THERAPY FOR HEPATOCELLULAR CARCINOMA

The present invention relates to the field of cancer treatment. More specifically, the present invention relates to the use of an anti-VEGFR-2 antibody, preferably ramucirumab, for the treatment of hepatocellular carcinoma (HCC) in patients having elevated levels of human vascular endothelial growth factor D (VEGF-D). VEGF-D is a ligand for the human vascular endothelial growth factor receptor-2 (VEGFR-2).

Hepatocellular cancer is the third most common cause of cancer death. The primary systemic treatment for HCC is sorafenib, a kinase inhibitor indicated for the treatment of unresectable hepatocellular carcinoma. Yet despite sorafenib’ s efficacy in some patients, sorafenib therapy exhibits a high resistance rate. Thus, there exists a need to distinguish amongst resistance mechanisms in order to find biomarkers that may identify which patients would benefit most from the diverse class of drugs being investigated for second line therapy.

Ramucirumab has been investigated in a Phase III (REACH) trial as a second-line treatment for individuals with hepatocellular carcinoma. The global, randomized, double- blind REACH trial compared ramucirumab plus best supportive care to placebo plus best supportive care as a second-line treatment in patients with HCC after being treated with sorafenib in the first-line setting. Median overall survival (OS) was 9.17 months on the ramucirumab arm compared to 7.62 months on the placebo arm (HR 0.866; 95% Cl: 0.717-1.046; P= 0.1391).

It has surprisingly been found that HCC patients who have previously been treated with sorafenib and display elevated plasma VEGF-D levels receive a greater survival benefit from treatment with ramucirumab than those patients with lower plasma VEGF-D levels. Despite the role that VEGF-D plays in the growth and lymphatic spread of HCC. (Int. J. Cancer: 122, 2471-2481 (2008)), a different, single-arm clinical study in patients with HCC treated with bevacizumab showed no correlation of VEGF-D levels with OS (HR 0.97; 95% Cl: 0.59-1.62; P= 0.91) or with PFS (HR 1.26; 95% Cl: 0.75-2.12; P= 0.39).

Despite the correlation of elevated VEGF-D levels and ramuncirumab efficacy observed in patients previously treated with sorafenib, no correlation between elevated VEGF-D levels and benefit from ramucirumab treatment was observed in a Phase III (RAINFALL) clinical trial in patients with metastatic gastric or gastroesophageal junction adenocarcinoma (NCT02314117).

The use of an anti-VEGFR-2 antibody, preferably ramucirumab, for the treatment of heaptocellular carcinoma (HCC) in patients having elevated levels of human vascular endothelial growth factor D was suprisingly discovered during a Phase 3 clinical trial of ramucirumab (“Randomized, Double-Blind, Phase 3 Study of Ramucirumab (IMC- 1121B) Drug Product and Best Supportive Care (BSC) Versus Placebo and BSC as Second-Line Treatment in Patients With Hepatocellular Carcinoma Following First-Line Therapy With Sorafenib”) (the“Study”).

As used herein, the term“about” means a deviation from a given value by no more or less than 10%. As a non-limiting example, when used in regards to weight, “about 100 mg” denotes a range from 90 mg (inclusive) to 110 mg (inclusive).

As used herein, the term “human VEGFR-2” refers to Human Vascular Endothelial Growth Factor Receptor 2 having the amino acid sequence of SEQ ID NO: 5. VEGFR-2 is also known as Fetal Liver Kinase-l (FLK1), and Kinase Insert Domain Receptor (KDR).

As used herein, the term“human VEGF-D” refers to human vascular endothelial growth factor- D having the amino acid sequence of SEQ ID NO: 6.

Ramucirumab is a human IgGl monoclonal antibody directed against human vascular endothelial growth factor receptor 2 (VEGFR-2). Ramucirumab and methods of making and using ramucirumab have been previously disclosed. Ramucirumab is approved by the United States Food and Drug Administration as a single agent or in combination with paclitaxel, for the treatment of advanced gastric or gastro-esophageal junction adenocarcinoma, with disease progression on or after prior fluoropyrimidine- or platinum-containing chemotherapy; in combination with docetaxel, for the treatment of metastatic non-small cell lung cancer with disease progression on or after platinum-based chemotherapy; and in combination with FOLFIRI (irinotecan, folinic acid, and 5- fluorouracil), for the treatment of metastatic colorectal cancer with disease progression on or after prior therapy with bevacizumab, oxaliplatin, and a fluoropyrimidine. A non-limiting example of ramucirumab is CYRAMZA® and has the CAS registry number 947687-13-0. Ramucirumab is an anti-human VEGFR-2 antibody comprising two light chains, each of the light chains having the amino acid sequence of SEQ ID NO: 3, and two heavy chains, each of the heavy chains having the amino acid sequence of SEQ ID NO: 4. The light chain variable region of ramucirumab is that given in SEQ ID NO: 1. The heavy chain variable region of ramucirumab is that given in SEQ ID NO: 2.

As used herein, “sorafenib” refers to 4-[4-({[4-chloro-3- (trifluoromethyl)phenyl]carbamoyl}amino)phenoxy]-N-methylpyr idine-2-carboxamide and salts thereof. Non-limiting examples of“sorafenib” include sorafenib tosylate and NEXAVAR®.

Sorafenib is a small molecule Raf kinase and VEGF receptor kinase inhibitor, and is approved by the United States Food and Drug Administration for patients with unresectable hepatocellular carcinoma.

