology to produce and screen libraries of polypeptides for binding to a selected analyte. See, e.g, Cwirla et al., Proc. Natl. Acad. Sci. USA 87, 6378-82, 1990; Devlin et al, Science 249, 404-6, 1990, Scott and Smith, Science 249, 386-88, 1990; and Ladner et al, U.S. Pat. No. 5,571,698. A basic concept of phage display methods is the establishment of a physical association between DNA encoding a polypeptide to be screened and the polypeptide. This physical association is provided by the phage particle, which displays a polypeptide as part of a capsid enclosing the phage genome which encodes the polypeptide. The establishment of a physical association between polypeptides and their genetic material allows simultaneous mass screening of very large numbers of phage bearing different polypeptides. Phage displaying a polypeptide with affinity to a target bind to the target and these phage are enriched by affinity screening to the target. The identity of polypeptides displayed from these phage can be determined from their respective genomes. Using these methods a polypeptide identified as having a binding affinity for a desired target can then be synthesized in bulk by conventional means. See, e.g., U.S. Patent No. 6,057,098, which is hereby incorporated in its entirety, including all tables, figures, and claims.

[0088] The antibodies that are generated by these methods may then be selected by first screening for affinity and specificity with the purified polypeptide of interest and, if required, comparing the results to the affinity and specificity of the antibodies with polypeptides that are desired to be excluded from binding. The screening procedure can involve immobilization of the purified polypeptides in separate wells of microtiter plates. The solution containing a potential antibody or groups of antibodies is then placed into the respective microtiter wells and incubated for about 30 min to 2 h. The microtiter wells are then washed and a labeled secondary antibody (for example, an anti-mouse antibody conjugated to alkaline phosphatase if the raised antibodies are mouse antibodies) is added to the wells and incubated for about 30 min and then washed. Substrate is added to the wells and a color reaction will appear where antibody to the immobilized polypeptide(s) are present.

[0089] The antibodies so identified may then be further analyzed for affinity and specificity in the assay design selected. In the development of immunoassays for a target protein, the purified target protein acts as a standard with which to judge the sensitivity and specificity of the immunoassay using the antibodies that have been selected. Because the binding affinity of various antibodies may differ; certain antibody pairs (e.g., in sandwich assays) may interfere with one another sterically, etc., assay performance of an antibody may be a more important measure than absolute affinity and specificity of an antibody.

[0090] Assay Correlations

[0091] The term "correlating" as used herein in reference to the use of biomarkers refers to comparing the presence or amount of the biomarker(s) in a patient to its presence or amount in persons known to suffer from, or known to be at risk of, a given condition; or in persons known to be free of a given condition. Often, this takes the form of comparing an assay result in the form of a biomarker concentration to a predetermined threshold selected to be indicative of the occurrence or nonoccurrence of a disease or the likelihood of some future outcome.

[0092] Selecting a diagnostic threshold involves, among other things, consideration of the probability of disease, distribution of true and false diagnoses at different test thresholds, and estimates of the consequences of treatment (or a failure to treat) based on the diagnosis. For example, when considering administering a specific therapy which is highly efficacious and has a low level of risk, few tests are needed because clinicians can accept substantial diagnostic uncertainty. On the other hand, in situations where treatment options are less effective and more risky, clinicians often need a higher degree of diagnostic certainty. Thus, cost/benefit analysis is involved in selecting a diagnostic threshold.

[0093] Suitable thresholds may be determined in a variety of ways. For example, one recommended diagnostic threshold for the diagnosis of acute myocardial infarction using cardiac troponin is the 97.5 ^th percentile of the concentration seen in a normal population.

Another method may be to look at serial samples from the same patient, where a prior

"baseline" result is used to monitor for temporal changes in a biomarker level.

[0094] Population studies may also be used to select a decision threshold. Reciever

Operating Characteristic ("ROC") arose from the field of signal dectection therory developed during World War II for the analysis of radar images, and ROC analysis is often used to select a threshold able to best distinguish a "diseased" subpopulation from a "nondiseased"

subpopulation. A false positive in this case occurs when the person tests positive, but actually does not have the disease. A false negative, on the other hand, occurs when the person tests negative, suggesting they are healthy, when they actually do have the disease. To draw a ROC curve, the true positive rate (TPR) and false positive rate (FPR) are determined as the decision threshold is varied continuously. Since TPR is equivalent with sensitivity and FPR is equal to 1 - specificity, the ROC graph is sometimes called the sensitivity vs (1 - specificity) plot. A perfect test will have an area under the ROC curve of 1.0; a random test will have an area of 0.5. A threshold is selected to provide an acceptable level of specificity and sensitivity.

[0095] In this context, "diseased" is meant to refer to a population having one characteristic (the presence of a disease or condition or the occurrence of some outcome) and "nondiseased" is meant to refer to a population lacking the characteristic. While a single decision threshold is the simplest application of such a method, multiple decision thresholds may be used. For example, below a first threshold, the absence of disease may be assigned with relatively high confidence, and above a second threshold the presence of disease may also be assigned with relatively high confidence. Between the two thresholds may be considered indeterminate. This is meant to be exemplary in nature only.

[0096] In addition to threshold comparisons, other methods for correlating assay results to a patient classification (occurrence or nonoccurrence of disease, likelihood of an outcome, etc.) include decision trees, rule sets, Bayesian methods, and neural network methods. These methods can produce probability values representing the degree to which a subject belongs to one classification out of a plurality of classifications.

[0097] Measures of test accuracy may be obtained as described in Fischer et ah, Intensive Care Med. 29: 1043-51, 2003, and used to determine the effectiveness of a given biomarker. These measures include sensitivity and specificity, predictive values, likelihood ratios, diagnostic odds ratios, and ROC curve areas. The area under the curve ("AUC") of a ROC plot is equal to the probability that a classifier will rank a randomly chosen positive instance higher than a randomly chosen negative one. The area under the ROC curve may be thought of as equivalent to the Mann- Whitney U test, which tests for the median difference between scores obtained in the two groups considered if the groups are of continuous data, or to the Wilcoxon test of ranks.

[0098] As discussed above, suitable tests may exhibit one or more of the following results on these various measures: a specificity of greater than 0.5, preferably at least 0.6, more preferably at least 0.7, still more preferably at least 0.8, even more preferably at least 0.9 and most preferably at least 0.95, with a corresponding sensitivity greater than 0.2, preferably greater than 0.3, more preferably greater than 0.4, still more preferably at least 0.5, even more preferably 0.6, yet more preferably greater than 0.7, still more preferably greater than 0.8, more preferably greater than 0.9, and most preferably greater than 0.95; a sensitivity of greater than 0.5, preferably at least 0.6, more preferably at least 0.7, still more preferably at least 0.8, even more preferably at least 0.9 and most preferably at least 0.95, with a corresponding specificity greater than 0.2, preferably greater than 0.3, more preferably greater than 0.4, still more preferably at least 0.5, even more preferably 0.6, yet more preferably greater than 0.7, still more preferably greater than 0.8, more preferably greater than 0.9, and most preferably greater than

0.95; at least 75% sensitivity, combined with at least 75% specificity; a ROC curve area of greater than 0.5, preferably at least 0.6, more preferably 0.7, still more preferably at least 0.8, even more preferably at least 0.9, and most preferably at least 0.95; an odds ratio different from

1, preferably at least about 2 or more or about 0.5 or less, more preferably at least about 3 or more or about 0.33 or less, still more preferably at least about 4 or more or about 0.25 or less, even more preferably at least about 5 or more or about 0.2 or less, and most preferably at least about 10 or more or about 0.1 or less; a positive likelihood ratio (calculated as sensitivity/(l- specificity)) of greater than 1, at least 2, more preferably at least 3, still more preferably at least 5, and most preferably at least 10; and or a negative likelihood ratio (calculated as (1- sensitivity)/specificity) of less than 1, less than or equal to 0.5, more preferably less than or equal to 0.3, and most preferably less than or equal to 0.1

[0099] Additional clinical indicia may be combined with the kidney injury marker assay result(s) of the present invention. These include other biomarkers related to renal status.

Examples include the following, which recite the common biomarker name, followed by the Swiss-Prot entry number for that biomarker or its parent: Actin (P68133); Adenosine deaminase binding protein (DPP4, P27487); Alpha-l-acid glycoprotein 1 (P02763); Alpha-1- microglobulin (P02760); Albumin (P02768); Angiotensinogenase (Renin, P00797); Annexin A2 (P07355); Beta-glucuronidase (P08236); B-2-microglobulin (P61769); Beta-galactosidase (P16278); BMP-7 (P18075); Brain natriuretic peptide (proBNP, BNP-32, NTproBNP; P16860); Calcium-binding protein Beta (S lOO-beta, P04271); Carbonic anhydrase 9 (Q16790); Casein Kinase 2 (P68400); Clusterin (P10909); Complement C3 (P01024); Cysteine-rich protein (CYR61, 000622); Cytochrome C (P99999); Epidermal growth factor (EGF, P01133);

Endothelin-1 (P05305); Exosomal Fetuin-A (P02765); Fatty acid-binding protein, heart (FABP3, P05413); Fatty acid-binding protein, liver (P07148); Ferritin (light chain, P02792; heavy chain P02794); Fructose- 1,6-biphosphatase (P09467); GRO-alpha (CXCL1, (P09341); Growth Hormone (P01241); Hepatocyte growth factor (PI 4210); Insulin-like growth factor I (P05019); Immunoglobulin G; Immunoglobulin Light Chains (Kappa and Lambda); Interferon gamma (P01579); Lysozyme (P61626); Interleukin-1 alpha (P01583); Interleukin-2 (P60568); Interleukin-4 (P05112); Interleukin-9 (P15248); Interleukin-12p40 (P29460); Interleukin-13 (P35225); Interleukin-16 (Q14005); LI cell adhesion molecule (P32004); Lactate

dehydrogenase (P00338); Leucine Aminopeptidase (P28838); Meprin A-alpha subunit

(Q16819); Meprin A-beta subunit (Q16820); Midkine (P21741); MJP2-alpha (CXCL2, P19875); MMP-2 (P08253); MMP-9 (P14780); Netrin-1 (095631); Neutral endopeptidase (P08473); Osteopontin (P10451); Renal papillary antigen 1 (RPAl); Renal papillary antigen 2 (RPA2); Retinol binding protein (P09455); Ribonuclease; S 100 calcium-binding protein A6 (P06703); Serum Amyloid P Component (P02743); Sodium/Hydrogen exchanger isoform (NHE3, P48764); Spermidine/spermine Nl-acetyltransferase (P21673); TGF-Betal (P01137); Transferrin (P02787); Trefoil factor 3 (TFF3, Q07654); Toll-Like protein 4 (000206); Total protein; Tubulointerstitial nephritis antigen (Q9UJW2); Uromodulin (Tamm-Horsfall protein, P07911).

[00100] For purposes of risk stratification, Adiponectin (Q15848); Alkaline phosphatase (P05186); Aminopeptidase N (P15144); CalbindinD28k (P05937); Cystatin C (P01034); 8 subunit of FIFO ATPase (P03928); Gamma-glutamyltransferase (P19440); GSTa (alpha- glutathione-S-transferase, P08263); GSTpi (Glutathione-S -transferase P; GST class-pi;

P09211); IGFBP-1 (P08833); IGFBP-2 (P18065); IGFBP-6 (P24592); Integral membrane protein 1 (Itml, P46977); Interleukin-6 (P05231); Interleukin-8 (P10145); Interleukin-18 (Q14116); IP-10 (10 kDa interferon-gamma-induced protein, P02778); IRPR (IFRD1,

000458); Isovaleryl-CoA dehydrogenase (IVD, P26440); I-TAC/CXCL11 (014625); Keratin 19 (P08727); Kim-1 (Hepatitis A virus cellular receptor 1, Q96D42); L-arginine:glycine amidinotransferase (P50440); Leptin (P41159); Lipocalin2 (NGAL, P80188); MCP-1

(P13500); MIG (Gamma-interferon-induced monokine Q07325); MIP-la (P10147); MIP-3a (P78556); MIP-lbeta (P13236); MIP-ld (Q16663); NAG (N-acetyl-beta-D-glucosaminidase, P54802); Organic ion transporter (OCT2, 015244); Osteoprotegerin (000300); P8 protein (060356); Plasminogen activator inhibitor 1 (PAI-1, P05121); ProANP(l-98) (P01160); Protein phosphatase 1-beta (PPI-beta, P62140); Rab GDI-beta (P50395); Renal kallikrein (P06870); RTl.B-1 (alpha) chain of the integral membrane protein (Q5Y7A8); soluble tumor necrosis factor receptor superfamily member 1 A (sTNFR-I, P19438); soluble tumor necrosis factor receptor superfamily member IB (sTNFR-II, P20333); Tissue inhibitor of metalloproteinases 3 (TIMP-3, P35625); uPAR (Q03405) may be combined with the kidney injury marker assay result(s) of the present invention. [00101] Other clinical indicia which may be combined with the kidney injury marker assay result(s) of the present invention includes demographic information (e.g., weight, sex, age, race), medical history (e.g., family history, type of surgery, pre-existing disease such as aneurism, congestive heart failure, preeclampsia, eclampsia, diabetes mellitus, hypertension, coronary artery disease, proteinuria, renal insufficiency, or sepsis, type of toxin exposure such as NSAIDs, cyclosporines, tacrolimus, aminoglycosides, foscarnet, ethylene glycol, hemoglobin, myoglobin, ifosfamide, heavy metals, methotrexate, radiopaque contrast agents, or streptozotocin), clinical variables (e.g., blood pressure, temperature, respiration rate), risk scores (APACHE score, PREDICT score, TIMI Risk Score for UA/NSTEMI, Framingham Risk Score), a urine total protein measurement, a glomerular filtration rate, an estimated glomerular filtration rate, a urine production rate, a serum or plasma creatinine concentration, a renal papillary antigen 1 (RPA1) measurement; a renal papillary antigen 2 (RPA2)

measurement; a urine creatinine concentration, a fractional excretion of sodium, a urine sodium concentration, a urine creatinine to serum or plasma creatinine ratio, a urine specific gravity, a urine osmolality, a urine urea nitrogen to plasma urea nitrogen ratio, a plasma BUN to creatnine ratio, and/or a renal failure index calculated as urine sodium / (urine creatinine / plasma creatinine). Other measures of renal function which may be combined with the kidney injury marker assay result(s) are described hereinafter and in Harrison' s Principles of Internal Medicine, 17 ^th Ed., McGraw Hill, New York, pages 1741-1830, and Current Medical Diagnosis & Treatment 2008, 47 ^th Ed, McGraw Hill, New York, pages 785-815, each of which are hereby incorporated by reference in their entirety.

[00102] Combining assay results/clinical indicia in this manner can comprise the use of multivariate logistical regression, loglinear modeling, neural network analysis, n-of-m analysis, decision tree analysis, etc. This list is not meant to be limiting.

[00103] Diagnosis of Acute Renal Failure

[00104] As noted above, the terms "acute renal (or kidney) injury" and "acute renal (or kidney) failure" as used herein are defined in part in terms of changes in serum creatinine from a baseline value. Most definitions of ARF have common elements, including the use of serum creatinine and, often, urine output. Patients may present with renal dysfunction without an available baseline measure of renal function for use in this comparison. In such an event, one may estimate a baseline serum creatinine value by assuming the patient initially had a normal GFR. Glomerular filtration rate (GFR) is the volume of fluid filtered from the renal (kidney) glomerular capillaries into the Bowman's capsule per unit time. Glomerular filtration rate (GFR) can be calculated by measuring any chemical that has a steady level in the blood, and is freely filtered but neither reabsorbed nor secreted by the kidneys. GFR is typically expressed in units of ml/min:

Urine Concentration x Urine Flow

G F li—

Plasma Coiieenir atkm

[00105] By normalizing the GFR to the body surface area, a GFR of approximately 75-100 ml/min per 1.73 m ² can be assumed. The rate therefore measured is the quantity of the substance in the urine that originated from a calculable volume of blood.

[00106] There are several different techniques used to calculate or estimate the glomerular filtration rate (GFR or eGFR). In clinical practice, however, creatinine clearance is used to measure GFR. Creatinine is produced naturally by the body (creatinine is a metabolite of creatine, which is found in muscle). It is freely filtered by the glomerulus, but also actively secreted by the renal tubules in very small amounts such that creatinine clearance overestimates actual GFR by 10-20%. This margin of error is acceptable considering the ease with which creatinine clearance is measured.

[00107] Creatinine clearance (CCr) can be calculated if values for creatinine's urine concentration (U&), urine flow rate (V), and creatinine's plasma concentration (P&) are known. Since the product of urine concentration and urine flow rate yields creatinine's excretion rate, creatinine clearance is also said to be its excretion rate (U&xV) divided by its plasma concentration. This is commonly represented mathematically as:

U _Cr x v

¹ C'r

[00108] Commonly a 24 hour urine collection is undertaken, from empty-bladder one morning to the contents of the bladder the following morning, with a comparative blood test then taken:

¾X- _> x 24-hour volume

^ ^• -C'r

X 24 X 60mm s

[00109] To allow comparison of results between people of different sizes, the CCr is often corrected for the body surface area (BSA) and expressed compared to the average sized man as ml/min/1.73 m2. While most adults have a BSA that approaches 1.7 (1.6-1.9), extremely obese or slim patients should have their CCr corrected for their actual BSA:

[00110] The accuracy of a creatinine clearance measurement (even when collection is complete) is limited because as glomerular filtration rate (GFR) falls creatinine secretion is increased, and thus the rise in serum creatinine is less. Thus, creatinine excretion is much greater than the filtered load, resulting in a potentially large overestimation of the GFR (as much as a twofold difference). However, for clinical purposes it is important to determine whether renal function is stable or getting worse or better. This is often determined by monitoring serum creatinine alone. Like creatinine clearance, the serum creatinine will not be an accurate reflection of GFR in the non- steady- state condition of ARF. Nonetheless, the degree to which serum creatinine changes from baseline will reflect the change in GFR. Serum creatinine is readily and easily measured and it is specific for renal function.

[00111] For purposes of determining urine output on a Urine output on a mL/kg/hr basis, hourly urine collection and measurement is adequate. In the case where, for example, only a cumulative 24-h output was available and no patient weights are provided, minor modifications of the RIFLE urine output criteria have been described. For example, Bagshaw et ah, Nephrol. Dial. Transplant. 23: 1203-1210, 2008, assumes an average patient weight of 70 kg, and patients are assigned a RIFLE classification based on the following: <35 mL/h (Risk), <21 mL/h (Injury) or <4 mL/h (Failure).

[00112] Selecting a Treatment Regimen

[00113] Once a diagnosis is obtained, the clinician can readily select a treatment regimen that is compatible with the diagnosis, such as initiating renal replacement therapy, withdrawing delivery of compounds that are known to be damaging to the kidney, kidney transplantation, delaying or avoiding procedures that are known to be damaging to the kidney, modifying diuretic administration, initiating goal directed therapy, etc. The skilled artisan is aware of appropriate treatments for numerous diseases discussed in relation to the methods of diagnosis described herein. See, e.g., Merck Manual of Diagnosis and Therapy, 17th Ed. Merck Research Laboratories, Whitehouse Station, NJ, 1999. In addition, since the methods and compositions described herein provide prognostic information, the markers of the present invention may be used to monitor a course of treatment. For example, improved or worsened prognostic state may indicate that a particular treatment is or is not efficacious.

[00114] One skilled in the art readily appreciates that the present invention is well adapted to carry out the objects and obtain the ends and advantages mentioned, as well as those inherent therein. The examples provided herein are representative of preferred embodiments, are exemplary, and are not intended as limitations on the scope of the invention.

[00115] Example 1: Contrast-induced nephropathy sample collection

[00116] The objective of this sample collection study is to collect samples of plasma and urine and clinical data from patients before and after receiving intravascular contrast media. Approximately 250 adults undergoing radiographic/angiographic procedures involving intravascular administration of iodinated contrast media are enrolled. To be enrolled in the study, each patient must meet all of the following inclusion criteria and none of the following exclusion criteria:

Inclusion Criteria males and females 18 years of age or older; undergoing a radiographic / angiographic procedure (such as a CT scan or coronary

intervention) involving the intravascular administration of contrast media; expected to be hospitalized for at least 48 hours after contrast administration. able and willing to provide written informed consent for study participation and to comply with all study procedures.

Exclusion Criteria renal transplant recipients; acutely worsening renal function prior to the contrast procedure; already receiving dialysis (either acute or chronic) or in imminent need of dialysis at enrollment; expected to undergo a major surgical procedure (such as involving cardiopulmonary bypass) or an additional imaging procedure with contrast media with significant risk for further renal insult within the 48 hrs following contrast administration; participation in an interventional clinical study with an experimental therapy within the previous 30 days; known infection with human immunodeficiency virus (HIV) or a hepatitis virus.

[00117] Immediately prior to the first contrast administration (and after any pre-procedure hydration), an EDTA anti-coagulated blood sample (10 mL) and a urine sample (10 mL) are collected from each patient. Blood and urine samples are then collected at 4 (±0.5), 8 (±1), 24 (±2) 48 (±2), and 72 (±2) hrs following the last administration of contrast media during the index contrast procedure. Blood is collected via direct venipuncture or via other available venous access, such as an existing femoral sheath, central venous line, peripheral intravenous line or hep-lock. These study blood samples are processed to plasma at the clinical site, frozen and shipped to Astute Medical, Inc., San Diego, CA. The study urine samples are frozen and shipped to Astute Medical, Inc.

[00118] Serum creatinine is assessed at the site immediately prior to the first contrast administration (after any pre-procedure hydration) and at 4 (±0.5), 8 (±1), 24 (±2) and 48 (±2) ), and 72 (±2) hours following the last administration of contrast (ideally at the same time as the study samples are obtained). In addition, each patient's status is evaluated through day 30 with regard to additional serum and urine creatinine measurements, a need for dialysis,

hospitalization status, and adverse clinical outcomes (including mortality).

[00119] Prior to contrast administration, each patient is assigned a risk based on the following assessment: systolic blood pressure <80 mm Hg = 5 points; intra- arterial balloon pump = 5 points; congestive heart failure (Class III- IV or history of pulmonary edema) = 5 points; age >75 yrs = 4 points; hematocrit level <39% for men, <35% for women = 3 points; diabetes = 3 points; contrast media volume = 1 point for each 100 mL; serum creatinine level >1.5 g/dL = 4 points OR estimated GFR 40-60 mL/min/1.73 m ² = 2 points, 20-40 mL/min/1.73 m ² = 4 points, < 20 mL/min/1.73 m ² = 6 points. The risks assigned are as follows: risk for CIN and dialysis: 5 or less total points = risk of CIN - 7.5%, risk of dialysis - 0.04%; 6-10 total points = risk of CIN - 14%, risk of dialysis - 0.12%; 11-16 total points = risk of CIN - 26.1%, risk of dialysis - 1.09%; >16 total points = risk of CIN - 57.3%, risk of dialysis - 12.8%.

[00120] Example 2: Cardiac surgery sample collection

[00121] The objective of this sample collection study is to collect samples of plasma and urine and clinical data from patients before and after undergoing cardiovascular surgery, a procedure known to be potentially damaging to kidney function. Approximately 900 adults undergoing such surgery are enrolled. To be enrolled in the study, each patient must meet all of the following inclusion criteria and none of the following exclusion criteria:

Inclusion Criteria males and females 18 years of age or older; undergoing cardiovascular surgery;

Toronto/Ottawa Predictive Risk Index for Renal Replacement risk score of at least 2

(Wijeysundera et al, JAMA 297: 1801-9, 2007); and able and willing to provide written informed consent for study participation and to comply with all study procedures.

Exclusion Criteria known pregnancy; previous renal transplantation; acutely worsening renal function prior to enrollment (e.g., any category of RIFLE criteria); already receiving dialysis (either acute or chronic) or in imminent need of dialysis at enrollment; currently enrolled in another clinical study or expected to be enrolled in another clinical study within 7 days of cardiac surgery that involves drug infusion or a therapeutic intervention for AKI; known infection with human immunodeficiency virus (HIV) or a hepatitis virus.

[00122] Within 3 hours prior to the first incision (and after any pre -procedure hydration), an EDTA anti-coagulated blood sample (10 mL), whole blood (3 mL), and a urine sample (35 mL) are collected from each patient. Blood and urine samples are then collected at 3 (±0.5), 6 (±0.5), 12 (±1), 24 (±2) and 48 (±2) hrs following the procedure and then daily on days 3 through 7 if the subject remains in the hospital. Blood is collected via direct venipuncture or via other available venous access, such as an existing femoral sheath, central venous line, peripheral intravenous line or hep-lock. These study blood samples are frozen and shipped to Astute Medical, Inc., San Diego, CA. The study urine samples are frozen and shipped to Astute Medical, Inc. [00123] Example 3: Acutely ill subject sample collection

[00124] The objective of this study is to collect samples from acutely ill patients.

Approximately 2200 adults expected to be in the ICU for at least 48 hours will be enrolled. To be enrolled in the study, each patient must meet all of the following inclusion criteria and none of the following exclusion criteria:

Inclusion Criteria males and females 18 years of age or older;

Study population 1 : approximately 300 patients that have at least one of: shock (SBP < 90 mmHg and/or need for vasopressor support to maintain MAP > 60 mmHg and/or documented drop in SBP of at least 40 mmHg); and sepsis;

Study population 2: approximately 300 patients that have at least one of:

IV antibiotics ordered in computerized physician order entry (CPOE) within 24 hours of enrollment; contrast media exposure within 24 hours of enrollment; increased Intra- Abdominal Pressure with acute decompensated heart failure; and severe trauma as the primary reason for ICU admission and likely to be hospitalized in the ICU for 48 hours after enrollment;

Study population 3: approximately 300 patients expected to be hospitalized through acute care setting (ICU or ED) with a known risk factor for acute renal injury (e.g. sepsis,

hypotension/shock (Shock = systolic BP < 90 mmHg and/or the need for vasopressor support to maintain a MAP > 60 mmHg and/or a documented drop in SBP > 40 mmHg), major trauma, hemorrhage, or major surgery); and/or expected to be hospitalized to the ICU for at least 24 hours after enrollment;

Study population 4: approximately 1000 patients that are 21 years of age or older, within 24 hours of being admitted into the ICU, expected to have an indwelling urinary catheter for at least 48 hours after enrollment, and have at least one of the following acute conditions within 24 hours prior to enrollment: (i) respiratory SOFA score of > 2 (Pa02/Fi02 <300), (ii) cardiovascular SOFA score of > 1 (MAP < 70 mm Hg and/or any vasopressor required).

Study population 5: approximately 300 patients that are 21 years of age or older, receiving care in the ICU, have an indwelling urinary catheter as standard care at the time of enrollment, have acute kidney injury (KDIGO stage 2 or stage 3) at the time of the first sample collection, and have their first sample collected within 36 hours of meeting KDIGO stage 2 criteria

Exclusion Criteria known pregnancy; prisoners or institutionalized individuals; previous renal transplantation; comfort-measures-only status for study population 5; known acutely worsening renal function prior to enrollment (e.g., any category of RIFLE criteria) for study populations 1, 2, 3, and 4; received dialysis (either acute or chronic) within 5 days prior to enrollment or in imminent need of dialysis at the time of enrollment; known infection with human immunodeficiency virus (HIV) or a hepatitis virus; meets any of the following in study populations 4 and 5:

(i) active bleeding with an anticipated need for > 4 units PRBC in a day;

(ii) hemoglobin < 7 g/dL;

(iii) any other condition that in the physician's opinion would contraindicate drawing serial blood samples for clinical study purposes; meets only the SBP < 90 mmHg inclusion criterion set forth above, and does not have shock in the attending physician's or principal investigator's opinion.

[00125] After obtaining informed consent, an EDTA anti-coagulated blood sample (10 mL) and a urine sample (25-50 mL) are collected from each patient. Blood and urine samples are then collected at 4 (+ 0.5) and 8 (+ 1) hours after contrast administration (if applicable); at 12 (± 1), 24 (± 2), 36 (± 2), 48 (± 2), 60 (± 2), 72 (± 2), and 84 (± 2) hours after enrollment, and thereafter daily up to day 7 to day 14 while the subject is hospitalized. Blood is collected via direct venipuncture or via other available venous access, such as an existing femoral sheath, central venous line, peripheral intravenous line or hep-lock. These study blood samples are processed to plasma at the clinical site, frozen and shipped to Astute Medical, Inc., San Diego, CA. The study urine samples are frozen and shipped to Astute Medical, Inc.

[00126] Example 4. Immunoassay format

[00127] Analytes are measured using standard sandwich enzyme immunoassay techniques. A first antibody which binds the analyte is immobilized in wells of a 96 well polystyrene microplate. Analyte standards and test samples are pipetted into the appropriate wells and any analyte present is bound by the immobilized antibody. After washing away any unbound substances, a horseradish peroxidase-conjugated second antibody which binds the analyte is added to the wells, thereby forming sandwich complexes with the analyte (if present) and the first antibody. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution comprising tetramethylbenzidine and hydrogen peroxide is added to the wells. Color develops in proportion to the amount of analyte present in the sample. The color development is stopped and the intensity of the color is measured at 540 nm or 570 nm. An analyte

concentration is assigned to the test sample by comparison to a standard curve determined from the analyte standards.

[00128] Table of Swiss-Prot numbers, preferred names and units for the analytes in the examples below.

[00129] Example 5. Apparently Healthy Donor and Chronic Disease Patient Samples

[00130] Human urine samples from donors with no known chronic or acute disease

("Apparently Healthy Donors") were purchased from two vendors (Golden West Biologicals, Inc., 27625 Commerce Center Dr., Temecula, CA 92590 and Virginia Medical Research, Inc., 915 First Colonial Rd., Virginia Beach, VA 23454). The urine samples were shipped and stored frozen at less than -20° C. The vendors supplied demographic information for the individual donors including gender, race (Black /White), smoking status and age. [00131] Human urine samples from donors with various chronic diseases ("Chronic Disease Patients") including congestive heart failure, coronary artery disease, chronic kidney disease, chronic obstructive pulmonary disease, diabetes mellitus and hypertension were purchased from Virginia Medical Research, Inc., 915 First Colonial Rd., Virginia Beach, VA 23454. The urine samples were shipped and stored frozen at less than -20 degrees centigrade. The vendor provided a case report form for each individual donor with age, gender, race (Black/White), smoking status and alcohol use, height, weight, chronic disease(s) diagnosis, current medications and previous surgeries.

[00132] Example 6. Use of Metalloproteinase inhibitor 1 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00133] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 1 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00134] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00135] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00136] Table 6.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

p Value 2.2E-16 2.7E-10 7.5E-7 6.6E-14 2.2E-9 8.9E-7 2.7E-13 8.0E-10 1.2E-7 nCohort Recovered 57 76 194 46 74 173 41 70 156 nCohort Non- recovered 260 240 121 271 242 142 276 246 159

Cutoff Quartile 2 6030 6020 6080 6030 6020 6080 6030 6020 6080

Sensitivity 82% 81 % 88% 80% 81 % 86% 79% 80% 85%

Specificity 54% 43% 34% 52% 43% 34% 54% 43% 35%

Cutoff Quartile 3 14200 14100 14200 14200 14100 14200 14200 14100 14200

Sensitivity 58% 58% 66% 56% 58% 63% 56% 58% 63%

Specificity 84% 76% 60% 87% 76% 61 % 88% 77% 63%

Cutoff Quartile 4 35900 35400 36300 35900 35400 36300 35900 35400 36300

Sensitivity 30% 31 % 35% 30% 31 % 33% 29% 31 % 33%

Specificity 98% 95% 81 % 100% 95% 82% 100% 96% 83%

OR Quartile 2 5.27 3.24 3.85 4.28 3.16 3.16 4.45 3.02 3.06 p Value 8.6E-8 3.5E-5 2.9E-5 1.1E-5 5.5E-5 7.2E-5 1.7E-5 1.4E-4 5.6E-5

Lower limit of 95%

CI 2.87 1.86 2.05 2.24 1.81 1.79 2.25 1.71 1.78

Upper limit of 95%

CI 9.67 5.64 7.24 8.21 5.53 5.57 8.78 5.32 5.28

OR Quartile 3 7.27 4.51 2.90 8.64 4.27 2.67 9.09 4.61 2.86 p Value 2.4E-7 5.1E-7 1.0E-5 2.1E-6 1.4E-6 2.6E-5 7.4E-6 1.0E-6 6.4E-6

Lower limit of 95%

CI 3.42 2.51 1.81 3.55 2.37 1.69 3.46 2.50 1.81

Upper limit of 95%

CI 15.4 8.12 4.66 21.1 7.70 4.22 23.9 8.50 4.52

OR Quartile 4 24.4 8.18 2.26 39.1 7.86 2.18 34.0 9.98 2.50 p Value 0.0017 7.8E-5 0.0021 0.010 1.1E-4 0.0033 0.014 1.5E-4 7.9E-4

Lower limit of 95%

CI 3.32 2.88 1.34 2.38 2.77 1.30 2.07 3.04 1.46

Upper limit of 95%

CI 180 23.2 3.79 642 22.3 3.66 559 32.7 4.27

[00137] Table 6.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 8310 20500 8310 20000 8210 19800

Average 15500 35500 15700 35200 14700 35200

Stdev 21000 36300 21100 36300 19000 36400 p (t-test) 2.9E-7 6.5E-7 2.4E-7

Min 634 297 634 297 634 297

Max 116000 146000 116000 146000 110000 146000 n (Patient) 106 211 104 213 99 218 O only

CI

Upper limit of 95%

CI 9.72 7.09 4.59 8.03 6.74 3.28 8.80 7.42 3.32

OR Quartile 4 5.62 4.77 1.58 4.55 4.60 1.39 3.86 4.83 1.68 p Value 3.7E-5 1.7E-5 0.089 3.1E-4 2.6E-5 0.21 0.0014 3.1E-5 0.046

Lower limit of 95%

CI 2.48 2.34 0.933 2.00 2.26 0.827 1.69 2.30 1.01

Upper limit of 95%

CI 12.8 9.71 2.68 10.4 9.38 2.33 8.82 10.1 2.81

[00138] Table 6.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO

sCr only

UO only

Stdev 32000 35500 32000 35400 31000 35900 p (t-test) 0.021 0.030 0.0057

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 127000 146000 n (Patient) 214 101 204 111 190 125

[00139] Table 6.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO Cohort recovered Cohort recovered Cohort recovered

Cohort Cohort Cohort

Median 8180 22000 8150 21100 7940 20000

Average 15800 38300 16200 37000 16600 35700

Stdev 21300 37200 21900 36900 22700 36300 p (t-test) 1.1E-9 2.9E-8 6.2E-7

Min 419 297 419 297 419 297

Max 111000 146000 111000 146000 111000 146000 n (Patient) 134 183 125 192 114 203 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 8710 21900 8740 21300 8730 21600

Average 17200 38000 17400 37700 16600 37900

Stdev 22900 37300 23000 37200 21600 37500 p (t-test) 1.6E-8 4.2E-8 9.3E-9

Min 419 297 419 297 419 297

Max 116000 146000 116000 146000 111000 146000 n (Patient) 140 177 138 179 135 182

UO only

Specificity 72% 69% 57% 71 % 69% 57% 72% 70% 58%

Cutoff Quartile 4 35900 35900 36300 35900 35900 36300 35900 35900 36300

Sensitivity 37% 36% 33% 35% 36% 31 % 33% 36% 33%

Specificity 90% 89% 79% 90% 88% 78% 89% 89% 80%

OR Quartile 2 5.42 4.46 3.26 4.70 4.28 3.42 4.35 4.19 2.87 p Value 3.8E-9 1.2E-7 5.3E-4 2.6E-8 2.1E-7 2.0E-4 7.9E-8 2.6E-7 5.1E-4

Lower limit of 95%

CI 3.09 2.57 1.67 2.73 2.47 1.79 2.54 2.43 1.58

Upper limit of 95%

CI 9.50 7.76 6.36 8.10 7.41 6.55 7.43 7.22 5.21

OR Quartile 3 5.24 4.29 2.53 4.41 4.07 2.21 4.28 4.23 2.31 p Value 2.8E-11 1.8E-9 2.4E-4 2.3E-9 6.2E-9 0.0011 1.0E-8 3.1E-9 4.4E-4

Lower limit of 95%

CI 3.22 2.67 1.54 2.71 2.53 1.37 2.60 2.62 1.45

Upper limit of 95%

CI 8.54 6.89 4.15 7.17 6.53 3.56 7.04 6.81 3.68

OR Quartile 4 5.38 4.39 1.85 4.62 4.24 1.65 3.83 4.44 1.96 p Value 3.4E-7 1.6E-6 0.023 3.5E-6 2.7E-6 0.060 4.8E-5 2.1E-6 0.011

Lower limit of 95%

CI 2.82 2.40 1.09 2.42 2.32 0.980 2.00 2.40 1.17

Upper limit of 95%

CI 10.3 8.03 3.14 8.81 7.76 2.79 7.31 8.23 3.29

[00140] Table 6.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 116000 146000 116000 146000 111000 146000 n (Patient) 160 157 157 160 156 161 O only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11300 21500 11600 20800 11300 21100

Average 24300 38300 24500 37500 24500 37300

Stdev 29900 38000 30100 37800 30200 37400 p (t-test) 4.4E-4 0.0010 0.0012

Min 297 399 297 399 297 399

Max 127000 146000 127000 146000 127000 146000 n (Patient) 211 104 208 107 205 110

[00141] Table 6.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.74 0.72 0.67 0.78 0.73 0.69 0.77 0.72 0.68

SE 0.032 0.031 0.031 0.032 0.031 0.030 0.034 0.032 0.030 p Value 7.8E-14 4.5E-13 8.1E-8 0 3.7E-14 5.0E-10 6.9E-15 1.3E-11 3.1E-9 nCohort Recovered 52 69 197 42 67 176 38 63 159 nCohort Non- recovered 275 257 128 285 259 149 289 263 166

Cutoff Quartile 2 260000 260000 262000 260000 260000 262000 260000 260000 262000

Sensitivity 80% 82% 84% 80% 82% 85% 79% 81 % 85%

Specificity 52% 51 % 31 % 57% 52% 33% 58% 51 % 35%

Cutoff Quartile 3 437000 433000 437000 437000 433000 437000 437000 433000 437000

Sensitivity 56% 57% 62% 55% 57% 63% 54% 56% 60%

Specificity 81 % 75% 58% 83% 76% 61 % 82% 75% 60%

Cutoff Quartile 4 707000 704000 708000 707000 704000 708000 707000 704000 708000

Sensitivity 28% 29% 41 % 28% 29% 40% 28% 29% 37%

Specificity 92% 88% 85% 98% 90% 88% 97% 89% 87%

OR Quartile 2 4.32 4.60 2.42 5.22 4.93 2.69 5.25 4.40 3.07 p Value 3.6E-6 1.4E-7 0.0021 1.7E-6 5.1E-8 3.6E-4 3.9E-6 6.1E-7 4.1E-5

Lower limit of 95%

CI 2.33 2.61 1.38 2.65 2.78 1.56 2.60 2.46 1.79

Upper limit of 95%

CI 8.02 8.12 4.26 10.3 8.76 4.64 10.6 7.87 5.24

OR Quartile 3 5.35 4.02 2.29 6.13 4.18 2.65 5.27 3.72 2.19 p Value 6.7E-6 5.6E-6 3.7E-4 2.6E-5 4.8E-6 2.2E-5 1.3E-4 3.0E-5 5.2E-4

Lower limit of 95%

CI 2.58 2.21 1.45 2.64 2.27 1.69 2.25 2.01 1.41

Upper limit of 95%

CI 11.1 7.34 3.61 14.3 7.72 4.16 12.4 6.90 3.42

OR Quartile 4 4.75 3.08 4.09 16.3 3.49 4.72 14.4 3.19 4.14 p Value 0.0037 0.0049 1.7E-7 0.0063 0.0030 4.0E-8 0.0090 0.0061 8.0E-7

Lower limit of 95%

CI 1.66 1.41 2.41 2.20 1.53 2.71 1.94 1.39 2.36

Upper limit of 95%

CI 13.6 6.76 6.94 120 7.99 8.21 107 7.32 7.29

[00142] Table 6.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 282000 528000 282000 528000 291000 521000

Average 380000 547000 376000 547000 382000 541000

Stdev 248000 273000 245000 273000 249000 274000 p (t-test) 3.0E-7 1.7E-7 1.8E-6

Min 66700 13600 66700 13600 66700 13600

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 101 226 99 228 94 233 O only

CI

Upper limit of 95%

CI 5.37 4.65 3.96 4.71 4.73 3.52 4.59 4.47 2.81

OR Quartile 4 3.79 2.98 4.01 3.55 3.21 3.37 3.07 2.93 3.05 p Value 4.4E-4 9.7E-4 2.2E-7 0.0015 5.9E-4 4.6E-6 0.0051 0.0016 2.8E-5

Lower limit of 95%

CI 1.80 1.56 2.37 1.63 1.65 2.00 1.40 1.51 1.81

Upper limit of 95%

CI 7.96 5.68 6.77 7.76 6.25 5.66 6.72 5.72 5.14

[00143] Table 6.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 247000 288000 249000 288000 243000 292000 p (t-test) 1.4E-9 2.8E-8 4.6E-8

Min 13600 135000 13600 135000 13600 135000

Max 1120000 1090000 1120000 1090000 1120000 1090000 n (Patient) 219 106 208 117 193 132

[00144] Table 6.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Cohort recovered Cohort recovered Cohort recovered

Cohort Cohort Cohort

Median 296000 564000 293000 536000 279000 529000

Average 383000 573000 381000 566000 372000 561000

Stdev 232000 278000 233000 277000 236000 274000 p (t-test) 3.3E-10 1.7E-9 1.6E-9

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 134 193 125 202 113 214 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 293000 567000 289000 567000 282000 565000

Average 381000 580000 377000 581000 376000 577000

Stdev 234000 275000 231000 275000 233000 274000 p (t-test) 3.6E-11 l .OE-11 2.7E-11

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 139 188 137 190 133 194

UO only

Cutoff Quartile 4 707000 707000 708000 707000 707000 708000 707000 707000 708000

Sensitivity 35% 36% 41 % 34% 36% 40% 33% 35% 39%

Specificity 90% 89% 82% 90% 90% 83% 89% 89% 84%

OR Quartile 2 3.98 3.90 3.99 4.08 4.05 3.32 4.51 4.05 3.39 p Value 3.1E-7 4.9E-7 7.7E-5 1.6E-7 2.5E-7 1.9E-4 2.2E-8 2.2E-7 7.1E-5