As used herein, “regorafenib” refers to 4-[4-({[4-chloro-3- (trifluoromethyl)phenyl]carbamoyl}amino)-3-fluorophenoxy]-N- methylpyridine-2- carboxamide monohydrate and salts thereof. Non-limiting examples of “regorafenib” include regorafenib monohydrate, regorafenib hydrate, and STIVARGA®.

Regorafenib is a small molecule inhibitor of VEGFR, Ret, Kit, PDGFR, and Raf and is approved by the United States Food and Drug Administration for the treatment of hepatocellular carcinoma in patients who have been previously treated with sorafenib.

As used herein, the terms“treating,”“treat,” or“treatment” refer to restraining, slowing, lessening, reducing, or reversing the progression or severity of an existing symptom, disorder, condition, or disease, or ameliorating clinical symptoms of a condition. Beneficial or desired clinical results include, but are not limited to, alleviation disease or disorder (i.e., where the disease or disorder does not worsen), delay or slowing of the progression of a disease or disorder, amelioration or palliation of the disease or disorder, and remission (whether partial or total) of the disease or disorder, whether detectable or undetectable. Treatment can also mean prolonging survival as compared to expected survival if not receiving treatment. Those in need of treatment include those already with the disease. In some embodiments, the present invention can be used as a medicament.

As used herein, the term “cancer” refers to or describes the physiological condition in mammals that is typically characterized by unregulated cell growth. Included in this definition are benign and malignant cancers.

In the methods of the present invention, a therapeutically effective amount of an anti-VEGFR-2 antibody described herein is administered to a mammal or patient in need thereof. Additionally, the pharmaceutical compositions of the invention may include a therapeutically effective amount of an anti-VEGFR-2 antibody described herein.

A“therapeutically effective amount,” refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic result. A therapeutically effective amount can be readily determined by the attending diagnostician, as one skilled in the art, by the use of known techniques and by observing results obtained under analogous circumstances. In determining the therapeutically effective amount for a patient, a number of factors are considered by the attending diagnostician, including, but not limited to: the species of patient; its size, age, and general health; the specific disease or disorder involved; the target site; the degree of the severity of the disease or disorder; the response of the individual patient; the particular compound administered; the mode of administration; the bioavailability characteristics of the preparation administered; the dose regimen selected; the use of concomitant medication; other medications administered; and other relevant circumstances. A therapeutically effective amount is also one in which any toxic or detrimental effects of the antibody or antibody portion are outweighed by the therapeutically beneficial effects.

Generally, dosage regimens may be adjusted to provide the optimum desired response ( e.g ., a therapeutic response). Treatment dosages may be titrated using routine methods known to those of skill in the art to optimize safety and efficacy. Dosing schedules will typically range from a single bolus dosage or continuous infusion, to multiple administrations per day (e.g., every 4-6 hours), or as indicated by the treating physician and the patient’s condition. Dosing frequencies of the antibody will be determined by the physicians treating the patient and may be given daily, three times per week, weekly, every two weeks, or less often, and more preferably every two-weeks. Dosing amounts of the antibody will also be determined by the physicians treating the patient and may fall within customary ranges, more preferably about 8 mg/kg every two or three weeks.

In some instances, dosage levels below the lower limit of the aforesaid dosing for ramucirumab may be more than adequate, while in other cases larger doses may be employed with acceptable side effects, and therefore the above dosage amount is not intended to limit the scope of the invention in any way.

The therapeutically effective amount of the treatment of the invention can be measured by various endpoints commonly used in evaluating cancer treatments, including, but not limited to: extending survival (including Overall Survival (OS) and Progression Free Survival (PFS)); resulting in an objective response (including a Complete Response (CR) or a Partial Response (PR)); tumor regression, tumor weight or size shrinkage, longer time to disease progression, increased duration of survival, longer PFS, improved OS rate, increased duration of response, and improved quality of life and/or improving signs or symptoms of cancer.

As used herein, the term“progressive disease” (PD) refers to least a 20% increase in the sum of diameters of target lesions, taking as reference the smallest sum on study (this includes the baseline sum if that is the smallest on study). In addition to the relative increase of 20%, the sum must also demonstrate an absolute increase of at least 5 mm. The appearance of one or more new lesions is also considered progression.

As used herein, the term“partial response,” (PR) refers to at least a 30% decrease in the sum of diameters of target lesions, taking as reference the baseline sum diameters.

As used herein, the term“complete response” (CR) refers to the disappearance of all non-nodal target lesions with the short axes of any target lymph nodes reduced to <10 mm .

As used herein, the term “stable disease” (SD) refers to neither sufficient shrinkage for PR nor sufficient increase to qualify for PD, taking as reference the smallest sum of diameters while on study.

As used herein, the term “objective response rate” (ORR) is equal to the proportion of patients achieving a best overall response of partial or complete response (PR and CR) according to RECIST 1.1. As used herein, the term“overall survival” (OS) refers to the percentage of patients remaining alive for a defined period of time, such as 1 year, 5 years, etc. from the time of diagnosis or treatment. In a preferred embodiment, OS refers to the time from the date of randomization in the Study to the date of death from any cause. If the patient is alive at the end of the follow-up period or is lost to follow-up, OS data is censored on the last date the patient is known to be alive. Overall survival is evaluated by the Kaplan- Meier method, and a 95% confidence interval (Cl) is provided for the median OS in each treatment arm.

As used herein, the term“progression-free survival” (PFS) refers to the patient remaining alive without the cancer progressing or getting worse. In a preferred aspect of the invention, PFS is defined as the time from randomization in the Study until the first radiographic documentation of objective progression as defined by RECIST (Version 1.1), or death from any cause. Patients who die without a reported prior progression areconsidered to have progressed on the day of their death. Patients who did not progress or are lost to follow-up are censored at the day of their last radiographic tumor assessment. In a preferred embodiment, progression-free survival is analyzed using the log-rank test, stratified by geographic region (North America versus Europe versus all other regions), and etiology of liver disease (hepatitis B vs hepatitis C vs other etiologies).

As used herein, the term“time to progression” (TTP) refers to the time from randomization until the date of radiographic progression.

As used herein, the term“disease control rate” (DCR) refers to lack of disease progression and rate thereof. It refers to the group of patients with a best overall response categorized as CR, PR or SD (specifically excluding the patients with PD), wherein the best overall response is the best response recorded from the start of treatment until PD.

As used herein, the term“clinical benefit rate,” refers to SD or better at 12 weeks. The tumor response rate of SD or better (i.e. CR+PR+SD) at 12 weeks is defined as the proportion of patients with a response of SD or better, as defined by RECIST 1.1, at 12 weeks following the first dose of study therapy. Patients are considered“failure” if they die or if radiographic evaluation indicates a response of PD at 12 weeks or before.

As used herein, the term“extending survival” or“prolonged survival” which are used interchangeably herein, is meant as increasing OS or PFS in a treated patient relative to i) an untreated patient, ii) a patient treated with less than all of the anti-tumor agents in a particular combination therapy, or iii) a control treatment protocol. Survival is monitored following the initiation of treatment or following the initial diagnosis of cancer.

In the present invention, any suitable method or route can be used to administer an anti-VEGFR-2 antibody described herein, although intravenous (i.v.) administration is the preferred route. It should be emphasized, however, that the present invention is not limited to any particular method or route of administration.

The anti-human VEGFR-2 antibodies, including but not limited to ramucirumab, where used in a patient for the purpose of treatment, is preferably formulated as a pharmaceutical composition. Such pharmaceutical compositions and processes for preparing the same are well known in the art. See, e.g. Remington: The Science and Practice of Pharmacy (Gennaro A., et al. , eds., 19^ ed., Mack Publishing Co., 1995).

VEGF-D can be collected from patient blood by techniques known in the art. Levels of VEGF-D are measured as described herein. Serum and plasma can be derived by methods known in the art. The level of VEGF-D has been observed to be predictive of or indicative of treatment efficacy with ramucirumab in patients with HCC.

ETnless indicated otherwise, the term“antibody” refers to an immunoglobulin molecule comprising two heavy chains (HC) and two light chains (LC) interconnected by disulfide bonds. The amino terminal portion of each chain includes a variable region of about 100 to about 110 amino acids primarily responsible for antigen recognition via the complementarity determining regions (CDRs) contained therein. The carboxy-terminal portion of each chain defines a constant region primarily responsible for effector function.

As used herein, the term“light chain variable region” or“LCVR” refers to a portion of a light chain of an antibody molecule that includes the amino acid sequences of CDRs and framework regions (FRs).

As used herein, the term“heavy chain variable region”“HCVR” refers to a portion of a heavy chain of an antibody molecule that includes the amino acid sequences of CDRs and FRs.

The antibodies described herein may readily be produced in mammalian cells, non-limiting examples of which includes CHO, NS0, HEK293 or COS cells. The host cells are cultured using techniques well known in the art. In this regard, an appropriate host cell can be either transiently or stably transfected with an expression system for secreting antibodies using an optimal predetermined HC:LC vector ratio or a single vector system encoding both HC (heavy chain) and LC (light chain). The vectors containing the polynucleotide sequences of interest (e.g., the polynucleotides encoding the polypeptides of the antibody and expression control sequences) can be transferred into the host cell by well-known methods, which may vary depending on the type of cellular host. Clarified media, into which the antibody has been secreted, may be purified using any of many commonly-used techniques. Various methods of protein purification may be employed and such methods are known in the art and described, for example, in Deutscher, Methods in Enzymology 182: 83-89 (1990) and Scopes, Protein Purification: Principles and Practice , 3rd Edition, Springer, NY (1994). In some examples, the medium may be conveniently applied to a column that has been equilibrated with a compatible buffer. The column may be washed to remove nonspecific binding components. The bound antibody may be eluted, for example, by pH gradient. Antibody fractions may be detected, such as by UV absorbance or SDS-PAGE, and then may be pooled. Further purification is optional, depending on the intended use. The antibody may be concentrated and/or sterile filtered using common techniques. Soluble aggregate and multimers may be effectively removed by common techniques, including size exclusion, hydrophobic interaction, ion exchange, multimodal, or hydroxyapatite chromatography. The purity of the antibody after these chromatography steps is typically greater than 95%. The product may be immediately frozen at -70°C or may be lyophilized.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody is ramucirumab.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab ; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib.