Lower limit of 95%

CI 2.34 2.30 2.01 2.41 2.38 1.77 2.66 2.39 1.86

Upper limit of 95%

CI 6.75 6.64 7.92 6.91 6.89 6.24 7.65 6.88 6.19

OR Quartile 3 2.70 3.05 2.42 2.69 3.24 2.39 3.00 3.28 2.23 p Value 2.0E-5 1.7E-6 3.2E-4 2.6E-5 5.4E-7 2.7E-4 6.8E-6 5.0E-7 5.5E-4

Lower limit of 95%

CI 1.71 1.93 1.49 1.70 2.05 1.50 1.86 2.06 1.42

Upper limit of 95%

CI 4.26 4.82 3.91 4.28 5.14 3.83 4.84 5.21 3.52

OR Quartile 4 4.66 4.58 3.22 4.47 4.90 3.29 4.09 4.59 3.37 p Value 1.5E-6 1.2E-6 1.1E-5 5.1E-6 6.9E-7 6.8E-6 3.1E-5 1.9E-6 4.6E-6

Lower limit of 95%

CI 2.49 2.48 1.91 2.35 2.61 1.96 2.11 2.45 2.00

Upper limit of 95%

CI 8.73 8.45 5.42 8.51 9.17 5.52 7.94 8.59 5.66

[00145] Table 6.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

n (Patient) 159 168 156 171 154 173 O only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 375000 582000 369000 586000 359000 596000

Average 436000 616000 433000 617000 428000 622000

Stdev 245000 296000 243000 294000 241000 292000 p (t-test) 1.4E-8 4.2E-9 3.7E-10

Min 13600 135000 13600 135000 13600 135000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 215 110 212 113 209 116

[00146] Example 7. Use of Metalloproteinase inhibitor 1 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00147] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 1 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00148] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00149] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00150] Table 7.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 116000 146000 110000 146000 110000 146000 n (Patient) 108 209 104 213 100 217 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 8650 20500 8510 20500 8310 20500

Average 16400 36800 16400 36600 16500 36400

Stdev 22300 36800 22400 36700 22600 36500 p (t-test) 5.2E-8 8.5E-8 1.3E-7

Min 475 297 475 297 475 297

Max 116000 146000 116000 146000 116000 146000 n (Patient) 126 190 124 192 122 194

UO only

Upper limit of 95%

CI 7.87 5.76 7.65 7.99 6.00 7.33 8.15 6.24 6.03

OR Quartile 3 4.80 4.22 2.81 4.99 4.26 2.74 5.62 4.31 2.47 p Value 2.2E-9 5.6E-9 7.2E-5 1.8E-9 5.3E-9 4.8E-5 3.9E-10 4.9E-9 1.7E-4

Lower limit of 95%

CI 2.87 2.60 1.69 2.96 2.62 1.69 3.27 2.64 1.54

Upper limit of 95%

CI 8.02 6.85 4.67 8.42 6.93 4.46 9.65 7.02 3.96

OR Quartile 4 3.86 4.63 2.10 4.60 4.47 2.41 4.29 4.32 2.27 p Value 7.1E-5 3.5E-6 0.0064 2.6E-5 5.7E-6 9.9E-4 6.3E-5 9.3E-6 0.0019

Lower limit of 95%

CI 1.98 2.42 1.23 2.26 2.34 1.43 2.10 2.26 1.35

Upper limit of 95%

CI 7.51 8.84 3.59 9.38 8.54 4.07 8.74 8.26 3.82

[00151] Table 7.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11900 21700 11800 21300 11200 20900

Average 25200 38600 25300 37400 24900 36700

Stdev 31100 37200 31400 36500 31200 36200 p (t-test) 0.0012 0.0030 0.0029

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 146000 140000 n (Patient) 226 89 220 95 207 108

[00152] Table 7.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 87% 86% 88% 87% 86% 87% 86% 86% 86% Specificity 37% 35% 30% 38% 36% 30% 38% 36% 31 %

Cutoff Quartile 3 14200 14200 14200 14200 14200 14200 14200 14200 14200

Sensitivity 66% 66% 65% 65% 66% 64% 65% 65% 63%

Specificity 67% 65% 56% 66% 65% 56% 67% 65% 56%

Cutoff Quartile 4 35900 35900 36300 35900 35900 36300 35900 35900 36300

Sensitivity 37% 38% 32% 36% 38% 32% 36% 37% 32%

Specificity 87% 87% 78% 87% 87% 78% 87% 87% 78%

OR Quartile 2 3.77 3.34 3.17 3.90 3.40 2.86 3.86 3.45 2.81 p Value 2.9E-6 2.0E-5 0.0011 1.6E-6 1.6E-5 0.0021 1.5E-6 1.2E-5 0.0014

Lower limit of 95%

CI 2.16 1.92 1.59 2.24 1.95 1.46 2.23 1.98 1.49

Upper limit of 95%

CI 6.57 5.81 6.33 6.80 5.91 5.60 6.70 6.02 5.30

OR Quartile 3 3.97 3.64 2.33 3.75 3.54 2.24 3.77 3.44 2.19 p Value 7.3E-9 4.6E-8 0.0010 2.5E-8 8.4E-8 0.0016 2.4E-8 1.5E-7 0.0015

Lower limit of 95%

CI 2.49 2.29 1.41 2.36 2.23 1.36 2.36 2.17 1.35

Upper limit of 95%

CI 6.33 5.79 3.86 5.97 5.63 3.70 6.00 5.46 3.55

OR Quartile 4 3.90 4.05 1.63 3.77 3.98 1.59 3.83 3.92 1.70 p Value 2.6E-6 1.0E-6 0.078 4.5E-6 1.4E-6 0.092 4.6E-6 1.8E-6 0.048

Lower limit of 95%

CI 2.21 2.31 0.947 2.14 2.27 0.927 2.16 2.23 1.00

Upper limit of 95%

CI 6.88 7.11 2.80 6.65 6.98 2.74 6.81 6.86 2.88

[00153] Table 7.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Average 19400 40000 19500 39700 19600 39500

Stdev 24900 38400 24900 38400 25000 38300 p (t-test) 2.4E-8 3.7E-8 5.7E-8

Min 419 297 419 297 419 297

Max 116000 146000 116000 146000 116000 146000 n (Patient) 172 145 171 146 170 147 O only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11800 21500 11900 21300 11800 20500

Average 25400 37400 25500 37200 25200 37000

Stdev 31500 36500 31500 36400 31100 36700 p (t-test) 0.0036 0.0046 0.0033

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 146000 140000 n (Patient) 223 92 222 93 214 101

CI

Upper limit of 95%

CI 6.23 6.17 3.41 5.90 6.06 3.34 5.98 5.95 3.23

[00154] Table 7.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.67 0.63 0.65 0.67 0.63 0.64 0.67 0.63

SE 0.033 0.031 0.035 0.033 0.031 0.035 0.033 0.031 0.035 p Value 4.4E-6 2.0E-8 1.3E-4 8.7E-6 2.0E-8 1.5E-4 9.7E-6 2.0E-8 2.5E-4 nCohort Recovered 203 185 217 201 185 216 200 185 214 nCohort Non- recovered 114 132 98 116 132 99 117 132 101

Cutoff Quartile 2 6030 6030 6080 6030 6030 6080 6030 6030 6080

Sensitivity 86% 87% 87% 86% 87% 87% 86% 87% 87%

Specificity 31 % 34% 30% 31 % 34% 31 % 32% 34% 31 %

Cutoff Quartile 3 14200 14200 14200 14200 14200 14200 14200 14200 14200

Sensitivity 66% 65% 64% 65% 65% 64% 64% 65% 62%

Specificity 59% 61 % 56% 58% 61 % 56% 58% 61 % 56%

Cutoff Quartile 4 35900 35900 36300 35900 35900 36300 35900 35900 36300

Sensitivity 37% 39% 35% 36% 39% 34% 36% 39% 34%

Specificity 81 % 84% 79% 81 % 84% 79% 81 % 84% 79%

OR Quartile 2 2.76 3.41 2.86 2.85 3.41 2.91 2.90 3.41 3.02 p Value 0.0010 5.1E-5 0.0016 7.0E-4 5.1E-5 0.0013 5.7E-4 5.1E-5 8.7E-4

Lower limit of 95%

CI 1.50 1.88 1.49 1.56 1.88 1.52 1.58 1.88 1.57

Upper limit of 95%

CI 5.05 6.17 5.48 5.23 6.17 5.58 5.32 6.17 5.79

OR Quartile 3 2.72 2.87 2.31 2.55 2.87 2.23 2.47 2.87 2.08 p Value 3.9E-5 8.5E-6 8.6E-4 1.1E-4 8.5E-6 0.0013 1.7E-4 8.5E-6 0.0031

Lower limit of 95%

CI 1.69 1.80 1.41 1.59 1.80 1.37 1.54 1.80 1.28

Upper limit of 95%

CI 4.39 4.56 3.78 4.09 4.56 3.64 3.95 4.56 3.37

OR Quartile 4 2.53 3.39 2.03 2.43 3.39 1.99 2.39 3.39 1.91 p Value 4.5E-4 6.2E-6 0.0088 7.6E-4 6.2E-6 0.011 9.7E-4 6.2E-6 0.017

Lower limit of 95%

CI 1.51 2.00 1.20 1.45 2.00 1.17 1.42 2.00 1.12

Upper limit of 95%

CI 4.26 5.75 3.45 4.09 5.75 3.37 4.00 5.75 3.23

[00155] Table 7.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 105 222 102 225 98 229 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 279000 566000 279000 566000 275000 566000

Average 365000 569000 362000 569000 358000 570000

Stdev 235000 269000 235000 268000 232000 268000 p (t-test) 2.1E-11 1.4E-11 4.4E-12

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 121 205 119 207 117 209

UO only

CI

OR Quartile 3 3.81 3.84 2.50 3.78 3.89 2.41 3.92 3.94 2.26 p Value 1.8E-7 4.1E-8 2.6E-4 2.7E-7 3.8E-8 2.6E-4 2.2E-7 3.5E-8 5.1E-4

Lower limit of 95%

CI 2.31 2.38 1.53 2.28 2.40 1.50 2.34 2.42 1.43

Upper limit of 95%

CI 6.29 6.22 4.08 6.26 6.31 3.87 6.56 6.41 3.57

OR Quartile 4 5.23 4.21 3.96 4.96 4.08 3.68 6.33 4.41 3.64 p Value 1.1E-5 1.2E-5 3.3E-7 2.1E-5 1.9E-5 1.0E-6 9.5E-6 1.2E-5 1.2E-6

Lower limit of 95%

CI 2.50 2.21 2.33 2.37 2.14 2.18 2.80 2.27 2.16

Upper limit of 95%

CI 10.9 8.03 6.71 10.4 7.77 6.20 14.3 8.55 6.13

[00156] Table 7.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort recovered Cohort recovered Cohort recovered

Cohort Cohort Cohort

Median 375000 598000 375000 598000 369000 596000

Average 448000 619000 443000 618000 432000 616000

Stdev 256000 287000 253000 288000 247000 288000 p (t-test) 2.5E-7 7.7E-8 4.4E-9

Min 13600 135000 13600 135000 13600 135000

Max 1120000 1090000 1120000 1090000 1120000 1090000 n (Patient) 231 94 225 100 210 115

[00157] Table 7.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 303000 573000 300000 576000 296000 576000

Average 394000 586000 390000 587000 384000 587000

Stdev 239000 276000 238000 275000 232000 276000 p (t-test) l.OE-10 2.8E-11 7.0E-12

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 154 173 152 175 148 179 sCr only

UO only

Specificity 39% 39% 30% 39% 39% 30% 40% 39% 32%

Cutoff Quartile 3 437000 437000 437000 437000 437000 437000 437000 437000 437000

Sensitivity 62% 63% 66% 62% 63% 66% 62% 64% 65%

Specificity 63% 63% 57% 64% 64% 57% 64% 64% 57%

Cutoff Quartile 4 707000 707000 708000 707000 707000 708000 707000 707000 708000

Sensitivity 36% 37% 42% 36% 37% 42% 36% 37% 42%

Specificity 87% 87% 82% 88% 87% 82% 89% 87% 83%

OR Quartile 2 4.38 4.57 3.34 4.54 4.65 3.07 4.50 4.73 3.45 p Value 1.6E-7 1.3E-7 5.9E-4 8.3E-8 9.8E-8 9.7E-4 7.4E-8 7.2E-8 1.7E-4

Lower limit of 95%

CI 2.52 2.60 1.68 2.61 2.64 1.58 2.60 2.69 1.81

Upper limit of 95%

CI 7.61 8.04 6.65 7.88 8.18 5.99 7.78 8.32 6.59

OR Quartile 3 2.76 2.98 2.56 2.91 3.06 2.53 2.93 3.14 2.43 p Value 9.2E-6 2.0E-6 2.2E-4 3.3E-6 1.2E-6 2.4E-4 3.2E-6 6.9E-7 2.4E-4

Lower limit of 95%

CI 1.76 1.90 1.55 1.86 1.95 1.54 1.86 2.00 1.51

Upper limit of 95%

CI 4.32 4.67 4.22 4.57 4.80 4.15 4.60 4.93 3.91

OR Quartile 4 3.74 3.87 3.26 3.94 3.81 3.34 4.39 4.07 3.57 p Value 4.4E-6 1.9E-6 1.1E-5 2.6E-6 2.5E-6 7.0E-6 8.8E-7 1.1E-6 1.7E-6

Lower limit of 95%

CI 2.13 2.22 1.92 2.22 2.18 1.97 2.44 2.31 2.12

Upper limit of 95%

CI 6.57 6.76 5.51 6.97 6.65 5.65 7.93 7.14 6.02

[00158] Table 7.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Stdev 242000 272000 242000 271000 240000 271000 p (t-test) 6.4E-12 3.5E-12 9.9E-13

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 169 158 168 159 167 160 O only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 377000 598000 375000 596000 371000 596000

Average 441000 626000 441000 624000 436000 621000

Stdev 251000 289000 251000 288000 249000 287000 p (t-test) 1.2E-8 1.7E-8 5.7E-9

Min 13600 135000 13600 135000 13600 135000

Max 1120000 1090000 1120000 1090000 1120000 1090000 n (Patient) 226 99 225 100 217 108

Upper limit of 95%

CI 6.20 6.53 5.39 6.41 6.41 5.26 7.14 6.86 5.85

[00159] Table 7.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.68 0.70 0.68 0.69 0.70 0.68 0.69 0.70 0.69

SE 0.031 0.030 0.033 0.031 0.030 0.033 0.031 0.030 0.032 p Value 3.2E-9 1.9E-11 3.8E-8 1.8E-9 1.9E-11 4.5E-8 5.7E-10 1.9E-11 8.2E-9 nCohort Recovered 204 184 220 202 184 219 201 184 217 nCohort Non- recovered 123 143 105 125 143 106 126 143 108

Cutoff Quartile 2 260000 260000 262000 260000 260000 262000 260000 260000 262000

Sensitivity 89% 88% 89% 89% 88% 89% 89% 88% 89%

Specificity 33% 35% 31 % 34% 35% 32% 34% 35% 32%

Cutoff Quartile 3 437000 437000 437000 437000 437000 437000 437000 437000 437000

Sensitivity 64% 64% 65% 64% 64% 64% 64% 64% 65%

Specificity 58% 61 % 57% 58% 61 % 57% 59% 61 % 57%

Cutoff Quartile 4 707000 707000 708000 707000 707000 708000 707000 707000 708000

Sensitivity 39% 38% 40% 39% 38% 40% 40% 38% 41 %

Specificity 83% 85% 82% 84% 85% 82% 84% 85% 82%

OR Quartile 2 3.89 4.05 3.54 4.02 4.05 3.60 4.09 4.05 3.73 p Value 2.2E-5 3.4E-6 1.9E-4 1.4E-5 3.4E-6 1.6E-4 1.1E-5 3.4E-6 1.0E-4

Lower limit of 95%

CI 2.08 2.24 1.82 2.15 2.24 1.85 2.18 2.24 1.92

Upper limit of 95%

CI 7.29 7.30 6.89 7.54 7.30 7.01 7.66 7.30 7.25

OR Quartile 3 2.51 2.81 2.42 2.50 2.81 2.34 2.56 2.81 2.46 p Value 9.2E-5 7.8E-6 3.2E-4 9.7E-5 7.8E-6 5.1E-4 6.2E-5 7.8E-6 2.3E-4

Lower limit of 95%

CI 1.58 1.78 1.49 1.58 1.78 1.45 1.62 1.78 1.52

Upper limit of 95%

CI 3.99 4.41 3.91 3.96 4.41 3.77 4.05 4.41 3.96

OR Quartile 4 3.20 3.63 3.00 3.30 3.63 2.94 3.47 3.63 3.24 p Value 1.0E-5 1.8E-6 3.4E-5 6.1E-6 1.8E-6 4.7E-5 2.6E-6 1.8E-6 9.3E-6

Lower limit of 95%

CI 1.91 2.14 1.78 1.97 2.14 1.75 2.07 2.14 1.93

Upper limit of 95%

CI 5.36 6.17 5.04 5.54 6.17 4.93 5.84 6.17 5.44

[00160] Example 8. Use of Metalloproteinase inhibitor 1 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00161] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 1 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00162] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00163] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00164] Table 8.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 12400 25100 12500 28600 12700 30000

Average 26300 47200 26300 48500 26300 49300

Stdev 31200 42100 31100 43100 31000 43400 p (t-test) 2.0E-4 1.3E-4 8.7E-5

Min 297 873 297 873 297 873

Max 146000 140000 146000 140000 146000 140000 n (Patient) 275 40 278 37 279 36

[00165] Table 8.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

Cohort Cohort Cohort Cohort Cohort Cohort

Median 10100 23400 10600 31500 10800 31500

Average 18900 42900 18900 46800 20700 46900

Stdev 23700 39500 23400 40500 26300 39900 p (t-test) 7.7E-11 9.3E-14 2.4E-11

Min 419 297 297 399 297 399

Max 121000 146000 121000 146000 121000 146000 n (Patient) 186 131 204 113 219 98 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11000 22400 10900 29300 11000 27300

Average 19500 42800 19400 46000 21400 44900

Stdev 23700 40400 23600 41200 26500 40600 p (t-test) 4.0E-10 2.3E-12 2.6E-9

Min 419 297 297 399 297 399

Max 121000 146000 121000 146000 121000 146000 n (Patient) 192 124 206 110 218 98

UO only

OR Quartile 2 3.68 3.00 5.70 4.64 3.94 5.17 6.37 4.68 5.04 p Value 2.4E-5 2.9E-4 0.0012 1.2E-5 5.6E-5 0.0022 9.4E-6 4.6E-5 0.0026

Lower limit of 95% CI 2.01 1.66 1.99 2.33 2.02 1.80 2.81 2.23 1.76 Upper limit of 95% CI 6.74 5.44 16.3 9.21 7.67 14.8 14.4 9.82 14.4

OR Quartile 3 3.51 2.98 2.46 4.36 3.60 2.36 4.52 3.45 2.28 p Value 1.8E-7 5.6E-6 0.0032 8.2E-9 3.8E-7 0.0061 2.3E-8 2.0E-6 0.0088

Lower limit of 95% CI 2.19 1.86 1.35 2.64 2.20 1.28 2.66 2.07 1.23 Upper limit of 95% CI 5.63 4.77 4.48 7.20 5.90 4.36 7.66 5.75 4.22

OR Quartile 4 5.04 4.41 2.28 6.27 5.09 2.37 5.19 3.95 2.46 p Value 8.7E-9 7.5E-8 0.0072 7.7E-11 4.4E-9 0.0058 2.6E-9 5.0E-7 0.0042

Lower limit of 95% CI 2.90 2.57 1.25 3.61 2.95 1.28 3.02 2.31 1.33 Upper limit of 95% CI 8.74 7.58 4.16 10.9 8.76 4.39 8.93 6.75 4.56

[00166] Table 8.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 31100 38700 30900 39700 30900 39700 p (t-test) 0.0014 8.6E-4 8.6E-4

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 146000 140000 n (Patient) 246 69 251 64 251 64

[00167] Table 8.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 116000 146000 116000 146000 116000 146000 n (Patient) 173 144 189 128 201 116 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 10800 22000 10200 27300 10800 22700

Average 19100 41400 18800 43900 20200 44100

Stdev 23100 39900 22900 40400 24400 41100 p (t-test) 1.4E-9 1.3E-11 3.8E-10

Min 419 297 297 399 297 399

Max 116000 146000 116000 146000 116000 146000 n (Patient) 180 136 192 124 204 112

UO only

Lower limit of 95% CI 2.19 1.92 1.36 2.40 2.20 1.31 2.24 2.05 1.31 Upper limit of 95% CI 5.56 4.84 3.99 6.26 5.73 3.96 5.94 5.44 3.96

OR Quartile 4 4.63 4.08 2.33 5.35 4.77 2.33 4.32 3.89 2.33 p Value 7.2E-8 4.1E-7 0.0029 2.7E-9 2.0E-8 0.0036 8.1E-8 5.7E-7 0.0036

Lower limit of 95% CI 2.65 2.37 1.33 3.08 2.76 1.32 2.53 2.28 1.32 Upper limit of 95% CI 8.08 7.04 4.08 9.29 8.22 4.12 7.37 6.62 4.12

[00168] Table 8.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.70 0.68 0.67 0.73 0.71 0.66 0.71 0.70 0.66

SE 0.030 0.031 0.037 0.029 0.030 0.037 0.031 0.032 0.037 p Value 1.4E-11 4.0E-9 5.2E-6 8.0E-15 8.8E-12 1.3E-5 5.9E-12 5.9E-10 1.3E-5 nCohort Non- persistent 169 176 234 184 188 237 197 200 237 nCohort Persistent 148 140 81 133 128 78 120 116 78

Cutoff Quartile 2 6030 6020 6080 6030 6020 6080 6030 6020 6080

Sensitivity 86% 86% 91 % 89% 88% 91 % 90% 90% 91 %

Specificity 35% 34% 31 % 35% 34% 30% 34% 34% 30%

Cutoff Quartile 3 14200 14100 14200 14200 14100 14200 14200 14100 14200

Sensitivity 66% 65% 68% 70% 68% 68% 70% 68% 68%

Specificity 64% 62% 56% 64% 62% 56% 62% 60% 56%

Cutoff Quartile 4 35900 35400 36300 35900 35400 36300 35900 35400 36300

Sensitivity 40% 39% 40% 44% 42% 40% 42% 41 % 40%

Specificity 88% 86% 80% 88% 87% 80% 85% 84% 80%

OR Quartile 2 3.43 3.03 4.70 4.20 3.89 4.43 4.64 4.37 4.43 p Value 2.1E-5 1.3E-4 2.3E-4 5.2E-6 1.6E-5 4.1E-4 6.2E-6 1.4E-5 4.1E-4

Lower limit of 95% CI 1.95 1.72 2.06 2.26 2.10 1.94 2.38 2.24 1.94

Upper limit of 95% CI 6.06 5.34 10.7 7.78 7.21 10.1 9.02 8.50 10.1

OR Quartile 3 3.47 3.02 2.69 4.16 3.50 2.67 3.80 3.27 2.67 p Value 1.4E-7 2.7E-6 2.7E-4 5.0E-9 2.3E-7 3.8E-4 7.0E-8 1.5E-6 3.8E-4

Lower limit of 95% CI 2.18 1.90 1.58 2.58 2.18 1.55 2.34 2.02 1.55

Upper limit of 95% CI 5.51 4.79 4.59 6.70 5.62 4.57 6.16 5.30 4.57

OR Quartile 4 4.67 4.10 2.60 5.69 4.76 2.60 4.28 3.85 2.60 p Value 8.1E-8 4.6E-7 6.4E-4 1.3E-9 2.4E-8 7.2E-4 1.0E-7 6.9E-7 7.2E-4

Lower limit of 95% CI 2.66 2.37 1.50 3.25 2.75 1.49 2.51 2.26 1.49

Upper limit of 95% CI 8.21 7.09 4.50 9.99 8.23 4.52 7.31 6.55 4.52

[00169] Table 8.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 350000 596000 359000 594000 375000 590000

Average 432000 602000 443000 604000 450000 607000

Stdev 255000 276000 261000 273000 261000 278000 p (t-test) 3.8E-8 5.1E-7 2.9E-6

Min 13600 105000 13600 151000 13600 154000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 208 118 223 103 236 90

UO only

[00170] Table 8.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.68 0.68 0.68 0.67 0.66 0.68 0.67 0.66 0.68

SE 0.030 0.031 0.040 0.031 0.032 0.040 0.033 0.033 0.041 p Value 1.3E-9 7.6E-9 6.7E-6 4.7E-8 4.9E-7 4.9E-6 1.3E-7 1.3E-6 8.7E-6 nCohort Non- persistent 187 192 260 204 205 263 219 217 265 nCohort Persistent 140 134 65 123 121 62 108 109 60

Cutoff Quartile 2 260000 260000 262000 260000 260000 262000 260000 260000 262000

Sensitivity 86% 86% 89% 86% 86% 89% 86% 86% 88%

Specificity 33% 33% 28% 32% 32% 28% 31 % 31 % 28%

Cutoff Quartile 3 437000 433000 437000 437000 433000 437000 437000 433000 437000

Sensitivity 66% 66% 68% 65% 64% 69% 66% 65% 70%

Specificity 61 % 61 % 54% 59% 59% 54% 58% 58% 54%

Cutoff Quartile 4 707000 704000 708000 707000 704000 708000 707000 704000 708000

Sensitivity 38% 37% 43% 38% 36% 45% 40% 37% 45%

Specificity 84% 83% 79% 83% 81 % 79% 82% 81 % 79%

OR Quartile 2 2.98 2.96 3.30 2.92 2.84 3.08 2.73 2.80 2.93 p Value 1.5E-4 2.0E-4 0.0048 3.8E-4 5.4E-4 0.0081 0.0014 0.0011 0.011

Lower limit of 95%

CI 1.70 1.67 1.44 1.62 1.57 1.34 1.48 1.51 1.28

Upper limit of 95%

CI 5.22 5.23 7.55 5.26 5.13 7.06 5.06 5.18 6.75

OR Quartile 3 3.06 3.15 2.48 2.66 2.56 2.70 2.60 2.54 2.78 p Value 1.6E-6 1.1E-6 0.0019 3.6E-5 7.3E-5 0.0010 9.3E-5 1.3E-4 9.0E-4

Lower limit of 95%

CI 1.94 1.99 1.40 1.67 1.61 1.49 1.61 1.58 1.52

Upper limit of 95%

CI 4.83 5.00 4.41 4.23 4.08 4.87 4.20 4.09 5.07

OR Quartile 4 3.32 2.98 2.89 2.99 2.51 3.19 3.05 2.42 3.12 p Value 6.9E-6 3.5E-5 3.0E-4 3.1E-5 4.1E-4 9.7E-5 2.4E-5 7.9E-4 1.5E-4

Lower limit of 95%

CI 1.97 1.78 1.62 1.78 1.51 1.78 1.82 1.44 1.73

Upper limit of 95%

CI 5.60 4.99 5.13 5.00 4.19 5.71 5.13 4.04 5.63

[00171] Table 8.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Min 13600 105000 13600 135000 13600 135000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 176 151 190 137 205 122 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 326000 594000 339000 582000 342000 586000

Average 411000 598000 422000 598000 432000 601000

Stdev 248000 272000 252000 274000 255000 275000 p (t-test) 3.0E-10 6.3E-9 4.2E-8

Min 13600 105000 13600 151000 13600 154000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 182 144 194 132 207 119

UO only

[00172] Table 8.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Average 449000 632000 451000 637000 453000 634000

Stdev 257000 285000 256000 289000 257000 290000 p (t-test) 8.9E-8 1.0E-7 2.4E-7

Min 13600 135000 13600 135000 13600 135000

Max 1120000 1090000 1120000 1090000 1120000 1090000 n (Patient) 240 85 245 80 246 79

[00173] Table 8.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 314000 577000 327000 572000 339000 578000

Average 407000 590000 417000 595000 424000 602000

Stdev 248000 273000 250000 276000 252000 277000 p (t-test) 6.5E-10 2.6E-9 5.7E-9

Min 13600 105000 13600 135000 13600 135000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 169 158 183 144 196 131 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 321000 586000 328000 577000 340000 582000

Average 406000 595000 416000 597000 424000 602000

Stdev 246000 272000 249000 274000 252000 275000 p (t-test) 1.8E-10 1.4E-9 5.1E-9

Min 13600 105000 13600 151000 13600 154000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 175 151 186 140 198 128

UO only

Specificity 85% 85% 81 % 84% 84% 81 % 83% 83% 81 %

OR Quartile 2 3.68 3.59 4.62 3.76 3.56 4.45 3.65 3.50 4.37 p Value 4.3E-6 8.6E-6 1.1E-4 7.4E-6 1.8E-5 1.7E-4 2.4E-5 4.5E-5 2.0E-4

Lower limit of 95%

CI 2.11 2.05 2.12 2.11 1.99 2.05 2.00 1.92 2.01

Upper limit of 95%

CI 6.43 6.31 10.0 6.72 6.35 9.68 6.66 6.39 9.51

OR Quartile 3 3.50 3.68 2.74 3.03 3.10 2.79 3.06 3.04 2.71 p Value 7.0E-8 2.6E-8 1.1E-4 1.7E-6 1.2E-6 1.0E-4 2.2E-6 2.7E-6 1.6E-4

Lower limit of 95%

CI 2.22 2.33 1.64 1.93 1.96 1.66 1.92 1.91 1.62

Upper limit of 95%

CI 5.52 5.83 4.58 4.78 4.91 4.67 4.85 4.84 4.56

OR Quartile 4 3.25 3.38 3.17 3.09 3.07 3.33 2.95 2.89 3.17 p Value 1.5E-5 7.2E-6 2.1E-5 2.2E-5 2.3E-5 9.9E-6 3.8E-5 5.1E-5 2.3E-5

Lower limit of 95%

CI 1.90 1.99 1.86 1.84 1.83 1.95 1.76 1.73 1.86

Upper limit of 95%

CI 5.55 5.75 5.40 5.21 5.17 5.69 4.94 4.84 5.42

[00174] Example 9. Use of Metalloproteinase inhibitor 1 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00175] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 1 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00176] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent". [00177] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00178] Table 9.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.73 0.70 0.68 0.68 0.68 0.68 0.70 0.70 0.69

SE 0.028 0.029 0.038 0.030 0.030 0.041 0.030 0.030 0.043 p Value 6.7E-16 7.2E-12 1.0E-6 1.7E-9 2.5E-9 7.7E-6 2.7E-11 4.0E-11 7.8E-6 nCohort Non- persistent 119 130 241 158 162 256 179 184 263 nCohort Persistent 198 186 74 159 154 59 138 132 52

Cutoff Quartile 2 6030 6020 6080 6030 6020 6080 6030 6020 6080

Sensitivity 86% 84% 92% 86% 85% 92% 88% 86% 90%

Specificity 43% 38% 30% 36% 35% 29% 35% 33% 28%

Cutoff Quartile 3 14200 14100 14200 14200 14100 14200 14200 14100 14200

Sensitivity 65% 65% 70% 64% 64% 71 % 69% 70% 73%

Specificity 74% 71 % 56% 64% 64% 55% 64% 64% 54%

Cutoff Quartile 4 35900 35400 36300 35900 35400 36300 35900 35400 36300

Sensitivity 34% 35% 39% 37% 38% 41 % 40% 41 % 42%

Specificity 90% 89% 79% 87% 88% 79% 86% 86% 78%

OR Quartile 2 4.55 3.38 4.92 3.51 3.01 4.39 3.77 3.14 3.68 p Value 3.8E-8 6.8E-6 3.8E-4 9.1E-6 8.3E-5 0.0024 1.2E-5 1.2E-4 0.0078

Lower limit of 95% CI 2.65 1.99 2.05 2.02 1.74 1.69 2.08 1.75 1.41

Upper limit of 95% CI 7.81 5.75 11.9 6.12 5.21 11.4 6.83 5.63 9.62

OR Quartile 3 5.19 4.40 3.01 3.17 3.14 2.98 3.97 4.11 3.23 p Value 1.3E-10 1.8E-9 1.1E-4 8.2E-7 1.0E-6 5.0E-4 1.1E-8 6.8E-9 4.8E-4

Lower limit of 95% CI 3.14 2.72 1.72 2.00 1.98 1.61 2.48 2.55 1.67

Upper limit of 95% CI 8.58 7.13 5.27 5.02 4.97 5.51 6.37 6.63 6.25

OR Quartile 4 4.66 4.45 2.46 3.85 4.41 2.51 4.08 4.40 2.65 p Value 5.6E-6 3.5E-6 0.0016 2.5E-6 3.3E-7 0.0026 3.8E-7 1.0E-7 0.0022

Lower limit of 95% CI 2.40 2.37 1.40 2.20 2.49 1.38 2.37 2.55 1.42

Upper limit of 95% CI 9.07 8.37 4.31 6.74 7.80 4.56 7.02 7.60 4.94

[00179] Table 9.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Median 8650 20500 9980 20500 9450 22000

Average 16600 36700 18300 38600 18900 39900

Stdev 22600 36700 22800 38400 24300 38400 p (t-test) 9.4E-8 3.1E-8 1.0E-8

Min 475 297 419 297 419 297

Max 116000 146000 116000 146000 116000 146000 n (Patient) 126 190 154 162 169 147

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11600 21900 12100 21900 12100 24700

Average 24400 39900 25100 41000 25200 41900

Stdev 30500 37400 30600 38700 30600 39300 p (t-test) 1.6E-4 2.6E-4 2.0E-4

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 146000 140000 n (Patient) 222 93 238 77 245 70

[00180] Table 9.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

p Value 6.7E-16 3.3E-11 6.2E-5 1.3E-11 5.2E-9 2.8E-4 3.1E-12 1.5E-9 4.3E-4 nCohort Non- persistent 111 124 212 144 151 225 158 163 232 nCohort Persistent 206 192 103 173 165 90 159 153 83

Cutoff Quartile 2 6030 6020 6080 6030 6020 6080 6030 6020 6080

Sensitivity 86% 84% 89% 87% 86% 89% 87% 86% 88%

Specificity 45% 40% 32% 40% 37% 31 % 37% 36% 30%

Cutoff Quartile 3 14200 14100 14200 14200 14100 14200 14200 14100 14200

Sensitivity 63% 63% 63% 64% 62% 63% 66% 65% 64%

Specificity 74% 70% 56% 66% 64% 55% 66% 64% 55%

Cutoff Quartile 4 35900 35400 36300 35900 35400 36300 35900 35400 36300

Sensitivity 33% 34% 34% 36% 36% 34% 38% 37% 35%

Specificity 90% 89% 79% 88% 87% 79% 87% 86% 78%

OR Quartile 2 5.00 3.53 3.95 4.50 3.64 3.54 4.14 3.47 3.09 p Value 5.9E-9 3.2E-6 9.2E-5 1.3E-7 4.2E-6 5.4E-4 9.8E-7 1.4E-5 0.0021

Lower limit of 95% CI 2.91 2.08 1.98 2.57 2.10 1.73 2.34 1.98 1.51 Upper limit of 95% CI 8.60 6.00 7.86 7.86 6.31 7.24 7.32 6.09 6.34

OR Quartile 3 4.84 4.01 2.19 3.39 2.90 2.12 3.74 3.23 2.14 p Value 1.3E-9 1.8E-8 0.0015 2.5E-7 5.0E-6 0.0034 2.5E-8 5.9E-7 0.0040

Lower limit of 95% CI 2.91 2.47 1.35 2.13 1.84 1.28 2.35 2.04 1.27 Upper limit of 95% CI 8.05 6.50 3.55 5.38 4.58 3.51 5.96 5.12 3.58

OR Quartile 4 4.58 4.02 1.97 4.28 3.65 1.94 4.18 3.51 1.95 p Value 1.4E-5 1.6E-5 0.012 1.6E-6 8.1E-6 0.016 8.0E-7 7.6E-6 0.017

Lower limit of 95% CI 2.30 2.14 1.16 2.36 2.07 1.13 2.37 2.03 1.13 Upper limit of 95% CI 9.10 7.56 3.32 7.74 6.43 3.32 7.38 6.09 3.39

[00181] Table 9.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Min 475 297 419 297 419 297

Max 116000 146000 116000 146000 116000 146000 n (Patient) 124 192 150 166 162 154

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11800 20500 11900 20500 11900 21300

Average 25100 36400 25300 37300 25500 37600

Stdev 31300 36300 31300 36700 31400 36900 p (t-test) 0.0044 0.0034 0.0038

Min 297 399 297 399 297 399

Max 146000 140000 146000 140000 146000 140000 n (Patient) 208 107 220 95 226 89

[00182] Table 9.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 6030 6020 6080 6030 6020 6080 6030 6020 6080

Sensitivity 85% 85% 88% 87% 86% 89% 87% 86% 89%

Specificity 45% 40% 33% 40% 38% 32% 38% 36% 31 %

Cutoff Quartile 3 14200 14100 14200 14200 14100 14200 14200 14100 14200

Sensitivity 63% 63% 64% 63% 62% 64% 65% 65% 64%

Specificity 75% 71 % 57% 66% 64% 57% 66% 65% 56%

Cutoff Quartile 4 35900 35400 36300 35900 35400 36300 35900 35400 36300

Sensitivity 34% 34% 35% 36% 36% 35% 37% 37% 35%

Specificity 92% 90% 80% 88% 88% 80% 87% 87% 79%

OR Quartile 2 4.81 3.74 3.73 4.28 3.68 3.95 4.04 3.49 3.61 p Value 1.2E-8 1.2E-6 6.8E-5 2.1E-7 3.0E-6 9.2E-5 8.8E-7 1.0E-5 2.6E-4

Lower limit of 95% CI 2.80 2.20 1.95 2.47 2.13 1.98 2.32 2.00 1.81 Upper limit of 95% CI 8.25 6.37 7.14 7.41 6.35 7.86 7.05 6.08 7.19

OR Quartile 3 4.89 4.20 2.37 3.24 2.99 2.33 3.55 3.32 2.31 p Value 2.0E-9 8.9E-9 3.6E-4 7.1E-7 2.9E-6 6.5E-4 8.2E-8 3.4E-7 8.6E-4

Lower limit of 95% CI 2.91 2.57 1.47 2.03 1.89 1.43 2.24 2.09 1.41 Upper limit of 95% CI 8.21 6.84 3.81 5.15 4.74 3.78 5.65 5.26 3.78

OR Quartile 4 5.47 4.75 2.13 4.24 3.98 2.11 3.97 3.78 2.03 p Value 6.9E-6 4.4E-6 0.0042 2.7E-6 3.6E-6 0.0053 2.6E-6 3.4E-6 0.0088

Lower limit of 95% CI 2.61 2.44 1.27 2.32 2.22 1.25 2.23 2.16 1.20 Upper limit of 95% CI 11.5 9.25 3.59 7.76 7.14 3.57 7.05 6.64 3.45

[00183] Table 9.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00184] Table 9.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 260000 260000 262000 260000 260000 262000 260000 260000 262000

Sensitivity 85% 86% 86% 86% 86% 89% 88% 88% 90%

Specificity 45% 44% 30% 39% 39% 30% 38% 37% 29%

Cutoff Quartile 3 437000 433000 437000 437000 433000 437000 437000 433000 437000

Sensitivity 61 % 63% 63% 62% 64% 65% 63% 64% 69%

Specificity 72% 72% 56% 65% 66% 55% 63% 63% 56%

Cutoff Quartile 4 707000 704000 708000 707000 704000 708000 707000 704000 708000

Sensitivity 33% 34% 42% 35% 36% 44% 37% 37% 45%

Specificity 92% 90% 82% 88% 87% 81 % 87% 86% 81 %

OR Quartile 2 4.68 4.83 2.60 3.84 4.06 3.55 4.35 4.17 3.60 p Value 1.1E-8 7.0E-9 0.0030 7.5E-7 4.0E-7 8.5E-4 3.4E-7 7.0E-7 0.0013

Lower limit of 95%

CI 2.76 2.83 1.38 2.25 2.36 1.69 2.47 2.37 1.65

Upper limit of 95%

CI 7.95 8.23 4.91 6.55 6.99 7.47 7.64 7.34 7.86

OR Quartile 3 4.10 4.24 2.13 3.11 3.50 2.34 2.90 3.03 2.77 p Value 3.4E-8 6.3E-9 0.0022 1.1E-6 8.4E-8 0.0013 3.4E-6 1.5E-6 2.4E-4

Lower limit of 95%

CI 2.48 2.60 1.31 1.97 2.21 1.39 1.85 1.93 1.61

Upper limit of 95%

CI 6.77 6.90 3.45 4.90 5.53 3.91 4.54 4.75 4.77

OR Quartile 4 5.50 4.63 3.35 3.76 3.78 3.43 4.07 3.63 3.56 p Value 6.0E-6 5.7E-6 6.1E-6 7.4E-6 5.0E-6 7.4E-6 7.8E-7 3.5E-6 5.8E-6

Lower limit of 95%

CI 2.63 2.39 1.98 2.11 2.14 2.00 2.33 2.11 2.06

Upper limit of 95%

CI 11.5 8.98 5.66 6.72 6.69 5.88 7.11 6.26 6.17

[00185] Table 9.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Stdev 223000 268000 233000 271000 235000 273000 p (t-test) 7.8E-14 2.2E-12 2.0E-12

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 118 208 146 180 160 166

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 371000 595000 373000 598000 371000 602000

Average 440000 609000 446000 620000 447000 633000

Stdev 253000 286000 256000 285000 256000 283000 p (t-test) 1.1E-7 1.5E-7 3.7E-8

Min 13600 135000 13600 135000 13600 135000

Max 1120000 1090000 1120000 1090000 1120000 1090000 n (Patient) 216 109 230 95 237 88

[00186] Table 9.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.74 0.75 0.66 0.71 0.72 0.67 0.71 0.72 0.68

SE 0.027 0.027 0.033 0.028 0.028 0.033 0.028 0.028 0.034 p Value 0 0 9.4E-7 1.7E-13 4.4E-16 2.2E-7 1.2E-13 2.2E-15 8.9E-8 nCohort Non- persistent 106 118 211 140 144 223 155 158 229 nCohort Persistent 221 208 114 187 182 102 172 168 96

Cutoff Quartile 2 260000 260000 262000 260000 260000 262000 260000 260000 262000

Sensitivity 85% 86% 86% 86% 87% 89% 88% 88% 90%

Specificity 45% 45% 31 % 40% 40% 31 % 39% 39% 31 %

Cutoff Quartile 3 437000 433000 437000 437000 433000 437000 437000 433000 437000

Sensitivity 61 % 63% 61 % 62% 64% 64% 62% 64% 66%

Specificity 73% 73% 56% 66% 67% 56% 63% 65% 56%

Cutoff Quartile 4 707000 704000 708000 707000 704000 708000 707000 704000 708000

Sensitivity 33% 34% 39% 34% 35% 39% 35% 36% 40%

Specificity 92% 91 % 82% 87% 88% 81 % 86% 87% 81 %

OR Quartile 2 4.55 5.03 2.73 4.13 4.44 3.78 4.67 4.78 3.86 p Value 2.0E-8 3.1E-9 0.0011 2.0E-7 7.6E-8 1.4E-4 6.6E-8 6.0E-8 2.0E-4

Lower limit of 95%

CI 2.68 2.95 1.49 2.42 2.58 1.90 2.67 2.71 1.90

Upper limit of 95%

CI 7.73 8.59 4.99 7.05 7.64 7.52 8.16 8.41 7.88

OR Quartile 3 4.17 4.57 2.02 3.13 3.63 2.24 2.83 3.19 2.46 p Value 3.1E-8 1.6E-9 0.0031 9.9E-7 4.3E-8 0.0010 5.6E-6 5.0E-7 3.6E-4

Lower limit of 95%

CI 2.51 2.79 1.27 1.98 2.29 1.38 1.81 2.03 1.50

Upper limit of 95%

CI 6.91 7.49 3.21 4.95 5.75 3.63 4.43 5.03 4.04

OR Quartile 4 5.32 5.04 2.86 3.53 3.80 2.78 3.51 3.72 2.75 p Value 9.2E-6 3.5E-6 6.3E-5 2.0E-5 6.6E-6 1.2E-4 9.8E-6 3.7E-6 1.5E-4

Lower limit of 95%

CI 2.54 2.54 1.71 1.97 2.13 1.65 2.01 2.13 1.63

Upper limit of 95%

CI 11.1 9.99 4.79 6.30 6.78 4.68 6.11 6.49 4.66

[00187] Table 9.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Persistence Period

Duration (hr) 24 48 72

Non- Non- Non- persistent Persistent persistent Persistent persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 275000 568000 293000 571000 303000 574000

Average 351000 571000 381000 579000 390000 586000

Stdev 225000 269000 236000 272000 238000 273000 p (t-test) 8.4E-13 3.2E-11 3.4E-11

Min 13600 105000 13600 105000 13600 105000

Max 1040000 1120000 1040000 1120000 1040000 1120000 n (Patient) 115 211 140 186 154 172 O only

CI

Upper limit of 95%

CI 6.73 6.96 3.40 5.17 5.50 4.01 4.72 4.77 4.27

OR Quartile 4 4.96 4.79 3.07 3.55 3.56 3.26 3.75 3.48 3.15 p Value 2.1E-5 7.0E-6 2.1E-5 2.5E-5 1.8E-5 7.9E-6 5.8E-6 1.1E-5 1.5E-5

Lower limit of 95%

CI 2.37 2.42 1.83 1.97 1.99 1.94 2.12 2.00 1.87

Upper limit of 95%

CI 10.4 9.50 5.14 6.41 6.35 5.48 6.64 6.07 5.30

[00188] Example 10. Use of Metalloproteinase inhibitor 2 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00189] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 2 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00190] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00191] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00192] Table 10.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 9920 16000 10100 15400 10000 15100

Average 12700 33200 13500 32200 13000 31900

Stdev 10100 58100 10900 57100 10500 56600 p (t-test) 0.0084 0.027 0.034

Min 200 658 200 658 200 658

Max 42400 465000 42400 465000 38600 465000 n (Patient) 57 260 46 271 41 276 sCr only

UO only

Specificity 42% 30% 37% 41 % 31 % 38% 44% 33% 39%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 54% 52% 69% 53% 52% 67% 53% 52% 66%

Specificity 68% 57% 62% 67% 57% 64% 68% 57% 66%

Cutoff Quartile 4 28900 28100 29400 28900 28100 29400 28900 28100 29400

Sensitivity 28% 28% 40% 27% 28% 37% 27% 28% 35%

Specificity 89% 84% 85% 87% 84% 85% 88% 86% 85%

OR Quartile 2 2.71 1.43 8.15 2.47 1.50 6.12 2.76 1.66 5.03 p Value 0.0012 0.23 1.1E-7 0.0066 0.17 4.2E-8 0.0034 0.087 6.8E-8

Lower limit of 95%

CI 1.48 0.803 3.76 1.29 0.842 3.20 1.40 0.929 2.80

Upper limit of 95% CI 4.96 2.53 17.7 4.76 2.66 11.7 5.44 2.97 9.04

OR Quartile 3 2.57 1.42 3.68 2.34 1.43 3.53 2.42 1.45 3.78 p Value 0.0024 0.19 1.2E-7 0.012 0.19 1.2E-7 0.013 0.18 2.3E-8

Lower limit of 95%

CI 1.40 0.842 2.27 1.21 0.844 2.21 1.20 0.847 2.37

Upper limit of 95% CI 4.72 2.38 5.97 4.54 2.41 5.63 4.87 2.47 6.02

OR Quartile 4 3.38 2.07 3.72 2.50 1.98 3.37 2.69 2.34 2.91 p Value 0.0071 0.036 1.3E-6 0.045 0.049 9.8E-6 0.046 0.022 1.2E-4

Lower limit of 95%

CI 1.39 1.05 2.19 1.02 1.00 1.97 1.02 1.13 1.69

Upper limit of 95% CI 8.22 4.07 6.33 6.15 3.90 5.77 7.10 4.83 5.01

[00193] Table 10.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 76000 465000 76000 465000 76000 465000 n (Patient) 106 211 104 213 99 218

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11400 21700 11200 21100 10900 19500

Average 20000 49400 19900 45100 19900 42300

Stdev 37100 73200 38400 68700 39800 65300 p (t-test) 3.4E-6 3.9E-5 2.0E-4

Min 200 2170 200 2170 200 2170

Max 465000 428000 465000 428000 465000 428000 n (Patient) 212 103 194 121 178 137

[00194] Table 10.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 114 127 214 107 124 204 97 119 190 nCohort Non- recovered 203 190 101 210 193 111 220 198 125

Cutoff Quartile 2 7990 7990 8020 7990 7990 8020 7990 7990 8020

Sensitivity 80% 79% 87% 80% 79% 86% 80% 79% 86%

Specificity 34% 31 % 31 % 35% 31 % 31 % 36% 32% 33%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 57% 55% 65% 57% 55% 64% 56% 55% 64%

Specificity 62% 57% 57% 63% 57% 57% 63% 58% 59%

Cutoff Quartile 4 28900 28900 29400 28900 28900 29400 28900 28900 29400

Sensitivity 33% 32% 41 % 31 % 32% 39% 31 % 32% 38%

Specificity 88% 85% 82% 87% 85% 82% 89% 87% 83%

OR Quartile 2 2.12 1.78 3.02 2.11 1.76 2.93 2.26 1.80 3.08 p Value 0.0046 0.028 8.7E-4 0.0050 0.032 6.8E-4 0.0026 0.026 2.1E-4

Lower limit of 95%

CI 1.26 1.06 1.57 1.25 1.05 1.57 1.33 1.07 1.70

Upper limit of 95% CI 3.56 2.98 5.79 3.57 2.94 5.44 3.84 3.01 5.58

OR Quartile 3 2.20 1.67 2.50 2.19 1.63 2.39 2.15 1.69 2.55 p Value 9.9E-4 0.027 2.5E-4 0.0013 0.035 3.5E-4 0.0022 0.025 8.0E-5

Lower limit of 95%

CI 1.38 1.06 1.53 1.36 1.04 1.48 1.32 1.07 1.60

Upper limit of 95% CI 3.52 2.63 4.09 3.53 2.57 3.84 3.51 2.68 4.07

OR Quartile 4 3.44 2.69 3.16 3.04 2.55 2.95 3.57 3.07 2.98 p Value 1.3E-4 7.5E-4 2.0E-5 5.7E-4 0.0014 5.3E-5 2.9E-4 2.7E-4 4.7E-5

Lower limit of 95%

CI 1.83 1.51 1.86 1.62 1.44 1.75 1.79 1.68 1.76

Upper limit of 95% CI 6.47 4.78 5.38 5.73 4.54 4.99 7.12 5.63 5.03

[00195] Table 10.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Median 11800 18200 11800 18100 11600 18100

Average 18000 38600 18100 38300 17800 38200

Stdev 22400 67300 22600 66900 22400 66500 p (t-test) 6.0E-4 7.8E-4 6.9E-4

Min 200 658 200 658 200 658

Max 215000 465000 215000 465000 215000 465000 n (Patient) 140 177 138 179 135 182

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11600 21400 11400 21500 11400 20600

Average 20700 49000 20600 46900 20200 44700

Stdev 37500 74400 38100 71700 38500 68700 p (t-test) 1.0E-5 2.8E-5 6.8E-5

Min 200 2170 200 2170 200 2170

Max 465000 428000 465000 428000 465000 428000 n (Patient) 216 99 207 108 194 121

Upper limit of 95% CI 5.38 4.96 5.63 5.08 4.79 5.32 5.17 5.47 5.06

[00196] Table 10.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.63 0.62 0.68 0.64 0.61 0.68 0.64 0.62 0.68

SE 0.032 0.031 0.033 0.032 0.031 0.033 0.032 0.031 0.033 p Value 3.2E-5 1.8E-4 9.2E-8 1.6E-5 4.8E-4 5.0E-8 1.6E-5 2.4E-4 2.9E-8 nCohort Recovered 188 160 211 185 157 208 185 156 205 nCohort Non- recovered 129 157 104 132 160 107 132 161 110

Cutoff Quartile 2 7990 7990 8020 7990 7990 8020 7990 7990 8020

Sensitivity 83% 82% 85% 83% 81 % 85% 83% 81 % 85%

Specificity 30% 31 % 30% 31 % 31 % 30% 31 % 31 % 30%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 60% 57% 65% 60% 56% 65% 60% 56% 65%

Specificity 56% 56% 57% 57% 55% 58% 57% 56% 58%

Cutoff Quartile 4 28900 28900 29400 28900 28900 29400 28900 28900 29400

Sensitivity 36% 34% 41 % 36% 34% 41 % 36% 34% 41 %

Specificity 82% 84% 83% 82% 83% 83% 82% 84% 83%

OR Quartile 2 2.12 2.01 2.34 2.23 1.97 2.47 2.23 2.00 2.37 p Value 0.0080 0.0092 0.0062 0.0046 0.011 0.0035 0.0046 0.0092 0.0046

Lower limit of 95%

CI 1.22 1.19 1.27 1.28 1.17 1.35 1.28 1.19 1.31

Upper limit of 95% CI 3.68 3.39 4.30 3.87 3.31 4.54 3.87 3.37 4.31

OR Quartile 3 1.91 1.68 2.54 1.96 1.56 2.58 1.96 1.60 2.62 p Value 0.0051 0.022 1.8E-4 0.0037 0.050 1.2E-4 0.0037 0.038 8.6E-5

Lower limit of 95%

CI 1.21 1.08 1.56 1.24 1.00 1.59 1.24 1.03 1.62

Upper limit of 95% CI 3.02 2.62 4.13 3.08 2.43 4.19 3.08 2.49 4.24

OR Quartile 4 2.51 2.70 3.43 2.55 2.57 3.45 2.55 2.72 3.48 p Value 4.9E-4 2.6E-4 5.3E-6 4.1E-4 5.3E-4 4.5E-6 4.1E-4 2.7E-4 3.8E-6

Lower limit of 95%

CI 1.50 1.58 2.02 1.52 1.51 2.03 1.52 1.59 2.05

Upper limit of 95% CI 4.21 4.61 5.82 4.28 4.38 5.86 4.28 4.65 5.91

[00197] Table 10.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only p Value 5.4E-5 1.3E-4 0.0038 5.1E-4 1.1E-4 0.0016 3.6E-4 2.4E-4 0.0024

Lower limit of 95%

CI 2.07 1.73 1.24 1.78 1.76 1.31 1.95 1.69 1.27

Upper limit of 95%

CI 8.19 5.46 3.06 7.94 5.68 3.19 9.92 5.59 3.08

OR Quartile 4 4.75 5.48 1.79 4.99 6.79 1.86 4.39 6.22 1.60 p Value 0.0037 4.4E-4 0.024 0.0088 3.3E-4 0.017 0.016 6.2E-4 0.070

Lower limit of 95%

CI 1.66 2.12 1.08 1.50 2.39 1.12 1.31 2.18 0.962

Upper limit of 95%

CI 13.6 14.1 2.97 16.6 19.3 3.08 14.7 17.7 2.66

[00198] Table 10.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Median 105000 123000 105000 120000 104000 120000

Average 117000 133000 116000 131000 116000 129000

Stdev 42700 45300 42100 45900 42900 44600 p (t-test) 0.0019 0.0029 0.0066

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 217 108 198 127 181 144

[00199] Table 10.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 98600 117000 98600 117000 98000 116000

Average 112000 127000 112000 126000 112000 126000

Stdev 40700 45000 40800 44900 41900 44400 p (t-test) 0.0049 0.0082 0.012

Min 57600 57000 57600 57000 57600 57000

Max 257000 299000 257000 299000 257000 299000 n (Patient) 111 216 104 223 94 233 sCr only

UO only

Specificity 32% 31 % 32% 31 % 31 % 32% 33% 31 % 33%

Cutoff Quartile 3 109000 109000 109000 109000 109000 109000 109000 109000 109000

Sensitivity 58% 59% 66% 57% 59% 65% 57% 59% 64%

Specificity 66% 65% 58% 65% 65% 58% 66% 66% 59%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 29% 31 % 39% 29% 31 % 37% 28% 31 % 34%

Specificity 83% 84% 81 % 83% 85% 81 % 82% 85% 81 %

OR Quartile 2 1.66 1.71 3.60 1.54 1.69 3.16 1.76 1.61 3.07 p Value 0.055 0.039 1.6E-4 0.11 0.044 2.5E-4 0.037 0.066 1.6E-4

Lower limit of 95%

CI 0.990 1.03 1.85 0.912 1.01 1.71 1.03 0.968 1.72

Upper limit of 95%

CI 2.77 2.83 7.01 2.59 2.80 5.85 2.98 2.69 5.49

OR Quartile 3 2.69 2.63 2.63 2.55 2.73 2.58 2.53 2.71 2.53 p Value 4.7E-5 4.2E-5 8.5E-5 1.5E-4 2.4E-5 7.6E-5 2.6E-4 3.4E-5 6.9E-5

Lower limit of 95%

CI 1.67 1.66 1.63 1.57 1.71 1.61 1.54 1.69 1.60

Upper limit of 95%

CI 4.33 4.17 4.27 4.13 4.35 4.12 4.17 4.34 3.99

OR Quartile 4 1.99 2.29 2.74 1.92 2.58 2.52 1.75 2.59 2.18 p Value 0.019 0.0041 1.4E-4 0.028 0.0014 4.1E-4 0.066 0.0016 0.0026

Lower limit of 95%

CI 1.12 1.30 1.63 1.07 1.44 1.51 0.964 1.43 1.31

Upper limit of 95%

CI 3.54 4.02 4.60 3.45 4.61 4.20 3.19 4.69 3.62

[00200] Table 10.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Stdev 39400 45800 38700 45900 38400 46000 p (t-test) 1.2E-4 3.3E-5 5.7E-5

Min 57600 57000 57600 57000 57600 57000

Max 269000 299000 269000 299000 269000 299000 n (Patient) 139 188 137 190 133 194

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 103000 128000 103000 124000 103000 123000

Average 114000 139000 114000 136000 114000 134000

Stdev 39800 47800 40400 47200 41200 46000 p (t-test) 9.4E-7 1.2E-5 7.2E-5

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 220 105 211 114 198 127

Upper limit of 95%

CI 3.66 4.32 5.04 3.74 4.98 4.47 4.03 5.11 3.71

[00201] Table 10.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.63 0.62 0.66 0.63 0.64 0.67 0.63 0.63 0.67

SE 0.031 0.031 0.033 0.031 0.030 0.032 0.031 0.031 0.032 p Value 5.0E-5 8.3E-5 4.7E-7 1.9E-5 8.0E-6 2.1E-7 3.2E-5 3.2E-5 2.0E-7 nCohort Recovered 187 159 215 184 156 212 183 154 209 nCohort Non- recovered 140 168 110 143 171 113 144 173 116

Cutoff Quartile 2 90200 90200 90200 90200 90200 90200 90200 90200 90200

Sensitivity 83% 80% 86% 83% 81 % 87% 83% 80% 87%

Specificity 31 % 31 % 31 % 32% 31 % 31 % 31 % 31 % 32%

Cutoff Quartile 3 109000 109000 109000 109000 109000 109000 109000 109000 109000

Sensitivity 61 % 60% 65% 61 % 60% 65% 60% 60% 66%

Specificity 58% 60% 58% 58% 61 % 58% 58% 60% 58%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 33% 32% 39% 34% 33% 39% 33% 33% 38%

Specificity 81 % 82% 82% 82% 84% 82% 81 % 84% 82%

OR Quartile 2 2.17 1.82 2.81 2.28 1.92 2.95 2.15 1.73 3.11 p Value 0.0047 0.021 0.0010 0.0026 0.012 5.7E-4 0.0048 0.033 3.1E-4

Lower limit of 95%

CI 1.27 1.10 1.51 1.33 1.15 1.59 1.26 1.04 1.68

Upper limit of 95%

CI 3.72 3.03 5.20 3.91 3.18 5.47 3.67 2.87 5.75

OR Quartile 3 2.11 2.18 2.58 2.16 2.36 2.62 2.10 2.24 2.66 p Value 0.0010 5.4E-4 9.8E-5 7.1E-4 1.5E-4 6.7E-5 0.0011 3.5E-4 4.6E-5

Lower limit of 95%

CI 1.35 1.40 1.60 1.38 1.51 1.63 1.35 1.44 1.66

Upper limit of 95%

CI 3.30 3.40 4.16 3.37 3.68 4.21 3.28 3.49 4.27

OR Quartile 4 2.05 2.22 2.90 2.23 2.62 2.92 2.19 2.54 2.75 p Value 0.0054 0.0028 5.5E-5 0.0020 4.0E-4 4.7E-5 0.0025 6.2E-4 1.1E-4

Lower limit of 95%

CI 1.24 1.32 1.73 1.34 1.54 1.74 1.32 1.49 1.64

Upper limit of 95%

CI 3.41 3.73 4.86 3.71 4.46 4.89 3.64 4.32 4.60

[00202] Example 11. Use of Metalloproteinase inhibitor 2 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00203] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 2 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00204] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00205] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00206] Table 11.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 11400 25100 11000 22700 11000 21700

Average 20500 51500 19900 48500 19900 46000

Stdev 38100 74800 38900 70600 39600 68200 p (t-test) 1.9E-6 5.3E-6 2.4E-5

Min 200 2170 200 2170 200 2170

Max 465000 428000 465000 428000 465000 428000 n (Patient) 222 93 208 107 198 117

[00207] Table 11.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 11300 19300 11100 19300 11000 19300

Average 15500 40000 15400 39800 14900 39500

Stdev 13800 67800 13800 67400 13300 66600 p (t-test) 4.4E-5 4.7E-5 4.7E-5

Min 200 658 200 658 200 658

Max 76000 465000 76000 465000 76000 465000 n (Patient) 136 181 134 183 129 188 sCr only

UO only

Specificity 33% 30% 31 % 34% 30% 31 % 34% 30% 32%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 59% 56% 69% 59% 56% 68% 60% 57% 68%

Specificity 61 % 56% 57% 62% 57% 58% 64% 58% 59%

Cutoff Quartile 4 28900 28900 29400 28900 28900 29400 28900 28900 29400

Sensitivity 34% 35% 44% 34% 34% 44% 35% 34% 43%

Specificity 87% 85% 82% 87% 85% 83% 88% 85% 84%

OR Quartile 2 2.14 1.59 3.47 2.22 1.62 3.88 2.26 1.68 3.42 p Value 0.0039 0.075 6.6E-4 0.0026 0.065 2.0E-4 0.0020 0.048 2.0E-4

Lower limit of 95%

CI 1.28 0.954 1.70 1.32 0.970 1.90 1.35 1.00 1.79

Upper limit of 95% CI 3.58 2.66 7.11 3.72 2.70 7.94 3.80 2.80 6.55

OR Quartile 3 2.21 1.64 2.90 2.34 1.68 2.96 2.57 1.78 2.99 p Value 6.1E-4 0.029 5.9E-5 2.6E-4 0.022 2.9E-5 6.2E-5 0.012 1.1E-5

Lower limit of 95%

CI 1.41 1.05 1.72 1.48 1.08 1.78 1.62 1.14 1.84

Upper limit of 95% CI 3.49 2.56 4.87 3.70 2.63 4.92 4.08 2.78 4.88

OR Quartile 4 3.42 3.04 3.63 3.61 2.99 3.92 4.02 2.89 3.91 p Value 3.7E-5 8.0E-5 3.0E-6 2.2E-5 1.0E-4 6.3E-7 9.9E-6 1.7E-4 5.2E-7

Lower limit of 95%

CI 1.91 1.75 2.11 2.00 1.72 2.29 2.17 1.66 2.30

Upper limit of 95% CI 6.12 5.29 6.23 6.54 5.20 6.71 7.44 5.03 6.67

[00208] Table 11.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 215000 465000 215000 465000 215000 465000 n (Patient) 161 156 160 157 159 158

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11900 21500 11900 21500 11500 21500

Average 21500 49500 21600 49100 21200 46900

Stdev 38600 75700 38600 75400 39400 71800 p (t-test) 2.1E-5 2.7E-5 5.4E-5

Min 200 2170 200 2170 200 2170

Max 465000 428000 465000 428000 465000 428000 n (Patient) 224 91 223 92 212 103

[00209] Table 11.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 168 172 223 165 171 222 161 170 214 nCohort Non- recovered 149 145 92 152 146 93 156 147 101

Cutoff Quartile 2 7990 7990 8020 7990 7990 8020 7990 7990 8020

Sensitivity 83% 81 % 85% 83% 82% 85% 83% 82% 86%

Specificity 32% 30% 29% 32% 30% 29% 33% 31 % 30%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 60% 58% 65% 60% 58% 66% 61 % 59% 66%

Specificity 58% 56% 56% 59% 57% 56% 60% 57% 57%

Cutoff Quartile 4 28900 28900 29400 28900 28900 29400 28900 28900 29400

Sensitivity 36% 37% 41 % 36% 36% 42% 36% 36% 41 %

Specificity 84% 84% 82% 84% 84% 82% 85% 84% 82%

OR Quartile 2 2.18 1.89 2.29 2.29 1.93 2.34 2.45 1.96 2.71 p Value 0.0042 0.018 0.011 0.0023 0.015 0.0091 9.9E-4 0.013 0.0021

Lower limit of 95%

CI 1.28 1.11 1.21 1.34 1.13 1.23 1.44 1.15 1.44

Upper limit of 95% CI 3.72 3.22 4.34 3.91 3.27 4.42 4.19 3.33 5.12

OR Quartile 3 2.08 1.78 2.39 2.13 1.83 2.46 2.36 1.87 2.66 p Value 0.0014 0.011 7.0E-4 9.7E-4 0.0082 4.7E-4 1.9E-4 0.0059 1.0E-4

Lower limit of 95%

CI 1.33 1.14 1.44 1.36 1.17 1.48 1.50 1.20 1.63

Upper limit of 95% CI 3.25 2.78 3.96 3.33 2.86 4.06 3.70 2.93 4.36

OR Quartile 4 2.88 3.09 3.12 2.95 3.04 3.29 3.20 2.99 3.16 p Value 9.3E-5 3.2E-5 3.1E-5 7.4E-5 4.2E-5 1.3E-5 2.8E-5 5.5E-5 2.0E-5

Lower limit of 95%

CI 1.70 1.82 1.83 1.73 1.79 1.92 1.86 1.76 1.86

Upper limit of 95% CI 4.90 5.27 5.34 5.03 5.17 5.61 5.50 5.08 5.38

[00210] Table 11.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Median 11900 19000 11900 19000 11900 19000

Average 19100 44000 19100 44000 19100 44000

Stdev 24400 75300 24400 75300 24400 75300 p (t-test) 3.4E-5 3.4E-5 3.4E-5

Min 200 658 200 658 200 658

Max 215000 465000 215000 465000 215000 465000 n (Patient) 185 132 185 132 185 132

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 11800 21500 11700 21500 11700 21500

Average 21700 47200 21600 47200 21600 46600

Stdev 39100 73500 39200 73100 39400 72500 p (t-test) 7.4E-5 6.6E-5 9.6E-5

Min 200 2170 200 2170 200 2170

Max 465000 428000 465000 428000 465000 428000 n (Patient) 217 98 216 99 214 101

Upper limit of 95% CI 4.91 5.75 5.36 5.07 5.75 5.63 4.96 5.75 5.38

[00211] Table 11.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.66 0.68 0.62 0.66 0.68 0.63 0.66 0.68 0.63

SE 0.031 0.030 0.035 0.031 0.030 0.033 0.031 0.030 0.032 p Value 7.3E-8 1.9E-9 9.0E-4 2.6E-7 1.6E-9 4.9E-5 7.9E-7 9.3E-10 5.1E-5 nCohort Recovered 105 121 226 102 119 213 98 117 202 nCohort Non- recovered 222 205 99 225 207 112 229 209 123

Cutoff Quartile 2 90200 90200 90200 90200 90200 90200 90200 90200 90200

Sensitivity 80% 80% 86% 80% 80% 87% 79% 80% 86%

Specificity 35% 34% 30% 35% 34% 31 % 35% 34% 32%

Cutoff Quartile 3 109000 109000 109000 109000 109000 109000 109000 109000 109000

Sensitivity 59% 60% 61 % 58% 60% 62% 58% 60% 60%

Specificity 68% 67% 54% 68% 67% 56% 67% 68% 56%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 32% 33% 35% 31 % 33% 37% 31 % 33% 34%

Specificity 89% 88% 79% 88% 88% 81 % 88% 89% 80%

OR Quartile 2 2.14 2.05 2.56 2.12 2.13 2.90 2.00 2.07 2.89 p Value 0.0039 0.0057 0.0036 0.0045 0.0037 7.0E-4 0.0093 0.0053 4.4E-4

Lower limit of 95%

CI 1.28 1.23 1.36 1.26 1.28 1.57 1.19 1.24 1.60

Upper limit of 95%

CI 3.59 3.41 4.82 3.57 3.54 5.38 3.38 3.44 5.23

OR Quartile 3 2.95 3.04 1.84 2.91 3.06 2.03 2.81 3.09 1.92 p Value 1.4E-5 3.7E-6 0.013 2.1E-5 3.5E-6 0.0029 4.7E-5 3.3E-6 0.0051

Lower limit of 95%

CI 1.81 1.90 1.14 1.78 1.91 1.27 1.71 1.92 1.22

Upper limit of 95%

CI 4.81 4.86 2.97 4.77 4.92 3.24 4.61 4.98 3.02

OR Quartile 4 3.57 3.79 2.08 3.39 3.67 2.42 3.16 3.94 2.10 p Value 1.7E-4 3.2E-5 0.0059 3.2E-4 5.0E-5 7.3E-4 7.2E-4 3.0E-5 0.0042

Lower limit of 95%

CI 1.84 2.02 1.24 1.74 1.96 1.45 1.62 2.07 1.26

Upper limit of 95%

CI 6.94 7.11 3.51 6.59 6.88 4.05 6.14 7.51 3.49

[00212] Table 11.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 299000 281000 299000 281000 299000 281000 n (Patient) 134 193 133 194 128 199 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 98600 121000 98600 121000 98600 121000

Average 110000 131000 110000 131000 110000 131000

Stdev 39300 45700 39400 45600 39500 45400 p (t-test) 1.9E-5 2.1E-5 2.0E-5

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 146 181 145 182 143 184

UO only

Upper limit of 95%

CI 4.06 3.77 6.53 4.14 3.84 7.26 3.69 3.71 5.66

OR Quartile 3 3.01 2.93 2.34 3.10 3.02 2.56 2.90 3.03 2.45 p Value 2.6E-6 3.1E-6 8.1E-4 1.5E-6 1.8E-6 1.7E-4 6.4E-6 1.8E-6 1.8E-4

Lower limit of 95%

CI 1.90 1.87 1.42 1.96 1.92 1.57 1.83 1.92 1.53

Upper limit of 95%

CI 4.76 4.61 3.84 4.92 4.75 4.18 4.60 4.77 3.92

OR Quartile 4 2.30 2.59 2.50 2.26 2.54 2.71 2.08 2.66 2.45 p Value 0.0030 6.0E-4 6.6E-4 0.0036 7.5E-4 1.8E-4 0.0091 4.7E-4 6.3E-4

Lower limit of 95%

CI 1.33 1.50 1.47 1.30 1.48 1.61 1.20 1.54 1.46

Upper limit of 95%

CI 3.98 4.45 4.23 3.91 4.38 4.56 3.60 4.61 4.08

[00213] Table 11.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 104000 124000 104000 125000 103000 127000

Average 115000 138000 115000 139000 114000 138000

Stdev 41100 47300 40900 47000 40200 47200 p (t-test) 2.2E-5 7.1E-6 2.8E-6

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 230 95 228 97 215 110

[00214] Table 11.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 83% 83% 89% 83% 83% 89% 82% 83% 87% Specificity 33% 33% 31 % 33% 33% 31 % 33% 33% 31 %

Cutoff Quartile 3 109000 109000 109000 109000 109000 109000 109000 109000 109000

Sensitivity 63% 63% 69% 63% 64% 68% 62% 64% 68%

Specificity 62% 62% 58% 63% 62% 58% 63% 63% 59%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 32% 33% 42% 31 % 33% 42% 31 % 33% 40%

Specificity 81 % 82% 82% 81 % 82% 82% 81 % 83% 82%

OR Quartile 2 2.29 2.34 3.59 2.41 2.38 3.65 2.24 2.42 3.00 p Value 0.0018 0.0015 2.7E-4 9.6E-4 0.0012 2.2E-4 0.0022 9.7E-4 6.5E-4

Lower limit of 95%

CI 1.36 1.39 1.81 1.43 1.41 1.84 1.34 1.43 1.60

Upper limit of 95%

CI 3.86 3.96 7.14 4.05 4.02 7.26 3.74 4.09 5.64

OR Quartile 3 2.75 2.83 3.02 2.82 2.90 2.91 2.82 2.98 2.94 p Value 9.3E-6 5.6E-6 1.4E-5 5.7E-6 3.3E-6 2.5E-5 5.7E-6 2.0E-6 1.3E-5

Lower limit of 95%

CI 1.76 1.81 1.83 1.80 1.85 1.77 1.80 1.90 1.81

Upper limit of 95%

CI 4.31 4.43 4.99 4.42 4.55 4.78 4.42 4.67 4.78

OR Quartile 4 2.02 2.27 3.43 1.92 2.24 3.35 1.93 2.36 3.02 p Value 0.0072 0.0018 4.3E-6 0.012 0.0022 6.1E-6 0.012 0.0012 2.9E-5

Lower limit of 95%

CI 1.21 1.36 2.03 1.15 1.34 1.98 1.15 1.40 1.80

Upper limit of 95%

CI 3.37 3.81 5.79 3.21 3.74 5.66 3.22 3.96 5.07

[00215] Table 11.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Average 114000 131000 114000 131000 114000 131000

Stdev 41500 45600 41500 45600 41500 45600 p (t-test) 4.5E-4 4.5E-4 4.5E-4

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 184 143 184 143 184 143

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 103000 125000 103000 125000 102000 126000

Average 114000 139000 114000 139000 114000 139000

Stdev 39400 48400 39500 48400 39600 47900 p (t-test) 6.0E-7 8.8E-7 6.3E-7

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 220 105 219 106 217 108

[00216] Example 12. Use of Metalloproteinase inhibitor 2 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00217] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 2 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00218] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00219] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00220] Table 12.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Stdev 21800 77800 22100 82600 35200 78100 p (t-test) 5.7E-8 3.8E-9 5.5E-7

Min 200 2940 200 2940 200 3000

Max 183000 428000 183000 428000 396000 428000 n (Patient) 205 112 221 96 236 81 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11800 21800 11800 25200 12000 24300

Average 19900 46400 20500 49500 23200 45800

Stdev 35400 70300 35500 73900 44300 65800 p (t-test) 1.1E-5 3.7E-6 5.9E-4

Min 200 2940 200 2940 200 2940

Max 428000 396000 428000 396000 428000 371000 n (Patient) 208 108 223 93 235 81

UO only

Upper limit of 95% CI 4.40 3.73 9.81 6.85 5.67 8.89 8.58 5.19 8.60

OR Quartile 3 2.75 2.37 5.66 3.03 2.83 5.02 3.12 2.52 5.90 p Value 3.6E-5 4.3E-4 6.4E-5 1.9E-5 6.3E-5 2.2E-4 3.9E-5 6.2E-4 1.3E-4

Lower limit of 95% CI 1.70 1.46 2.42 1.82 1.70 2.13 1.81 1.48 2.38 Upper limit of 95% CI 4.44 3.82 13.2 5.03 4.71 11.8 5.36 4.28 14.6

OR Quartile 4 5.07 4.24 7.78 5.46 4.82 7.49 5.18 3.86 8.15 p Value 4.0E-9 1.2E-7 1.9E-8 1.0E-9 1.4E-8 8.5E-8 6.3E-9 1.5E-6 4.8E-8

Lower limit of 95% CI 2.95 2.48 3.80 3.17 2.80 3.58 2.97 2.23 3.84 Upper limit of 95% CI 8.72 7.24 15.9 9.42 8.31 15.6 9.02 6.70 17.3

[00221] Table 12.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Max 396000 428000 396000 428000 396000 428000 n (Patient) 256 59 260 55 261 54

[00222] Table 12.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 183000 428000 183000 428000 396000 428000 n (Patient) 175 142 190 127 205 112 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11800 19600 11600 20600 11800 20900

Average 19700 41600 19900 43600 23100 40400

Stdev 35700 65400 36300 67100 46100 59200 p (t-test) 1.6E-4 6.2E-5 0.0044

Min 200 658 200 2170 200 2170

Max 428000 396000 428000 396000 428000 371000 n (Patient) 182 134 195 121 208 108

UO only

p Value 6.9E-5 0.0032 3.6E-4 1.3E-5 3.8E-4 0.0027 3.6E-5 1.0E-3 0.0027

Lower limit of 95%

CI 1.60 1.26 1.60 1.77 1.46 1.36 1.70 1.38 1.36

Upper limit of 95% CI 3.97 3.11 5.00 4.51 3.71 4.32 4.44 3.59 4.32

OR Quartile 4 4.10 3.38 3.99 4.67 3.74 3.97 4.05 3.16 3.97 p Value 4.3E-7 7.4E-6 2.1E-6 2.7E-8 1.2E-6 3.5E-6 2.4E-7 1.9E-5 3.5E-6

Lower limit of 95%

CI 2.37 1.98 2.25 2.71 2.20 2.22 2.38 1.87 2.22

Upper limit of 95% CI 7.08 5.76 7.05 8.03 6.37 7.11 6.90 5.35 7.11

[00223] Table 12.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Max 396000 428000 396000 428000 396000 428000 n (Patient) 239 76 245 70 245 70

[00224] Table 12.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 183000 428000 183000 428000 215000 428000 n (Patient) 169 148 184 133 197 120 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 11600 19200 11500 20000 11600 20400

Average 19800 40600 19300 43300 20700 43300

Stdev 36300 64200 35200 66400 37400 67300 p (t-test) 3.4E-4 3.9E-5 1.5E-4

Min 200 658 200 2170 200 2170

Max 428000 396000 428000 396000 428000 396000 n (Patient) 176 140 188 128 200 116

UO only

p Value 7.3E-5 0.0034 1.3E-5 5.2E-6 2.8E-4 5.3E-5 1.7E-5 5.2E-4 5.3E-5

Lower limit of 95%

CI 1.59 1.25 1.95 1.85 1.48 1.79 1.76 1.44 1.79

Upper limit of 95% CI 3.93 3.08 5.85 4.68 3.73 5.39 4.52 3.67 5.39

OR Quartile 4 3.98 3.26 4.16 4.48 3.78 3.87 3.98 3.33 3.87 p Value 8.9E-7 1.5E-5 4.0E-7 6.7E-8 1.1E-6 1.7E-6 3.6E-7 7.9E-6 1.7E-6

Lower limit of 95%

CI 2.29 1.91 2.40 2.60 2.22 2.22 2.34 1.96 2.22

Upper limit of 95% CI 6.89 5.56 7.22 7.73 6.46 6.73 6.77 5.63 6.73

[00225] Table 12.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Max 299000 281000 299000 281000 299000 281000 n (Patient) 281 44 283 42 285 40

[00226] Table 12.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Stdev 41800 45200 43100 44400 41900 46200 p (t-test) 5.6E-4 0.0070 0.0013

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 187 140 204 123 219 108 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 103000 123000 104000 120000 105000 120000

Average 115000 132000 117000 130000 117000 131000

Stdev 41500 45800 43100 44800 42100 46400 p (t-test) 4.2E-4 0.013 0.0051

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 192 134 205 121 217 109

UO only

[00227] Table 12.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort Cohort Cohort Cohort Cohort Cohort

Median 104000 133000 104000 133000 104000 133000

Average 115000 145000 115000 146000 115000 146000

Stdev 40400 48000 40200 48400 40200 48700 p (t-test) 1.6E-7 4.8E-8 7.4E-8

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 249 76 253 72 254 71

[00228] Table 12.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 99600 120000 103000 120000 103000 123000

Average 113000 132000 115000 131000 114000 134000

Stdev 40000 46100 41400 45700 40300 47100 p (t-test) 4.6E-5 5.8E-4 1.0E-4

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 173 154 188 139 200 127 sCr only

UO only

Specificity 61 % 61 % 58% 59% 58% 58% 57% 57% 58%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 34% 35% 47% 33% 33% 49% 36% 36% 48%

Specificity 83% 83% 82% 81 % 81 % 82% 82% 82% 82%

OR Quartile 2 2.54 2.28 4.97 2.14 2.05 4.51 2.05 1.96 4.43 p Value 6.2E-4 0.0024 1.3E-4 0.0056 0.0089 3.3E-4 0.011 0.017 3.9E-4

Lower limit of 95%

CI 1.49 1.34 2.19 1.25 1.20 1.98 1.18 1.13 1.94

Upper limit of 95%

CI 4.32 3.89 11.3 3.66 3.51 10.3 3.55 3.40 10.1

OR Quartile 3 2.69 2.68 3.71 2.29 2.15 4.14 2.23 2.09 4.03 p Value 1.5E-5 1.7E-5 2.2E-6 3.0E-4 8.2E-4 8.9E-7 5.7E-4 0.0015 1.4E-6

Lower limit of 95%

CI 1.72 1.71 2.16 1.46 1.37 2.35 1.41 1.33 2.29

Upper limit of 95%

CI 4.21 4.21 6.38 3.58 3.37 7.28 3.51 3.30 7.10

OR Quartile 4 2.43 2.54 4.19 2.09 2.04 4.47 2.59 2.54 4.25 p Value 7.5E-4 4.0E-4 2.1E-7 0.0044 0.0056 8.7E-8 2.7E-4 3.5E-4 2.3E-7

Lower limit of 95%

CI 1.45 1.52 2.44 1.26 1.23 2.58 1.55 1.52 2.46

Upper limit of 95%

CI 4.07 4.24 7.20 3.47 3.39 7.73 4.31 4.23 7.37

[00229] Table 12.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00230] Example 13. Use of Metalloproteinase inhibitor 2 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F [00231] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 2 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00232] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00233] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00234] Table 13.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 2.99 2.15 7.64 3.41 2.88 7.78 4.00 3.30 8.16

OR Quartile 4 3.39 2.56 5.15 3.28 3.19 5.27 3.78 3.26 5.96 p Value 1.1E-4 0.0012 1.4E-8 2.2E-5 2.9E-5 6.3E-8 1.3E-6 1.2E-5 3.1E-8

Lower limit of 95%

CI 1.83 1.45 2.92 1.90 1.85 2.89 2.21 1.92 3.17

Upper limit of 95% CI 6.28 4.51 9.07 5.68 5.51 9.62 6.48 5.54 11.2

[00235] Table 13.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.59 0.73 0.65 0.62 0.72 0.67 0.64 0.71

SE 0.031 0.032 0.033 0.031 0.031 0.036 0.030 0.031 0.038 p Value 1.6E-6 0.0039 1.9E-12 7.9E-7 1.1E-4 1.3E-9 1.7E-8 9.4E-6 1.8E-8 nCohort Non- persistent 112 126 222 147 154 238 165 169 245 nCohort Persistent 205 190 93 170 162 77 152 147 70

Cutoff Quartile 2 7990 7980 8020 7990 7980 8020 7990 7980 8020

Sensitivity 81 % 79% 89% 81 % 80% 87% 84% 82% 86%

Specificity 36% 31 % 31 % 32% 30% 29% 33% 31 % 28%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 57% 54% 71 % 60% 57% 70% 62% 59% 70%

Specificity 62% 56% 59% 61 % 58% 56% 61 % 58% 56%

Cutoff Quartile 4 28900 28100 29400 28900 28100 29400 28900 28100 29400

Sensitivity 33% 32% 46% 35% 35% 47% 38% 37% 49%

Specificity 88% 85% 84% 86% 85% 82% 86% 85% 82%

OR Quartile 2 2.36 1.68 3.74 2.03 1.66 2.74 2.67 2.13 2.35 p Value 0.0012 0.048 3.0E-4 0.0075 0.052 0.0062 4.0E-4 0.0056 0.021

Lower limit of 95%

CI 1.40 1.01 1.83 1.21 0.995 1.33 1.55 1.25 1.14

Upper limit of 95% CI 3.98 2.81 7.65 3.40 2.79 5.63 4.59 3.63 4.86

OR Quartile 3 2.22 1.45 3.45 2.37 1.85 3.03 2.63 2.00 2.91 p Value 9.5E-4 0.11 3.2E-6 1.8E-4 0.0071 8.3E-5 3.0E-5 0.0024 2.4E-4

Lower limit of 95%

CI 1.38 0.921 2.05 1.51 1.18 1.74 1.67 1.28 1.65

Upper limit of 95% CI 3.55 2.28 5.82 3.72 2.88 5.25 4.14 3.14 5.15

OR Quartile 4 3.70 2.60 4.44 3.19 3.01 3.98 3.70 3.34 4.20 p Value 7.6E-5 0.0012 6.8E-8 4.9E-5 8.3E-5 1.1E-6 3.0E-6 1.2E-5 7.9E-7

Lower limit of 95% CI 1.93 1.46 2.58 1.82 1.74 2.28 2.14 1.95 2.38

Upper limit of 95% CI 7.07 4.62 7.64 5.58 5.21 6.95 6.42 5.75 7.42

[00236] Table 13.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 11500 16000 11600 18600 11600 18800