The present disclosure provides a method of treating hepatocellular carcinoma in a patient in need of therapy, comprising measuring an amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient and subsequently administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with regorafenib.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody is ramucirumab.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of anh anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib. The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a test measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from the patient; wherein the anti -human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with regorafenib.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti -human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3. The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody is ramucirumab.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti -human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides a method of identifying a patient with hepatocellular carcinoma for treatment with an anti-human VEGFR-2 (SEQ ID NO: 5) antibody, comprising testing a biological sample from the patient for the presence of human VEGF- D (SEQ ID NO: 6); wherein the patient is selected for treatment with the anti-human VEGFR-2 antibody if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti-human VEGFR- 2 (SEQ ID NO: 5) antibody.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody is ramucirumab.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more prefereably from 115 pg/ml to 1290 pg/ml .

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti-human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib.

The present disclosure provides a therapeutic regimen for treating hepatocellular carcinoma comprising: a) selecting a patient having hepatocellular carcinoma and whose levels of VEGF-D (SEQ ID NO: 6) in a biological sample is 75 pg/ml or greater and b) administering to the patient a therapeutically effective amount of an anti -human VEGFR- 2 (SEQ ID NO: 5) antibody; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with regorafenib.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma , comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if the levels of human VEGF-D present in the biological sample are at an elevated amount.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma , comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma , comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma , comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody is ramucirumab.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma , comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more prefereably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib.

The present disclosure provides an anti-human VEGFR-2 (SEQ ID NO: 5) antibody for use in treating hepatocellular carcinoma, comprising: (a) performing an immunoassay for measuring the amount of human VEGF-D (SEQ ID NO: 6) in a biological sample from a patient; (b) determining the amount of human VEGF-D in the biological sample; and (c) administering a therapeutically effective amount of the anti-human VEGFR-2 antibody to the patient if human VEGF-D is present in the biological sample; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with regorafenib.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 1290 pg/ml, more preferably from 75 pg/ml to 2630 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more prefereably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 1290 pg/ml, more preferably from 75 pg/ml to 2630 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more prefereably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma , wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody further comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3.

A pharmaceutical composition comprising an anti-human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody is ramucirumab. The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the biological sample comprises serum derived from the patient or plasma derived from the patient; optionally wherein the biological sample further comprises heparin or ethylenediaminetetraacetic acid.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the anti-human VEGFR-2 antibody is administered at a dose of 8 mg/kg every 2 weeks. The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D. The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and/or a detection antibody that is a mouse anti-human VEGF-D antibody.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the amount of human VEGF-D is measured using an immunoassay; optionally, wherein the immunoassay comprises a standard comprising human VEGF-D or recombinant human VEGF-D; wherein the immunoassay further comprises a capture antibody that is a mouse anti-human VEGF-D antibody and a detection antibody that is a mouse anti-human VEGF-D antibody; wherein the capture antibody is labeled with biotin and the detection antibody is labeled with ruthenium or horseradish peroxide.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with sorafenib.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody with one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma, wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the anti-human VEGFR-2 antibody: (a) comprises a heavy chain variable region having the amino acid sequence of SEQ ID NO: 2 and a light chain variable region having the amino acid sequence of SEQ ID NO: 1, (b) comprises a heavy chain having the amino acid sequence of SEQ ID NO: 4 and a light chain having the amino acid sequence of SEQ ID NO: 3, or (c) is ramucirumab; wherein the patient was previously treated with regorafenib.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of an anti-human VEGFR-2 (SEQ ID NO: 5) antibody to a patient in need thereof, provided that the patient is selected for treatment after receiving a result from a measurement of the amount of human VEGF- D (SEQ ID NO: 6) in a biological sample from the patient; wherein the amount of human VEGF-D is measured using an immunoassay; wherein the immunoassay comprises a capture antibody comprising a heavy chain having the amino acid sequence of SEQ ID NO: 9 and a light chain having the amino acid sequence of SEQ ID NO: 10 and a detection antibody comprising a heavy chain having the amino acid sequence of SEQ ID NO: 11 and a light chain having the amino acid sequence of SEQ ID NO: 12.

The present disclosure provides a method of treating hepatocellular carcinoma, comprising administering a therapeutically effective amount of ramucirumab to a patient in need thereof, provided that the patient is selected for treatment if the level of human VEGF-D (SEQ ID NO: 6) in a biological sample derived from the patient is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml.

The present disclosure provides a pharmaceutical composition comprising an anti human VEGFR-2 (SEQ ID NO: 5) antibody and one or more pharmaceutically acceptable carriers, diluents, or excipients for use in the treatment of a patient having hepatocellular carcinoma , wherein a biological sample from the patient is measured for the level of human VEGF-D (SEQ ID NO: 6); wherein the anti-human VEGFR-2 antibody is administered if the level of VEGF-D in the biological sample is 75 pg/ml or greater, preferably from 75 pg/ml to 2630 pg/ml, more preferably from 75 pg/ml to 1290 pg/ml, more preferably from 115 pg/ml or greater, more preferably from 115 pg/ml to 2630 pg/ml, or more preferably from 115 pg/ml to 1290 pg/ml; wherein the amount of human VEGF-D is measured using an immunoassay comprising a capture antibody and a detection antibody; wherein the capture antibody comprises a heavy chain having the amino acid sequence of SEQ ID NO: 9 and a light chain having the amino acid sequence of SEQ ID NO: 10 and the detection antibody comprises a heavy chain having the amino acid sequence of SEQ ID NO: 11 and a light chain having the amino acid sequence of SEQ ID NO: 12.