Average 17200 36600 16900 40100 18100 40600

Stdev 15000 63900 14800 68100 21500 68900 p (t-test) 9.8E-4 5.3E-5 8.5E-5

Min 200 658 200 658 200 658

Max 76000 428000 76000 428000 215000 428000 n (Patient) 124 192 151 165 163 153

UO only

p Value 9.5E-5 0.0017 1.0E-6 3.8E-5 1.7E-4 4.4E-6 7.4E-6 6.3E-5 3.7E-6

Lower limit of 95%

CI 1.90 1.41 2.21 1.87 1.65 2.06 2.04 1.75 2.11

Upper limit of 95% CI 6.95 4.47 6.43 5.80 4.95 6.06 6.18 5.17 6.29

[00237] Table 13.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.60 0.69 0.65 0.62 0.69 0.65 0.63 0.68

SE 0.031 0.032 0.033 0.031 0.031 0.034 0.031 0.031 0.035 p Value 3.2E-6 0.0030 3.4E-9 9.4E-7 1.8E-4 4.8E-8 3.8E-7 6.6E-5 4.3E-7 nCohort Non- persistent 110 124 208 143 150 220 156 162 226 nCohort Persistent 207 192 107 174 166 95 161 154 89

Cutoff Quartile 2 7990 7980 8020 7990 7980 8020 7990 7980 8020

Sensitivity 81 % 79% 86% 81 % 80% 85% 83% 81 % 84%

Specificity 35% 31 % 31 % 32% 30% 30% 33% 31 % 29%

Cutoff Quartile 3 13500 13500 13500 13500 13500 13500 13500 13500 13500

Sensitivity 57% 54% 66% 60% 57% 66% 61 % 58% 66%

Specificity 63% 56% 58% 62% 58% 57% 61 % 58% 56%

Cutoff Quartile 4 28900 28100 29400 28900 28100 29400 28900 28100 29400

Sensitivity 32% 31 % 41 % 34% 34% 42% 36% 35% 43%

Specificity 88% 85% 83% 86% 85% 82% 86% 85% 82%

OR Quartile 2 2.29 1.74 2.73 2.03 1.66 2.43 2.31 1.92 2.16 p Value 0.0018 0.034 0.0015 0.0074 0.052 0.0064 0.0019 0.014 0.018

Lower limit of 95%

CI 1.36 1.04 1.47 1.21 0.995 1.28 1.36 1.14 1.14

Upper limit of 95% CI 3.86 2.92 5.07 3.40 2.78 4.59 3.91 3.25 4.10

OR Quartile 3 2.23 1.53 2.74 2.38 1.85 2.59 2.42 1.94 2.52 p Value 9.1E-4 0.066 4.8E-5 1.8E-4 0.0071 2.0E-4 1.2E-4 0.0036 4.0E-4

Lower limit of 95%

CI 1.39 0.972 1.69 1.51 1.18 1.57 1.54 1.24 1.51

Upper limit of 95% CI 3.59 2.41 4.46 3.74 2.89 4.28 3.80 3.04 4.21

OR Quartile 4 3.57 2.51 3.45 3.24 2.81 3.38 3.43 2.96 3.36 p Value 1.2E-4 0.0017 4.5E-6 4.8E-5 2.2E-4 8.1E-6 1.3E-5 8.1E-5 1.1E-5

Lower limit of 95%

CI 1.87 1.41 2.03 1.84 1.62 1.98 1.97 1.73 1.96

Upper limit of 95% CI 6.83 4.47 5.86 5.70 4.86 5.76 5.97 5.08 5.77

[00238] Table 13.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 3.44 2.36 5.19 3.88 2.90 4.91 3.81 3.04 4.95

OR Quartile 4 3.72 2.60 3.78 3.53 3.08 3.50 3.64 3.22 3.39 p Value 1.1E-4 0.0013 9.3E-7 2.2E-5 8.3E-5 3.7E-6 7.8E-6 3.0E-5 7.1E-6

Lower limit of 95%

CI 1.91 1.45 2.22 1.97 1.76 2.06 2.07 1.86 1.99

Upper limit of 95% CI 7.25 4.68 6.44 6.33 5.40 5.96 6.43 5.58 5.77

[00239] Table 13.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.67 0.62 0.64 0.66 0.66 0.62 0.62 0.69

SE 0.031 0.030 0.037 0.030 0.030 0.040 0.031 0.031 0.042 p Value 4.4E-7 4.0E-9 0.0013 2.8E-6 6.2E-8 9.2E-5 1.6E-4 2.2E-4 5.8E-6 nCohort Non- persistent 114 124 245 155 157 261 179 182 269 nCohort Persistent 213 202 80 172 169 64 148 144 56

Cutoff Quartile 2 90400 90300 90600 90400 90300 90600 90400 90300 90600

Sensitivity 80% 80% 85% 83% 83% 89% 83% 82% 91 %

Specificity 34% 34% 28% 34% 34% 28% 32% 31 % 28%

Cutoff Quartile 3 109000 109000 109000 109000 109000 109000 109000 109000 109000

Sensitivity 58% 60% 62% 61 % 63% 66% 63% 63% 71 %

Specificity 65% 66% 54% 62% 64% 54% 60% 60% 54%

Cutoff Quartile 4 137000 137000 137000 137000 137000 137000 137000 137000 137000

Sensitivity 31 % 33% 38% 31 % 33% 41 % 31 % 32% 45%

Specificity 87% 88% 79% 81 % 83% 79% 80% 80% 79%

OR Quartile 2 2.06 2.07 2.22 2.56 2.46 3.22 2.30 2.02 4.02 p Value 0.0058 0.0049 0.020 3.8E-4 6.7E-4 0.0057 0.0022 0.0093 0.0044

Lower limit of 95%

CI 1.23 1.25 1.13 1.52 1.46 1.41 1.35 1.19 1.54

Upper limit of 95%

CI 3.43 3.45 4.36 4.31 4.13 7.39 3.92 3.42 10.4

OR Quartile 3 2.58 2.92 1.95 2.55 2.95 2.21 2.57 2.62 2.97 p Value 8.2E-5 6.8E-6 0.012 3.9E-5 2.5E-6 0.0065 3.6E-5 2.7E-5 6.8E-4

Lower limit of 95%

CI 1.61 1.83 1.16 1.63 1.88 1.25 1.64 1.67 1.58

Upper limit of 95%

CI 4.13 4.65 3.27 3.98 4.63 3.91 4.03 4.12 5.56

OR Quartile 4 3.03 3.61 2.23 1.94 2.50 2.50 1.79 1.90 3.00 p Value 4.2E-4 4.2E-5 0.0041 0.012 7.0E-4 0.0019 0.024 0.013 3.5E-4

Lower limit of 95%

CI 1.64 1.95 1.29 1.15 1.47 1.40 1.08 1.15 1.64

Upper limit of 95%

CI 5.60 6.66 3.85 3.25 4.24 4.47 2.97 3.16 5.48

[00240] Table 13.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

n (Patient) 108 219 144 183 164 163 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 97500 120000 98700 121000 100000 121000

Average 105000 132000 111000 131000 115000 129000

Stdev 31200 47500 41100 44700 43800 43300 p (t-test) 8.4E-8 6.2E-5 0.0032

Min 57600 57000 57600 57000 57600 57000

Max 252000 299000 299000 281000 299000 281000 n (Patient) 120 206 149 177 166 160

UO only

p Value 3.3E-6 7.1E-7 9.9E-4 8.4E-6 2.4E-6 5.4E-4 9.3E-6 1.2E-5 9.0E-5

Lower limit of 95%

CI 1.95 2.07 1.39 1.78 1.89 1.49 1.76 1.74 1.73

Upper limit of 95%

CI 5.18 5.36 3.68 4.38 4.68 4.21 4.30 4.26 5.18

OR Quartile 4 3.76 4.15 2.52 2.15 2.54 2.73 1.95 2.04 3.05 p Value 9.3E-5 1.5E-5 5.1E-4 0.0048 6.9E-4 2.5E-4 0.010 0.0065 7.0E-5

Lower limit of 95%

CI 1.93 2.18 1.50 1.26 1.48 1.60 1.17 1.22 1.76

Upper limit of 95%

CI 7.30 7.90 4.25 3.67 4.35 4.68 3.26 3.40 5.27

[00241] Table 13.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

p (t-test) 1.6E-4 1.6E-5 2.7E-6

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 216 109 230 95 237 88

[00242] Table 13.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Median 96500 118000 98000 120000 98700 120000

Average 106000 129000 111000 130000 113000 129000

Stdev 33600 46300 41000 44600 42500 44100 p (t-test) 3.6E-6 1.1E-4 9.9E-4

Min 57600 57000 57600 57000 57600 57000

Max 252000 299000 299000 281000 299000 281000 n (Patient) 106 221 140 187 155 172 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 97100 120000 98300 122000 98800 122000

Average 104000 132000 110000 131000 112000 131000

Stdev 31100 47300 39100 45500 41100 44900 p (t-test) 4.6E-8 7.0E-6 1.0E-4

Min 57600 57000 57600 57000 57600 57000

Max 252000 299000 299000 281000 299000 281000 n (Patient) 118 208 144 182 158 168

UO only

p Value 8.5E-4 0.0012 0.0011 1.6E-4 4.8E-4 1.4E-4 3.8E-4 8.4E-4 2.0E-4

Lower limit of 95%

CI 1.44 1.39 1.49 1.61 1.49 1.90 1.52 1.44 1.90

Upper limit of 95%

CI 4.04 3.87 4.99 4.50 4.17 7.52 4.31 4.06 7.88

OR Quartile 3 3.22 3.57 2.39 2.97 3.25 2.87 2.83 3.03 3.20 p Value 3.0E-6 2.1E-7 2.7E-4 2.9E-6 4.3E-7 2.7E-5 5.6E-6 1.5E-6 7.4E-6

Lower limit of 95%

CI 1.97 2.21 1.50 1.88 2.06 1.75 1.81 1.93 1.92

Upper limit of 95%

CI 5.26 5.78 3.83 4.68 5.13 4.70 4.43 4.76 5.32

OR Quartile 4 3.63 4.48 2.67 2.17 2.73 2.98 1.94 2.36 3.18 p Value 1.4E-4 9.2E-6 1.8E-4 0.0047 3.4E-4 3.9E-5 0.012 0.0013 1.6E-5

Lower limit of 95%

CI 1.87 2.31 1.60 1.27 1.57 1.77 1.15 1.40 1.88

Upper limit of 95%

CI 7.06 8.69 4.47 3.72 4.72 5.02 3.25 3.99 5.38

[00243] Table 13.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 104000 123000 103000 125000 103000 125000

Average 115000 134000 114000 137000 114000 138000

Stdev 40800 46800 40500 46800 40200 47400 p (t-test) 1.3E-4 5.9E-6 3.3E-6

Min 57600 57000 57600 57000 57600 57000

Max 299000 281000 299000 281000 299000 281000 n (Patient) 204 121 214 111 219 106

[00244] Example 14. Use of Metalloproteinase inhibitor 4 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00245] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 4 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00246] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00247] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00248] Table 14.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Cohort Cohort Cohort

Median 13.0 29.0 13.3 28.8 13.3 28.8

Average 22.6 203 23.1 201 21.3 199

Stdev 27.4 884 27.6 880 21.5 873 p (t-test) 0.077 0.083 0.091

Min 0.00110 0.00167 0.00167 0.00110 0.00167 0.00110

Max 173 9030 173 9030 108 9030 n (Patient) 76 240 74 242 70 246

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 17.0 38.8 16.9 34.6 15.7 33.7

Average 97.5 261 98.2 236 98.8 220

Stdev 655 931 691 863 725 819 p (t-test) 0.069 0.12 0.16

Min 0.00110 0.00277 0.00110 0.00273 0.00110 0.00273

Max 9030 7570 9030 7570 9030 7570 n (Patient) 194 121 173 142 156 159

CI

Upper limit of 95% CI 49.5 23.2 4.07 37.6 22.3 3.66 32.6 32.7 3.95

[00249] Table 14.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.71 0.68 0.64 0.68 0.67 0.61 0.69 0.67 0.62

SE 0.030 0.030 0.034 0.032 0.031 0.033 0.032 0.031 0.032 p Value 2.8E-12 7.0E-9 7.9E-5 2.9E-8 5.4E-8 0.0012 2.0E-9 3.6E-8 1.9E-4 nCohort Recovered 90 106 212 79 104 194 71 99 178 nCohort Non- recovered 227 211 103 238 213 121 246 218 137

Cutoff Quartile 2 8.33 8.33 8.36 8.33 8.33 8.36 8.33 8.33 8.36

Sensitivity 83% 81 % 89% 81 % 81 % 86% 81 % 80% 85%

Specificity 44% 37% 32% 42% 37% 32% 45% 36% 33%

Cutoff Quartile 3 24.1 24.1 24.1 24.1 24.1 24.1 24.1 24.1 24.1

Sensitivity 58% 57% 62% 56% 57% 59% 57% 57% 60%

Specificity 70% 64% 56% 68% 63% 55% 73% 65% 57%

Cutoff Quartile 4 60.9 60.9 61.8 60.9 60.9 61.8 60.9 60.9 61.8

Sensitivity 33% 34% 33% 31 % 33% 30% 30% 33% 31 %

Specificity 93% 92% 79% 92% 91 % 78% 93% 92% 79%

OR Quartile 2 3.86 2.49 3.95 2.99 2.42 2.87 3.47 2.33 2.90 p Value 9.7E-7 6.5E-4 9.2E-5 9.5E-5 9.9E-4 5.1E-4 1.6E-5 0.0017 2.4E-4

Lower limit of 95% CI 2.25 1.47 1.98 1.73 1.43 1.58 1.97 1.37 1.64 Upper limit of 95% CI 6.62 4.20 7.86 5.19 4.08 5.21 6.11 3.94 5.12

OR Quartile 3 3.24 2.41 2.06 2.78 2.28 1.75 3.61 2.41 2.00 p Value 1.0E-5 3.5E-4 0.0033 2.0E-4 7.9E-4 0.017 1.6E-5 4.5E-4 0.0027

Lower limit of 95% CI 1.92 1.49 1.27 1.62 1.41 1.10 2.02 1.48 1.27 Upper limit of 95% CI 5.47 3.89 3.33 4.77 3.70 2.76 6.47 3.94 3.15

OR Quartile 4 6.77 5.47 1.83 5.49 5.28 1.49 5.79 5.61 1.68 p Value 1.8E-5 6.9E-6 0.025 1.4E-4 1.1E-5 0.13 2.9E-4 1.3E-5 0.046

Lower limit of 95% CI 2.83 2.61 1.08 2.29 2.52 0.887 2.24 2.58 1.01 Upper limit of 95% CI 16.2 11.5 3.10 13.2 11.1 2.49 15.0 12.2 2.81

[00250] Table 14.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 12.7 35.5 13.3 33.4 13.3 34.0

Average 27.3 248 27.8 244 26.9 239

Stdev 36.2 989 36.5 982 34.6 970 p (t-test) 0.013 0.015 0.018

Min 0.00110 0.00167 0.00167 0.00110 0.00167 0.00110

Max 223 9030 223 9030 223 9030 n (Patient) 127 190 124 193 119 198

UO only

p Value 2.7E-6 2.1E-6 0.065 1.4E-5 4.5E-6 0.16 2.0E-5 5.6E-6 0.022

Lower limit of 95% CI 2.69 2.50 0.970 2.38 2.38 0.861 2.49 2.40 1.09

Upper limit of 95% CI 11.1 9.12 2.80 9.88 8.67 2.45 11.8 9.07 3.05

[00251] Table 14.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.71 0.69 0.64 0.70 0.68 0.63 0.70 0.69 0.65

SE 0.028 0.029 0.035 0.029 0.030 0.034 0.029 0.029 0.032 p Value 6.0E-14 5.9E-11 7.2E-5 9.9E-12 5.2E-10 8.2E-5 2.5E-11 2.4E-10 5.1E-6 nCohort Recovered 134 140 216 125 138 207 114 135 194 nCohort Non- recovered 183 177 99 192 179 108 203 182 121

Cutoff Quartile 2 8.33 8.33 8.36 8.33 8.33 8.36 8.33 8.33 8.36

Sensitivity 85% 84% 86% 83% 83% 86% 83% 83% 86%

Specificity 38% 36% 30% 38% 36% 31 % 39% 36% 32%

Cutoff Quartile 3 24.1 24.1 24.1 24.1 24.1 24.1 24.1 24.1 24.1

Sensitivity 62% 60% 64% 61 % 60% 64% 61 % 60% 65%

Specificity 66% 63% 56% 67% 62% 57% 69% 63% 59%

Cutoff Quartile 4 60.9 60.9 61.8 60.9 60.9 61.8 60.9 60.9 61.8

Sensitivity 37% 37% 34% 35% 36% 32% 34% 36% 33%

Specificity 90% 89% 79% 90% 89% 79% 90% 90% 80%

OR Quartile 2 3.40 2.84 2.61 3.01 2.73 2.77 3.02 2.69 2.87 p Value 6.6E-6 1.1E-4 0.0031 3.8E-5 1.7E-4 0.0013 3.6E-5 2.1E-4 5.1E-4

Lower limit of 95% CI 2.00 1.67 1.38 1.78 1.62 1.49 1.79 1.59 1.58 Upper limit of 95% CI 5.79 4.80 4.94 5.09 4.62 5.16 5.10 4.53 5.21

OR Quartile 3 3.27 2.59 2.23 3.27 2.46 2.35 3.54 2.54 2.74 p Value 6.6E-7 4.5E-5 0.0013 9.4E-7 1.1E-4 4.9E-4 3.7E-7 6.7E-5 2.8E-5

Lower limit of 95% CI 2.05 1.64 1.37 2.04 1.56 1.45 2.18 1.61 1.71 Upper limit of 95% CI 5.21 4.08 3.64 5.24 3.88 3.79 5.77 4.01 4.39

OR Quartile 4 5.38 4.84 1.99 4.62 4.68 1.78 4.82 4.92 1.96 p Value 3.4E-7 5.4E-7 0.011 3.5E-6 9.4E-7 0.031 7.0E-6 7.4E-7 0.011

Lower limit of 95% CI 2.82 2.61 1.17 2.42 2.52 1.05 2.43 2.62 1.17 Upper limit of 95% CI 10.3 8.96 3.37 8.81 8.66 3.00 9.58 9.24 3.29

[00252] Table 14.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 15.1 38.6 15.4 37.8 15.1 38.6

Average 36.5 284 36.8 280 35.9 279

Stdev 67.1 1080 67.5 1070 66.9 1070 p (t-test) 0.0041 0.0050 0.0050

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 453 9030 453 9030 453 9030 n (Patient) 160 157 157 160 156 161

UO only

p Value 6.7E-5 1.4E-6 0.0029 5.5E-5 1.1E-6 0.0057 5.5E-5 4.6E-7 0.0049

Lower limit of 95% CI 1.71 2.27 1.31 1.74 2.33 1.24 1.74 2.47 1.25

Upper limit of 95% CI 4.87 6.98 3.76 4.95 7.25 3.52 4.95 7.82 3.55

[00253] Table 14.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.56 0.56 0.62 0.56 0.57 0.62 0.56 0.56 0.61

SE 0.042 0.038 0.032 0.046 0.038 0.031 0.048 0.039 0.031 p Value 0.13 0.12 2.5E-4 0.16 0.062 1.5E-4 0.22 0.12 5.5E-4 nCohort Recovered 52 69 197 42 67 176 38 63 159 nCohort Non- recovered 275 257 128 285 259 149 289 263 166

Cutoff Quartile 2 2320 2320 2330 2320 2320 2330 2320 2320 2330

Sensitivity 76% 76% 83% 76% 76% 82% 76% 76% 82%

Specificity 33% 30% 30% 33% 31 % 31 % 32% 30% 32%

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 51 % 51 % 60% 51 % 51 % 59% 51 % 51 % 57%

Specificity 52% 52% 56% 52% 54% 57% 53% 54% 57%

Cutoff Quartile 4 4720 4710 4720 4720 4710 4720 4720 4710 4720

Sensitivity 27% 28% 32% 27% 28% 31 % 26% 27% 30%

Specificity 85% 84% 79% 86% 85% 80% 84% 84% 79%

OR Quartile 2 1.57 1.41 2.06 1.60 1.48 2.00 1.44 1.37 2.14 p Value 0.17 0.26 0.010 0.19 0.19 0.0098 0.33 0.31 0.0039

Lower limit of 95% CI 0.825 0.781 1.19 0.795 0.821 1.18 0.692 0.746 1.28 Upper limit of 95% CI 2.98 2.53 3.57 3.20 2.67 3.38 3.01 2.52 3.59

OR Quartile 3 1.10 1.12 1.95 1.12 1.21 1.94 1.13 1.22 1.70 p Value 0.74 0.68 0.0038 0.73 0.49 0.0033 0.72 0.48 0.017

Lower limit of 95% CI 0.610 0.656 1.24 0.587 0.704 1.25 0.576 0.702 1.10 Upper limit of 95% CI 2.00 1.90 3.06 2.15 2.07 3.02 2.23 2.11 2.64

OR Quartile 4 2.02 2.01 1.79 2.18 2.19 1.74 1.90 2.00 1.60 p Value 0.084 0.050 0.024 0.090 0.034 0.032 0.17 0.062 0.070

Lower limit of 95% CI 0.911 0.999 1.08 0.884 1.06 1.05 0.766 0.965 0.962 Upper limit of 95% CI 4.50 4.05 2.97 5.38 4.53 2.88 4.73 4.14 2.66

[00254] Table 14.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 3040 3390 3040 3440 2980 3430

Average 3380 4090 3310 4110 3330 4090

Stdev 1640 2680 1520 2700 1530 2680 p (t-test) 0.015 0.0059 0.011

Min 1080 863 1080 863 1080 863

Max 9820 19700 9100 19700 9100 19700 n (Patient) 101 226 99 228 94 233

UO only

p Value 0.012 0.023 0.018 0.036 0.016 0.020 0.052 0.035 0.027

Lower limit of 95% CI 1.20 1.10 1.11 1.05 1.15 1.10 0.994 1.05 1.07

Upper limit of 95% CI 4.41 3.62 3.11 4.05 3.88 3.03 4.03 3.54 2.93

[00255] Table 14.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.58 0.59 0.60 0.57 0.60 0.59 0.58 0.60 0.61

SE 0.033 0.032 0.034 0.033 0.032 0.033 0.034 0.032 0.032 p Value 0.013 0.0032 0.0029 0.034 0.0020 0.0070 0.013 0.0028 8.2E-4 nCohort Recovered 111 124 219 104 121 208 94 116 193 nCohort Non- recovered 216 203 106 223 206 117 233 211 132

Cutoff Quartile 2 2320 2320 2330 2320 2320 2330 2320 2320 2330

Sensitivity 79% 80% 83% 78% 80% 81 % 79% 80% 82%

Specificity 32% 34% 29% 33% 34% 28% 34% 34% 30%

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 53% 54% 58% 52% 54% 56% 54% 55% 58%

Specificity 56% 56% 53% 55% 57% 53% 59% 58% 55%

Cutoff Quartile 4 4720 4720 4720 4720 4720 4720 4720 4720 4720

Sensitivity 29% 30% 31 % 28% 30% 30% 27% 29% 31 %

Specificity 83% 83% 78% 82% 83% 77% 81 % 83% 79%

OR Quartile 2 1.77 2.09 1.97 1.77 2.06 1.71 1.89 1.98 1.89 p Value 0.029 0.0045 0.023 0.031 0.0053 0.057 0.018 0.0088 0.021

Lower limit of 95% CI 1.06 1.26 1.10 1.05 1.24 0.983 1.11 1.19 1.10 Upper limit of 95% CI 2.97 3.47 3.55 2.98 3.43 2.97 3.21 3.30 3.24

OR Quartile 3 1.44 1.53 1.55 1.34 1.58 1.48 1.63 1.64 1.65 p Value 0.12 0.063 0.064 0.22 0.047 0.091 0.047 0.034 0.027

Lower limit of 95% CI 0.909 0.978 0.974 0.839 1.01 0.939 1.01 1.04 1.06 Upper limit of 95% CI 2.28 2.40 2.48 2.14 2.49 2.34 2.65 2.59 2.58

OR Quartile 4 1.99 2.11 1.57 1.76 2.17 1.46 1.60 2.00 1.67 p Value 0.019 0.0087 0.090 0.054 0.0070 0.15 0.12 0.017 0.046

Lower limit of 95% CI 1.12 1.21 0.933 0.990 1.24 0.876 0.887 1.13 1.01 Upper limit of 95% CI 3.54 3.68 2.64 3.14 3.82 2.44 2.88 3.52 2.77

[00256] Table 14.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 3130 3450 3130 3450 3110 3470

Average 3470 4170 3430 4190 3430 4170

Stdev 1860 2740 1790 2760 1800 2740 p (t-test) 0.0099 0.0054 0.0061

Min 1080 863 1080 863 1080 863

Max 14500 19700 14500 19700 14500 19700 n (Patient) 139 188 137 190 133 194

UO only

p Value 0.027 0.023 0.041 0.097 0.016 0.096 0.15 0.031 0.038

Lower limit of 95% CI 1.07 1.09 1.02 0.921 1.13 0.925 0.862 1.05 1.03

Upper limit of 95% CI 3.12 3.13 2.88 2.68 3.27 2.58 2.57 3.06 2.83

[00257] Table 14.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.58 0.56 0.62 0.59 0.56 0.62 0.59 0.55 0.63

SE 0.032 0.032 0.034 0.032 0.032 0.033 0.032 0.032 0.033 p Value 0.0081 0.071 3.9E-4 0.0063 0.072 2.7E-4 0.0052 0.094 4.6E-5 nCohort Recovered 187 159 215 184 156 212 183 154 209 nCohort Non- recovered 140 168 110 143 171 113 144 173 116

Cutoff Quartile 2 2320 2320 2330 2320 2320 2330 2320 2320 2330

Sensitivity 81 % 79% 85% 82% 79% 85% 82% 79% 85%

Specificity 30% 30% 30% 30% 29% 30% 31 % 29% 31 %

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 55% 52% 57% 55% 52% 58% 56% 52% 59%

Specificity 53% 52% 53% 54% 52% 54% 54% 52% 55%

Cutoff Quartile 4 4720 4720 4720 4720 4720 4720 4720 4720 4720

Sensitivity 31 % 29% 32% 31 % 29% 32% 31 % 29% 34%

Specificity 79% 79% 78% 79% 79% 78% 79% 79% 79%

OR Quartile 2 1.87 1.59 2.32 1.97 1.57 2.44 2.00 1.52 2.57 p Value 0.020 0.070 0.0055 0.012 0.080 0.0032 0.010 0.10 0.0018

Lower limit of 95%

CI 1.10 0.963 1.28 1.16 0.948 1.35 1.18 0.918 1.42

Upper limit of 95% CI 3.18 2.64 4.20 3.34 2.59 4.42 3.39 2.51 4.65

OR Quartile 3 1.40 1.20 1.54 1.44 1.17 1.58 1.47 1.17 1.70 p Value 0.13 0.41 0.067 0.10 0.47 0.053 0.084 0.47 0.023

Lower limit of 95%

CI 0.905 0.778 0.970 0.927 0.759 0.994 0.950 0.759 1.07

Upper limit of 95% CI 2.18 1.85 2.45 2.23 1.81 2.50 2.28 1.81 2.69

OR Quartile 4 1.68 1.57 1.67 1.71 1.60 1.69 1.68 1.55 1.96 p Value 0.043 0.081 0.052 0.037 0.070 0.046 0.044 0.092 0.010

Lower limit of 95%

CI 1.02 0.946 0.996 1.03 0.962 1.01 1.01 0.931 1.17

Upper limit of 95% CI 2.78 2.61 2.79 2.83 2.67 2.82 2.78 2.58 3.26

[00258] Example 15. Use of Metalloproteinase inhibitor 4 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00259] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 4 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00260] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00261] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00262] Table 15.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort Cohort Cohort

Median 18.5 39.3 18.1 39.3 18.1 36.7

Average 99.9 304 101 275 104 255

Stdev 623 1040 643 976 658 936 p (t-test) 0.033 0.059 0.094

Min 0.00110 0.00277 0.00110 0.00277 0.00110 0.00277

Max 9030 7570 9030 7570 9030 7570 n (Patient) 222 93 208 107 198 117

[00263] Table 15.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Cohort Cohort Cohort

Median 13.3 36.5 13.3 33.4 13.3 33.4

Average 29.4 257 29.6 254 28.6 249

Stdev 47.4 1010 47.7 1010 46.7 994 p (t-test) 0.0093 0.010 0.012

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 409 9030 409 9030 409 9030 n (Patient) 136 181 134 183 129 188 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 15.4 36.9 15.9 36.7 15.9 36.7

Average 30.5 274 30.7 272 30.6 269

Stdev 47.1 1050 47.2 1050 47.4 1040 p (t-test) 0.0050 0.0054 0.0060

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 409 9030 409 9030 409 9030 n (Patient) 149 168 148 169 146 171

UO only

Sensitivity 36% 38% 36% 36% 38% 35% 36% 37% 33% Specificity 90% 89% 79% 90% 89% 79% 90% 89% 79%

OR Quartile 2 2.83 2.10 3.05 2.94 2.14 3.03 3.00 2.21 2.77 p Value 1.0E-4 0.0051 0.0016 5.9E-5 0.0042 0.0012 4.1E-5 0.0028 0.0013

Lower limit of 95%

CI 1.68 1.25 1.53 1.74 1.27 1.55 1.77 1.31 1.49

Upper limit of 95% CI 4.79 3.53 6.10 4.98 3.59 5.92 5.07 3.72 5.16

OR Quartile 3 2.47 2.24 2.36 2.47 2.19 2.13 2.57 2.19 1.96 p Value 1.1E-4 4.4E-4 9.6E-4 1.1E-4 6.5E-4 0.0027 6.2E-5 6.5E-4 0.0053

Lower limit of 95%

CI 1.56 1.43 1.42 1.57 1.39 1.30 1.62 1.40 1.22

Upper limit of 95% CI 3.89 3.52 3.92 3.91 3.43 3.50 4.08 3.43 3.16

OR Quartile 4 5.00 5.12 2.14 4.84 5.03 2.01 4.94 4.86 1.91 p Value 5.6E-7 1.2E-7 0.0057 9.6E-7 1.7E-7 0.010 1.3E-6 3.0E-7 0.015

Lower limit of 95%

CI 2.66 2.79 1.25 2.57 2.75 1.18 2.59 2.65 1.13

Upper limit of 95% CI 9.39 9.37 3.67 9.08 9.21 3.43 9.43 8.90 3.22

[00264] Table 15.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 18.1 38.8 18.2 37.8 18.1 33.4

Average 109 286 110 283 112 259

Stdev 628 1050 629 1040 645 986 p (t-test) 0.066 0.071 0.11

Min 0.00110 0.00277 0.00110 0.00277 0.00110 0.00277

Max 9030 7570 9030 7570 9030 7570 n (Patient) 224 91 223 92 212 103

[00265] Table 15.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 15.1 38.8 15.4 37.0 15.4 36.9

Average 36.0 298 36.4 293 35.9 287

Stdev 62.4 1110 62.9 1100 62.7 1090 p (t-test) 0.0024 0.0030 0.0037

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 453 9030 453 9030 453 9030 n (Patient) 168 149 165 152 161 156 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 15.9 37.0 16.3 37.0 16.3 37.0

Average 36.4 305 36.6 303 36.7 301

Stdev 61.8 1130 61.9 1120 62.1 1120 p (t-test) 0.0019 0.0021 0.0023

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 453 9030 453 9030 453 9030 n (Patient) 172 145 171 146 170 147

UO only

Specificity 61 % 60% 56% 61 % 60% 56% 61 % 59% 56%

Cutoff Quartile 4 60.9 60.9 61.8 60.9 60.9 61.8 60.9 60.9 61.8

Sensitivity 38% 38% 35% 37% 38% 34% 37% 37% 34%

Specificity 86% 85% 79% 85% 85% 79% 86% 85% 79%

OR Quartile 2 2.54 2.20 2.55 2.67 2.23 2.60 2.64 2.27 2.45 p Value 7.8E-4 0.0041 0.0049 4.0E-4 0.0034 0.0041 4.0E-4 0.0028 0.0048

Lower limit of 95%

CI 1.47 1.28 1.33 1.55 1.30 1.36 1.54 1.33 1.31

Upper limit of 95% CI 4.36 3.76 4.91 4.59 3.83 5.00 4.53 3.89 4.55

OR Quartile 3 2.71 2.44 2.24 2.63 2.38 2.30 2.63 2.31 2.21 p Value 1.8E-5 1.1E-4 0.0016 3.0E-5 1.8E-4 0.0011 3.0E-5 2.8E-4 0.0014

Lower limit of 95%

CI 1.72 1.55 1.36 1.67 1.51 1.39 1.67 1.47 1.36

Upper limit of 95% CI 4.27 3.84 3.70 4.14 3.73 3.80 4.13 3.63 3.59

OR Quartile 4 3.61 3.59 2.00 3.43 3.53 1.95 3.45 3.47 1.91 p Value 3.8E-6 3.5E-6 0.011 9.1E-6 4.8E-6 0.014 9.5E-6 6.4E-6 0.017

Lower limit of 95%

CI 2.10 2.09 1.17 1.99 2.06 1.14 2.00 2.02 1.12

Upper limit of 95% CI 6.23 6.17 3.41 5.90 6.06 3.34 5.98 5.95 3.23

[00266] Table 15.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00267] Table 15.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 82% 81 % 85% 81 % 81 % 86% 81 % 81 % 84% Specificity 39% 36% 29% 39% 36% 31 % 39% 37% 30%

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 55% 55% 65% 56% 55% 64% 55% 56% 61 %

Specificity 61 % 58% 56% 62% 59% 57% 61 % 60% 56%

Cutoff Quartile 4 4720 4710 4720 4720 4710 4720 4720 4710 4720

Sensitivity 30% 31 % 35% 30% 31 % 35% 30% 31 % 34%

Specificity 86% 84% 79% 86% 85% 80% 86% 85% 80%

OR Quartile 2 2.83 2.35 2.31 2.81 2.44 2.64 2.66 2.53 2.23 p Value 8.4E-5 0.0010 0.0081 9.8E-5 6.4E-4 0.0017 2.4E-4 3.8E-4 0.0055

Lower limit of 95%

CI 1.68 1.41 1.24 1.67 1.46 1.44 1.58 1.52 1.27

Upper limit of 95% CI 4.75 3.91 4.29 4.73 4.06 4.82 4.49 4.23 3.92

OR Quartile 3 1.94 1.65 2.35 2.02 1.75 2.41 1.93 1.86 2.02 p Value 0.0061 0.030 6.3E-4 0.0040 0.016 2.6E-4 0.0075 0.0082 0.0025

Lower limit of 95%

CI 1.21 1.05 1.44 1.25 1.11 1.50 1.19 1.17 1.28

Upper limit of 95% CI 3.11 2.60 3.82 3.26 2.76 3.87 3.13 2.94 3.20

OR Quartile 4 2.59 2.38 2.08 2.72 2.51 2.11 2.53 2.66 2.10 p Value 0.0025 0.0030 0.0059 0.0019 0.0019 0.0043 0.0040 0.0012 0.0042

Lower limit of 95%

CI 1.40 1.34 1.24 1.45 1.40 1.26 1.35 1.47 1.26

Upper limit of 95% CI 4.81 4.22 3.51 5.12 4.49 3.53 4.77 4.80 3.49

[00268] Table 15.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Min 1080 863 1080 863 1080 863

Max 9820 19700 9820 19700 9100 19700 n (Patient) 146 181 145 182 143 184

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 3060 3890 3040 3930 3050 3920

Average 3530 4670 3500 4670 3460 4600

Stdev 2060 2970 2060 2910 2040 2840 p (t-test) 9.7E-5 5.0E-5 3.9E-5

Min 893 863 893 863 893 863

Max 19700 18800 19700 18800 19700 18800 n (Patient) 231 94 225 100 210 115

[00269] Table 15.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Sensitivity 80% 81 % 85% 81 % 81 % 86% 80% 81 % 85% Specificity 31 % 31 % 29% 32% 31 % 29% 31 % 31 % 30%

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 53% 53% 59% 54% 53% 60% 53% 54% 58%

Specificity 53% 53% 53% 54% 53% 54% 53% 53% 54%

Cutoff Quartile 4 4720 4720 4720 4720 4720 4720 4720 4720 4720

Sensitivity 29% 30% 32% 30% 30% 33% 29% 30% 33%

Specificity 79% 80% 77% 80% 80% 78% 80% 81 % 79%

OR Quartile 2 1.85 1.89 2.38 1.91 1.92 2.47 1.79 1.95 2.32 p Value 0.017 0.015 0.0075 0.012 0.012 0.0052 0.024 0.010 0.0055

Lower limit of 95% CI 1.12 1.13 1.26 1.15 1.15 1.31 1.08 1.17 1.28 Upper limit of 95% CI 3.07 3.14 4.49 3.18 3.19 4.65 2.97 3.25 4.20

OR Quartile 3 1.29 1.26 1.65 1.36 1.29 1.74 1.23 1.33 1.63 p Value 0.25 0.29 0.042 0.17 0.25 0.024 0.35 0.20 0.039

Lower limit of 95% CI 0.837 0.817 1.02 0.879 0.837 1.08 0.797 0.858 1.02 Upper limit of 95% CI 2.00 1.95 2.68 2.10 2.00 2.82 1.91 2.05 2.59

OR Quartile 4 1.55 1.62 1.58 1.72 1.71 1.75 1.61 1.80 1.79 p Value 0.092 0.061 0.092 0.039 0.039 0.037 0.069 0.024 0.027

Lower limit of 95% CI 0.931 0.978 0.928 1.03 1.03 1.03 0.963 1.08 1.07 Upper limit of 95% CI 2.58 2.70 2.69 2.88 2.85 2.97 2.69 3.00 2.99

[00270] Table 15.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00271] Table 15.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 3100 3510 3080 3520 3090 3510

Average 3560 4390 3550 4390 3550 4380

Stdev 2090 2830 2100 2810 2100 2800 p (t-test) 0.0026 0.0020 0.0025

Min 1080 863 1080 863 1080 863

Max 19700 18800 19700 18800 19700 18800 n (Patient) 204 123 202 125 201 126 sCr only

UO only

Specificity 30% 30% 30% 30% 30% 31 % 30% 30% 31 %

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 55% 55% 58% 56% 55% 58% 56% 55% 58%

Specificity 53% 53% 54% 53% 53% 54% 53% 53% 54%

Cutoff Quartile 4 4720 4720 4720 4720 4720 4720 4720 4720 4720

Sensitivity 29% 30% 32% 30% 30% 32% 29% 30% 32%

Specificity 77% 79% 78% 78% 79% 78% 78% 79% 78%

OR Quartile 2 2.07 1.97 2.85 2.14 1.97 2.90 2.18 1.97 3.00 p Value 0.010 0.012 0.0012 0.0073 0.012 9.6E-4 0.0061 0.012 6.5E-4

Lower limit of 95% CI 1.19 1.16 1.51 1.23 1.16 1.54 1.25 1.16 1.60 Upper limit of 95% CI 3.62 3.34 5.35 3.74 3.34 5.45 3.80 3.34 5.64

OR Quartile 3 1.39 1.37 1.60 1.46 1.37 1.65 1.42 1.37 1.64 p Value 0.15 0.16 0.049 0.097 0.16 0.037 0.12 0.16 0.038

Lower limit of 95% CI 0.887 0.882 1.00 0.934 0.882 1.03 0.910 0.882 1.03 Upper limit of 95% CI 2.18 2.12 2.57 2.29 2.12 2.63 2.23 2.12 2.61

OR Quartile 4 1.42 1.60 1.72 1.47 1.60 1.68 1.44 1.60 1.73 p Value 0.18 0.067 0.041 0.14 0.067 0.049 0.16 0.067 0.037

Lower limit of 95% CI 0.855 0.967 1.02 0.883 0.967 1.00 0.868 0.967 1.03 Upper limit of 95% CI 2.36 2.64 2.88 2.44 2.64 2.83 2.39 2.64 2.91

[00272] Example 16. Use of Metalloproteinase inhibitor 4 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00273] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 4 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00274] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00275] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis. [00276] Table 16.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 205 208 275 221 223 278 236 235 279 nCohort Persistent 112 108 40 96 93 37 81 81 36

Cutoff Quartile 2 8.33 8.31 8.36 8.33 8.31 8.36 8.33 8.31 8.36

Sensitivity 86% 84% 88% 89% 87% 86% 89% 86% 86%

Specificity 31 % 30% 27% 31 % 30% 27% 30% 29% 27%

Cutoff Quartile 3 24.1 23.6 24.1 24.1 23.6 24.1 24.1 23.6 24.1

Sensitivity 71 % 69% 80% 74% 72% 78% 77% 73% 78%

Specificity 61 % 60% 54% 60% 59% 54% 59% 58% 53%

Cutoff Quartile 4 60.9 60.8 61.8 60.9 60.8 61.8 60.9 60.8 61.8

Sensitivity 46% 45% 45% 50% 48% 46% 48% 46% 47%

Specificity 86% 86% 78% 86% 85% 78% 83% 82% 78%

OR Quartile 2 2.66 2.27 2.58 3.43 2.90 2.32 3.37 2.59 2.24 p Value 0.0016 0.0070 0.057 4.6E-4 0.0019 0.092 0.0014 0.0073 0.11

Lower limit of 95%

CI 1.45 1.25 0.973 1.72 1.48 0.872 1.60 1.29 0.839

Upper limit of 95% CI 4.88 4.13 6.83 6.85 5.67 6.18 7.12 5.19 5.97

OR Quartile 3 3.74 3.21 4.73 4.29 3.74 4.19 4.68 3.68 4.01 p Value 1.6E-7 3.3E-6 1.7E-4 6.9E-8 8.9E-7 6.0E-4 1.5E-7 3.9E-6 9.0E-4

Lower limit of 95%

CI 2.28 1.97 2.10 2.53 2.21 1.85 2.63 2.12 1.77

Upper limit of 95% CI 6.13 5.25 10.6 7.29 6.32 9.48 8.32 6.41 9.11

OR Quartile 4 5.07 4.93 2.87 5.91 5.21 2.96 4.42 3.86 3.13 p Value 4.0E-9 7.8E-9 0.0025 2.1E-10 3.2E-9 0.0026 1.3E-7 1.5E-6 0.0017

Lower limit of 95%

CI 2.95 2.87 1.45 3.41 3.02 1.46 2.55 2.23 1.54

Upper limit of 95% CI 8.72 8.47 5.69 10.2 9.00 6.00 7.66 6.70 6.39

[00277] Table 16.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Stdev 60.8 1210 59.2 1280 612 1030 p (t-test) 4.7E-4 1.0E-4 0.0075

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 409 9030 409 9030 9030 7570 n (Patient) 192 124 206 110 218 98

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 20.0 43.9 20.4 50.8 20.5 51.9

Average 101 416 100 443 100 451

Stdev 588 1280 584 1330 583 1340 p (t-test) 0.0048 0.0027 0.0023

Min 0.00110 0.00277 0.00110 0.00277 0.00110 0.00277

Max 9030 7570 9030 7570 9030 7570 n (Patient) 256 59 260 55 261 54

[00278] Table 16.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 175 182 246 190 195 251 205 208 251 nCohort Persistent 142 134 69 127 121 64 112 108 64

Cutoff Quartile 2 8.33 8.31 8.36 8.33 8.31 8.36 8.33 8.31 8.36

Sensitivity 85% 83% 86% 87% 85% 84% 87% 85% 84%

Specificity 33% 31 % 28% 33% 31 % 27% 31 % 30% 27%

Cutoff Quartile 3 24.1 23.6 24.1 24.1 23.6 24.1 24.1 23.6 24.1

Sensitivity 65% 64% 70% 69% 68% 70% 70% 67% 70%

Specificity 62% 60% 55% 63% 61 % 55% 60% 59% 55%

Cutoff Quartile 4 60.9 60.8 61.8 60.9 60.8 61.8 60.9 60.8 61.8

Sensitivity 41 % 41 % 39% 45% 44% 41 % 44% 42% 41 %

Specificity 87% 87% 79% 88% 87% 79% 85% 84% 79%

OR Quartile 2 2.63 2.14 2.30 3.13 2.60 2.05 2.94 2.50 2.05 p Value 6.0E-4 0.0064 0.024 1.6E-4 0.0013 0.054 6.5E-4 0.0031 0.054

Lower limit of 95% CI 1.51 1.24 1.11 1.73 1.45 0.987 1.58 1.36 0.987 Upper limit of 95% CI 4.58 3.71 4.75 5.68 4.68 4.25 5.45 4.59 4.25

OR Quartile 3 3.13 2.74 2.83 3.78 3.29 2.89 3.51 2.84 2.89 p Value 1.2E-6 1.9E-5 3.6E-4 4.9E-8 1.0E-6 4.3E-4 5.2E-7 2.6E-5 4.3E-4

Lower limit of 95% CI 1.98 1.73 1.60 2.34 2.04 1.60 2.15 1.75 1.60 Upper limit of 95% CI 4.97 4.34 5.00 6.10 5.31 5.22 5.74 4.61 5.22

OR Quartile 4 4.80 4.58 2.40 5.91 5.07 2.56 4.37 3.66 2.56 p Value 3.6E-8 5.9E-8 0.0027 4.6E-10 6.2E-9 0.0016 6.4E-8 1.7E-6 0.0016

Lower limit of 95% CI 2.75 2.64 1.35 3.38 2.93 1.43 2.56 2.15 1.43 Upper limit of 95% CI 8.39 7.95 4.25 10.3 8.76 4.58 7.45 6.21 4.58

[00279] Table 16.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 409 9030 409 9030 409 9030 n (Patient) 180 136 192 124 204 112

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 18.2 42.1 18.8 47.4 18.8 47.4

Average 104 337 102 363 102 363

Stdev 608 1140 601 1180 601 1180 p (t-test) 0.022 0.013 0.013

Min 0.00110 0.00277 0.00110 0.00277 0.00110 0.00277

Max 9030 7570 9030 7570 9030 7570 n (Patient) 239 76 245 70 245 70

[00280] Table 16.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 65% 63% 70% 69% 66% 71 % 68% 66% 71 % Specificity 63% 60% 57% 64% 61 % 57% 61 % 59% 57%

Cutoff Quartile 4 60.9 60.8 61.8 60.9 60.8 61.8 60.9 60.8 61.8

Sensitivity 41 % 40% 41 % 44% 43% 41 % 42% 41 % 41 %

Specificity 88% 87% 80% 89% 87% 80% 85% 84% 80%

OR Quartile 2 2.49 2.04 2.61 2.90 2.47 2.45 2.78 2.39 2.45 p Value 9.7E-4 0.0096 0.0069 2.8E-4 0.0018 0.012 7.4E-4 0.0036 0.012

Lower limit of 95% CI 1.45 1.19 1.30 1.63 1.40 1.22 1.54 1.33 1.22 Upper limit of 95% CI 4.29 3.49 5.23 5.16 4.36 4.92 5.04 4.29 4.92

OR Quartile 3 3.11 2.56 3.13 3.92 3.11 3.11 3.36 2.73 3.11 p Value 1.3E-6 5.4E-5 3.8E-5 1.7E-8 2.1E-6 5.3E-5 7.1E-7 3.4E-5 5.3E-5

Lower limit of 95% CI 1.96 1.62 1.82 2.44 1.95 1.79 2.08 1.70 1.79 Upper limit of 95% CI 4.92 4.05 5.38 6.30 4.98 5.39 5.43 4.40 5.39

OR Quartile 4 5.08 4.43 2.81 6.19 5.15 2.81 4.28 3.85 2.81 p Value 2.4E-8 1.3E-7 2.2E-4 3.4E-10 6.4E-9 2.5E-4 1.0E-7 6.9E-7 2.5E-4

Lower limit of 95% CI 2.87 2.55 1.62 3.50 2.96 1.62 2.51 2.26 1.62 Upper limit of 95% CI 8.99 7.71 4.86 10.9 8.95 4.89 7.31 6.55 4.89

[00281] Table 16.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3140 4580 3150 4610 3140 4750

Average 3580 5670 3580 5740 3570 5940

Stdev 2020 3670 2010 3740 2010 3720 p (t-test) 5.0E-8 3.4E-8 2.5E-9

Min 893 863 893 863 893 863

Max 19700 18800 19700 18800 19700 18800 n (Patient) 281 44 283 42 285 40

[00282] Table 16.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Stdev 1590 3130 1800 3120 2090 2930 p (t-test) 9.5E-5 4.3E-4 0.0035

Min 1080 863 1080 863 1080 863

Max 9820 19700 14500 19700 19700 18800 n (Patient) 187 140 204 123 219 108 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3100 3620 3090 3810 3130 3520

Average 3420 4490 3490 4480 3600 4380

Stdev 1580 3180 1790 3140 2090 2930 p (t-test) 8.1E-5 3.4E-4 0.0059

Min 1080 863 1080 863 1080 863

Max 9820 19700 14500 19700 19700 18800 n (Patient) 192 134 205 121 217 109

UO only

Upper limit of 95% CI 2.73 2.69 6.12 3.05 2.97 7.06 3.07 2.87 11.5

OR Quartile 3 1.48 1.43 1.78 1.47 1.49 1.75 1.46 1.36 1.93 p Value 0.082 0.12 0.042 0.096 0.086 0.053 0.11 0.20 0.025

Lower limit of 95% CI 0.951 0.916 1.02 0.935 0.945 0.993 0.919 0.854 1.09 Upper limit of 95% CI 2.29 2.22 3.11 2.30 2.33 3.08 2.32 2.15 3.45

OR Quartile 4 1.92 1.85 1.87 1.86 1.79 2.04 1.64 1.49 2.17 p Value 0.011 0.017 0.037 0.017 0.025 0.018 0.062 0.13 0.011

Lower limit of 95% CI 1.16 1.12 1.04 1.12 1.08 1.13 0.976 0.887 1.20 Upper limit of 95% CI 3.18 3.07 3.35 3.09 2.98 3.68 2.75 2.50 3.94

[00283] Table 16.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 249 76 253 72 254 71

[00284] Table 16.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 3090 3510 3080 3620 3100 3620

Average 3390 4430 3450 4430 3480 4480

Stdev 1520 3110 1740 3060 1730 3170 p (t-test) 9.8E-5 2.9E-4 2.7E-4

Min 1080 863 1080 863 1080 863

Max 9100 19700 14500 19700 14500 19700 n (Patient) 179 147 190 136 202 124

UO only

p Value 0.033 0.021 0.014 0.019 0.024 0.0097 0.062 0.075 0.0076

Lower limit of 95%

CI 1.04 1.09 1.15 1.10 1.08 1.19 0.976 0.955 1.22

Upper limit of 95% CI 2.87 3.00 3.39 3.03 2.97 3.56 2.68 2.64 3.66

[00285] Table 16.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.59 0.60 0.66 0.60 0.60 0.66 0.59 0.59 0.67

SE 0.031 0.032 0.035 0.032 0.032 0.035 0.032 0.033 0.035 p Value 0.0032 0.0022 7.1E-6 0.0024 0.0019 3.7E-6 0.0084 0.0044 1.3E-6 nCohort Non- persistent 169 175 235 183 186 237 196 198 238 nCohort Persistent 158 151 90 144 140 88 131 128 87

Cutoff Quartile 2 2320 2320 2330 2320 2320 2330 2320 2320 2330

Sensitivity 80% 80% 89% 82% 81 % 90% 82% 81 % 91 %

Specificity 30% 30% 30% 31 % 30% 30% 30% 29% 31 %

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 54% 55% 62% 55% 56% 62% 55% 56% 63%

Specificity 54% 54% 54% 54% 54% 54% 53% 54% 55%

Cutoff Quartile 4 4720 4710 4720 4720 4710 4720 4720 4710 4720

Sensitivity 30% 32% 36% 31 % 32% 36% 30% 31 % 37%

Specificity 80% 81 % 79% 80% 80% 79% 78% 79% 79%

OR Quartile 2 1.77 1.71 3.46 2.00 1.89 3.83 1.87 1.80 4.37 p Value 0.029 0.042 6.5E-4 0.010 0.018 4.0E-4 0.022 0.033 2.0E-4

Lower limit of 95%

CI 1.06 1.02 1.70 1.18 1.11 1.82 1.09 1.05 2.01

Upper limit of 95% CI 2.95 2.85 7.07 3.39 3.20 8.05 3.21 3.08 9.51

OR Quartile 3 1.39 1.45 1.97 1.40 1.49 1.99 1.38 1.51 2.07 p Value 0.14 0.096 0.0075 0.13 0.074 0.0071 0.16 0.070 0.0048

Lower limit of 95%

CI 0.902 0.936 1.20 0.904 0.962 1.21 0.885 0.967 1.25

Upper limit of 95% CI 2.15 2.24 3.24 2.17 2.32 3.29 2.15 2.36 3.43

OR Quartile 4 1.73 1.93 2.04 1.79 1.91 2.14 1.51 1.69 2.19 p Value 0.033 0.011 0.0087 0.023 0.012 0.0054 0.11 0.042 0.0042

Lower limit of 95%

CI 1.04 1.16 1.20 1.08 1.15 1.25 0.911 1.02 1.28

Upper limit of 95% CI 2.87 3.21 3.48 2.97 3.16 3.65 2.50 2.80 3.74

[00286] Example 17. Use of Metalloproteinase inhibitor 4 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00287] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized.

Metalloproteinase inhibitor 4 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00288] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00289] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00290] Table 17.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 598 1160 588 1280 581 1360 p (t-test) 0.0081 0.0038 0.0017

Min 0.00110 1.76 0.00110 1.76 0.00110 1.76

Max 9030 7570 9030 7570 9030 7570 n (Patient) 241 74 256 59 263 52

[00291] Table 17.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

n (Patient) 112 205 147 170 165 152 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 13.3 34.9 16.3 36.9 15.4 40.1

Average 29.5 246 31.6 281 32.9 305

Stdev 47.4 989 47.8 1070 52.3 1120 p (t-test) 0.015 0.0041 0.0017

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 409 9030 409 9030 409 9030 n (Patient) 126 190 154 162 169 147

UO only

Upper limit of 95% CI 4.53 3.75 4.36 3.93 3.38 4.83 4.63 3.89 5.65

OR Quartile 4 5.29 4.63 2.44 4.94 4.56 2.46 4.73 4.23 2.54 p Value 4.4E-6 3.5E-6 0.0010 2.2E-7 3.7E-7 0.0015 9.0E-8 3.9E-7 0.0013

Lower limit of 95% CI 2.60 2.42 1.43 2.70 2.54 1.41 2.68 2.42 1.44

Upper limit of 95% CI 10.8 8.84 4.16 9.05 8.19 4.29 8.36 7.39 4.48

[00292] Table 17.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.69 0.66 0.64 0.68 0.66 0.64 0.69 0.67 0.64

SE 0.030 0.030 0.034 0.030 0.030 0.036 0.030 0.030 0.037 p Value 1.8E-10 8.1E-8 5.9E-5 1.0E-9 1.4E-7 1.5E-4 1.2E-10 2.2E-8 1.6E-4 nCohort Non- persistent 111 124 212 144 151 225 158 163 232 nCohort Persistent 206 192 103 173 165 90 159 153 83

Cutoff Quartile 2 8.33 8.31 8.36 8.33 8.31 8.36 8.33 8.31 8.36

Sensitivity 82% 81 % 85% 83% 82% 86% 85% 84% 86%

Specificity 38% 34% 30% 35% 32% 29% 35% 33% 29%

Cutoff Quartile 3 24.1 23.6 24.1 24.1 23.6 24.1 24.1 23.6 24.1

Sensitivity 59% 58% 63% 61 % 59% 64% 63% 61 % 66%

Specificity 67% 63% 56% 63% 60% 56% 63% 60% 56%

Cutoff Quartile 4 60.9 60.8 61.8 60.9 60.8 61.8 60.9 60.8 61.8

Sensitivity 34% 34% 35% 37% 37% 36% 38% 37% 36%

Specificity 91 % 90% 80% 89% 88% 79% 87% 87% 79%

OR Quartile 2 2.78 2.15 2.54 2.64 2.16 2.46 3.00 2.54 2.40 p Value 1.3E-4 0.0038 0.0033 3.0E-4 0.0038 0.0070 7.4E-5 7.1E-4 0.011

Lower limit of 95% CI 1.65 1.28 1.36 1.56 1.28 1.28 1.74 1.48 1.22

Upper limit of 95% CI 4.69 3.60 4.72 4.47 3.64 4.73 5.17 4.35 4.72

OR Quartile 3 2.90 2.37 2.19 2.72 2.22 2.27 2.84 2.34 2.46 p Value 1.5E-5 2.6E-4 0.0015 1.8E-5 5.2E-4 0.0015 6.9E-6 2.3E-4 7.5E-4

Lower limit of 95% CI 1.79 1.49 1.35 1.72 1.41 1.37 1.80 1.49 1.46

Upper limit of 95% CI 4.71 3.77 3.55 4.29 3.48 3.76 4.48 3.67 4.15

OR Quartile 4 5.20 4.47 2.11 4.70 4.33 2.09 4.18 3.81 2.11 p Value 5.5E-6 5.7E-6 0.0053 5.3E-7 9.0E-7 0.0073 8.0E-7 2.5E-6 0.0074

Lower limit of 95% CI 2.55 2.34 1.25 2.57 2.41 1.22 2.37 2.18 1.22

Upper limit of 95% CI 10.6 8.54 3.57 8.60 7.78 3.58 7.38 6.64 3.66

[00293] Table 17.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 13.3 34.7 16.3 34.9 15.9 36.9

Average 29.7 243 32.0 275 33.7 292

Stdev 47.7 984 48.4 1060 53.3 1090 p (t-test) 0.016 0.0052 0.0029

Min 0.00110 0.00167 0.00110 0.00167 0.00110 0.00167

Max 409 9030 409 9030 409 9030 n (Patient) 124 192 150 166 162 154

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 18.2 36.7 18.2 37.0 18.2 38.8

Average 104 270 108 281 108 294

Stdev 643 977 633 1030 625 1060 p (t-test) 0.071 0.068 0.055

Min 0.00110 0.00277 0.00110 0.00277 0.00110 0.00277

Max 9030 7570 9030 7570 9030 7570 n (Patient) 208 107 220 95 226 89

[00294] Table 17.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 106 121 202 138 146 212 150 158 217 nCohort Persistent 211 195 113 179 170 103 167 158 98

Cutoff Quartile 2 8.33 8.31 8.36 8.33 8.31 8.36 8.33 8.31 8.36

Sensitivity 82% 81 % 85% 83% 82% 86% 84% 84% 87%

Specificity 38% 35% 31 % 35% 33% 31 % 35% 34% 30%

Cutoff Quartile 3 24.1 23.6 24.1 24.1 23.6 24.1 24.1 23.6 24.1

Sensitivity 59% 58% 63% 60% 59% 63% 62% 61 % 64%

Specificity 67% 64% 57% 63% 61 % 56% 63% 61 % 56%

Cutoff Quartile 4 60.9 60.8 61.8 60.9 60.8 61.8 60.9 60.8 61.8

Sensitivity 34% 34% 35% 36% 36% 35% 37% 37% 35%

Specificity 92% 90% 81 % 89% 88% 80% 87% 87% 79%

OR Quartile 2 2.67 2.27 2.50 2.55 2.20 2.81 2.96 2.56 2.86 p Value 2.4E-4 0.0019 0.0026 4.5E-4 0.0030 0.0014 7.1E-5 5.6E-4 0.0016

Lower limit of 95% CI 1.58 1.35 1.38 1.51 1.31 1.49 1.73 1.50 1.49 Upper limit of 95% CI 4.52 3.81 4.54 4.29 3.70 5.30 5.06 4.37 5.48

OR Quartile 3 2.89 2.46 2.23 2.59 2.29 2.19 2.70 2.40 2.31 p Value 2.1E-5 1.6E-4 8.5E-4 4.4E-5 3.4E-4 0.0015 1.8E-5 1.5E-4 8.6E-4

Lower limit of 95% CI 1.77 1.54 1.39 1.64 1.45 1.35 1.71 1.53 1.41 Upper limit of 95% CI 4.71 3.93 3.58 4.10 3.59 3.55 4.26 3.77 3.78

OR Quartile 4 5.47 4.75 2.29 4.68 4.36 2.11 3.97 3.78 2.03 p Value 6.9E-6 4.4E-6 0.0018 9.4E-7 1.2E-6 0.0053 2.6E-6 3.4E-6 0.0088

Lower limit of 95% CI 2.61 2.44 1.36 2.52 2.40 1.25 2.23 2.16 1.20 Upper limit of 95% CI 11.5 9.25 3.85 8.66 7.89 3.57 7.05 6.64 3.45

[00295] Table 17.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 9820 19700 9820 19700 14500 19700 n (Patient) 124 202 157 169 182 144

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3040 4350 3100 4340 3100 4390

Average 3520 4910 3570 5040 3570 5240

Stdev 2050 3060 2050 3280 2050 3390 p (t-test) 6.0E-6 1.0E-5 1.8E-6

Min 893 863 893 863 893 863

Max 19700 18800 19700 18800 19700 18800 n (Patient) 245 80 261 64 269 56

[00296] Table 17.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 2320 2320 2330 2320 2320 2330 2320 2320 2330

Sensitivity 82% 81 % 86% 82% 82% 87% 80% 80% 88%

Specificity 39% 36% 30% 34% 34% 29% 30% 30% 29%

Cutoff Quartile 3 3280 3280 3280 3280 3280 3280 3280 3280 3280

Sensitivity 57% 55% 64% 57% 56% 62% 55% 54% 62%

Specificity 63% 59% 56% 58% 57% 54% 54% 54% 54%

Cutoff Quartile 4 4720 4710 4720 4720 4710 4720 4720 4710 4720

Sensitivity 31 % 31 % 36% 33% 32% 33% 31 % 30% 34%

Specificity 87% 85% 80% 85% 83% 78% 80% 80% 77%

OR Quartile 2 2.85 2.39 2.60 2.34 2.29 2.72 1.68 1.72 3.09 p Value 7.2E-5 8.2E-4 0.0030 0.0011 0.0015 0.0047 0.046 0.036 0.0031

Lower limit of 95% CI 1.70 1.44 1.38 1.41 1.37 1.36 1.01 1.04 1.46 Upper limit of 95% CI 4.78 3.98 4.91 3.91 3.82 5.43 2.79 2.87 6.53

OR Quartile 3 2.22 1.80 2.26 1.84 1.69 1.90 1.43 1.34 1.91 p Value 9.6E-4 0.012 9.9E-4 0.0067 0.020 0.013 0.11 0.18 0.015

Lower limit of 95% CI 1.38 1.14 1.39 1.18 1.09 1.15 0.924 0.869 1.13 Upper limit of 95% CI 3.56 2.83 3.68 2.87 2.62 3.16 2.21 2.08 3.23

OR Quartile 4 3.02 2.55 2.19 2.71 2.36 1.73 1.83 1.66 1.75 p Value 5.8E-4 0.0016 0.0032 3.8E-4 0.0016 0.049 0.021 0.049 0.050

Lower limit of 95% CI 1.61 1.43 1.30 1.56 1.38 1.00 1.10 1.00 1.00 Upper limit of 95% CI 5.68 4.57 3.69 4.68 4.01 2.99 3.04 2.76 3.05

[00297] Table 17.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00298] Table 17.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 2820 3520 2910 3560 3090 3500

Average 3130 4230 3240 4340 3440 4250

Stdev 1540 2680 1520 2840 1810 2820 p (t-test) 1.1E-4 3.6E-5 0.0024

Min 1080 863 1080 863 1080 863

Max 9100 19700 9100 19700 14500 19700 n (Patient) 106 221 140 187 155 172 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 2870 3510 2930 3520 3100 3480

Average 3210 4230 3260 4340 3450 4250

Stdev 1540 2740 1520 2870 1800 2850 p (t-test) 2.1E-4 5.4E-5 0.0027

Min 1080 863 1080 863 1080 863

Max 9100 19700 9100 19700 14500 19700 n (Patient) 118 208 144 182 158 168

UO only

Specificity 87% 86% 80% 84% 83% 78% 80% 80% 78%

OR Quartile 2 2.77 2.48 2.48 2.35 2.33 2.44 1.82 1.84 2.70 p Value 1.1E-4 4.9E-4 0.0026 0.0010 0.0012 0.0048 0.020 0.019 0.0028

Lower limit of 95%

CI 1.65 1.49 1.37 1.41 1.40 1.31 1.10 1.11 1.41

Upper limit of 95% CI 4.65 4.14 4.50 3.91 3.88 4.53 3.02 3.06 5.16

OR Quartile 3 2.11 1.80 1.81 1.76 1.65 1.57 1.33 1.28 1.60 p Value 0.0020 0.012 0.012 0.012 0.026 0.062 0.20 0.27 0.057

Lower limit of 95%

CI 1.31 1.14 1.14 1.13 1.06 0.978 0.858 0.828 0.986

Upper limit of 95% CI 3.39 2.85 2.87 2.74 2.56 2.52 2.05 1.98 2.58

OR Quartile 4 2.92 2.70 2.03 2.53 2.34 1.82 1.69 1.67 1.79 p Value 8.7E-4 1.0E-3 0.0066 9.0E-4 0.0020 0.024 0.045 0.049 0.031

Lower limit of 95%

CI 1.55 1.49 1.22 1.46 1.37 1.08 1.01 1.00 1.06

Upper limit of 95% CI 5.49 4.88 3.39 4.39 4.00 3.07 2.81 2.78 3.03

[00299] Table 17.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort Cohort Cohort Cohort Cohort Cohort

Median 3010 3910 3050 3860 3040 3890

Average 3430 4580 3470 4620 3470 4670

Stdev 2030 2810 2010 2900 2020 2910 p (t-test) 2.7E-5 3.4E-5 2.0E-5

Min 893 863 893 863 893 863

Max 19700 18800 19700 18800 19700 18800 n (Patient) 204 121 214 111 219 106

[00300] Example 18. Use of C-C motif chemokine 15 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00301] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 15 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00302] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00303] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00304] Table 18.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

p (t-test) 0.0046 0.0054 0.0076

Min 0.0116 0.0116 0.0116 0.0116 0.0187 0.0116

Max 1300 53500 1300 53500 1300 53500 n (Patient) 76 239 74 241 70 245

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 118 248 104 275 103 237

Average 1880 1470 1750 1690 1460 1980

Stdev 6590 3060 6510 3970 5360 5640 p (t-test) 0.53 0.92 0.41

Min 0.0116 0.0116 0.0187 0.0116 0.0187 0.0116

Max 53500 19200 53500 30000 52500 53500 n (Patient) 193 121 172 142 155 159

[00305] Table 18.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.72 0.73 0.55 0.73 0.73 0.53 0.73 0.73 0.54

SE 0.029 0.028 0.035 0.030 0.028 0.034 0.030 0.029 0.033 p Value 3.0E-14 0 0.18 3.1E-14 6.7E-16 0.33 8.2E-15 3.1E-15 0.23 nCohort Recovered 90 106 211 79 104 193 71 99 177 nCohort Non- recovered 226 210 103 237 212 121 245 217 137

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 81 % 84% 79% 81 % 83% 78% 82% 83% 79%

Specificity 36% 38% 24% 38% 38% 24% 41 % 37% 25%

Cutoff Quartile 3 155 155 155 155 155 155 155 155 155

Sensitivity 61 % 63% 57% 60% 62% 56% 58% 61 % 55%

Specificity 78% 75% 54% 80% 75% 54% 79% 75% 54%

Cutoff Quartile 4 909 909 902 909 909 902 909 909 902

Sensitivity 33% 35% 28% 32% 34% 26% 31 % 34% 26%

Specificity 96% 94% 76% 95% 94% 75% 96% 95% 76%

OR Quartile 2 2.42 3.14 1.17 2.69 3.03 1.09 3.07 2.90 1.23 p Value 0.0015 3.1E-5 0.58 5.5E-4 5.2E-5 0.76 1.3E-4 1.1E-4 0.44

Lower limit of 95%

CI 1.40 1.83 0.666 1.53 1.77 0.634 1.73 1.69 0.723

Upper limit of 95% CI 4.17 5.37 2.07 4.70 5.19 1.87 5.44 4.98 2.10

OR Quartile 3 5.49 5.21 1.55 5.89 4.95 1.50 5.23 4.69 1.40 p Value 3.3E-9 6.5E-10 0.072 1.0E-8 2.1E-9 0.083 2.0E-7 1.1E-8 0.14

Lower limit of 95%

CI 3.12 3.08 0.962 3.21 2.93 0.949 2.80 2.76 0.896

Upper limit of 95% CI 9.65 8.79 2.49 10.8 8.36 2.37 9.77 7.96 2.19

OR Quartile 4 10.7 8.88 1.26 8.68 8.58 1.04 10.2 9.73 1.11 p Value 8.1E-6 9.0E-7 0.39 4.8E-5 1.3E-6 0.88 1.3E-4 2.2E-6 0.69

Lower limit of 95%

CI 3.77 3.72 0.740 3.06 3.59 0.617 3.11 3.79 0.665

Upper limit of 95% CI 30.2 21.2 2.15 24.6 20.5 1.75 33.4 25.0 1.85

[00306] Table 18.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 36.6 423 39.4 403 36.6 366

Average 365 2620 373 2580 370 2530

Stdev 1060 6900 1070 6850 1080 6770 p (t-test) 3.0E-4 4.3E-4 6.4E-4

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 7940 53500 7940 53500 7940 53500 n (Patient) 127 189 124 192 119 197

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 118 338 118 237 115 222

Average 1870 1420 1950 1310 1740 1700

Stdev 6330 3100 6480 2970 5520 5490 p (t-test) 0.50 0.33 0.95

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 19200 53500 19200 52500 53500 n (Patient) 213 101 203 111 189 125

p Value 1.4E-6 3.7E-7 0.32 6.7E-6 7.9E-7 0.77 2.4E-5 1.0E-6 0.68

Lower limit of 95%

CI 2.95 2.98 0.769 2.61 2.84 0.637 2.45 2.89 0.664

Upper limit of 95% CI 13.0 11.8 2.24 11.5 11.2 1.84 11.6 12.0 1.87

[00307] Table 18.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.70 0.71 0.55 0.69 0.70 0.54 0.69 0.70 0.54

SE 0.029 0.029 0.035 0.030 0.029 0.034 0.030 0.029 0.034 p Value 2.3E-12 2.8E-13 0.13 2.5E-10 6.1E-12 0.25 2.2E-10 7.9E-12 0.19 nCohort Recovered 134 140 215 125 138 206 114 135 193 nCohort Non- recovered 182 176 99 191 178 108 202 181 121

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 84% 85% 81 % 83% 84% 81 % 84% 83% 81 %

Specificity 33% 34% 25% 34% 33% 25% 36% 33% 26%

Cutoff Quartile 3 155 155 155 155 155 155 155 155 155

Sensitivity 64% 65% 58% 62% 64% 56% 61 % 64% 55%

Specificity 69% 69% 53% 69% 68% 53% 69% 68% 53%

Cutoff Quartile 4 909 909 902 909 909 902 909 909 902

Sensitivity 37% 38% 28% 35% 37% 26% 34% 37% 26%

Specificity 91 % 91 % 76% 90% 91 % 75% 90% 91 % 76%

OR Quartile 2 2.48 2.79 1.41 2.51 2.49 1.40 2.88 2.43 1.49 p Value 8.4E-4 1.9E-4 0.25 6.7E-4 8.3E-4 0.25 1.1E-4 0.0010 0.16

Lower limit of 95%

CI 1.45 1.63 0.785 1.48 1.46 0.790 1.69 1.43 0.854

Upper limit of 95% CI 4.22 4.78 2.54 4.28 4.24 2.48 4.91 4.14 2.60

OR Quartile 3 4.08 4.01 1.56 3.64 3.81 1.40 3.51 3.73 1.42 p Value 7.1E-9 8.1E-9 0.069 1.2E-7 2.7E-8 0.15 4.5E-7 4.6E-8 0.13

Lower limit of 95%

CI 2.54 2.50 0.965 2.26 2.38 0.880 2.16 2.33 0.899

Upper limit of 95% CI 6.57 6.43 2.52 5.88 6.10 2.24 5.73 5.98 2.24

OR Quartile 4 5.92 5.86 1.27 5.09 5.67 1.06 4.75 6.02 1.12 p Value 1.6E-7 8.5E-8 0.39 1.6E-6 1.5E-7 0.82 8.7E-6 1.2E-7 0.68

Lower limit of 95%

CI 3.05 3.07 0.740 2.62 2.97 0.624 2.39 3.10 0.663

Upper limit of 95% CI 11.5 11.2 2.17 9.90 10.8 1.81 9.44 11.7 1.88

[00308] Table 18.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95%

CI 1.69 2.39 1.11 1.64 2.33 1.01 1.64 2.27 1.04

Upper limit of 95% CI 4.29 6.08 2.88 4.13 5.91 2.60 4.13 5.74 2.65

OR Quartile 4 2.25 4.98 1.54 2.13 4.72 1.45 2.13 4.64 1.47 p Value 0.0021 8.1E-8 0.11 0.0041 2.0E-7 0.16 0.0041 2.8E-7 0.15

Lower limit of 95%

CI 1.34 2.77 0.909 1.27 2.63 0.858 1.27 2.59 0.872

Upper limit of 95% CI 3.77 8.95 2.61 3.57 8.49 2.46 3.57 8.34 2.48

[00309] Table 18.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 62.8 513 62.8 276 62.8 276

Max 13800 21200 13800 21200 11800 21200 n (Patient) 91 59 84 66 77 73

[00310] Table 18.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95% CI 1.96 1.41 0.725 1.55 1.33 0.767 1.66 1.11 0.950 Upper limit of 95% CI 9.89 6.20 2.84 7.93 5.87 2.89 9.63 4.97 3.51

OR Quartile 4 3.97 3.48 0.899 3.36 3.34 0.936 4.03 2.96 1.24 p Value 0.015 0.016 0.79 0.033 0.020 0.86 0.029 0.037 0.56

Lower limit of 95% CI 1.31 1.26 0.409 1.10 1.21 0.437 1.15 1.07 0.594 Upper limit of 95% CI 12.0 9.62 1.98 10.2 9.26 2.00 14.1 8.23 2.61

[00311] Table 18.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 98 52 94 56 86 64

[00312] Table 18.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 5.02 3.88 3.99 4.96 3.68 3.86 6.26 3.31 4.19

OR Quartile 4 1.49 1.60 1.05 1.45 1.54 1.05 1.63 1.43 1.29 p Value 0.32 0.24 0.90 0.36 0.28 0.91 0.25 0.37 0.50

Lower limit of 95% CI 0.682 0.734 0.489 0.654 0.708 0.492 0.702 0.657 0.616

Upper limit of 95% CI 3.25 3.49 2.25 3.23 3.36 2.22 3.80 3.13 2.71

[00313] Table 18.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.50 0.56 0.54 0.51 0.55 0.53 0.51 0.55 0.53

SE 0.047 0.047 0.049 0.047 0.047 0.049 0.047 0.047 0.049 p Value 0.92 0.23 0.44 0.91 0.31 0.59 0.91 0.31 0.52 nCohort Recovered 81 67 95 80 65 93 80 65 91 nCohort Non- recovered 70 84 55 71 86 57 71 86 59

Cutoff Quartile 2 2740 2740 2740 2740 2740 2740 2740 2740 2740

Sensitivity 73% 77% 75% 73% 77% 74% 73% 77% 75%

Specificity 23% 28% 25% 24% 28% 25% 24% 28% 25%

Cutoff Quartile 3 4230 4230 4260 4230 4230 4260 4230 4230 4260

Sensitivity 53% 57% 56% 54% 57% 56% 54% 57% 56%

Specificity 52% 58% 54% 52% 58% 54% 52% 58% 54%

Cutoff Quartile 4 6010 6010 6060 6010 6010 6060 6010 6010 6060

Sensitivity 24% 26% 24% 24% 26% 23% 24% 26% 24%

Specificity 74% 76% 74% 74% 75% 73% 74% 75% 74%

OR Quartile 2 0.823 1.35 0.990 0.852 1.26 0.920 0.852 1.26 0.992 p Value 0.60 0.42 0.98 0.67 0.53 0.83 0.67 0.53 0.98

Lower limit of 95% CI 0.394 0.648 0.462 0.408 0.604 0.433 0.408 0.604 0.467 Upper limit of 95% CI 1.72 2.83 2.12 1.78 2.65 1.96 1.78 2.65 2.11

OR Quartile 3 1.21 1.86 1.50 1.27 1.86 1.49 1.27 1.86 1.48 p Value 0.56 0.062 0.24 0.46 0.061 0.24 0.46 0.061 0.24

Lower limit of 95% CI 0.636 0.970 0.767 0.671 0.971 0.767 0.671 0.971 0.766 Upper limit of 95% CI 2.29 3.56 2.92 2.41 3.58 2.89 2.41 3.58 2.86

OR Quartile 4 0.916 1.13 0.867 0.884 1.05 0.804 0.884 1.05 0.869 p Value 0.82 0.75 0.72 0.74 0.89 0.58 0.74 0.89 0.72

Lower limit of 95% CI 0.438 0.538 0.401 0.423 0.500 0.372 0.423 0.500 0.406 Upper limit of 95% CI 1.92 2.38 1.87 1.85 2.21 1.74 1.85 2.21 1.86

[00314] Example 19. Use of C-C motif chemokine 15 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00315] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 15 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00316] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00317] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00318] Table 19.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Average 1780 1580 1880 1420 1670 1810

Stdev 6210 3290 6400 3100 5420 5670 p (t-test) 0.77 0.49 0.83

Min 0.0116 0.0116 0.0187 0.0116 0.0187 0.0116

Max 53500 19200 53500 19200 52500 53500 n (Patient) 221 93 207 107 197 117

[00319] Table 19.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Average 874 2340 888 2320 908 2270

Stdev 4000 6310 4030 6280 4090 6220 p (t-test) 0.018 0.022 0.029

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 34200 53500 34200 53500 34200 53500 n (Patient) 135 181 133 183 129 187 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 48.7 436 49.4 434 51.3 434

Average 836 2490 841 2480 847 2460

Stdev 3820 6530 3840 6510 3850 6490 p (t-test) 0.0072 0.0080 0.0089

Min 0.01 16 0.0116 0.0116 0.0116 0.0116 0.0116

Max 34200 53500 34200 53500 34200 53500 n (Patient) 148 168 147 169 146 170

UO only

OR Quartile 2 2.62 3.06 1.40 2.52 3.11 1.31 2.34 2.93 1.18 p Value 4.1E-4 6.7E-5 0.28 6.7E-4 5.2E-5 0.37 0.0017 1.1E-4 0.55

Lower limit of 95%

CI 1.54 1.77 0.763 1.48 1.80 0.726 1.38 1.70 0.677

Upper limit of 95% CI 4.47 5.31 2.58 4.29 5.40 2.36 3.97 5.06 2.07

OR Quartile 3 3.95 3.73 1.95 3.75 3.63 1.78 3.59 3.54 1.57 p Value 1.4E-8 3.1E-8 0.0090 4.4E-8 5.6E-8 0.020 1.3E-7 1.0E-7 0.058

Lower limit of 95%

CI 2.46 2.34 1.18 2.34 2.28 1.09 2.23 2.22 0.985

Upper limit of 95% CI 6.35 5.95 3.22 6.02 5.79 2.91 5.77 5.63 2.52

OR Quartile 4 4.38 4.51 1.45 4.23 4.43 1.28 3.95 4.36 1.23 p Value 2.8E-6 6.9E-7 0.18 4.6E-6 9.2E-7 0.38 1.3E-5 1.2E-6 0.44

Lower limit of 95%

CI 2.36 2.49 0.838 2.28 2.45 0.740 2.13 2.40 0.726

Upper limit of 95% CI 8.11 8.17 2.51 7.84 8.03 2.20 7.33 7.89 2.09

[00320] Table 19.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 115 354 113 351 115 261

Average 1790 1550 1800 1530 1640 1900

Stdev 6200 3230 6210 3220 5260 5980 p (t-test) 0.72 0.69 0.69

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 19200 53500 19200 52500 53500 n (Patient) 223 91 222 92 211 103

[00321] Table 19.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 53.2 555 54.2 544 57.7 492

Average 856 2680 871 2630 893 2560

Stdev 3670 6870 3700 6810 3740 6740 p (t-test) 0.0030 0.0043 0.0067

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 34200 53500 34200 53500 34200 53500 n (Patient) 167 149 164 152 160 156 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 55.3 555 55.7 553 56.1 550

Average 847 2740 852 2720 857 2700

Stdev 3620 6950 3630 6930 3640 6910 p (t-test) 0.0021 0.0024 0.0027

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 34200 53500 34200 53500 34200 53500 n (Patient) 171 145 170 146 169 147

UO only

Specificity 66% 65% 55% 65% 65% 54% 64% 64% 54%

Cutoff Quartile 4 909 909 902 909 909 902 909 909 902

Sensitivity 38% 39% 29% 38% 38% 29% 37% 38% 28%

Specificity 87% 87% 77% 87% 86% 76% 86% 86% 76%

OR Quartile 2 2.41 2.65 1.49 2.34 2.70 1.38 2.14 2.53 1.34 p Value 0.0018 7.1E-4 0.20 0.0024 5.8E-4 0.29 0.0057 0.0012 0.32

Lower limit of 95%

CI 1.39 1.51 0.811 1.35 1.53 0.759 1.25 1.44 0.751

Upper limit of 95% CI 4.19 4.67 2.73 4.05 4.74 2.51 3.68 4.41 2.40

OR Quartile 3 4.06 3.86 1.86 3.72 3.74 1.80 3.32 3.63 1.55 p Value 4.9E-9 1.6E-8 0.014 3.2E-8 3.1E-8 0.020 3.4E-7 5.6E-8 0.071

Lower limit of 95%

CI 2.54 2.41 1.14 2.33 2.35 1.10 2.09 2.28 0.963

Upper limit of 95% CI 6.49 6.16 3.06 5.92 5.97 2.94 5.26 5.79 2.50

OR Quartile 4 4.08 4.05 1.36 3.87 3.98 1.33 3.61 3.91 1.22 p Value 7.4E-7 6.9E-7 0.27 1.8E-6 9.4E-7 0.31 5.9E-6 1.3E-6 0.47

Lower limit of 95%

CI 2.34 2.33 0.787 2.22 2.29 0.771 2.07 2.25 0.712

Upper limit of 95% CI 7.13 7.03 2.34 6.76 6.90 2.29 6.29 6.78 2.09

[00322] Table 19.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00323] Table 19.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 79% 78% 76% 79% 78% 75% 78% 77% 76% Specificity 35% 31 % 26% 33% 31 % 26% 33% 29% 26%

Cutoff Quartile 3 4230 4260 4260 4230 4260 4260 4230 4260 4260

Sensitivity 58% 57% 52% 57% 57% 53% 57% 57% 56%

Specificity 65% 63% 51 % 65% 63% 52% 65% 63% 54%

Cutoff Quartile 4 6010 6060 6060 6010 6060 6060 6010 6060 6060

Sensitivity 31 % 33% 24% 31 % 33% 23% 30% 32% 25%

Specificity 88% 88% 74% 88% 88% 73% 87% 88% 75%

OR Quartile 2 2.05 1.54 1.12 1.84 1.54 1.07 1.73 1.38 1.15 p Value 0.064 0.27 0.79 0.12 0.27 0.87 0.17 0.41 0.72

Lower limit of 95%

CI 0.959 0.721 0.498 0.858 0.721 0.493 0.798 0.643 0.539

Upper limit of 95% CI 4.38 3.27 2.50 3.95 3.27 2.32 3.73 2.95 2.46

OR Quartile 3 2.58 2.32 1.13 2.45 2.32 1.19 2.50 2.19 1.48 p Value 0.0086 0.017 0.72 0.013 0.017 0.61 0.013 0.026 0.24

Lower limit of 95%

CI 1.27 1.16 0.566 1.20 1.16 0.609 1.22 1.10 0.766

Upper limit of 95% CI 5.24 4.63 2.27 4.97 4.63 2.33 5.13 4.39 2.86

OR Quartile 4 3.28 3.72 0.896 3.15 3.72 0.799 2.92 3.58 1.01 p Value 0.014 0.0067 0.79 0.018 0.0067 0.58 0.027 0.0085 0.98

Lower limit of 95%

CI 1.27 1.44 0.400 1.22 1.44 0.365 1.13 1.38 0.475

Upper limit of 95% CI 8.48 9.61 2.01 8.17 9.61 1.75 7.58 9.26 2.14

[00324] Table 19.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Min 703 62.8 703 62.8 703 62.8

Max 11800 21200 11800 21200 11800 21200 n (Patient) 67 84 67 84 66 85

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 4060 4710 4050 4710 3930 4780

Average 4550 5020 4540 5010 4420 5170

Stdev 2710 3580 2730 3500 2620 3520 p (t-test) 0.37 0.37 0.14

Min 62.8 276 62.8 276 62.8 276

Max 13800 21200 13800 21200 13800 21200 n (Patient) 105 45 103 47 95 55

[00325] Table 19.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 73 74 105 72 74 103 70 74 96 nCohort Non- recovered 78 77 45 79 77 47 81 77 54