A Multicenter, Randomized, Double-Blind, Phase 3 Study of Ramucirumab (IMC- 1121B) Drug Product and Best Supportive Care (BSC) Versus Placebo and BSC as Second-Line Treatment in Patients With Hepatocellular Carcinoma Following First- Line Therapy With Sorafenib

Study Design:

The Study NCT01140347 is a multicenter, randomized, double-blind, phase 3 study of ramucirumab (IMC-l 121B) drug product and best supportive care (BSC) versus and BSC as second-line treatment in patients with hepatocellular carcinoma following first-line therapy with sorafenib.

The primary objective is to compare the overall survival (time from randomization to death) in patients with HCC disease progression during or following sorafenib therapy, or are intolerant to this agent.

Approximately 565 patients Child-Pugh Class A score at baseline who meet all eligibility criteria are randomized into two Arms. Arm A patients receive placebo and BSC (hereafter called placebo arm) and Arm B patients receive ramucirumab drug product (DP) and best supportive care (BSC), (hereafter called ramucirumab arm). Patients receive ramucirumab DP every 14 days at a dose of 8 mg/kg by IV. infusion, and BSC as determined appropriate by the investigator(s) or placebo every 14 days or 2 weeks at an equivalent volume by IV. infusion, and BSC as determined appropriate by the investigator(s). At randomization, patients are stratified by geographic region (North America versus Europe versus all other regions), and etiology of liver disease (hepatitis B vs hepatitis C vs other etiologies). Ramucirumab DP is a sterile, preservative-free solution for infusion and is formulated in an aqueous solution at a concentration of 10 mg/mL (500 mg/50-mL vial), administered as an intravenous (IV.) infusion at a dose of 8 mg/kg every 2 weeks. The infusion is delivered in approximately 60 minutes. The infusion rate does not exceed 25 mg/minute. Placebo is a sterile, preservative-free solution for injection supplied in 50-mL vials. The volume of placebo to be administered is calculated as if it were active product formulated at 10 mg/mL (administered at a dose of 8 mg/kg every 2 weeks).

Treatment is continued until disease progression, the development of unacceptable toxicity, noncompliance or withdrawal of consent by the patient, or investigator decision. Following discontinuation of study treatment, all patients are followed for survival at regularly scheduled intervals (every 2 months [±14 days]) as long as the patient remains alive or until study completion. Study completion occurs when 438 events are observed and the OS analysis is performed and fully interpreted.

Primary Efficacy Analysis:

The primary efficacy analysis is performed in the ITT population (patients with Child- Pugh Class A score), consisting of all randomized patients. The primary analysis compares the OS between the 2 treatment groups (with versus without ramucirumab DP) using the p-value from a log-rank test stratified by geographic region (North America versus Europe versus East Asia), and etiology of liver disease (hepatitis B vs hepatitis C vs other etiologies) The estimation of survival curves for the 2 treatment groups are generated using the Kaplan-Meier methodology. A stratified Cox regression model to compare the treatments is performed to generate the HR and its 95% confidence limit. A group sequential analysis for OS, the comparison of OS between the 2 study arms, is conducted. The first interim analysis for OS is performed when approximately 109 deaths are observed in all randomized patients (patients with Child-Pugh Class A score and patients with Child-Pugh Class B score). The expected time to observe 109 events is approximately 12.7 months after the first patient’s enrollment. The second interim analysis for futility is conducted when 50% of the planned OS events, approximately 219 deaths, are observed in the ITT population (patients with Child-Pugh Class A score). The expected time to observe 219 events is approximately 26 months from the first patient’s enrollment. The interim analysis for unequivocal efficacy is conducted when 75% of the planned OS events, approximately 328 deaths, are observed in the ITT population (patients with Child-Pugh Class A score). The expected time to observe 328 events is approximately 33 months from the first patient’s enrollment. If unequivocal efficacy is not declared, then the final, inferential OS analysis is conducted once 438 OS events have been observed in the ITT population (patients with Child-Pugh Class A score). The expected time to observe 438 events is approximately 43 months from the enrollment of the first patient.

Secondary Efficacy Analysis:

The secondary efficacy endpoints analyzed are PFS, ORR and TTP. Secondary efficacy endpoints are analyzed at the same time as OS and at the same level of significance. However, if statistical significance at the 0.0050 level is not observed for OS at the interim analysis, the secondary efficacy endpoints are then analyzed at the end of the study with the same level of significance as OS (l-sided 0.0241), provided that OS is significant.

Progression-free survival is analyzed using the Kaplan-Meier method, using the log-rank test, stratified by the same stratification factors used in the analysis of the primary endpoint, OS. Progression-free survival is defined as the time from the date of randomization until the date of objectively determined progressive disease (according to RECIST v. 1.1) or death due to any cause, whichever is first. The ORR in each treatment group is compared using the Cochran-Mantel-Haenszel test adjusting for the stratification variables. Exact confidence bounds (confidence interval: 95%) are determined. The ORR (per RECIST v. 1.1) is defined as the proportion of patients with a best overall response of partial response or complete response. TTP Imaging is done every 6 weeks after first dose for the first 6 months and every 9 weeks thereafter. Patients who do not progress and are subsequently lost to follow-up are censored at the date of the last adequate tumor assessment before loss to follow-up. The treatment groups are compared using the stratified log-rank test and the survival curves are estimated using the Kaplan- Meier methodology, together with corresponding summary statistics. TTP is defined as the time from randomization until the date of radiographic progression according to RECIST v 1.1.

Pharmacodynamic analysis is conducted on blood samples from all patients to assess ramucirumab pharmacokinetics. Tumor tissue, plasma, and whole blood are collected and samples are assayed to determine circulating levels of markers that include but not limited to: VEGF, soluble VEGFR-l, and soluble VEGFR-2. Results are presented using descriptive statistics and correlative analyses with outcome.

Biomarker Statistical Analysis for Clinical Study NCT01140347:

VEGF-D levels are measured in baseline plasma samples collected from 322 patients enrolled in the study. Plasma protein levels are reported as continuous variables. If the protein level in a sample is below the level that is quantifiable by the assay, then a BQL (below the limit of quantitation) result is reported for that sample. The relationship between plasma VEGF-D levels and clinical outcomes are evaluated by creating OS and PFS STEPP figures and diagnostic plots for dichotomization across the range of plasma VEGF-D levels measured in the trial. These analyses are used to narrow the potential range of plasma VEGF-D outpoints at which to divide patients into high and low VEGF- D subgroups and to identify the subgroup with the best efficacy derived from ramucirumab treatment. Individual Cox regression interaction analyses are then performed using outpoints at intervals of 5 pg/ml within the selected range to dichotomize the patients and further define specific optimal cutpoints.

Overall Trial Results and Biomarker Analysis Results for Clinical Study NCT01140347

Observed Treatment Effect with Ramucirumab

In the ITT patient population, regardless of VEGF-D level, the median OS in the ramucirumab arm is 9.17 months (95% Cl: 8.05, 10.64) versus 7.62 months (95% Cl: 6.01, 9.33) in the placebo arm. The stratified HR is 0.866 (95% Cl: 0.717, 1.046, ? = 0.1391). Additionally, treatment with ramucirumab reduced the risk of disease progression or death by 37.5% with a stratified HR of 0.625 (95% Cl: 0.522, 0.750; p < 0.0001), representing a 33.3% (0.7 months) longer median PFS in the ramucirumab arm over the placebo arm (2.8 months vs. 2.1 months). The PFS rates for ramucirumab vs. placebo, respectively, was 81.7% vs. 75.7% at 6 weeks (p = 0.0854), 47.4% vs. 32.6% at 3 months (p = 0.0004), 32.1% vs. 12.9% at 6 months (p < 0.0001), and 20.7% vs. 8.3% at 9 months (p < 0.0001). Furthemore, treatment with ramucirumab reduced the risk of radiographic progression by 40.7% with a stratified HR of 0.593 (95% Cl: 0.487, 0.722; p < 0.0001), representing a 35% (0.9 months) longer median TTP in the ramucirumab arm over the placebo arm (3.5 vs. 2.6 months, respectively). The 6- and 9-month progression- free rates were (ramucirumab vs. placebo) 37.0% vs. 14.9% and 27.0% vs. 10.8%, respectively. Furthermore, treatment with Ramucirumab improved the ORR by approximately lO-fold over placebo; ramucirumab 7.1% (95% Cl: 4.6, 10.7) vs. placebo 0.7 % (95% Cl: 0.2, 2.5) (p = 0.0001). The DCR with Ramucirumab was 56.2% (95% Cl: 50.4, 61.8) vs. placebo 45.7% [95% Cl: 40.0, 51.6]) (p = 0.0110).

Association of VEGF-D levels and Observed Treatment Effect with Ramucirumab

Analyses of the relationship between the benefit of ramucirumab treatment and plasma VEGF-D levels across the range of plasma VEGF-D levels measured in the trial, reveal that patients with higher baseline plasma VEGF-D levels tended to have a greater benefit from ramucirumab in both OS and PFS, when compared to patients with lower baseline plama VEGF-D levels (Table 1 - Table 4). The highest plasma VEGF-D level measured in this study was 2630 pg/ml. The VEGF-D outpoint analysis of > 75 pg/ml vs.

< 75 pg/ml showed a clear differentiation between the patients who had a greater benefit from Ramucirumab treatment and those with a lesser benefit. Furthermore, the VEGF-D cutpoint of > H5pg/ml vs. < 115 pg/ml demonstrated a slightly better OS hazard ratio in the patients with > 1 l5pg/ml VEGF-D levels when compared to the patients with VEGF- D levels > 75 pg/ml. However, the PFS HR in patients with > 115 pg/ml VEGF-D levels, was slightly less favorable when compared to patients with VEGF-D levels > 75 pg/ml cutpoint.

Furthermore, the PFS HR in the < 115 pg/ml cutpoint VEGF-D group is less than 1 (Table 4), whereas the PFS HR in the < 75 pg/ml cutpoint VEGF-D group is above 1 (Table 2). Similarly, the OS HR was lower in the < 115 pg/ml cutpoint VEGF-D group (Table 3), when compared to the OS HR in the < 75 pg/ml cutpoint VEGF-D group (Table 1). These results suggests that some patients classified in the low VEGF-D group using the 115 pg/ml cutpoint actually benefited from ramuciumab treatment, and that the 75 pg/ml cutpoint more appropriately assigns them to the high VEGF-D group, with the greater apparent benefit from ramucirumab treatment. Table 1: OS Analysis by VEGF-D 75 pg/mL cutpoint.

ap-value=from separate interaction analyses.

Table 2: PFS Analysis by 75 pg/mL VEGF-D cutpoint.

aP-value=from separate interaction analyses.