Cutoff Quartile 2 2740 2740 2740 2740 2740 2740 2740 2740 2740

Sensitivity 77% 75% 73% 76% 75% 72% 77% 75% 74%

Specificity 27% 26% 25% 26% 26% 24% 27% 26% 25%

Cutoff Quartile 3 4230 4230 4260 4230 4230 4260 4230 4230 4260

Sensitivity 60% 58% 62% 59% 58% 62% 60% 58% 63%

Specificity 60% 58% 55% 60% 58% 55% 61 % 58% 57%

Cutoff Quartile 4 6010 6010 6060 6010 6010 6060 6010 6010 6060

Sensitivity 29% 30% 29% 29% 30% 28% 28% 30% 28%

Specificity 79% 80% 76% 79% 80% 76% 79% 80% 76%

OR Quartile 2 1.26 1.05 0.905 1.13 1.05 0.838 1.22 1.05 0.952 p Value 0.54 0.89 0.81 0.74 0.89 0.66 0.60 0.89 0.90

Lower limit of 95% CI 0.602 0.506 0.408 0.543 0.506 0.384 0.582 0.506 0.444 Upper limit of 95% CI 2.63 2.20 2.01 2.36 2.20 1.83 2.54 2.20 2.04

OR Quartile 3 2.30 1.95 2.03 2.18 1.95 2.00 2.44 1.95 2.28 p Value 0.012 0.043 0.052 0.019 0.043 0.055 0.0077 0.043 0.018

Lower limit of 95% CI 1.20 1.02 0.994 1.14 1.02 0.987 1.27 1.02 1.15 Upper limit of 95% CI 4.42 3.73 4.15 4.18 3.73 4.04 4.70 3.73 4.52

OR Quartile 4 1.62 1.68 1.30 1.56 1.68 1.19 1.45 1.68 1.22 p Value 0.21 0.18 0.51 0.24 0.18 0.66 0.33 0.18 0.61

Lower limit of 95% CI 0.765 0.793 0.593 0.739 0.793 0.546 0.688 0.793 0.572 Upper limit of 95% CI 3.42 3.54 2.85 3.30 3.54 2.61 3.07 3.54 2.60

[00326] Table 19.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

p (t-test) 0.15 0.15 0.15

Min 703 62.8 703 62.8 703 62.8

Max 11800 21200 11800 21200 11800 21200 n (Patient) 77 74 77 74 77 74

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 3930 4820 3930 4820 3920 4840

Average 4460 5160 4460 5160 4380 5230

Stdev 2670 3540 2670 3540 2600 3530 p (t-test) 0.18 0.18 0.092

Min 62.8 276 62.8 276 62.8 276

Max 13800 21200 13800 21200 13800 21200 n (Patient) 100 50 100 50 95 55

[00327] Table 19.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 89 79 98 88 79 98 87 79 96 nCohort Non- recovered 62 72 52 63 72 52 64 72 54

Cutoff Quartile 2 2740 2740 2740 2740 2740 2740 2740 2740 2740

Sensitivity 74% 78% 75% 73% 78% 75% 73% 78% 74%

Specificity 25% 28% 26% 24% 28% 26% 24% 28% 25%

Cutoff Quartile 3 4230 4230 4260 4230 4230 4260 4230 4230 4260

Sensitivity 56% 58% 60% 56% 58% 60% 56% 58% 59%

Specificity 54% 57% 55% 53% 57% 55% 54% 57% 55%

Cutoff Quartile 4 6010 6010 6060 6010 6010 6060 6010 6010 6060

Sensitivity 24% 28% 25% 24% 28% 25% 23% 28% 24%

Specificity 74% 77% 74% 74% 77% 74% 74% 77% 74%

OR Quartile 2 0.944 1.35 1.03 0.848 1.35 1.03 0.880 1.35 0.952 p Value 0.88 0.43 0.95 0.66 0.43 0.95 0.73 0.43 0.90

Lower limit of 95% CI 0.448 0.643 0.473 0.404 0.643 0.473 0.419 0.643 0.444 Upper limit of 95% CI 1.99 2.84 2.23 1.78 2.84 2.23 1.85 2.84 2.04

OR Quartile 3 1.52 1.85 1.81 1.43 1.85 1.81 1.51 1.85 1.79 p Value 0.21 0.061 0.088 0.28 0.061 0.088 0.21 0.061 0.090

Lower limit of 95% CI 0.790 0.971 0.916 0.748 0.971 0.916 0.789 0.971 0.912 Upper limit of 95% CI 2.91 3.54 3.58 2.74 3.54 3.58 2.89 3.54 3.52

OR Quartile 4 0.916 1.30 0.973 0.883 1.30 0.973 0.852 1.30 0.900 p Value 0.82 0.48 0.95 0.75 0.48 0.95 0.67 0.48 0.79

Lower limit of 95% CI 0.432 0.624 0.449 0.417 0.624 0.449 0.403 0.624 0.416 Upper limit of 95% CI 1.94 2.72 2.11 1.87 2.72 2.11 1.80 2.72 1.95

[00328] Example 20. Use of C-C motif chemokine 15 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00329] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 15 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00330] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent". [00331] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00332] Table 20.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

UO UO UO

AUC 0.70 0.71 0.58 0.73 0.74 0.60 0.72 0.72 0.61

SE 0.032 0.032 0.050 0.033 0.033 0.052 0.035 0.035 0.052 p Value 3.6E-10 2.6E-11 0.12 4.4E-12 2.8E-13 0.065 1.0E-9 6.9E-10 0.034 nCohort Non- persistent 204 207 274 220 222 277 235 234 278 nCohort Persistent 112 108 40 96 93 37 81 81 36

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 88% 90% 82% 90% 91 % 84% 90% 93% 86%

Specificity 29% 30% 24% 29% 30% 24% 28% 29% 24%

Cutoff Quartile 3 155 154 155 155 154 155 155 154 155

Sensitivity 71 % 71 % 68% 75% 75% 70% 77% 74% 72%

Specificity 61 % 61 % 53% 61 % 60% 53% 59% 58% 53%

Cutoff Quartile 4 909 912 902 909 912 902 909 912 902

Sensitivity 42% 44% 32% 46% 48% 35% 46% 47% 36%

Specificity 84% 85% 76% 84% 85% 76% 82% 82% 76%

OR Quartile 2 2.92 3.86 1.50 3.53 4.50 1.65 3.56 5.12 2.01 p Value 9.7E-4 1.3E-4 0.36 5.6E-4 1.6E-4 0.29 0.0015 2.7E-4 0.16

Lower limit of 95%

CI 1.54 1.93 0.632 1.72 2.06 0.659 1.63 2.13 0.751

Upper limit of 95% CI 5.51 7.69 3.54 7.22 9.81 4.12 7.80 12.3 5.37

OR Quartile 3 3.79 3.86 2.30 4.67 4.63 2.63 4.72 3.97 2.92 p Value 1.3E-7 1.3E-7 0.020 1.7E-8 3.0E-8 0.011 1.3E-7 1.5E-6 0.0062

Lower limit of 95%

CI 2.31 2.34 1.14 2.74 2.69 1.25 2.66 2.26 1.36

Upper limit of 95% CI 6.21 6.38 4.65 7.99 7.97 5.54 8.41 6.95 6.28

OR Quartile 4 3.89 4.21 1.52 4.47 5.18 1.73 3.86 4.16 1.82 p Value 5.7E-7 1.4E-7 0.25 5.4E-8 3.6E-9 0.14 1.5E-6 4.0E-7 0.11

Lower limit of 95%

CI 2.28 2.47 0.741 2.61 3.00 0.835 2.23 2.40 0.871

Upper limit of 95% CI 6.62 7.20 3.11 7.68 8.95 3.59 6.70 7.22 3.78

[00333] Table 20.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 60.0 577 59.2 717 65.2 616

Average 1140 2610 1090 2900 1460 2300

Stdev 5190 5840 5020 6140 6090 3820 p (t-test) 0.021 0.0050 0.21

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 52500 53500 52500 53500 19200 n (Patient) 191 124 205 110 217 98

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 118 406 118 488 118 521

Average 1790 1450 1760 1530 1760 1560

Stdev 5950 2870 5910 2960 5900 2980 p (t-test) 0.67 0.78 0.81

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 18400 53500 18400 53500 18400 n (Patient) 255 59 259 55 260 54

CI

Upper limit of 95% CI 6.59 7.54 2.61 7.56 8.71 2.97 5.80 6.22 3.08

[00334] Table 20.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.71 0.72 0.59 0.74 0.74 0.61 0.71 0.72 0.61

SE 0.029 0.029 0.040 0.029 0.030 0.041 0.031 0.032 0.041 p Value 3.3E-13 4.0E-14 0.018 4.4E-16 2.2E-16 0.0070 1.2E-11 l . lE-11 0.0070 nCohort Non- persistent 174 181 245 189 194 250 204 207 250 nCohort Persistent 142 134 69 127 121 64 112 108 64

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 87% 89% 84% 89% 91 % 86% 88% 91 % 86%

Specificity 32% 33% 25% 32% 32% 26% 30% 31 % 26%

Cutoff Quartile 3 155 154 155 155 154 155 155 154 155

Sensitivity 70% 70% 65% 72% 73% 67% 72% 71 % 67%

Specificity 66% 65% 54% 65% 64% 54% 62% 61 % 54%

Cutoff Quartile 4 909 912 902 909 912 902 909 912 902

Sensitivity 40% 43% 30% 44% 46% 33% 42% 44% 33%

Specificity 87% 88% 76% 88% 88% 77% 84% 85% 77%

OR Quartile 2 2.99 3.84 1.79 3.75 4.81 2.10 3.25 4.39 2.10 p Value 2.1E-4 2.2E-5 0.11 4.3E-5 7.9E-6 0.055 3.9E-4 5.0E-5 0.055

Lower limit of 95%

CI 1.68 2.06 0.882 1.99 2.42 0.984 1.69 2.15 0.984

Upper limit of 95% CI 5.34 7.14 3.62 7.08 9.58 4.50 6.23 8.96 4.50

OR Quartile 3 4.49 4.30 2.23 4.90 4.72 2.44 4.31 3.86 2.44 p Value 6.4E-10 2.5E-9 0.0047 2.2E-10 8.4E-10 0.0025 1.1E-8 1.3E-7 0.0025

Lower limit of 95%

CI 2.79 2.66 1.28 3.00 2.88 1.37 2.61 2.34 1.37

Upper limit of 95% CI 7.22 6.94 3.88 8.00 7.76 4.35 7.12 6.38 4.35

OR Quartile 4 4.63 5.35 1.41 5.69 6.41 1.62 3.89 4.21 1.62 p Value 7.6E-8 5.0E-9 0.25 1.1E-9 l .OE-10 0.12 5.7E-7 1.4E-7 0.12

Lower limit of 95%

CI 2.65 3.05 0.781 3.25 3.65 0.888 2.28 2.47 0.888

Upper limit of 95% CI 8.10 9.39 2.55 9.96 11.2 2.94 6.62 7.20 2.94

[00335] Table 20.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 56.1 585 55.3 701 61.1 616

Average 1080 2560 1030 2790 1150 2740

Stdev 5270 5680 5110 5900 5060 6090 p (t-test) 0.017 0.0053 0.014

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 52500 53500 52500 53500 52500 n (Patient) 179 136 191 124 203 112

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 116 351 116 380 116 380

Average 1750 1650 1710 1760 1710 1760

Stdev 6010 3470 5940 3590 5940 3590 p (t-test) 0.90 0.95 0.95

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 19200 53500 19200 53500 19200 n (Patient) 238 76 244 70 244 70

Upper limit of 95% CI 8.53 9.03 2.50 9.76 10.5 2.70 6.55 7.11 2.70

[00336] Table 20.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.72 0.72 0.62 0.75 0.74 0.62 0.72 0.72 0.62

SE 0.029 0.029 0.037 0.029 0.029 0.038 0.031 0.031 0.038 p Value 3.4E-14 1.0E-13 0.0019 0 2.2E-16 0.0012 2.3E-12 3.5E-12 0.0012 nCohort Non- persistent 168 175 233 183 187 236 196 199 236 nCohort Persistent 148 140 81 133 128 78 120 116 78

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 86% 87% 85% 89% 89% 86% 89% 89% 86%

Specificity 32% 32% 26% 33% 32% 26% 31 % 31 % 26%

Cutoff Quartile 3 155 154 155 155 154 155 155 154 155

Sensitivity 69% 69% 68% 72% 71 % 69% 71 % 70% 69%

Specificity 67% 65% 56% 66% 64% 56% 63% 61 % 56%

Cutoff Quartile 4 909 912 902 909 912 902 909 912 902

Sensitivity 41 % 42% 32% 44% 45% 33% 42% 43% 33%

Specificity 89% 89% 77% 89% 89% 78% 85% 85% 78%

OR Quartile 2 3.03 3.19 2.04 4.15 3.85 2.17 3.72 3.50 2.17 p Value 1.4E-4 1.1E-4 0.040 1.1E-5 3.1E-5 0.030 7.5E-5 1.6E-4 0.030

Lower limit of 95%

CI 1.71 1.77 1.03 2.20 2.04 1.08 1.94 1.83 1.08

Upper limit of 95% CI 5.37 5.74 4.02 7.82 7.25 4.37 7.13 6.71 4.37

OR Quartile 3 4.43 3.98 2.72 5.06 4.41 2.91 4.09 3.67 2.91 p Value 6.9E-10 1.0E-8 2.4E-4 6.9E-11 2.1E-9 1.3E-4 1.6E-8 1.8E-7 1.3E-4

Lower limit of 95%

CI 2.76 2.48 1.59 3.11 2.71 1.68 2.51 2.25 1.68

Upper limit of 95% CI 7.12 6.38 4.63 8.24 7.16 5.01 6.67 5.98 5.01

OR Quartile 4 5.35 5.65 1.61 6.50 6.55 1.73 4.11 4.44 1.73 p Value 1.4E-8 3.4E-9 0.096 2.0E-10 1.2E-10 0.057 2.3E-7 5.8E-8 0.057

Lower limit of 95%

CI 3.00 3.18 0.919 3.65 3.70 0.985 2.41 2.59 0.985

Upper limit of 95% CI 9.54 10.0 2.81 11.6 11.6 3.03 7.03 7.61 3.03

[00337] Table 20.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 4150 4490 4120 4500 4230 4300

Average 4520 4930 4480 5090 4660 4720

Stdev 2700 3470 2660 3650 2780 3560 p (t-test) 0.42 0.26 0.91

Min 276 62.8 276 62.8 276 62.8

Max 11800 21200 11800 21200 13800 21200 n (Patient) 94 56 103 47 108 42

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 4170 4800 4170 4800 4150 4880

Average 4560 5570 4560 5570 4540 5820

Stdev 2670 4660 2670 4660 2680 4660 p (t-test) 0.17 0.17 0.089

Min 62.8 1120 62.8 1120 62.8 1630

Max 13800 21200 13800 21200 13800 21200 n (Patient) 131 19 131 19 132 18

[00338] Table 20.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 88 88 123 94 93 123 101 99 124 nCohort Persistent 63 62 27 57 57 27 50 51 26

Cutoff Quartile 2 2740 2730 2740 2740 2730 2740 2740 2730 2740

Sensitivity 76% 74% 70% 79% 77% 70% 78% 75% 73%

Specificity 26% 25% 24% 28% 27% 24% 27% 25% 25%

Cutoff Quartile 3 4230 4260 4260 4230 4260 4260 4230 4260 4260

Sensitivity 57% 55% 59% 58% 56% 59% 54% 51 % 62%

Specificity 55% 53% 52% 54% 54% 52% 51 % 51 % 52%

Cutoff Quartile 4 6010 6060 6060 6010 6060 6060 6010 6060 6060

Sensitivity 27% 27% 22% 26% 28% 22% 20% 22% 23%

Specificity 76% 76% 74% 76% 76% 74% 72% 73% 74%

OR Quartile 2 1.13 0.958 0.766 1.43 1.24 0.766 1.29 0.988 0.905 p Value 0.75 0.91 0.57 0.37 0.58 0.57 0.53 0.97 0.84

Lower limit of 95% CI 0.535 0.455 0.304 0.657 0.576 0.304 0.581 0.455 0.347 Upper limit of 95% CI 2.40 2.02 1.93 3.13 2.69 1.93 2.88 2.15 2.36

OR Quartile 3 1.60 1.39 1.58 1.63 1.49 1.58 1.25 1.06 1.76 p Value 0.16 0.32 0.29 0.15 0.24 0.29 0.53 0.86 0.20

Lower limit of 95% CI 0.834 0.725 0.678 0.839 0.767 0.678 0.632 0.540 0.742 Upper limit of 95% CI 3.07 2.67 3.67 3.17 2.89 3.67 2.46 2.09 4.19

OR Quartile 4 1.18 1.21 0.812 1.10 1.26 0.812 0.652 0.733 0.862 p Value 0.66 0.62 0.68 0.80 0.55 0.68 0.31 0.45 0.77

Lower limit of 95% CI 0.562 0.574 0.301 0.519 0.595 0.301 0.287 0.329 0.318 Upper limit of 95% CI 2.47 2.53 2.19 2.34 2.67 2.19 1.48 1.63 2.34

[00339] Table 20.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 11800 21200 11800 21200 13800 21200 n (Patient) 82 68 86 64 93 57

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 4060 4780 4060 4780 4050 4790

Average 4550 5150 4550 5150 4520 5270

Stdev 2750 3690 2750 3690 2750 3680 p (t-test) 0.30 0.30 0.20

Min 62.8 513 62.8 513 62.8 513

Max 13800 21200 13800 21200 13800 21200 n (Patient) 115 35 115 35 116 34

[00340] Table 20.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 60% 58% 63% 59% 58% 62% 57% 55% 64% Specificity 58% 57% 55% 56% 56% 55% 54% 53% 55%

Cutoff Quartile 4 6010 6060 6060 6010 6060 6060 6010 6060 6060

Sensitivity 27% 28% 24% 27% 27% 25% 25% 25% 26%

Specificity 77% 77% 74% 76% 76% 75% 75% 74% 75%

OR Quartile 2 1.45 1.24 1.07 1.46 1.28 1.02 1.37 1.19 1.18 p Value 0.33 0.58 0.87 0.33 0.52 0.95 0.42 0.65 0.71

Lower limit of 95% CI 0.688 0.587 0.466 0.686 0.606 0.445 0.636 0.559 0.500 Upper limit of 95% CI 3.07 2.60 2.46 3.11 2.71 2.36 2.95 2.55 2.78

OR Quartile 3 2.07 1.81 2.12 1.87 1.72 2.00 1.53 1.40 2.18 p Value 0.028 0.073 0.046 0.059 0.10 0.067 0.21 0.32 0.043

Lower limit of 95% CI 1.08 0.947 1.01 0.977 0.899 0.952 0.792 0.725 1.03 Upper limit of 95% CI 3.97 3.47 4.44 3.60 3.31 4.20 2.94 2.69 4.63

OR Quartile 4 1.22 1.24 0.933 1.22 1.20 0.976 0.986 0.971 1.02 p Value 0.60 0.57 0.87 0.60 0.63 0.95 0.97 0.94 0.96

Lower limit of 95% CI 0.582 0.593 0.406 0.583 0.573 0.424 0.465 0.458 0.443 Upper limit of 95% CI 2.54 2.59 2.14 2.55 2.51 2.25 2.09 2.06 2.36

[00341] Table 20.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3930 4790 3930 4790 3930 4800

Average 4500 5170 4500 5170 4470 5270

Stdev 2780 3480 2780 3480 2780 3460 p (t-test) 0.22 0.22 0.15

Min 62.8 513 62.8 513 62.8 513

Max 13800 21200 13800 21200 13800 21200 n (Patient) 108 42 108 42 109 41

[00342] Example 21. Use of C-C motif chemokine 15 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00343] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 15 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents. [00344] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00345] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00346] Table 21.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

n (Patient) 129 186 161 154 183 132

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 108 420 127 434 127 585

Average 1750 1620 1740 1630 1710 1780

Stdev 6070 2990 5930 3050 5860 3210 p (t-test) 0.86 0.88 0.94

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 18400 53500 18400 53500 18400 n (Patient) 240 74 255 59 262 52

[00347] Table 21.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 84% 86% 84% 84% 86% 82% 85% 86% 83%

Specificity 38% 38% 26% 32% 33% 25% 31 % 32% 25%

Cutoff Quartile 3 155 154 155 155 154 155 155 154 155

Sensitivity 64% 65% 63% 64% 65% 65% 68% 69% 67%

Specificity 76% 73% 56% 66% 65% 55% 66% 66% 55%

Cutoff Quartile 4 909 912 902 909 912 902 909 912 902

Sensitivity 35% 36% 31 % 36% 38% 31 % 39% 40% 33%

Specificity 93% 92% 77% 88% 88% 77% 88% 88% 77%

OR Quartile 2 3.29 3.64 1.85 2.51 3.02 1.49 2.53 3.01 1.61 p Value 1.4E-5 3.4E-6 0.055 7.9E-4 9.3E-5 0.23 0.0010 1.6E-4 0.17

Lower limit of 95%

CI 1.92 2.11 0.987 1.47 1.74 0.779 1.46 1.70 0.811

Upper limit of 95% CI 5.64 6.27 3.47 4.31 5.26 2.86 4.40 5.33 3.20

OR Quartile 3 5.51 5.03 2.18 3.54 3.48 2.25 4.17 4.28 2.49 p Value 1.2E-10 1.5E-10 0.0022 1.0E-7 1.4E-7 0.0029 2.6E-9 1.7E-9 0.0014

Lower limit of 95%

CI 3.28 3.07 1.32 2.22 2.19 1.32 2.60 2.66 1.42

Upper limit of 95% CI 9.25 8.24 3.59 5.63 5.53 3.84 6.66 6.86 4.35

OR Quartile 4 6.82 6.56 1.55 4.36 4.53 1.50 4.57 4.96 1.64 p Value 1.2E-6 2.1E-7 0.11 1.2E-6 4.3E-7 0.16 1.8E-7 4.0E-8 0.094

Lower limit of 95%

CI 3.14 3.22 0.903 2.40 2.52 0.848 2.58 2.80 0.919

Upper limit of 95% CI 14.8 13.3 2.66 7.89 8.14 2.65 8.08 8.79 2.94

[00348] Table 21.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 34200 53500 34200 53500 34200 53500 n (Patient) 123 192 150 165 162 153

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 91.3 347 116 351 118 406

Average 1810 1540 1780 1570 1740 1660

Stdev 6340 3150 6180 3250 6100 3360 p (t-test) 0.67 0.76 0.91

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 19200 53500 19200 53500 19200 n (Patient) 211 103 224 90 231 83

[00349] Table 21.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489 0.0489

Sensitivity 84% 85% 83% 83% 85% 81 % 84% 86% 82%

Specificity 38% 37% 27% 32% 33% 25% 32% 32% 25%

Cutoff Quartile 3 155 154 155 155 154 155 155 154 155

Sensitivity 64% 65% 60% 63% 64% 60% 66% 66% 62%

Specificity 76% 72% 55% 66% 65% 54% 66% 65% 55%

Cutoff Quartile 4 909 912 902 909 912 902 909 912 902

Sensitivity 34% 36% 30% 36% 37% 29% 37% 38% 30%

Specificity 93% 92% 77% 88% 88% 77% 88% 88% 77%

OR Quartile 2 3.18 3.24 1.79 2.32 2.76 1.43 2.51 2.86 1.55 p Value 2.5E-5 1.9E-5 0.055 0.0020 2.6E-4 0.24 9.0E-4 2.3E-4 0.17

Lower limit of 95%

CI 1.86 1.89 0.989 1.36 1.60 0.790 1.46 1.64 0.834

Upper limit of 95% CI 5.44 5.56 3.24 3.95 4.77 2.61 4.34 5.01 2.87

OR Quartile 3 5.62 4.77 1.82 3.37 3.30 1.78 3.82 3.68 1.95 p Value l . lE-10 5.4E-10 0.013 3.1E-7 4.3E-7 0.020 1.8E-8 4.2E-8 0.0090

Lower limit of 95%

CI 3.33 2.91 1.14 2.12 2.08 1.09 2.40 2.31 1.18

Upper limit of 95% CI 9.49 7.82 2.93 5.36 5.23 2.91 6.08 5.86 3.22

OR Quartile 4 6.59 6.34 1.45 4.07 4.23 1.38 4.25 4.38 1.45 p Value 1.9E-6 3.5E-7 0.16 3.6E-6 1.4E-6 0.25 8.7E-7 3.8E-7 0.18

Lower limit of 95%

CI 3.04 3.11 0.858 2.25 2.36 0.801 2.39 2.48 0.838

Upper limit of 95% CI 14.3 12.9 2.46 7.37 7.59 2.36 7.57 7.74 2.51

[00350] Table 21.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 34200 53500 34200 53500 34200 53500 n (Patient) 120 195 145 170 157 158

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 81.4 347 88.0 341 97.4 344

Average 1830 1530 1820 1530 1790 1570

Stdev 6480 3070 6350 3120 6280 3180 p (t-test) 0.65 0.66 0.74

Min 0.0116 0.0116 0.0116 0.0116 0.0116 0.0116

Max 53500 19200 53500 19200 53500 19200 n (Patient) 201 113 211 103 216 98

[00351] Table 21.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 2740 2730 2740 2740 2730 2740 2740 2730 2740

Sensitivity 79% 78% 76% 75% 73% 80% 76% 77% 80%

Specificity 34% 31 % 26% 25% 23% 27% 26% 27% 26%

Cutoff Quartile 3 4230 4260 4260 4230 4260 4260 4230 4260 4260

Sensitivity 58% 57% 58% 57% 56% 67% 56% 58% 64%

Specificity 66% 63% 53% 56% 56% 54% 54% 56% 53%

Cutoff Quartile 4 6010 6060 6060 6010 6060 6060 6010 6060 6060

Sensitivity 32% 33% 24% 29% 30% 30% 29% 31 % 28%

Specificity 88% 88% 74% 79% 79% 76% 78% 79% 75%

OR Quartile 2 1.96 1.54 1.13 1.02 0.810 1.45 1.09 1.19 1.43 p Value 0.081 0.27 0.79 0.96 0.58 0.45 0.82 0.65 0.50

Lower limit of 95% CI 0.920 0.721 0.477 0.488 0.387 0.545 0.519 0.563 0.498 Upper limit of 95% CI 4.18 3.27 2.66 2.12 1.70 3.88 2.29 2.52 4.13

OR Quartile 3 2.73 2.32 1.53 1.66 1.62 2.36 1.50 1.73 1.99 p Value 0.0054 0.017 0.26 0.12 0.14 0.045 0.22 0.100 0.13

Lower limit of 95% CI 1.35 1.16 0.728 0.872 0.850 1.02 0.788 0.900 0.818 Upper limit of 95% CI 5.53 4.63 3.22 3.16 3.09 5.47 2.87 3.33 4.84

OR Quartile 4 3.40 3.72 0.888 1.50 1.65 1.34 1.40 1.72 1.18 p Value 0.012 0.0067 0.79 0.28 0.19 0.51 0.37 0.15 0.74

Lower limit of 95% CI 1.31 1.44 0.376 0.715 0.779 0.555 0.671 0.818 0.450 Upper limit of 95% CI 8.80 9.61 2.10 3.16 3.48 3.26 2.94 3.60 3.09

[00352] Table 21.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00353] Table 21.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3620 4600 3930 4580 3920 4690

Average 3690 5130 4300 4980 4240 5090

Stdev 2220 3210 2550 3310 2530 3360 p (t-test) 0.0053 0.17 0.081

Min 62.8 276 62.8 276 62.8 276

Max 10700 21200 11800 21200 11800 21200 n (Patient) 49 102 69 82 75 76 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3730 4670 3940 4500 3930 4670

Average 3820 5120 4370 4930 4300 5040

Stdev 2120 3300 2500 3370 2490 3430 p (t-test) 0.012 0.25 0.13

Min 703 62.8 703 62.8 703 62.8

Max 10700 21200 11800 21200 11800 21200 n (Patient) 51 99 69 81 75 75

UO only

Specificity 88% 88% 77% 80% 80% 78% 80% 80% 77%

OR Quartile 2 1.76 1.38 1.20 0.950 0.810 1.53 1.17 1.00 1.50 p Value 0.14 0.41 0.64 0.89 0.58 0.33 0.67 1.0 0.37

Lower limit of 95% CI 0.824 0.643 0.549 0.454 0.385 0.655 0.561 0.479 0.621 Upper limit of 95% CI 3.77 2.95 2.64 1.99 1.70 3.56 2.45 2.09 3.62

OR Quartile 3 2.58 2.19 2.05 1.85 1.62 3.07 2.06 1.81 2.68 p Value 0.0086 0.026 0.041 0.062 0.14 0.0030 0.029 0.073 0.011

Lower limit of 95% CI 1.27 1.10 1.03 0.969 0.851 1.46 1.08 0.945 1.25 Upper limit of 95% CI 5.24 4.39 4.07 3.54 3.10 6.44 3.94 3.45 5.74

OR Quartile 4 3.28 3.58 1.32 1.63 1.65 1.78 1.74 1.77 1.64 p Value 0.014 0.0085 0.47 0.21 0.19 0.14 0.15 0.14 0.23

Lower limit of 95% CI 1.27 1.38 0.618 0.764 0.777 0.823 0.822 0.837 0.737 Upper limit of 95% CI 8.48 9.26 2.83 3.46 3.52 3.86 3.67 3.74 3.64

[00354] Table 21.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 2750 3330 2710 3410 2680 3530 p (t-test) 0.11 0.041 0.037

Min 62.8 513 62.8 513 62.8 513

Max 13800 21200 13800 21200 13800 21200 n (Patient) 95 55 100 50 105 45

[00355] Table 21.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 10700 21200 11800 21200 11800 21200 n (Patient) 47 104 65 86 70 81 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 3700 4680 3930 4580 3930 4690

Average 3790 5120 4360 4920 4290 5030

Stdev 2130 3280 2540 3320 2520 3370 p (t-test) 0.010 0.26 0.14

Min 703 62.8 703 62.8 703 62.8

Max 10700 21200 11800 21200 11800 21200 n (Patient) 50 100 66 84 72 78

UO only

Lower limit of 95% CI 1.29 1.17 0.906 0.971 0.899 1.08 1.02 0.998 0.974 Upper limit of 95% CI 5.42 4.75 3.43 3.58 3.31 4.20 3.74 3.65 3.85

OR Quartile 4 3.04 3.45 1.05 1.41 1.49 1.17 1.45 1.59 1.01 p Value 0.022 0.011 0.91 0.37 0.31 0.68 0.33 0.23 0.97

Lower limit of 95% CI 1.17 1.33 0.492 0.662 0.697 0.551 0.688 0.752 0.466 Upper limit of 95% CI 7.87 8.93 2.22 3.00 3.17 2.50 3.07 3.36 2.20

[00356] Example 22. Use of C-C motif chemokine 18 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00357] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 18 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00358] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00359] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00360] Table 22.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Median 0.870 1.22 0.716 1.23 0.716 1.18

Average 0.858 2.90 0.745 2.86 0.739 2.83

Stdev 0.658 9.70 0.561 9.57 0.588 9.49 p (t-test) 0.32 0.33 0.35

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 2.61 60.0 2.01 60.0 2.01 60.0 n (Patient) 23 109 20 112 18 114 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.797 1.27 0.819 1.26 0.797 1.24

Average 0.783 3.07 0.794 3.04 0.786 3.02

Stdev 0.513 10.1 0.519 10.0 0.527 9.96 p (t-test) 0.22 0.23 0.24

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 2.01 60.0 2.01 60.0 2.01 60.0 n (Patient) 30 101 29 102 28 103

UO only

Specificity 87% 93% 78% 90% 93% 83% 89% 93% 84%

OR Quartile 2 1.79 2.08 4.38 2.32 1.81 5.83 2.85 1.93 4.95 p Value 0.24 0.10 0.0050 0.099 0.20 8.0E-4 0.046 0.15 7.0E-4

Lower limit of 95% CI 0.681 0.862 1.56 0.854 0.738 2.08 1.02 0.784 1.96 Upper limit of 95% CI 4.71 5.01 12.3 6.30 4.43 16.4 7.98 4.76 12.5

OR Quartile 3 2.70 3.02 2.88 3.59 2.81 3.83 2.99 2.62 3.21 p Value 0.044 0.013 0.0052 0.020 0.021 3.6E-4 0.050 0.033 0.0014

Lower limit of 95% CI 1.03 1.26 1.37 1.22 1.17 1.83 1.00 1.08 1.57 Upper limit of 95% CI 7.08 7.25 6.06 10.5 6.78 8.01 8.95 6.33 6.55

OR Quartile 4 2.53 6.20 1.64 3.44 5.89 2.77 2.99 5.60 2.78 p Value 0.16 0.017 0.23 0.11 0.020 0.014 0.16 0.024 0.015

Lower limit of 95% CI 0.701 1.39 0.735 0.755 1.32 1.23 0.649 1.25 1.22 Upper limit of 95% CI 9.14 27.7 3.66 15.7 26.3 6.23 13.8 25.1 6.36

[00361] Table 22.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort Cohort Cohort

Median 0.978 1.39 0.974 1.42 0.983 1.36

Average 1.08 5.69 1.06 5.25 1.04 4.66

Stdev 0.846 15.3 0.838 14.5 0.843 13.4 p (t-test) 0.0051 0.0091 0.020

Min 0.00242 0.273 0.00242 0.273 0.00242 0.273

Max 4.23 60.0 4.23 60.0 4.23 60.0 n (Patient) 89 42 84 47 76 55

[00362] Table 22.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Cohort Cohort Cohort

Median 0.739 1.30 0.700 1.30 0.700 1.28

Average 0.865 3.50 0.851 3.45 0.842 3.34

Stdev 0.690 11.0 0.700 10.9 0.702 10.6 p (t-test) 0.100 0.11 0.13

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 3.02 60.0 3.02 60.0 3.02 60.0 n (Patient) 48 84 46 86 42 90 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.797 1.34 0.819 1.33 0.819 1.28

Average 0.857 3.71 0.864 3.67 0.874 3.59

Stdev 0.614 11.4 0.617 11.3 0.622 11.2 p (t-test) 0.068 0.074 0.086

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 2.07 60.0 2.07 60.0 2.07 60.0 n (Patient) 54 78 53 79 51 81

UO only

Sensitivity 32% 35% 32% 31 % 34% 31 % 31 % 33% 32% Specificity 88% 89% 78% 87% 89% 78% 88% 88% 79%

OR Quartile 2 3.28 2.94 2.50 3.62 2.59 2.97 3.75 2.40 3.29 p Value 0.0043 0.0092 0.066 0.0021 0.020 0.028 0.0017 0.033 0.012

Lower limit of 95%

CI 1.45 1.31 0.942 1.59 1.16 1.12 1.64 1.08 1.30

Upper limit of 95% CI 7.40 6.63 6.64 8.22 5.81 7.87 8.56 5.36 8.28

OR Quartile 3 2.55 2.44 2.17 2.60 2.30 2.40 2.35 2.04 2.25 p Value 0.012 0.014 0.046 0.012 0.022 0.021 0.027 0.051 0.026

Lower limit of 95%

CI 1.23 1.20 1.01 1.24 1.13 1.14 1.10 0.998 1.10

Upper limit of 95% CI 5.32 4.99 4.63 5.47 4.69 5.06 5.02 4.16 4.60

OR Quartile 4 3.32 4.24 1.62 3.05 4.07 1.59 3.34 3.75 1.83 p Value 0.015 0.0035 0.25 0.024 0.0045 0.26 0.022 0.0075 0.14

Lower limit of 95%

CI 1.26 1.61 0.713 1.15 1.54 0.708 1.19 1.42 0.825

Upper limit of 95% CI 8.75 11.2 3.71 8.06 10.7 3.58 9.41 9.88 4.06

[00363] Table 22.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 1.02 1.36 1.00 1.36 1.02 1.35

Average 1.09 5.57 1.08 5.20 1.08 4.67

Stdev 0.825 15.1 0.840 14.5 0.864 13.6 p (t-test) 0.0063 0.010 0.022

Min 0.00242 0.233 0.00242 0.233 0.00242 0.233

Max 4.23 60.0 4.23 60.0 4.23 60.0 n (Patient) 88 43 84 47 77 54

[00364] Table 22.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 1.02 1.35 1.02 1.33 1.02 1.33

Average 1.04 4.93 1.04 4.78 1.04 4.78

Stdev 0.797 13.9 0.805 13.7 0.805 13.7 p (t-test) 0.013 0.017 0.017

Min 0.00242 0.197 0.00242 0.197 0.00242 0.197

Max 4.23 60.0 4.23 60.0 4.23 60.0 n (Patient) 81 51 79 53 79 53 sCr only

UO only

Specificity 32% 34% 30% 33% 34% 31 % 33% 34% 31 %

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 59% 61 % 61 % 58% 61 % 61 % 58% 61 % 62%

Specificity 56% 60% 55% 56% 60% 55% 56% 60% 56%

Cutoff Quartile 4 1.65 1.65 1.66 1.65 1.65 1.66 1.65 1.65 1.66

Sensitivity 35% 34% 34% 34% 34% 33% 34% 34% 32%

Specificity 81 % 84% 79% 81 % 84% 79% 81 % 84% 79%

OR Quartile 2 2.97 2.76 2.25 3.22 2.76 2.46 3.22 2.76 2.56 p Value 0.021 0.018 0.087 0.013 0.018 0.058 0.013 0.018 0.047

Lower limit of 95% CI 1.18 1.19 0.890 1.28 1.19 0.971 1.28 1.19 1.01 Upper limit of 95% CI 7.49 6.40 5.71 8.12 6.40 6.21 8.12 6.40 6.47

OR Quartile 3 1.79 2.37 1.95 1.77 2.37 1.92 1.77 2.37 2.05 p Value 0.11 0.016 0.076 0.11 0.016 0.079 0.11 0.016 0.054

Lower limit of 95% CI 0.879 1.18 0.933 0.876 1.18 0.927 0.876 1.18 0.987 Upper limit of 95% CI 3.63 4.76 4.09 3.58 4.76 3.99 3.58 4.76 4.24

OR Quartile 4 2.40 2.71 1.98 2.19 2.71 1.80 2.19 2.71 1.72 p Value 0.033 0.018 0.098 0.054 0.018 0.15 0.054 0.018 0.19

Lower limit of 95% CI 1.08 1.19 0.881 0.986 1.19 0.804 0.986 1.19 0.769 Upper limit of 95% CI 5.36 6.20 4.46 4.88 6.20 4.04 4.88 6.20 3.84

[00365] Example 23. Use of C-C motif chemokine 18 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00366] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 18 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00367] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00368] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis. [00369] Table 23.1:Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

AUC 0.66 0.67 0.66 0.66 0.67 0.69 0.65 0.66 0.66

SE 0.048 0.047 0.055 0.048 0.047 0.051 0.049 0.047 0.051 p Value 8.0E-4 2.0E-4 0.0031 8.0E-4 2.0E-4 2.0E-4 0.0020 8.6E-4 0.0019 nCohort Recovered 46 55 93 46 55 88 44 53 84 nCohort Non- recovered 86 76 38 86 76 43 88 78 47

Cutoff Quartile 2 0.561 0.557 0.569 0.561 0.557 0.569 0.561 0.557 0.569

Sensitivity 81 % 83% 87% 81 % 83% 88% 81 % 81 % 87%

Specificity 37% 36% 30% 37% 36% 32% 36% 34% 32%

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 58% 61 % 68% 58% 61 % 70% 57% 59% 64%

Specificity 65% 64% 57% 65% 64% 59% 64% 62% 57%

Cutoff Quartile 4 1.65 1.64 1.66 1.65 1.64 1.66 1.65 1.64 1.66

Sensitivity 29% 33% 34% 29% 33% 37% 28% 32% 34%

Specificity 83% 85% 78% 83% 85% 81 % 82% 85% 80%

OR Quartile 2 2.56 2.77 2.84 2.56 2.77 3.55 2.39 2.16 3.24 p Value 0.022 0.014 0.049 0.022 0.014 0.016 0.035 0.059 0.018

Lower limit of 95%

CI 1.14 1.23 1.01 1.14 1.23 1.26 1.06 0.970 1.23

Upper limit of 95% CI 5.76 6.23 8.04 5.76 6.23 9.98 5.37 4.81 8.55

OR Quartile 3 2.60 2.68 2.87 2.60 2.68 3.33 2.30 2.37 2.35 p Value 0.012 0.0069 0.0096 0.012 0.0069 0.0024 0.028 0.018 0.023

Lower limit of 95%

CI 1.24 1.31 1.29 1.24 1.31 1.53 1.09 1.16 1.13

Upper limit of 95% CI 5.47 5.49 6.37 5.47 5.49 7.25 4.85 4.85 4.91

OR Quartile 4 1.95 2.88 1.90 1.95 2.88 2.47 1.79 2.65 2.03 p Value 0.14 0.020 0.13 0.14 0.020 0.029 0.20 0.032 0.084

Lower limit of 95%

CI 0.797 1.18 0.825 0.797 1.18 1.10 0.729 1.09 0.910

Upper limit of 95% CI 4.76 7.01 4.37 4.76 7.01 5.59 4.37 6.46 4.55

[00370] Table 23.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.905 1.36 0.905 1.36 0.950 1.35

Average 0.928 4.06 0.928 4.06 0.943 3.96

Stdev 0.731 12.1 0.731 12.1 0.738 12.0 p (t-test) 0.041 0.041 0.050

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 4.23 60.0 4.23 60.0 4.23 60.0 n (Patient) 64 68 64 68 62 70

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.983 1.36 0.974 1.42 1.00 1.36

Average 1.71 4.72 1.69 4.60 1.75 4.10

Stdev 6.13 13.4 6.20 13.1 6.41 12.2 p (t-test) 0.080 0.086 0.15

Min 0.00242 0.310 0.00242 0.310 0.00242 0.310

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 94 37 92 39 86 45

p Value 0.014 0.0020 0.23 0.014 0.0020 0.069 0.022 0.0036 0.26

Lower limit of 95%

CI 1.26 1.67 0.719 1.26 1.67 0.942 1.16 1.54 0.708

Upper limit of 95% CI 7.44 9.91 3.87 7.44 9.91 4.92 6.87 9.12 3.58

[00371] Table 23.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.68 0.64 0.65 0.68 0.64 0.65 0.67 0.63

SE 0.048 0.047 0.056 0.048 0.047 0.056 0.047 0.047 0.053 p Value 0.0014 1.1E-4 0.012 0.0014 1.1E-4 0.012 0.0014 2.5E-4 0.012 nCohort Recovered 65 67 95 65 67 95 63 66 89 nCohort Non- recovered 67 65 36 67 65 36 69 66 42