Table 3: OS Analysis by 115 pg/mL VEGF-D cutpoint.

aP-value=from separate interaction analyses.

Table 4: PFS Analysis by 115 pg/mL VEGF-D cutpoint.

aP-value=from separate interaction analyses.

VEGF-D Assay

The purpose of this study is to assess the levels of human VEGF-D in samples derived from human blood samples utilizing the Meso Scale Discovery® Electrochemiluminescence (MSD-ECL) platform. The following MSD-ECL method is validated for the determination of VEGF-D in human serum, ethyl enediaminetetraacetic acid (EDTA) treated plasma, and heparin-treated plasma samples from clinical trials in a regulatory compliant manner. Validation of the method includes assessment of the minimum required dilution (MRD), standard curve accuracy and precision, intra- and inter assay precision, lower limit of quantification (LLOQ), dilutional linearity, normal human serum (NITS) sample reference range, spiked recovery, and short-term, long-term and freeze/thaw stability. Blood samples to determine human VEGF-D concentrations are collected at the specified time points and analyzed using the following immunoassay. The signal agent used with the detection antibody below may be altered as known in the art and in some examples an unlabeled capture antibody may be substituted; non-limiting examples of signal agents include the use of a colorimetric or a chemiluminescent conjugate instead of a ruthenium conjugate (e.g. horseradish peroxidase conjugate, phycoerythrin conjugate, biotin-conjugate, etc.). However, such modifications may affect the sensitivity of the assay, and as a result, the modified assay should be calibrated with the assay described below for best results. In this assay, plates (streptavidin MSD ELISA plates, Cat. L15SA-1) are washed three times with 350 pL/well of wash buffer (IX TBST buffer), and then blocked with 200 pL/well blocking buffer (IX TBST buffer containing 1% bovine serum albumin). TBST is obtained from Boston Bioproducts Cat. IBB-181X (20X TBST). Plates are sealed and incubated for approximately 1 hour at room temperature (RT) on a Titramax™ 1000 plate shaker set at approximately 600 rpm. Blocked plates are washed three times with 350 pL/well of wash buffer. Wells are coated with 5 pg/mL biotin-labeled mouse anti-human VEGF-D antibody (Bio-anti-VEGF-D) (R&D Systems, Cat. MAB2861) in Coating Buffer (IX TBST Buffer + 0.1% BSA) at 50 pL/well. Alternatively, IBA111 that has a heavy chain and a light chain having the amino acid sequence as shown in SEQ ID NO: 9 and SEQ ID NO: 10, respectively, may be used in place of MAB2861. Plates are sealed and incubated for approximately 1 hour at RT on a Titramax™ 1000 plate shaker set at approximately 600 rpm. During the bio-anti -VEGF-D incubation, all plasma samples are spun at approximately 14,000 rpm for 10 minutes and serum samples are spun if necessary.

Before the end of the incubation, a standard curve is prepared in a volume appropriate for 2 replicates for each point of the standard curve by diluting VEGF-D standard in dilution buffer (assay buffer + 0.1% BSA + 100 pg/mL heterophilic blocking reagent 1). Recombinant human VEGF-D protein is used for the standard (R&D Systems, 622-VD-025). The standard curve is prepared as a 12 step dilution curve starting at 300 ng/mL and diluting down 1 to 3 (1 part standard, 2 parts dilution buffer) for a total of 11 dilutions with the l2th dilution being a zero VEGF-D point. Controls are pre-diluted 5-fold in serum. Samples and pre-diluted controls are diluted 2-fold in dilution buffer. Sufficient volume is prepared for two 50 pL/well duplicates. Samples are vortexed prior to addition to plates. After the incubation is complete, the plates are washed three times with 350 pL/well of wash buffer. 50 pL of standards, diluted controls and diluted samples are added to duplicate wells of the blocked and bio-anti -VEGF-D coated plates, unless otherwise specified. Plates are sealed and incubated for approximately 2 hours at RT on a Titramax 1000 plate shaker set at approximately 600 rpm. Plates are washed three times with 350 pL/well of wash buffer. 50 pL of 0.5 pg/mL ruthenium -labeled mouse anti-human VEGF-D antibody (Ru-anti-VEGF-D) (R&D Systems, Cat. MAB286) in assay buffer + 0.1% BSA are added to each well. Alternatively, IBA112 that has a heavy chain and a light chain having the amino acid sequence as shown in SEQ ID NO: 11 and SEQ ID NO: 12, respectively, may be used in place of MAB286. Plates are sealed and incubated for approximately 1 hour at RT on a Titramax 1000 plate shaker set at approximately 600 rpm. Plates are washed three times with 350 pL/well of wash buffer. 2X MSD Read Buffer T (Meso Scale Diagnostics, Cat. R92TC-2) is prepared, and 150 pL is added to each well. Plates are shaken briefly and read on a SECTOR Imager 6000 (620 nm) within 5 minutes of read buffer addition.