Cutoff Quartile 2 0.561 0.561 0.569 0.561 0.561 0.569 0.561 0.561 0.569

Sensitivity 84% 85% 86% 84% 85% 86% 84% 83% 88%

Specificity 34% 34% 29% 34% 34% 29% 35% 33% 31 %

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 58% 60% 64% 58% 60% 64% 58% 59% 62%

Specificity 58% 60% 55% 58% 60% 55% 59% 59% 55%

Cutoff Quartile 4 1.65 1.65 1.66 1.65 1.65 1.66 1.65 1.65 1.66

Sensitivity 34% 37% 33% 34% 37% 33% 33% 36% 31 %

Specificity 85% 87% 78% 85% 87% 78% 84% 86% 78%

OR Quartile 2 2.60 2.88 2.59 2.60 2.88 2.59 2.83 2.50 3.40 p Value 0.023 0.014 0.073 0.023 0.014 0.073 0.014 0.030 0.021

Lower limit of 95%

CI 1.14 1.24 0.914 1.14 1.24 0.914 1.24 1.10 1.21

Upper limit of 95% CI 5.95 6.67 7.35 5.95 6.67 7.35 6.47 5.71 9.57

OR Quartile 3 1.96 2.22 2.14 1.96 2.22 2.14 1.96 2.09 1.99 p Value 0.057 0.025 0.059 0.057 0.025 0.059 0.056 0.038 0.072

Lower limit of 95%

CI 0.981 1.11 0.970 0.981 1.11 0.970 0.982 1.04 0.940

Upper limit of 95% CI 3.92 4.46 4.72 3.92 4.46 4.72 3.92 4.18 4.21

OR Quartile 4 2.87 3.77 1.76 2.87 3.77 1.76 2.65 3.62 1.55 p Value 0.014 0.0026 0.19 0.014 0.0026 0.19 0.023 0.0035 0.30

Lower limit of 95%

CI 1.24 1.59 0.756 1.24 1.59 0.756 1.14 1.53 0.680

Upper limit of 95% CI 6.67 8.95 4.10 6.67 8.95 4.10 6.14 8.58 3.52

[00372] Table 23.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95%

CI 0.982 1.04 0.949 0.924 1.04 0.880 0.923 0.982 0.933

Upper limit of 95% CI 3.92 4.19 4.35 3.68 4.19 3.96 3.67 3.92 4.09

OR Quartile 4 2.83 3.54 2.04 2.71 3.54 1.94 2.50 3.39 1.67 p Value 0.014 0.0034 0.090 0.018 0.0034 0.11 0.030 0.0046 0.22

Lower limit of 95%

CI 1.24 1.52 0.896 1.19 1.52 0.853 1.10 1.46 0.741

Upper limit of 95% CI 6.47 8.24 4.65 6.20 8.24 4.40 5.71 7.89 3.77

[00373] Table 23.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 0.00242 0.233 0.00242 0.233 0.00242 0.233

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 91 40 90 41 89 42

[00374] Example 24. Use of C-C motif chemokine 18 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00375] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 18 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00376] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00377] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00378] Table 24.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Median 1.06 1.58 1.06 1.74 1.06 1.74

Average 1.65 8.26 1.64 9.16 1.64 9.16

Stdev 5.60 18.9 5.55 19.9 5.55 19.9 p (t-test) 0.0030 0.0013 0.0013

Min 0.00242 0.310 0.00242 0.310 0.00242 0.310

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 113 18 115 16 115 16

[00379] Table 24.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Average 1.73 3.75 1.67 4.12 1.68 4.32

Stdev 6.68 11.3 6.45 12.0 6.30 12.5 p (t-test) 0.20 0.13 0.11

Min 0.0156 0.00242 0.00242 0.233 0.00242 0.310

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 79 53 85 47 89 43 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.965 1.36 0.965 1.43 0.978 1.39

Average 1.72 3.84 1.68 4.09 1.71 4.20

Stdev 6.64 11.5 6.48 12.0 6.37 12.4 p (t-test) 0.18 0.14 0.13

Min 0.0156 0.00242 0.00242 0.233 0.00242 0.233

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 80 51 84 47 87 44

UO only

Lower limit of 95%

CI 1.28 1.20 0.995 1.20 1.23 0.876 1.24 1.08 0.876

Upper limit of 95% CI 8.12 7.63 20.2 8.40 8.55 18.0 9.81 7.54 18.0

OR Quartile 3 2.30 2.61 2.28 3.20 2.71 2.44 3.40 2.61 2.44 p Value 0.022 0.0096 0.082 0.0024 0.0085 0.073 0.0020 0.013 0.073

Lower limit of 95%

CI 1.13 1.26 0.900 1.51 1.29 0.921 1.56 1.23 0.921

Upper limit of 95% CI 4.69 5.40 5.78 6.78 5.70 6.45 7.38 5.56 6.45

OR Quartile 4 3.08 2.80 2.61 3.44 2.86 3.12 2.98 2.35 3.12 p Value 0.0068 0.013 0.044 0.0030 0.011 0.020 0.0087 0.039 0.020

Lower limit of 95%

CI 1.36 1.25 1.03 1.52 1.27 1.20 1.32 1.05 1.20

Upper limit of 95% CI 6.94 6.29 6.63 7.79 6.42 8.11 6.75 5.30 8.11

[00380] Table 24.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 5.89 15.1 5.80 15.9 5.80 15.9 p (t-test) 0.035 0.020 0.020

Min 0.00242 0.310 0.00242 0.310 0.00242 0.310

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 102 29 105 26 105 26

[00381] Table 24.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

n (Patient) 73 59 78 54 80 52 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.960 1.36 0.960 1.39 0.970 1.39

Average 1.77 3.59 1.73 3.79 1.74 3.86

Stdev 6.86 11.0 6.68 11.4 6.60 11.6 p (t-test) 0.25 0.19 0.18

Min 0.0156 0.00242 0.00242 0.233 0.00242 0.233

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 75 56 79 52 81 50

UO only

Upper limit of 95% CI 4.52 4.81 3.88 5.73 4.97 4.59 5.85 5.08 4.59

OR Quartile 4 2.81 2.65 2.54 2.94 2.67 3.10 2.71 2.47 3.10 p Value 0.013 0.018 0.033 0.0092 0.017 0.011 0.015 0.027 0.011

Lower limit of 95% CI 1.24 1.18 1.08 1.31 1.19 1.29 1.21 1.11 1.29

Upper limit of 95% CI 6.36 5.96 5.95 6.63 5.99 7.41 6.08 5.53 7.41

[00382] Table 24.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.67 0.65 0.64 0.69 0.67 0.65 0.67 0.65 0.65

SE 0.048 0.049 0.057 0.048 0.049 0.058 0.049 0.050 0.058 p Value 5.6E-4 0.0016 0.013 1.0E-4 7.0E-4 0.0093 3.5E-4 0.0020 0.0093 nCohort Non- persistent 73 74 96 78 78 98 80 80 98 nCohort Persistent 59 57 35 54 53 33 52 51 33

Cutoff Quartile 2 0.561 0.557 0.569 0.561 0.557 0.569 0.561 0.557 0.569

Sensitivity 85% 82% 89% 85% 83% 88% 85% 82% 88%

Specificity 33% 31 % 30% 32% 31 % 30% 31 % 30% 30%

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 61 % 61 % 60% 65% 62% 61 % 65% 63% 61 %

Specificity 59% 58% 53% 60% 58% 53% 60% 57% 53%

Cutoff Quartile 4 1.65 1.64 1.66 1.65 1.64 1.66 1.65 1.64 1.66

Sensitivity 36% 35% 37% 37% 36% 39% 37% 35% 39%

Specificity 84% 82% 79% 83% 82% 80% 82% 81 % 80%

OR Quartile 2 2.72 2.12 3.35 2.71 2.17 3.05 2.50 2.00 3.05 p Value 0.023 0.080 0.036 0.028 0.078 0.054 0.043 0.12 0.054

Lower limit of 95% CI 1.15 0.914 1.08 1.12 0.916 0.983 1.03 0.843 0.983

Upper limit of 95% CI 6.44 4.92 10.4 6.60 5.15 9.45 6.08 4.75 9.45

OR Quartile 3 2.24 2.21 1.70 2.79 2.25 1.74 2.83 2.28 1.74 p Value 0.024 0.028 0.19 0.0051 0.026 0.18 0.0049 0.025 0.18

Lower limit of 95% CI 1.11 1.09 0.775 1.36 1.10 0.779 1.37 1.11 0.779

Upper limit of 95% CI 4.52 4.47 3.73 5.73 4.60 3.88 5.85 4.68 3.88

OR Quartile 4 2.81 2.54 2.25 2.94 2.55 2.54 2.71 2.36 2.54 p Value 0.013 0.024 0.060 0.0092 0.023 0.033 0.015 0.036 0.033

Lower limit of 95% CI 1.24 1.13 0.965 1.31 1.14 1.08 1.21 1.06 1.08

Upper limit of 95% CI 6.36 5.70 5.22 6.63 5.72 5.95 6.08 5.28 5.95

[00383] Example 25. Use of C-C motif chemokine 18 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00384] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 18 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00385] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00386] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00387] Table 25.1:Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 0.00242 0.310 0.00242 0.310 0.00242 0.310

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 102 29 108 23 110 21

[00388] Table 25.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 6.56 5.94 8.76 3.68 4.35 7.43 4.18 4.10 6.51

OR Quartile 4 2.74 3.42 1.67 2.30 2.56 1.95 2.71 2.90 2.22 p Value 0.033 0.0090 0.23 0.047 0.026 0.13 0.018 0.012 0.077

Lower limit of 95%

CI 1.09 1.36 0.719 1.01 1.12 0.813 1.19 1.27 0.916

Upper limit of 95% CI 6.90 8.62 3.87 5.26 5.85 4.68 6.20 6.64 5.39

[00389] Table 25.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Persistence Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.69 0.68 0.64 0.65 0.66 0.62 0.64 0.66 0.62

SE 0.046 0.046 0.053 0.048 0.047 0.057 0.048 0.048 0.058 p Value 4.1E-5 1.1E-4 0.0096 0.0023 6.6E-4 0.031 0.0028 8.3E-4 0.046 nCohort Non- persistent 49 53 89 64 65 95 66 67 97 nCohort Persistent 83 78 42 68 66 36 66 64 34

Cutoff Quartile 2 0.561 0.557 0.569 0.561 0.557 0.569 0.561 0.557 0.569

Sensitivity 83% 82% 88% 84% 83% 86% 83% 83% 85%

Specificity 39% 36% 31 % 34% 34% 29% 33% 33% 29%

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 60% 60% 64% 57% 59% 61 % 58% 58% 59%

Specificity 67% 64% 56% 58% 58% 54% 58% 57% 53%

Cutoff Quartile 4 1.65 1.64 1.66 1.65 1.64 1.66 1.65 1.64 1.66

Sensitivity 31 % 33% 29% 32% 33% 31 % 33% 34% 32%

Specificity 86% 87% 76% 83% 83% 77% 83% 84% 77%

OR Quartile 2 3.12 2.55 3.40 2.71 2.56 2.59 2.50 2.36 2.35 p Value 0.0060 0.023 0.021 0.018 0.026 0.073 0.030 0.042 0.11

Lower limit of 95% CI 1.38 1.14 1.21 1.19 1.12 0.914 1.10 1.03 0.827

Upper limit of 95% CI 7.03 5.72 9.57 6.20 5.85 7.35 5.71 5.38 6.70

OR Quartile 3 3.12 2.71 2.31 1.84 2.03 1.82 1.84 1.80 1.58 p Value 0.0026 0.0067 0.030 0.083 0.046 0.13 0.083 0.098 0.25

Lower limit of 95% CI 1.49 1.32 1.08 0.924 1.01 0.833 0.923 0.898 0.718

Upper limit of 95% CI 6.56 5.58 4.92 3.68 4.08 3.98 3.67 3.59 3.49

OR Quartile 4 2.74 3.29 1.30 2.30 2.45 1.46 2.50 2.67 1.63 p Value 0.033 0.012 0.54 0.047 0.033 0.39 0.030 0.020 0.27

Lower limit of 95% CI 1.09 1.30 0.565 1.01 1.07 0.621 1.10 1.17 0.689

Upper limit of 95% CI 6.90 8.28 2.97 5.26 5.61 3.43 5.71 6.10 3.86

[00390] Table 25.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.819 1.34 0.950 1.35 0.965 1.36

Average 0.911 3.66 0.949 4.07 0.955 4.16

Stdev 0.755 11.4 0.736 12.2 0.726 12.4 p (t-test) 0.082 0.044 0.038

Min 0.0264 0.00242 0.0264 0.00242 0.0264 0.00242

Max 4.23 60.0 4.23 60.0 4.23 60.0 n (Patient) 53 78 64 67 66 65

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 1.00 1.36 1.05 1.35 1.06 1.34

Average 1.75 4.11 1.76 4.39 1.75 4.55

Stdev 6.41 12.2 6.23 13.0 6.16 13.3 p (t-test) 0.15 0.12 0.10

Min 0.00242 0.233 0.00242 0.233 0.00242 0.233

Max 60.0 60.0 60.0 60.0 60.0 60.0 n (Patient) 86 45 91 40 93 38

[00391] Table 25.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 49 53 84 62 64 88 64 66 90 nCohort Persistent 83 78 47 70 67 43 68 65 41

Cutoff Quartile 2 0.561 0.557 0.569 0.561 0.557 0.569 0.561 0.557 0.569

Sensitivity 83% 82% 87% 84% 84% 86% 84% 83% 85%

Specificity 39% 36% 32% 35% 34% 31 % 34% 33% 30%

Cutoff Quartile 3 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07 1.07

Sensitivity 60% 60% 64% 59% 60% 60% 59% 58% 59%

Specificity 67% 64% 57% 60% 59% 55% 59% 58% 53%

Cutoff Quartile 4 1.65 1.64 1.66 1.65 1.64 1.66 1.65 1.64 1.66

Sensitivity 31 % 33% 30% 33% 34% 30% 34% 35% 32%

Specificity 86% 87% 77% 84% 84% 77% 84% 85% 78%

OR Quartile 2 3.12 2.55 3.24 2.95 2.67 2.73 2.71 2.45 2.50 p Value 0.0060 0.023 0.018 0.010 0.020 0.043 0.018 0.033 0.066

Lower limit of 95% CI 1.38 1.14 1.23 1.29 1.17 1.03 1.19 1.07 0.942 Upper limit of 95% CI 7.03 5.72 8.55 6.75 6.10 7.23 6.20 5.61 6.64

OR Quartile 3 3.12 2.71 2.35 2.09 2.17 1.84 2.09 1.91 1.61 p Value 0.0026 0.0067 0.023 0.037 0.030 0.11 0.038 0.068 0.21

Lower limit of 95% CI 1.49 1.32 1.13 1.04 1.08 0.874 1.04 0.954 0.765 Upper limit of 95% CI 6.56 5.58 4.91 4.19 4.35 3.85 4.18 3.82 3.40

OR Quartile 4 2.74 3.29 1.45 2.54 2.82 1.47 2.76 3.07 1.62 p Value 0.033 0.012 0.37 0.029 0.016 0.35 0.018 0.0092 0.25

Lower limit of 95% CI 1.09 1.30 0.647 1.10 1.22 0.649 1.19 1.32 0.713 Upper limit of 95% CI 6.90 8.28 3.25 5.90 6.55 3.34 6.40 7.13 3.71

[00392] Example 26. Use of C-C motif chemokine 23 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00393] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 23 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00394] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered".

[00395] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00396] Table 26.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 194 121 173 142 156 159

[00397] Table 26.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

p (t-test) 7.3E-4 0.0019 0.0039

Min 7.09E-7 3.28E-9 7.09E-7 3.28E-9 7.09E-7 3.28E-9

Max 0.102 2.47 0.102 2.47 0.102 2.47 n (Patient) 90 227 79 238 71 246 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00328 0.0222 0.00328 0.0220 0.00329 0.0213

Average 0.00870 0.163 0.00881 0.161 0.00872 0.158

Stdev 0.0153 0.408 0.0154 0.406 0.0155 0.402 p (t-test) 1.3E-4 1.6E-4 2.7E-4

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 0.102 2.47 0.102 2.47 0.102 2.47 n (Patient) 106 211 104 213 99 218

UO only

Lower limit of 95%

CI 1.55 1.82 0.717 1.73 1.77 0.863 1.54 1.70 0.861

Upper limit of 95%

CI 4.53 5.23 2.18 5.19 5.08 2.54 4.77 4.91 2.46

OR Quartile 3 4.71 6.07 1.72 4.51 5.77 1.48 3.95 5.48 1.46 p Value 3.3E-8 3.9E-11 0.026 3.1E-7 1.3E-10 0.091 5.2E-6 7.1E-10 0.098

Lower limit of 95%

CI 2.72 3.56 1.07 2.53 3.38 0.939 2.19 3.19 0.932

Upper limit of 95%

CI 8.15 10.4 2.77 8.03 9.85 2.34 7.14 9.41 2.28

OR Quartile 4 14.9 19.7 2.27 18.8 19.1 1.96 16.0 17.5 1.93 p Value 7.7E-6 7.6E-7 0.0022 5.8E-5 1.0E-6 0.011 1.5E-4 2.1E-6 0.012

Lower limit of 95%

CI 4.56 6.05 1.34 4.49 5.85 1.17 3.83 5.36 1.15

Upper limit of 95%

CI 48.6 64.3 3.84 78.4 62.1 3.29 67.0 57.0 3.24

[00398] Table 26.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered

Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00730 0.0207 0.00764 0.0182 0.00724 0.0182

Average 0.0934 0.151 0.0971 0.139 0.0964 0.135

Stdev 0.335 0.354 0.342 0.340 0.350 0.328 p (t-test) 0.16 0.30 0.32

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 214 101 204 111 190 125

[00399] Table 26.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 134 140 216 125 138 207 114 135 194 nCohort Non- recovered 183 177 99 192 179 108 203 182 121

Cutoff Quartile 2 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207

Sensitivity 83% 84% 80% 83% 84% 80% 82% 84% 79%

Specificity 35% 36% 27% 37% 36% 28% 38% 37% 28%

Cutoff Quartile 3 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104

Sensitivity 64% 66% 65% 62% 65% 61 % 61 % 65% 60%

Specificity 69% 70% 56% 68% 70% 56% 68% 70% 56%

Cutoff Quartile 4 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478

Sensitivity 38% 40% 34% 36% 40% 32% 34% 39% 32%

Specificity 93% 94% 79% 92% 93% 79% 91 % 93% 79%

OR Quartile 2 2.55 3.05 1.48 2.81 2.94 1.49 2.81 3.10 1.48 p Value 4.3E-4 3.8E-5 0.18 1.1E-4 6.2E-5 0.17 1.1E-4 2.7E-5 0.15

Lower limit of 95%

CI 1.51 1.79 0.836 1.66 1.73 0.849 1.67 1.83 0.863

Upper limit of 95%

CI 4.29 5.18 2.64 4.73 4.98 2.60 4.74 5.27 2.54

OR Quartile 3 4.12 4.55 2.37 3.46 4.31 1.96 3.33 4.49 1.85 p Value 5.7E-9 4.8E-10 5.8E-4 3.0E-7 1.7E-9 0.0053 1.2E-6 8.4E-10 0.0091

Lower limit of 95%

CI 2.56 2.82 1.45 2.15 2.68 1.22 2.05 2.78 1.16

Upper limit of 95%

CI 6.63 7.33 3.88 5.57 6.94 3.16 5.41 7.24 2.92

OR Quartile 4 7.68 9.75 1.99 6.60 9.42 1.78 5.47 8.95 1.83 p Value 1.8E-8 1.6E-9 0.011 1.9E-7 2.7E-9 0.031 2.7E-6 6.1E-9 0.022

Lower limit of 95%

CI 3.78 4.66 1.17 3.24 4.50 1.05 2.69 4.28 1.09

Upper limit of 95%

CI 15.6 20.4 3.37 13.4 19.7 3.00 11.1 18.7 3.07

[00400] Table 26.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00460 0.0298 0.00458 0.0284 0.00443 0.0284

Average 0.0332 0.191 0.0336 0.187 0.0338 0.186

Stdev 0.161 0.443 0.163 0.439 0.163 0.438 p (t-test) 3.2E-5 4.9E-5 5.7E-5

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 1.92 2.47 1.92 2.47 1.92 2.47 n (Patient) 160 157 157 160 156 161

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00709 0.0213 0.00710 0.0207 0.00709 0.0209

Average 0.0861 0.164 0.0872 0.160 0.0868 0.159

Stdev 0.328 0.363 0.330 0.359 0.332 0.355 p (t-test) 0.056 0.074 0.075

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 211 104 208 107 205 110

OR Quartile 4 2.51 6.19 2.07 2.38 5.88 1.95 2.38 5.77 1.97 p Value 4.9E-4 3.8E-9 0.0066 0.0010 1.0E-8 0.013 0.0010 1.4E-8 0.011

Lower limit of 95%

CI 1.50 3.38 1.22 1.42 3.20 1.15 1.42 3.15 1.17

Upper limit of 95%

CI 4.21 11.4 3.50 3.98 10.8 3.28 3.98 10.6 3.31

[00401] Example 27. Use of C-C motif chemokine 23 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00402] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 23 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00403] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00404] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00405] Table 27.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Median 0.00335 0.0207 0.00326 0.0192 0.00296 0.0192

Average 0.0505 0.143 0.0522 0.140 0.0531 0.138

Stdev 0.299 0.357 0.305 0.354 0.311 0.351 p (t-test) 0.022 0.031 0.039

Min 6.85E-7 3.28E-9 6.85E-7 3.28E-9 6.85E-7 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 108 209 104 213 100 217 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00335 0.0249 0.00335 0.0235 0.00335 0.0231

Average 0.0443 0.156 0.0449 0.154 0.0456 0.153

Stdev 0.277 0.372 0.279 0.370 0.282 0.368 p (t-test) 0.0042 0.0052 0.0063

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 126 190 124 192 122 194

UO only

Specificity 95% 95% 81 % 95% 95% 82% 96% 95% 82%

OR Quartile 2 2.22 2.94 1.03 2.42 2.84 1.35 2.63 2.75 1.49 p Value 0.0027 5.6E-5 0.93 9.9E-4 9.0E-5 0.29 3.3E-4 1.4E-4 0.15

Lower limit of 95%

CI 1.32 1.74 0.586 1.43 1.68 0.773 1.55 1.63 0.863

Upper limit of 95%

CI 3.75 4.96 1.80 4.08 4.79 2.34 4.46 4.63 2.57

OR Quartile 3 4.19 5.42 1.57 3.79 5.14 1.52 3.92 4.89 1.58 p Value 2.7E-8 2.8E-11 0.070 2.4E-7 l .OE-10 0.082 2.0E-7 3.6E-10 0.053

Lower limit of 95%

CI 2.53 3.30 0.963 2.29 3.13 0.949 2.34 2.98 0.994

Upper limit of 95%

CI 6.95 8.91 2.56 6.28 8.45 2.43 6.55 8.02 2.50

OR Quartile 4 11.5 12.5 2.63 10.8 12.1 2.78 12.9 11.7 2.81 p Value 3.5E-7 1.3E-8 3.9E-4 7.5E-7 2.1E-8 1.4E-4 1.4E-6 3.2E-8 1.1E-4

Lower limit of 95%

CI 4.50 5.23 1.54 4.20 5.05 1.64 4.58 4.88 1.66

Upper limit of 95%

CI 29.5 29.8 4.48 27.6 28.8 4.70 36.5 27.9 4.73

[00406] Table 27.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00407] Table 27.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Recovered 154 161 224 152 160 223 148 159 212 nCohort Non- recovered 163 156 91 165 157 92 169 158 103

Cutoff Quartile 2 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207

Sensitivity 84% 85% 81 % 84% 85% 82% 84% 85% 83%

Specificity 34% 35% 28% 34% 35% 28% 35% 35% 29%

Cutoff Quartile 3 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104

Sensitivity 65% 67% 67% 64% 67% 66% 63% 66% 62%

Specificity 66% 66% 57% 65% 66% 57% 65% 66% 56%

Cutoff Quartile 4 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478

Sensitivity 39% 41 % 37% 39% 41 % 37% 38% 41 % 35%

Specificity 90% 90% 80% 89% 90% 80% 89% 90% 80%

OR Quartile 2 2.77 3.08 1.67 2.66 3.14 1.70 2.85 2.95 1.91 p Value 1.9E-4 5.7E-5 0.097 3.1E-4 4.4E-5 0.085 1.2E-4 9.5E-5 0.032

Lower limit of 95%

CI 1.62 1.78 0.912 1.56 1.81 0.930 1.67 1.71 1.06

Upper limit of 95%

CI 4.72 5.34 3.04 4.52 5.43 3.10 4.85 5.09 3.44

OR Quartile 3 3.54 4.08 2.66 3.36 3.96 2.56 3.19 3.85 2.06 p Value 8.4E-8 3.9E-9 1.7E-4 2.7E-7 7.3E-9 2.9E-4 8.0E-7 1.4E-8 0.0033

Lower limit of 95%

CI 2.23 2.56 1.60 2.12 2.49 1.54 2.01 2.42 1.27

Upper limit of 95%

CI 5.63 6.51 4.44 5.32 6.32 4.24 5.05 6.14 3.33

OR Quartile 4 5.58 6.30 2.37 5.39 6.19 2.32 5.03 6.09 2.11 p Value 2.7E-8 2.7E-9 0.0016 5.0E-8 3.8E-9 0.0021 1.7E-7 5.4E-9 0.0053

Lower limit of 95%

CI 3.04 3.44 1.39 2.94 3.38 1.36 2.75 3.32 1.25

Upper limit of 95%

CI 10.2 11.6 4.06 9.87 11.4 3.96 9.21 11.2 3.57

[00408] Table 27.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00443 0.0336 0.00428 0.0317 0.00443 0.0298

Average 0.0468 0.188 0.0471 0.186 0.0473 0.185

Stdev 0.243 0.417 0.243 0.416 0.244 0.415 p (t-test) 2.2E-4 2.5E-4 2.9E-4

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 172 145 171 146 170 147

UO only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00711 0.0227 0.00710 0.0222 0.00710 0.0215

Average 0.0898 0.165 0.0902 0.164 0.0927 0.153

Stdev 0.328 0.368 0.329 0.366 0.335 0.354 p (t-test) 0.074 0.081 0.15

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 223 92 222 93 214 101

OR Quartile 4 5.44 6.39 2.32 5.15 6.27 2.27 4.80 6.16 2.05 p Value 9.7E-9 6.6E-10 0.0021 2.7E-8 9.5E-10 0.0027 9.8E-8 1.4E-9 0.0077

Lower limit of 95%

CI 3.05 3.55 1.36 2.89 3.48 1.33 2.70 3.42 1.21

Upper limit of 95% CI 9.69 11.5 3.96 9.18 11.3 3.87 8.54 11.1 3.47

[00409] Table 27.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.71 0.64 0.64 0.71 0.64 0.64 0.71 0.63

SE 0.033 0.030 0.035 0.033 0.030 0.035 0.033 0.030 0.035 p Value 4.7E-6 7.9E-12 6.6E-5 1.1E-5 7.9E-12 7.0E-5 1.5E-5 7.9E-12 1.9E-4 nCohort Recovered 203 185 217 201 185 216 200 185 214 nCohort Non- recovered 114 132 98 116 132 99 117 132 101

Cutoff Quartile 2 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207 0.00202 0.00202 0.00207

Sensitivity 83% 84% 83% 83% 84% 83% 83% 84% 82%

Specificity 30% 31 % 29% 29% 31 % 29% 30% 31 % 29%

Cutoff Quartile 3 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104

Sensitivity 69% 71 % 69% 68% 71 % 69% 68% 71 % 67%

Specificity 61 % 65% 59% 60% 65% 58% 60% 65% 58%

Cutoff Quartile 4 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478 0.0477 0.0477 0.0478

Sensitivity 37% 42% 37% 36% 42% 36% 36% 42% 36%

Specificity 81 % 87% 80% 81 % 87% 80% 81 % 87% 80%

OR Quartile 2 2.10 2.41 1.91 1.99 2.41 1.94 2.03 2.41 1.84 p Value 0.012 0.0021 0.035 0.017 0.0021 0.030 0.015 0.0021 0.043

Lower limit of 95%

CI 1.18 1.38 1.05 1.13 1.38 1.07 1.15 1.38 1.02

Upper limit of 95%

CI 3.74 4.23 3.47 3.52 4.23 3.54 3.59 4.23 3.32

OR Quartile 3 3.47 4.57 3.20 3.23 4.57 3.07 3.12 4.57 2.84 p Value 5.7E-7 7.0E-10 7.1E-6 1.9E-6 7.0E-10 1.3E-5 3.3E-6 7.0E-10 3.8E-5

Lower limit of 95%

CI 2.13 2.82 1.93 1.99 2.82 1.86 1.93 2.82 1.73

Upper limit of 95%

CI 5.65 7.40 5.31 5.23 7.40 5.08 5.04 7.40 4.66

OR Quartile 4 2.53 4.94 2.35 2.43 4.94 2.30 2.39 4.94 2.20 p Value 4.5E-4 1.3E-8 0.0016 7.6E-4 1.3E-8 0.0020 9.7E-4 1.3E-8 0.0033

Lower limit of 95%

CI 1.51 2.85 1.38 1.45 2.85 1.36 1.42 2.85 1.30

Upper limit of 95%

CI 4.26 8.57 3.99 4.09 8.57 3.90 4.00 8.57 3.73

[00410] Example 28. Use of C-C motif chemokine 23 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00411] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 23 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00412] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00413] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00414] Table 28.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00870 0.0349 0.00864 0.0832 0.00859 0.0861

Average 0.0897 0.265 0.0887 0.286 0.0884 0.294

Stdev 0.311 0.484 0.309 0.497 0.309 0.502 p (t-test) 0.0023 9.0E-4 6.2E-4

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 275 40 278 37 279 36

[00415] Table 28.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 72% 72% 71 % 76% 75% 75% 74% 74% 76% Specificity 65% 64% 55% 64% 64% 55% 61 % 61 % 55%

Cutoff Quartile 4 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478

Sensitivity 45% 44% 46% 50% 47% 47% 50% 46% 48%

Specificity 89% 87% 80% 88% 87% 80% 86% 84% 80%

OR Quartile 2 2.37 2.51 1.58 2.99 3.16 1.88 2.53 2.84 2.15 p Value 0.0025 0.0017 0.21 5.3E-4 3.7E-4 0.10 0.0042 0.0017 0.061

Lower limit of 95%

CI 1.36 1.41 0.775 1.61 1.68 0.876 1.34 1.48 0.967

Upper limit of 95%

CI 4.16 4.47 3.22 5.54 5.95 4.05 4.78 5.45 4.78

OR Quartile 3 4.73 4.53 2.98 5.72 5.37 3.58 4.52 4.57 3.88 p Value 3.5E-10 1.5E-9 5.0E-4 4.4E-11 2.2E-10 1.3E-4 2.3E-8 1.9E-8 7.2E-5

Lower limit of 95%

CI 2.91 2.78 1.61 3.40 3.20 1.86 2.66 2.69 1.99

Upper limit of 95%

CI 7.68 7.40 5.51 9.60 9.02 6.88 7.66 7.76 7.58

OR Quartile 4 6.44 5.15 3.31 7.37 5.94 3.50 6.06 4.59 3.64 p Value 1.5E-10 5.2E-9 8.2E-5 3.7E-12 2.3E-10 5.5E-5 1.2E-10 3.1E-8 3.6E-5

Lower limit of 95%

CI 3.64 2.97 1.82 4.19 3.43 1.90 3.50 2.68 1.97

Upper limit of 95%

CI 11.4 8.93 6.01 12.9 10.3 6.44 10.5 7.89 6.73

[00416] Table 28.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

n (Patient) 182 134 195 121 208 108

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00765 0.0238 0.00751 0.0272 0.00751 0.0272

Average 0.0863 0.203 0.0849 0.218 0.0849 0.218

Stdev 0.314 0.415 0.311 0.428 0.311 0.428 p (t-test) 0.012 0.0053 0.0053

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 246 69 251 64 251 64

[00417] Table 28.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 173 180 239 189 192 245 201 204 245 nCohort Persistent 144 136 76 128 124 70 116 112 70

Cutoff Quartile 2 0.00202 0.00200 0.00207 0.00202 0.00200 0.00207 0.00202 0.00200 0.00207

Sensitivity 84% 85% 83% 87% 87% 86% 86% 87% 86%

Specificity 32% 32% 28% 33% 33% 28% 31 % 31 % 28%

Cutoff Quartile 3 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104 0.0104

Sensitivity 70% 71 % 67% 75% 74% 71 % 74% 73% 71 %

Specificity 66% 66% 55% 67% 66% 56% 64% 63% 56%

Cutoff Quartile 4 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478

Sensitivity 44% 43% 41 % 48% 47% 43% 47% 46% 43%

Specificity 90% 89% 80% 90% 89% 80% 87% 86% 80%

OR Quartile 2 2.52 2.60 1.85 3.19 3.30 2.35 2.85 2.96 2.35 p Value 9.5E-4 8.2E-4 0.068 1.3E-4 1.1E-4 0.021 7.0E-4 6.0E-4 0.021

Lower limit of 95%

CI 1.46 1.49 0.955 1.76 1.80 1.14 1.56 1.59 1.14

Upper limit of 95%

CI 4.36 4.56 3.58 5.77 6.04 4.86 5.23 5.49 4.86

OR Quartile 3 4.66 4.57 2.52 6.00 5.49 3.17 5.03 4.60 3.17 p Value 2.5E-10 5.6E-10 8.5E-4 2.5E-12 2.6E-11 8.8E-5 3.8E-10 3.2E-9 8.8E-5

Lower limit of 95%

CI 2.89 2.83 1.46 3.63 3.33 1.78 3.03 2.78 1.78

Upper limit of 95%

CI 7.50 7.38 4.33 9.91 9.05 5.65 8.33 7.63 5.65

OR Quartile 4 7.14 6.13 2.74 8.15 7.16 3.00 5.86 5.26 3.00 p Value 1.3E-10 6.6E-10 3.8E-4 2.6E-12 1.9E-11 1.5E-4 3.0E-10 2.5E-9 1.5E-4

Lower limit of 95%

CI 3.92 3.45 1.57 4.53 4.03 1.70 3.38 3.05 1.70

Upper limit of 95%

CI 13.0 10.9 4.78 14.7 12.7 5.29 10.2 9.07 5.29

[00418] Table 28.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00552 0.0317 0.00552 0.0369 0.00599 0.0351

Average 0.0493 0.190 0.0469 0.206 0.0516 0.215

Stdev 0.243 0.423 0.235 0.438 0.231 0.458 p (t-test) 2.5E-4 3.8E-5 3.5E-5

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 176 140 188 128 200 116

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00717 0.0285 0.00722 0.0291 0.00722 0.0291

Average 0.0798 0.205 0.0791 0.212 0.0791 0.212

Stdev 0.312 0.402 0.311 0.409 0.311 0.409 p (t-test) 0.0045 0.0028 0.0028

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 234 81 237 78 237 78

Lower limit of 95%

CI 3.64 3.20 1.76 4.10 3.72 1.75 3.11 2.80 1.75

Upper limit of 95%

CI 12.0 10.1 5.26 13.2 11.7 5.29 9.30 8.29 5.29

[00419] Example 29. Use of C-C motif chemokine 23 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00420] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 23 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00421] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00422] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00423] Table 29.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Average 0.0467 0.150 0.0475 0.175 0.0486 0.192

Stdev 0.285 0.365 0.250 0.402 0.239 0.426 p (t-test) 0.0087 8.3E-4 1.7E-4

Min 6.85E-7 3.28E-9 6.85E-7 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 119 198 158 159 179 138 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00367 0.0249 0.00428 0.0259 0.00460 0.0351

Average 0.0479 0.156 0.0473 0.179 0.0487 0.199

Stdev 0.277 0.374 0.250 0.406 0.238 0.433 p (t-test) 0.0055 5.6E-4 9.8E-5

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 130 186 162 154 184 132

UO only

Lower limit of 95%

CI 1.51 1.62 0.688 1.23 1.40 0.564 1.53 1.62 0.632

Upper limit of 95%

CI 4.28 4.60 2.39 3.50 4.08 2.12 4.68 5.12 2.67

OR Quartile 3 4.56 4.98 1.76 3.35 3.93 2.24 3.54 3.88 2.60 p Value 1.9E-9 1.2E-10 0.037 2.7E-7 9.5E-9 0.0075 1.2E-7 2.3E-8 0.0033

Lower limit of 95%

CI 2.78 3.05 1.03 2.11 2.46 1.24 2.22 2.41 1.38

Upper limit of 95%

CI 7.48 8.12 2.99 5.31 6.27 4.06 5.65 6.24 4.92

OR Quartile 4 11.2 9.42 2.67 4.97 5.75 3.63 5.17 5.60 4.38 p Value 5.0E-8 1.4E-8 5.9E-4 7.8E-8 9.3E-9 2.2E-5 8.7E-9 2.0E-9 3.3E-6

Lower limit of 95%

CI 4.71 4.34 1.53 2.77 3.16 2.00 2.96 3.19 2.35

Upper limit of 95% CI 26.8 20.4 4.68 8.93 10.4 6.59 9.06 9.83 8.17

[00424] Table 29.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Median 0.00730 0.0215 0.00744 0.0285 0.00737 0.0326

Average 0.0880 0.169 0.0843 0.197 0.0833 0.212

Stdev 0.328 0.367 0.318 0.396 0.314 0.412 p (t-test) 0.055 0.011 0.0053

Min 6.85E-7 3.28E-9 6.85E-7 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 222 93 238 77 245 70

[00425] Table 29.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00321 0.0217 0.00424 0.0232 0.00424 0.0285

Average 0.0488 0.145 0.0472 0.164 0.0478 0.174

Stdev 0.295 0.359 0.261 0.388 0.252 0.401 p (t-test) 0.016 0.0022 8.8E-4

Min 6.85E-7 3.28E-9 6.85E-7 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 111 206 144 173 158 159 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00352 0.0235 0.00427 0.0259 0.00427 0.0293

Average 0.0493 0.152 0.0490 0.169 0.0500 0.177

Stdev 0.283 0.369 0.259 0.394 0.252 0.407 p (t-test) 0.0091 0.0018 8.9E-4

Min 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9 3.28E-9

Max 2.47 2.43 2.47 2.43 2.47 2.43 n (Patient) 124 192 151 165 163 153

UO only

Specificity 95% 94% 81 % 90% 90% 81 % 90% 90% 81 %

OR Quartile 2 2.78 2.84 1.60 2.29 2.50 1.49 3.00 2.96 1.73 p Value 1.3E-4 9.0E-5 0.11 0.0018 6.9E-4 0.19 7.4E-5 1.1E-4 0.089

Lower limit of 95%

CI 1.65 1.68 0.902 1.36 1.47 0.821 1.74 1.71 0.921

Upper limit of 95%

CI 4.69 4.79 2.84 3.85 4.23 2.70 5.17 5.12 3.24

OR Quartile 3 4.52 4.83 2.19 3.79 4.05 2.59 3.96 4.05 2.84 p Value 4.7E-9 3.9E-10 0.0015 2.3E-8 5.0E-9 2.6E-4 7.4E-9 5.0E-9 1.2E-4

Lower limit of 95%

CI 2.73 2.95 1.35 2.37 2.54 1.55 2.48 2.53 1.67

Upper limit of 95%

CI 7.50 7.90 3.55 6.05 6.48 4.32 6.32 6.46 4.84

OR Quartile 4 12.1 10.0 2.44 5.73 5.75 2.82 5.98 5.85 3.12 p Value 2.0E-7 3.2E-8 8.9E-4 5.9E-8 3.0E-8 1.5E-4 7.4E-9 6.6E-9 4.4E-5

Lower limit of 95%

CI 4.74 4.43 1.44 3.05 3.10 1.65 3.26 3.22 1.81

Upper limit of 95%

CI 31.1 22.7 4.12 10.8 10.7 4.83 11.0 10.6 5.38

[00426] Table 29.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 0.00710 0.0207 0.00717 0.0222 0.00717 0.0232

Average 0.0925 0.150 0.0897 0.163 0.0888 0.170

Stdev 0.339 0.345 0.330 0.363 0.326 0.373 p (t-test) 0.16 0.079 0.056

Min 6.85E-7 3.28E-9 6.85E-7 3.28E-9 3.28E-9 3.28E-9

Max 2.47 1.82 2.47 1.82 2.47 1.82 n (Patient) 208 107 220 95 226 89

[00427] Table 29.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 63% 65% 65% 64% 66% 67% 65% 68% 68% Specificity 75% 74% 58% 68% 69% 58% 67% 68% 58%

Cutoff Quartile 4 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478 0.0477 0.0468 0.0478

Sensitivity 36% 37% 35% 37% 38% 37% 38% 39% 38%

Specificity 95% 94% 81 % 90% 90% 81 % 89% 89% 81 %

OR Quartile 2 3.09 3.01 1.92 2.55 2.72 1.75 2.96 3.22 1.91 p Value 2.7E-5 3.8E-5 0.025 4.5E-4 2.1E-4 0.060 7.1E-5 3.0E-5 0.035

Lower limit of 95%

CI 1.82 1.78 1.08 1.51 1.60 0.976 1.73 1.86 1.05

Upper limit of 95%

CI 5.23 5.09 3.39 4.29 4.62 3.12 5.06 5.58 3.47

OR Quartile 3 4.89 5.42 2.51 3.84 4.45 2.80 3.76 4.40 2.99 p Value 2.0E-9 4.6E-11 1.5E-4 2.1E-8 6.8E-10 4.1E-5 2.5E-8 7.3E-10 2.0E-5

Lower limit of 95%

CI 2.91 3.28 1.56 2.40 2.77 1.71 2.36 2.75 1.81

Upper limit of 95%

CI 8.21 8.97 4.04 6.15 7.15 4.59 5.99 7.05 4.95

OR Quartile 4 11.1 9.53 2.29 5.17 5.27 2.44 5.20 5.36 2.53 p Value 5.2E-7 6.3E-8 0.0018 3.2E-7 1.3E-7 8.9E-4 9.1E-8 3.4E-8 6.0E-4

Lower limit of 95%

CI 4.35 4.21 1.36 2.75 2.84 1.44 2.84 2.95 1.49

Upper limit of 95%

CI 28.6 21.6 3.85 9.72 9.78 4.12 9.53 9.72 4.29

[00428] Example 30. Use of C-C motif chemokine 24 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 from RIFLE I and F

[00429] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 24 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00430] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is risk of injury (R), injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non-recovered". [00431] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00432] Table 30.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.65 0.62 0.66 0.64 0.63 0.66 0.63 0.65 0.64

SE 0.037 0.035 0.032 0.041 0.035 0.031 0.043 0.035 0.031 p Value 8.8E-5 4.2E-4 3.1E-7 8.5E-4 1.5E-4 1.3E-7 0.0027 1.7E-5 8.6E-6 nCohort Recovered 57 76 193 46 74 172 41 70 155 nCohort Non- recovered 259 239 121 270 241 142 275 245 159

Cutoff Quartile 2 4.17 4.15 4.13 4.17 4.15 4.13 4.17 4.15 4.13

Sensitivity 78% 79% 87% 77% 79% 85% 77% 79% 82%

Specificity 40% 37% 33% 39% 38% 34% 39% 40% 33%

Cutoff Quartile 3 29.6 29.3 28.8 29.6 29.3 28.8 29.6 29.3 28.8

Sensitivity 54% 55% 65% 53% 56% 63% 53% 56% 60%

Specificity 68% 66% 60% 67% 68% 61 % 68% 70% 61 %

Cutoff Quartile 4 74.9 74.4 74.6 74.9 74.4 74.6 74.9 74.4 74.6

Sensitivity 28% 28% 39% 27% 28% 36% 27% 28% 35%

Specificity 88% 84% 83% 87% 85% 84% 85% 86% 85%

OR Quartile 2 2.45 2.15 3.18 2.20 2.27 2.93 2.15 2.54 2.29 p Value 0.0037 0.0073 1.8E-4 0.019 0.0043 1.7E-4 0.029 0.0013 0.0021

Lower limit of 95%

CI 1.34 1.23 1.73 1.14 1.29 1.67 1.08 1.44 1.35

Upper limit of 95% CI 4.50 3.76 5.83 4.25 3.98 5.14 4.28 4.48 3.89

OR Quartile 3 2.55 2.37 2.77 2.33 2.61 2.71 2.40 2.96 2.35 p Value 0.0026 0.0017 2.3E-5 0.012 6.2E-4 2.0E-5 0.014 1.9E-4 2.2E-4

Lower limit of 95%

CI 1.39 1.39 1.73 1.20 1.51 1.71 1.19 1.67 1.49

Upper limit of 95% CI 4.69 4.06 4.45 4.51 4.52 4.29 4.83 5.23 3.69

OR Quartile 4 2.75 2.08 3.20 2.47 2.25 2.88 2.11 2.35 2.89 p Value 0.018 0.035 1.5E-5 0.049 0.023 9.2E-5 0.11 0.021 1.3E-4

Lower limit of 95%

CI 1.19 1.05 1.89 1.01 1.12 1.70 0.852 1.14 1.68

Upper limit of 95% CI 6.35 4.09 5.41 6.07 4.53 4.90 5.22 4.86 4.97

[00433] Table 30.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95%

CI 1.47 1.39 1.63 1.22 1.48 1.55 1.26 1.65 1.36

Upper limit of 95% CI 4.06 3.63 4.35 3.48 3.91 3.95 3.78 4.47 3.37

OR Quartile 4 4.70 2.95 3.75 3.80 3.16 3.20 3.81 3.65 3.16 p Value 9.9E-5 8.4E-4 1.2E-6 8.3E-4 5.1E-4 1.5E-5 0.0015 2.3E-4 2.3E-5

Lower limit of 95%

CI 2.16 1.56 2.20 1.74 1.65 1.89 1.66 1.83 1.85

Upper limit of 95% CI 10.2 5.55 6.39 8.30 6.06 5.41 8.70 7.27 5.37

[00434] Table 30.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 213 101 203 111 189 125

[00435] Table 30.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

p (t-test) 0.032 0.070 0.0073

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 670 1150 n (Patient) 134 182 125 191 114 202 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 17.2 40.6 16.4 40.6 15.8 41.4

Average 51.7 92.3 51.8 91.8 49.7 92.7

Stdev 119 163 120 162 120 161 p (t-test) 0.014 0.016 0.0097

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 140 176 138 178 135 181

UO only

Lower limit of 95% CI 1.40 1.45 1.68 1.44 1.50 1.50 1.44 1.58 1.60 Upper limit of 95% CI 3.97 4.11 6.41 4.07 4.26 5.19 4.08 4.50 5.25

OR Quartile 3 2.20 2.30 3.32 2.06 2.31 2.85 2.31 2.52 2.62 p Value 7.0E-4 3.2E-4 3.6E-6 0.0020 3.2E-4 2.5E-5 5.0E-4 8.1E-5 6.0E-5

Lower limit of 95% CI 1.39 1.46 2.00 1.30 1.46 1.75 1.44 1.59 1.64 Upper limit of 95% CI 3.47 3.62 5.51 3.26 3.64 4.63 3.71 3.98 4.19

OR Quartile 4 2.73 2.57 3.82 2.35 2.69 3.36 3.06 3.30 3.20 p Value 5.1E-4 8.4E-4 9.2E-7 0.0033 5.3E-4 7.1E-6 3.9E-4 6.1E-5 1.5E-5

Lower limit of 95% CI 1.55 1.48 2.24 1.33 1.54 1.98 1.65 1.84 1.89 Upper limit of 95% CI 4.82 4.47 6.52 4.14 4.72 5.70 5.68 5.93 5.41

[00436] Table 30.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 135 164 136 163 137 161 p (t-test) 0.0068 0.011 0.012

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 210 104 207 107 204 110

[00437] Table 30.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 1070 4690 1070 4690 1070 4690 n (Patient) 25 126 21 130 19 132 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 330 374 287 375 304 374

Average 428 535 423 535 433 531

Stdev 402 536 408 534 422 530 p (t-test) 0.32 0.30 0.38

Min 46.8 50.8 46.8 50.8 46.8 50.8

Max 1880 4690 1880 4690 1880 4690 n (Patient) 29 121 28 122 26 124

UO only

OR Quartile 3 1.36 1.29 1.18 1.42 1.42 1.24 1.15 1.20 1.17 p Value 0.49 0.54 0.62 0.46 0.40 0.51 0.78 0.67 0.62

Lower limit of 95% CI 0.572 0.573 0.614 0.559 0.622 0.651 0.437 0.516 0.618 Upper limit of 95% CI 3.22 2.92 2.28 3.59 3.26 2.37 3.00 2.81 2.23

OR Quartile 4 1.94 1.38 1.35 1.51 1.30 1.04 1.30 1.16 1.07 p Value 0.25 0.52 0.43 0.49 0.60 0.92 0.66 0.77 0.85

Lower limit of 95% CI 0.621 0.515 0.640 0.473 0.485 0.496 0.404 0.428 0.514 Upper limit of 95% CI 6.06 3.69 2.84 4.79 3.50 2.18 4.19 3.14 2.24

[00438] Table 30.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 50.8 46.8 50.8 46.8 50.8 46.8

Max 4690 1880 4690 1880 1540 4690 n (Patient) 100 50 93 57 85 65

[00439] Table 30.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95% CI 0.518 0.660 0.815 0.653 0.740 0.909 0.647 0.785 0.725 Upper limit of 95% CI 2.01 2.51 3.17 2.62 2.85 3.47 2.76 3.10 2.66

OR Quartile 4 1.29 1.51 1.78 1.31 1.66 1.52 1.22 1.77 1.29 p Value 0.53 0.31 0.13 0.52 0.23 0.28 0.65 0.19 0.50

Lower limit of 95% CI 0.581 0.679 0.838 0.575 0.732 0.717 0.518 0.761 0.616 Upper limit of 95% CI 2.88 3.35 3.79 2.98 3.75 3.20 2.86 4.10 2.71

[00440] Table 30.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 96 54 92 58 86 64

[00441] Table 30.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 2.29 2.29 3.72 2.41 2.29 3.66 2.41 2.29 3.61

OR Quartile 4 1.06 1.31 1.58 1.17 1.41 1.68 1.17 1.41 1.80 p Value 0.89 0.48 0.23 0.67 0.37 0.17 0.67 0.37 0.12

Lower limit of 95% CI 0.506 0.618 0.745 0.562 0.662 0.799 0.562 0.662 0.855

Upper limit of 95% CI 2.20 2.76 3.34 2.45 3.00 3.55 2.45 3.00 3.79

[00442] Example 31. Use of C-C motif chemokine 24 for evaluating renal status in patients admitted to the ICU: Recovery to RIFLE 0 and R from RIFLE I and F

[00443] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 24 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00444] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "recovered" and a "non-recovered" population. "Recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-recovered" indicates those patients whose maximum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the recovery period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "non- recovered".

[00445] The ability to distinguish the "recovered" and "non-recovered" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00446] Table 31.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Average 40.8 91.5 39.2 91.3 40.0 90.2

Stdev 87.7 166 88.2 165 89.4 164 p (t-test) 0.0035 0.0029 0.0044

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 670 1150 670 1150 670 1150 n (Patient) 107 209 103 213 100 216 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 15.8 39.8 14.5 40.3 14.5 40.4

Average 51.4 88.2 51.4 87.7 51.2 87.7

Stdev 125 157 126 156 127 156 p (t-test) 0.029 0.031 0.031

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 125 190 123 192 122 193

UO only

OR Quartile 2 2.26 2.24 3.31 2.45 2.33 4.27 2.43 2.37 3.62 p Value 0.0023 0.0022 6.8E-4 8.2E-4 0.0014 3.6E-5 9.6E-4 0.0011 6.6E-5

Lower limit of 95% CI 1.34 1.34 1.66 1.45 1.39 2.15 1.44 1.41 1.92 Upper limit of 95% CI 3.81 3.76 6.61 4.15 3.91 8.50 4.12 3.99 6.82

OR Quartile 3 2.94 2.61 3.04 3.21 2.78 3.81 2.98 2.87 3.37 p Value 1.5E-5 5.5E-5 2.2E-5 4.5E-6 2.0E-5 1.8E-7 1.9E-5 1.2E-5 8.4E-7

Lower limit of 95% CI 1.80 1.64 1.82 1.95 1.74 2.31 1.80 1.79 2.08 Upper limit of 95% CI 4.80 4.16 5.08 5.29 4.44 6.30 4.90 4.60 5.46

OR Quartile 4 3.34 2.81 3.52 3.92 2.72 3.98 3.72 2.92 3.46 p Value 2.7E-4 5.3E-4 4.3E-6 1.0E-4 8.2E-4 4.0E-7 1.9E-4 4.1E-4 4.2E-6

Lower limit of 95% CI 1.74 1.57 2.06 1.97 1.51 2.33 1.87 1.61 2.04 Upper limit of 95% CI 6.39 5.04 6.02 7.81 4.88 6.78 7.40 5.30 5.88

[00447] Table 31.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Median 20.4 69.8 19.9 69.8 19.7 55.0

Average 53.3 127 51.1 127 51.6 117

Stdev 117 195 115 192 118 183 p (t-test) 4.9E-5 1.9E-5 1.5E-4

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 225 89 219 95 206 108

[00448] Table 31.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

p (t-test) 0.0031 0.0040 0.0042

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 153 163 151 165 147 169 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 17.2 44.7 16.9 44.6 16.4 44.7

Average 47.8 102 47.8 101 47.6 101

Stdev 112 171 112 170 113 170 p (t-test) 0.0010 0.0011 0.0011

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 160 156 159 157 158 158

UO only

Lower limit of 95% CI 1.42 1.56 1.31 1.47 1.58 1.34 1.58 1.61 1.37 Upper limit of 95% CI 4.09 4.58 4.85 4.23 4.65 4.94 4.54 4.74 4.75

OR Quartile 3 2.90 2.97 2.88 2.90 3.06 2.96 2.91 3.14 2.84 p Value 5.1E-6 3.0E-6 5.6E-5 5.0E-6 1.8E-6 3.6E-5 4.9E-6 1.0E-6 3.5E-5

Lower limit of 95% CI 1.83 1.88 1.72 1.84 1.93 1.77 1.84 1.98 1.73 Upper limit of 95% CI 4.57 4.70 4.82 4.58 4.83 4.95 4.60 4.97 4.65

OR Quartile 4 3.47 3.34 3.99 3.36 3.28 3.89 3.41 3.49 3.48 p Value 1.4E-5 1.7E-5 5.2E-7 2.3E-5 2.3E-5 7.9E-7 2.3E-5 1.0E-5 4.1E-6

Lower limit of 95% CI 1.98 1.93 2.32 1.91 1.89 2.27 1.93 2.00 2.05 Upper limit of 95% CI 6.09 5.78 6.86 5.88 5.68 6.68 6.01 6.08 5.93

[00449] Table 31.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Stdev 138 161 138 161 140 156 p (t-test) 0.0057 0.0072 0.016

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 222 92 221 93 213 101

[00450] Table 31.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Max 1140 1150 1140 1150 1140 1150 n (Patient) 202 114 200 116 199 117 sCr only

Recovery Period

Duration (hr) 24 48 72

Non- Non- Non-

Recovered recovered Recovered recovered Recovered recovered Cohort Cohort Cohort Cohort Cohort Cohort

Median 18.8 46.0 18.8 46.0 18.8 46.0

Average 53.6 103 53.6 103 53.6 103

Stdev 120 173 120 173 120 173 p (t-test) 0.0028 0.0028 0.0028

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 184 132 184 132 184 132

UO only

OR Quartile 3 3.11 2.91 3.03 2.90 2.91 2.91 2.98 2.91 2.87 p Value 4.1E-6 6.7E-6 1.7E-5 1.2E-5 6.7E-6 2.9E-5 7.7E-6 6.7E-6 3.2E-5

Lower limit of 95% CI 1.92 1.83 1.83 1.80 1.83 1.76 1.85 1.83 1.75 Upper limit of 95% CI 5.04 4.63 5.01 4.68 4.63 4.80 4.80 4.63 4.72

OR Quartile 4 3.22 3.51 3.37 3.09 3.51 3.29 3.03 3.51 3.15 p Value 1.3E-5 4.0E-6 7.8E-6 2.5E-5 4.0E-6 1.1E-5 3.4E-5 4.0E-6 2.2E-5

Lower limit of 95% CI 1.90 2.06 1.98 1.83 2.06 1.93 1.80 2.06 1.85 Upper limit of 95% CI 5.45 5.98 5.74 5.23 5.98 5.60 5.12 5.98 5.35

[00451] Table 31.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 50.8 46.8 50.8 46.8 50.8 46.8

Max 4690 1880 4690 1880 4690 1880 n (Patient) 104 46 97 53 91 59

[00452] Table 31.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Lower limit of 95% CI 0.742 0.783 0.682 0.742 0.783 0.682 0.665 0.742 0.685 Upper limit of 95% CI 2.71 2.85 2.77 2.71 2.85 2.73 2.44 2.70 2.60

OR Quartile 4 1.58 1.76 1.78 1.58 1.76 1.90 1.47 1.70 1.58 p Value 0.24 0.15 0.14 0.24 0.15 0.10 0.32 0.17 0.23

Lower limit of 95% CI 0.736 0.819 0.823 0.736 0.819 0.884 0.684 0.790 0.745 Upper limit of 95% CI 3.40 3.78 3.86 3.40 3.78 4.09 3.17 3.65 3.34

[00453] Table 31.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 105 45 103 47 96 54

[00454] Table 31.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Upper limit of 95% CI 2.41 2.83 3.65 2.28 2.83 3.65 2.28 2.83 2.92

OR Quartile 4 1.35 1.61 1.43 1.30 1.61 1.43 1.40 1.61 1.36 p Value 0.43 0.21 0.35 0.48 0.21 0.35 0.37 0.21 0.42

Lower limit of 95% CI 0.645 0.768 0.669 0.623 0.768 0.669 0.666 0.768 0.642

Upper limit of 95% CI 2.82 3.39 3.08 2.73 3.39 3.08 2.94 3.39 2.89

[00455] Table 31.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "recovered" and "non-recovered" cohorts where recovery starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Recovery Period

Duration (hr) 24 48 72

sCr or sCr or sCr or

UO sCr only UO only UO sCr only UO only UO sCr only UO only

AUC 0.56 0.58 0.59 0.55 0.58 0.59 0.56 0.58 0.59

SE 0.048 0.047 0.050 0.048 0.047 0.050 0.048 0.047 0.049 p Value 0.22 0.096 0.068 0.34 0.096 0.068 0.23 0.096 0.072 nCohort Recovered 89 79 98 88 79 98 87 79 96 nCohort Non- recovered 62 72 52 63 72 52 64 72 54

Cutoff Quartile 2 218 218 217 218 218 217 218 218 217

Sensitivity 77% 79% 81 % 76% 79% 81 % 77% 79% 80%

Specificity 27% 29% 29% 26% 29% 29% 26% 29% 28%

Cutoff Quartile 3 373 373 372 373 373 372 373 373 372

Sensitivity 56% 56% 62% 56% 56% 62% 56% 56% 61 %

Specificity 54% 54% 56% 53% 54% 56% 54% 54% 56%

Cutoff Quartile 4 674 674 661 674 674 661 674 674 661

Sensitivity 27% 29% 31 % 27% 29% 31 % 28% 29% 31 %

Specificity 76% 78% 78% 76% 78% 78% 77% 78% 78%

OR Quartile 2 1.27 1.56 1.68 1.13 1.56 1.68 1.17 1.56 1.53 p Value 0.54 0.24 0.21 0.75 0.24 0.21 0.67 0.24 0.30

Lower limit of 95% CI 0.594 0.739 0.742 0.535 0.739 0.742 0.555 0.739 0.689 Upper limit of 95% CI 2.70 3.30 3.80 2.40 3.30 3.80 2.48 3.30 3.40

OR Quartile 3 1.52 1.49 2.05 1.43 1.49 2.05 1.51 1.49 2.02 p Value 0.21 0.22 0.041 0.28 0.22 0.041 0.21 0.22 0.043

Lower limit of 95% CI 0.790 0.786 1.03 0.748 0.786 1.03 0.789 0.786 1.02 Upper limit of 95% CI 2.91 2.84 4.07 2.74 2.84 4.07 2.89 2.84 3.99

OR Quartile 4 1.22 1.50 1.54 1.18 1.50 1.54 1.31 1.50 1.64 p Value 0.59 0.28 0.27 0.66 0.28 0.27 0.47 0.28 0.20

Lower limit of 95% CI 0.582 0.717 0.721 0.562 0.717 0.721 0.626 0.717 0.774 Upper limit of 95% CI 2.57 3.14 3.27 2.47 3.14 3.27 2.75 3.14 3.48

[00456] Example 32. Use of C-C motif chemokine 24 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE F

[00457] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 24 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00458] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non- persistent" indicates those patients who are not persistent at failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0), risk of injury (R), or injury (I) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00459] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00460] Table 32.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 274 40 277 37 278 36

[00461] Table 32.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only p Value 2.6E-7 3.6E-6 1.4E-6 1.5E-8 1.1E-6 8.5E-7 1.3E-7 7.1E-6 5.0E-7

Lower limit of 95% CI 2.42 2.06 2.39 2.77 2.20 2.53 2.49 1.99 2.63

Upper limit of 95% CI 7.14 5.96 7.91 8.18 6.37 8.60 7.29 5.77 9.03

[00462] Table 32.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

UO UO UO

AUC 0.67 0.65 0.67 0.70 0.67 0.67 0.67 0.65 0.67

SE 0.031 0.032 0.039 0.031 0.032 0.040 0.033 0.033 0.040 p Value 4.2E-8 1.1E-6 1.3E-5 1.2E-10 5.1E-8 1.5E-5 1.2E-7 8.7E-6 1.5E-5 nCohort Non- persistent 174 181 245 189 194 250 204 207 250 nCohort Persistent 142 134 69 127 121 64 112 108 64

Cutoff Quartile 2 4.17 4.15 4.13 4.17 4.15 4.13 4.17 4.15 4.13

Sensitivity 82% 82% 86% 85% 85% 86% 85% 85% 86%

Specificity 30% 30% 28% 32% 31 % 28% 30% 30% 28%

Cutoff Quartile 3 29.6 29.3 28.8 29.6 29.3 28.8 29.6 29.3 28.8

Sensitivity 64% 63% 68% 68% 66% 69% 67% 65% 69%

Specificity 61 % 60% 55% 62% 60% 55% 59% 57% 55%

Cutoff Quartile 4 74.9 74.4 74.6 74.9 74.4 74.6 74.9 74.4 74.6

Sensitivity 39% 38% 46% 42% 40% 47% 41 % 39% 47%

Specificity 86% 85% 81 % 86% 84% 80% 84% 82% 80%

OR Quartile 2 1.95 2.00 2.31 2.64 2.62 2.38 2.44 2.52 2.38 p Value 0.014 0.012 0.024 9.4E-4 0.0012 0.025 0.0034 0.0029 0.025

Lower limit of 95% CI 1.15 1.16 1.12 1.49 1.46 1.11 1.34 1.37 1.11 Upper limit of 95% CI 3.33 3.44 4.78 4.70 4.71 5.07 4.43 4.62 5.07

OR Quartile 3 2.85 2.57 2.62 3.41 2.90 2.67 2.96 2.49 2.67 p Value 7.8E-6 6.0E-5 8.3E-4 3.9E-7 1.0E-5 1.0E-3 1.1E-5 2.0E-4 1.0E-3

Lower limit of 95% CI 1.80 1.62 1.49 2.12 1.81 1.49 1.82 1.54 1.49 Upper limit of 95% CI 4.51 4.07 4.62 5.47 4.66 4.78 4.79 4.03 4.78

OR Quartile 4 3.95 3.36 3.64 4.49 3.46 3.62 3.61 2.92 3.62 p Value 8.7E-7 8.4E-6 8.8E-6 6.2E-8 4.4E-6 1.5E-5 2.0E-6 6.3E-5 1.5E-5

Lower limit of 95% CI 2.29 1.97 2.06 2.61 2.04 2.02 2.13 1.73 2.02 Upper limit of 95% CI 6.83 5.72 6.44 7.73 5.87 6.48 6.13 4.94 6.48

[00463] Table 32.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Average 58.8 93.0 56.9 99.2 65.2 88.8

Stdev 146 144 142 149 160 116 p (t-test) 0.039 0.012 0.17

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1150 1140 1150 1140 1150 679 n (Patient) 179 136 191 124 203 112

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 23.8 71.6 24.1 73.6 24.1 73.6

Average 62.3 111 61.9 117 61.9 117

Stdev 137 169 135 175 135 175 p (t-test) 0.011 0.0053 0.0053

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 238 76 244 70 244 70

[00464] Table 32.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

nCohort Non- persistent 168 175 233 183 187 236 196 199 236 nCohort Persistent 148 140 81 133 128 78 120 116 78

Cutoff Quartile 2 4.17 4.15 4.13 4.17 4.15 4.13 4.17 4.15 4.13

Sensitivity 82% 81 % 86% 85% 84% 87% 86% 84% 87%

Specificity 31 % 30% 29% 32% 32% 29% 32% 31 % 29%

Cutoff Quartile 3 29.6 29.3 28.8 29.6 29.3 28.8 29.6 29.3 28.8

Sensitivity 64% 64% 68% 67% 66% 69% 67% 66% 69%

Specificity 62% 61 % 56% 62% 61 % 56% 60% 59% 56%

Cutoff Quartile 4 74.9 74.4 74.6 74.9 74.4 74.6 74.9 74.4 74.6

Sensitivity 38% 38% 44% 41 % 40% 45% 41 % 40% 45%

Specificity 86% 85% 82% 86% 85% 81 % 85% 83% 81 %

OR Quartile 2 2.01 1.90 2.62 2.69 2.49 2.81 2.80 2.41 2.81 p Value 0.0099 0.018 0.0066 6.4E-4 0.0017 0.0050 6.8E-4 0.0033 0.0050

Lower limit of 95% CI 1.18 1.12 1.31 1.52 1.41 1.37 1.55 1.34 1.37 Upper limit of 95% CI 3.41 3.25 5.26 4.74 4.39 5.78 5.08 4.32 5.78

OR Quartile 3 2.83 2.68 2.72 3.34 3.09 2.91 3.03 2.71 2.91 p Value 8.3E-6 2.5E-5 2.4E-4 4.6E-7 2.6E-6 1.3E-4 5.0E-6 4.0E-5 1.3E-4

Lower limit of 95% CI 1.79 1.69 1.59 2.09 1.93 1.68 1.88 1.68 1.68 Upper limit of 95% CI 4.47 4.24 4.63 5.34 4.94 5.01 4.87 4.36 5.01

OR Quartile 4 3.84 3.49 3.53 4.32 3.76 3.55 3.82 3.31 3.55 p Value 1.7E-6 5.4E-6 6.6E-6 1.5E-7 1.2E-6 7.2E-6 8.2E-7 8.8E-6 7.2E-6

Lower limit of 95% CI 2.21 2.04 2.04 2.50 2.20 2.04 2.24 1.95 2.04 Upper limit of 95% CI 6.66 5.98 6.12 7.46 6.42 6.18 6.50 5.60 6.18

[00465] Table 32.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

Max 1880 4690 1880 4690 4690 1700 n (Patient) 94 56 103 47 108 42

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 364 392 364 392 350 394

Average 518 435 518 435 517 441

Stdev 531 260 531 260 529 265 p (t-test) 0.50 0.50 0.55

Min 46.8 66.1 46.8 66.1 46.8 66.1

Max 4690 1050 4690 1050 4690 1050 n (Patient) 131 19 131 19 132 18

[00466] Table 32.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria. sCr or UO sCr only

UO only Sensitivity 54% 55% 56% 54% 56% 56% 50% 53% 58% Specificity 52% 53% 51 % 52% 54% 51 % 50% 52% 52%

Cutoff Quartile 4 674 661 661 674 661 661 674 661 661

Sensitivity 30% 31 % 26% 32% 32% 26% 26% 27% 27%

Specificity 78% 78% 75% 79% 78% 75% 75% 76% 75%

OR Quartile 2 1.53 1.50 1.61 1.69 1.71 1.61 1.53 1.62 1.52 p Value 0.28 0.30 0.37 0.20 0.19 0.37 0.31 0.25 0.43

Lower limit of 95% CI 0.709 0.695 0.565 0.761 0.772 0.565 0.677 0.714 0.531 Upper limit of 95% CI 3.28 3.22 4.61 3.73 3.79 4.61 3.48 3.66 4.37

OR Quartile 3 1.28 1.39 1.31 1.30 1.49 1.31 0.980 1.20 1.45 p Value 0.45 0.32 0.52 0.44 0.24 0.52 0.95 0.61 0.39

Lower limit of 95% CI 0.671 0.725 0.568 0.671 0.767 0.568 0.498 0.608 0.619 Upper limit of 95% CI 2.46 2.67 3.03 2.51 2.89 3.03 1.93 2.35 3.42

OR Quartile 4 1.57 1.60 1.04 1.71 1.68 1.04 1.07 1.18 1.11 p Value 0.23 0.21 0.94 0.16 0.17 0.94 0.87 0.67 0.84

Lower limit of 95% CI 0.748 0.765 0.401 0.810 0.799 0.401 0.491 0.549 0.424 Upper limit of 95% CI 3.29 3.37 2.69 3.60 3.55 2.69 2.32 2.55 2.88

[00467] Table 32.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 332 396 332 396 331 397

Average 498 540 498 540 496 547

Stdev 538 383 538 383 536 386 p (t-test) 0.66 0.66 0.61

Min 46.8 66.1 46.8 66.1 46.8 66.1

Max 4690 1510 4690 1510 4690 1510 n (Patient) 115 35 115 35 116 34

[00468] Table 32.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Cohort Cohort Cohort Cohort Cohort Cohort

Median 332 394 330 397 330 397

Average 455 585 447 603 498 541

Stdev 363 638 360 650 576 398 p (t-test) 0.12 0.063 0.61

Min 50.8 46.8 46.8 50.8 46.8 63.9

Max 1880 4690 1880 4690 4690 1700 n (Patient) 81 70 85 66 91 60 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 330 396 329 397 329 397

Average 442 598 440 608 492 547

Stdev 359 642 356 652 576 403 p (t-test) 0.064 0.045 0.52

Min 46.8 50.8 46.8 50.8 46.8 63.9

Max 1880 4690 1880 4690 4690 1700 n (Patient) 81 69 84 66 90 60

UO only

OR Quartile 2 1.26 1.43 1.57 1.46 1.49 1.85 1.88 1.91 1.77 p Value 0.54 0.35 0.32 0.33 0.31 0.19 0.12 0.11 0.22

Lower limit of 95% CI 0.599 0.675 0.650 0.686 0.697 0.741 0.850 0.863 0.708 Upper limit of 95% CI 2.64 3.02 3.77 3.11 3.17 4.62 4.16 4.22 4.43

OR Quartile 3 1.34 1.46 1.61 1.50 1.54 1.74 1.53 1.56 1.88 p Value 0.37 0.25 0.20 0.22 0.19 0.14 0.21 0.18 0.096

Lower limit of 95% CI 0.707 0.765 0.777 0.788 0.807 0.832 0.792 0.809 0.893 Upper limit of 95% CI 2.55 2.78 3.32 2.87 2.95 3.62 2.94 3.02 3.97

OR Quartile 4 1.61 1.65 1.32 1.62 1.59 1.38 1.32 1.30 1.45 p Value 0.21 0.19 0.50 0.20 0.22 0.43 0.47 0.49 0.37

Lower limit of 95% CI 0.770 0.785 0.591 0.773 0.761 0.618 0.626 0.617 0.647 Upper limit of 95% CI 3.38 3.46 2.95 3.39 3.34 3.10 2.77 2.73 3.26

[00469] Table 32.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

p (t-test) 0.93 0.93 0.88

Min 50.8 46.8 50.8 46.8 50.8 46.8

Max 4690 1510 4690 1510 4690 1510 n (Patient) 108 42 108 42 109 41

[00470] Example 33. Use of C-C motif chemokine 24 for evaluating renal status in patients admitted to the ICU: Persistent at RIFLE I or F

[00471] Patients from the intensive care unit (ICU) with RIFLE stage of injury (I) or failure (F) are enrolled in the following study. EDTA anti-coagulated blood samples (10 mL) and urine samples (50 mL) are collected from each patient at enrollment, and at every 12 hours up to day 3, and then every 24 hours thereafter up to day 7 while the subject is hospitalized. C-C motif chemokine 24 is measured in the enrollment samples by standard immunoassay methods using commercially available assay reagents.

[00472] Kidney status is assessed by RIFLE criteria based on serum creatinine, urine output, or both serum creatinine and urine output. Two cohorts are defined to represent a "persistent" and a "non-persistent" population. "Persistent" indicates those patients whose minimum RIFLE stage during a period of 24, 48 or 72 hours is injury (I) or failure (F) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. "Non-persistent" indicates those patients who are not persistent at injury (I) or failure (F) and whose minimum RIFLE stage during a period of 24, 48 or 72 hours is non-injury (RIFLE 0) or risk of injury (R) where the persistence period can start from the time of sample collection to 24, 48, 72, 96 or 168 hours after sample collection. If a patient dies after injury (I) or failure (F) or is placed on renal replacement therapy (RRT) at any time from sample collection to 24, 48, 72, 96 or 168 hours after sample collection, the patient is considered "persistent".

[00473] The ability to distinguish the "persistent" and "non-persistent" cohorts is determined using receiver operating characteristic (ROC) analysis.

[00474] Table 33.1: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 22.9 73.9 24.4 89.6 25.6 97.6

Average 57.8 127 59.3 138 60.8 142

Stdev 131 181 130 191 131 197 p (t-test) 3.2E-4 1.7E-4 2.6E-4

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 240 74 255 59 262 52

[00475] Table 33.2: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 14.3 40.4 17.9 42.7 17.9 44.7

Average 47.1 89.0 47.9 97.0 49.1 101

Stdev 125 155 115 166 114 171 p (t-test) 0.015 0.0028 0.0014

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 111 205 146 170 164 152 sCr only

UO only

Specificity 67% 64% 59% 62% 61 % 57% 62% 62% 56%

Cutoff Quartile 4 74.9 74.4 74.6 74.9 74.4 74.6 74.9 74.4 74.6

Sensitivity 32% 32% 46% 35% 35% 49% 37% 37% 50%

Specificity 88% 86% 84% 86% 86% 83% 86% 85% 82%

OR Quartile 2 2.23 2.24 3.31 2.20 2.39 3.17 2.49 2.70 3.18 p Value 0.0025 0.0022 6.8E-4 0.0030 0.0012 0.0026 8.8E-4 4.0E-4 0.0039

Lower limit of 95% CI 1.33 1.34 1.66 1.31 1.41 1.50 1.46 1.56 1.45 Upper limit of 95% CI 3.76 3.76 6.61 3.70 4.06 6.70 4.27 4.67 6.98

OR Quartile 3 2.88 2.61 3.75 2.54 2.50 3.59 2.82 2.88 3.20 p Value 1.8E-5 5.5E-5 8.6E-7 5.7E-5 7.3E-5 8.8E-6 8.5E-6 6.1E-6 7.7E-5

Lower limit of 95% CI 1.78 1.64 2.22 1.61 1.59 2.04 1.79 1.82 1.80 Upper limit of 95% CI 4.67 4.16 6.35 4.01 3.94 6.31 4.45 4.56 5.70

OR Quartile 4 3.58 2.81 4.42 3.35 3.23 4.66 3.58 3.32 4.55 p Value 1.2E-4 5.3E-4 7.7E-8 3.0E-5 3.4E-5 7.0E-8 5.7E-6 1.4E-5 2.0E-7

Lower limit of 95% CI 1.87 1.57 2.57 1.90 1.86 2.66 2.06 1.93 2.57 Upper limit of 95% CI 6.85 5.04 7.60 5.91 5.63 8.15 6.20 5.71 8.05

[00476] Table 33.3: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only Cohort Cohort Cohort Cohort Cohort Cohort

Median 20.3 54.7 22.9 64.0 23.7 69.8

Average 58.5 106 60.1 109 61.7 109

Stdev 138 159 138 163 138 164 p (t-test) 0.0064 0.0069 0.012

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1140 1150 1140 1150 1140 1150 n (Patient) 211 103 224 90 231 83

[00477] Table 33.4: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

Stdev 126 155 117 164 117 167 p (t-test) 0.019 0.0036 0.0032

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 109 207 142 174 155 161 sCr only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 14.5 39.7 16.9 42.7 16.9 44.6

Average 48.7 89.5 45.9 98.4 48.1 100

Stdev 120 159 111 168 112 171 p (t-test) 0.015 0.0014 0.0014

Min 0.00127 0.00127 0.00127 0.00127 0.00127 0.00127

Max 1070 1150 1070 1150 1070 1150 n (Patient) 123 192 149 166 161 154

UO only

Upper limit of 95% CI 3.65 3.91 4.57 3.97 4.35 5.22 4.26 4.46 5.33

OR Quartile 3 2.74 2.62 3.14 2.70 2.65 3.20 3.06 2.95 2.93 p Value 4.4E-5 5.3E-5 5.2E-6 2.1E-5 2.8E-5 8.7E-6 1.8E-6 3.8E-6 5.1E-5

Lower limit of 95% CI 1.69 1.64 1.92 1.71 1.68 1.92 1.93 1.86 1.74 Upper limit of 95% CI 4.44 4.19 5.14 4.26 4.17 5.35 4.83 4.66 4.92

OR Quartile 4 3.46 2.97 3.69 3.41 3.27 3.90 3.31 3.17 3.61 p Value 1.8E-4 3.3E-4 1.5E-6 3.0E-5 3.4E-5 7.0E-7 2.3E-5 3.3E-5 3.3E-6

Lower limit of 95% CI 1.81 1.64 2.17 1.92 1.87 2.28 1.90 1.84 2.10 Upper limit of 95% CI 6.61 5.39 6.28 6.06 5.74 6.67 5.77 5.47 6.19

[00478] Table 33.5: Comparison of marker levels and the area under the ROC curve (AUC) in urine samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

n (Patient) 201 113 211 103 216 98

[00479] Table 33.6: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 24 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent

Cohort Cohort Cohort Cohort Cohort Cohort

Median 327 386 371 374 376 332

Average 437 555 481 545 491 544

Stdev 379 569 387 610 393 642 p (t-test) 0.18 0.45 0.54

Min 46.8 50.8 46.8 50.8 46.8 50.8

Max 1880 4690 1880 4690 1880 4690 n (Patient) 52 98 73 77 86 64

UO only

Upper limit of 95% CI 4.25 3.80 4.04 3.67 3.00 3.92 2.55 2.04 3.80

[00480] Table 33.7: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 48 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

SE 0.049 0.048 0.051 0.047 0.047 0.054 0.047 0.047 0.057 p Value 0.22 0.097 0.39 0.58 0.32 0.28 0.69 0.36 0.33 nCohort Non- persistent 49 51 103 70 70 111 79 79 116 nCohort Persistent 102 99 47 81 80 39 72 71 34

Cutoff Quartile 2 218 217 217 218 217 217 218 217 217

Sensitivity 75% 77% 77% 74% 75% 79% 75% 76% 79%

Specificity 27% 29% 26% 24% 26% 27% 25% 27% 27%

Cutoff Quartile 3 373 372 372 373 372 372 373 372 372

Sensitivity 53% 54% 53% 52% 52% 56% 50% 51 % 59%

Specificity 55% 57% 51 % 51 % 53% 52% 49% 51 % 53%

Cutoff Quartile 4 674 661 661 674 661 661 674 661 661

Sensitivity 28% 28% 34% 30% 30% 33% 28% 28% 32%

Specificity 82% 80% 79% 80% 80% 77% 77% 77% 77%

OR Quartile 2 1.11 1.38 1.16 0.916 1.04 1.44 1.02 1.15 1.41 p Value 0.79 0.41 0.71 0.82 0.92 0.42 0.96 0.71 0.47

Lower limit of 95% CI 0.511 0.643 0.520 0.438 0.497 0.594 0.487 0.549 0.556 Upper limit of 95% CI 2.42 2.95 2.60 1.92 2.17 3.47 2.12 2.41 3.56

OR Quartile 3 1.38 1.52 1.20 1.14 1.24 1.42 0.975 1.05 1.58 p Value 0.36 0.23 0.60 0.69 0.51 0.35 0.94 0.87 0.24

Lower limit of 95% CI 0.697 0.769 0.604 0.601 0.652 0.679 0.515 0.556 0.731 Upper limit of 95% CI 2.74 3.00 2.40 2.16 2.36 2.95 1.85 2.00 3.44

OR Quartile 4 1.77 1.62 1.90 1.68 1.71 1.72 1.30 1.33 1.58 p Value 0.19 0.25 0.10 0.18 0.16 0.18 0.48 0.45 0.29

Lower limit of 95% CI 0.761 0.714 0.884 0.791 0.805 0.772 0.624 0.636 0.682 Upper limit of 95% CI 4.10 3.66 4.09 3.58 3.65 3.83 2.72 2.78 3.64

[00481] Table 33.8: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 72 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

p (t-test) 0.17 0.16 0.13

Min 46.8 50.8 46.8 50.8 46.8 50.8

Max 1880 4690 1880 4690 1880 4690 n (Patient) 51 99 69 81 75 75

UO only

Persistence Period

Duration (hr) 24 48 72

Non-persistent Persistent Non-persistent Persistent Non-persistent Persistent Cohort Cohort Cohort Cohort Cohort Cohort

Median 334 394 332 396 331 397

Average 491 538 495 538 501 527

Stdev 551 408 544 403 546 376 p (t-test) 0.59 0.63 0.78

Min 50.8 46.8 50.8 46.8 50.8 46.8

Max 4690 1700 4690 1700 4690 1510 n (Patient) 98 52 105 45 110 40

[00482] Table 33.9: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 96 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

Cutoff Quartile 2 218 217 217 218 217 217 218 217 217

Sensitivity 75% 77% 78% 75% 76% 80% 77% 78% 80%

Specificity 27% 29% 27% 25% 26% 28% 27% 28% 28%

Cutoff Quartile 3 373 372 372 373 372 372 373 372 372

Sensitivity 53% 54% 58% 53% 54% 60% 53% 54% 62%

Specificity 55% 57% 55% 53% 54% 55% 53% 54% 55%

Cutoff Quartile 4 674 661 661 674 661 661 674 661 661

Sensitivity 28% 28% 33% 29% 29% 32% 27% 28% 31 %

Specificity 82% 80% 79% 79% 79% 78% 77% 77% 77%

OR Quartile 2 1.11 1.38 1.35 0.984 1.12 1.56 1.21 1.38 1.53 p Value 0.79 0.41 0.45 0.97 0.77 0.29 0.61 0.40 0.33

Lower limit of 95% CI 0.511 0.643 0.617 0.470 0.534 0.686 0.582 0.657 0.655 Upper limit of 95% CI 2.42 2.95 2.95 2.06 2.33 3.53 2.53 2.88 3.56

OR Quartile 3 1.38 1.52 1.68 1.27 1.38 1.83 1.27 1.38 2.03 p Value 0.36 0.23 0.13 0.47 0.33 0.085 0.47 0.33 0.052

Lower limit of 95% CI 0.697 0.769 0.860 0.668 0.725 0.920 0.670 0.725 0.994 Upper limit of 95% CI 2.74 3.00 3.29 2.41 2.63 3.65 2.40 2.62 4.15

OR Quartile 4 1.77 1.62 1.82 1.57 1.60 1.67 1.26 1.28 1.52 p Value 0.19 0.25 0.12 0.24 0.23 0.19 0.54 0.51 0.29

Lower limit of 95% CI 0.761 0.714 0.863 0.737 0.749 0.781 0.601 0.611 0.700 Upper limit of 95% CI 4.10 3.66 3.86 3.34 3.40 3.57 2.63 2.68 3.32

[00483] Table 33.10: Comparison of marker levels and the area under the ROC curve (AUC) in EDTA samples for the "persistent" and "non-persistent" cohorts where persistence starts within 168 hours after sample collection and renal status is assessed by serum creatinine (sCr) only, urine output (UO) only, or serum creatinine or urine output RIFLE criteria.

sCr or UO

sCr only

UO only

[00484] While the invention has been described and exemplified in sufficient detail for those skilled in this art to make and use it, various alternatives, modifications, and improvements should be apparent without departing from the spirit and scope of the invention. The examples provided herein are representative of preferred embodiments, are exemplary, and are not intended as limitations on the scope of the invention. Modifications therein and other uses will occur to those skilled in the art. These modifications are encompassed within the spirit of the invention and are defined by the scope of the claims.

[00485] It will be readily apparent to a person skilled in the art that varying substitutions and modifications may be made to the invention disclosed herein without departing from the scope and spirit of the invention.

[00486] All patents and publications mentioned in the specification are indicative of the levels of those of ordinary skill in the art to which the invention pertains. All patents and publications are herein incorporated by reference to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference.

[00487] The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. Thus, for example, in each instance herein any of the terms "comprising", "consisting essentially of and "consisting of may be replaced with either of the other two terms. The terms and expressions which have been employed are used as terms of description and not of limitation, and there is no intention that in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments and optional features, modification and variation of the concepts herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention as defined by the appended claims.

[00488] Other embodiments are set forth within the following claims.