SEQUENCE LISTING

SEQ ID NO: 1 (Anti-Human VEGFR-2 Antibody, LCVR) (Artificial Sequence)

DIQMTQSPSSVSASIGDRVTITCRASQGIDNWLGWYQQKPGKAPKLLIYDASNLD

TGVPSRFSGSGSGTYFTLTISSLQAEDFAVYFCQQAKAFPPTFGGGTKVDIK

SEQ ID NO: 2 (Anti-Human VEGFR-2 Antibody, HCVR) (Artificial Sequence)

EVQLVQSGGGLVKPGGSLRLSC AASGFTFS S YSMNWVRQAPGKGLEWVS SIS SS S S YIYY AD S VKGRFTISRDNAKN SLYLQMN SLRAEDT AVYYC ARVTD AFDIW GQG TMVTVSS

SEQ ID NO: 3 (Anti-Human VEGFR-2 Antibody, LC) (Artificial Sequence)

DIQMTQSPSSVSASIGDRVTITCRASQGIDNWLGWYQQKPGKAPKLLIYDASNLD TGVPSRFSGSGSGTYFTLTISSLQAEDFAVYFCQQAKAFPPTFGGGTKVDIKRTVA APSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQ D SKD STYSLSSTLTL SK AD YEKHK V Y ACE VTHQGLS SP VTK SFNRGEC

SEQ ID NO: 4 (Anti-Human VEGFR-2 Antibody, HC) (Artificial Sequence)

EVQLVQSGGGLVKPGGSLRLSC AASGFTFS S YSMNWVRQAPGKGLEWVS SISS S S S YIYY AD S VKGRFTISRDNAKN SLYLQMN SLRAEDT AVYY C ARVTD AFDIWGQG TM VT V S S ASTKGP S VLPL AP S SK S T S GGT AALGCL VKD YFPEP VT V S WN S GALT S GVHTFP AVLQS SGLYSLS S VVTVPS S SLGT Q T YICNVNHKP SNTK VDKRVEPK S C DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN W YVDGVEVHNAKTKPREEQYNSTYRVV S VLTVLHQDWLNGKEYKCKV SNKAL PAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESN GQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYT QKSLSLSPGK

SEQ ID NO: 5 (Human VEGFR-2) (Homo Sapiens)

MQSKVLLAVALWLCVETRAASVGLPSVSLDLPRLSIQKDILTIKANTTLQITCRGQ

RDLDWLWPNNQSGSEQRVEVTECSDGLFCKTLTIPKVIGNDTGAYKCFYRETDL AS VIYVYVQDYRSPFIAS V SDQHGVVYITENKNKT VVIPCLGSISNLNV SLC ARYP

EKRFVPDGNRISWDSKKGFTIPSYMISYAGMVFCEAKINDESYQSIMYIVVVVGY

RI YD VVL SP SHGIELS V GEKL VLNCT ARTELN V GIDFNWE YP S SKHQHKKL VNRD

LKTQSGSEMKKFLSTLTIDGVTRSDQGLYTCAASSGLMTKKNSTFVRVHEKPFV

AFGSGMESLVEATVGERVRIPAKYLGYPPPEIKWYKNGIPLESNHTIKAGHVLTIM

E V SERDTGNYT VILTNPISKEKQ SHVV SL VVYVPPQIGEKSLISP VD S Y Q Y GTTQT

LTCT VY AIPPPHHIHWYW QLEEEC ANEP SQ AV S VTNP YPCEEWRS VEDF QGGNKI

EVNKNQFALIEGKNKTVSTLVIQAANVSALYKCEAVNKVGRGERVISFHVTRGPE

ITLQPDMQPTEQESVSLWCTADRSTFENLTWYKLGPQPLPIHVGELPTPVCKNLD

TLWKLN ATMF SN S TNDILIMELKN ASLQDQ GD Y V CL AQDRKTKKRHC VVRQLT

VLERVAPTITGNLENQTTSIGESIEVSCTASGNPPPQIMWFKDNETLVEDSGIVLKD

GNRNT TIRR VRK FDF.GI ,YTC,QAC,SVT .GCA KVFA FFIIFGAOFKTNI ETTTT VGTAV1

AMFFWLLLVIILRTVKRANGGELKTGYLSIVMDPDELPLDEHCERLPYDASKWEF

PRDRLKLGKPLGRGAF GQ VIE AD AF GIDKT AT CRT V A VKMLKEGATHSEHR ALM

SELKILIHIGHHLNVVNLLGACTKPGGPLMVIVEFCKFGNLSTYLRSKRNEFVPYK

TKGARFRQGKD YVGAIPVDLKRRLDSITS SQS S AS SGFVEEKSLSD VEEEEAPEDL

YKDFLTLEHLICYSFQVAKGMEFLASRKCIHRDLAARNILLSEKNVVKICDFGLA

RDIYKDPD YVRKGD ARLPLKWMAPETIFDRVYTIQ SD VW SF GVLLWEIF SLGASP

YPGVKIDEEF CRRLKEGTRMRAPD YTTPEMY QTMLDCWHGEP SQRPTF SELVEH

LGNLLQANAQQDGKDYIVLPISETLSMEEDSGLSLPTSPVSCMEEEEVCDPKFHY

DNTAGISQYLQNSKRKSRPVSVKTFEDIPLEEPEVKVIPDDNQTDSGMVLASEELK

TLEDRTKLSPSFGGMVPSKSRESVASEGSNQTSGYQSGYHSDDTDTTVYSSEEAE

LLKLIEIGV QTGS T AQILQPD S GTTL S SPP V

SEQ ID NO: 6 (Human VEGF-D) (Homo Sapiens)

M YREW V VVNVFMML Y V QL VQGS SNEHGPVKRS SQSTLERSEQQIRAAS SLEELL RITHSEDWKLWRCRLRLKSFTSMDSRSASHRSTRFAATFYDIETLKVIDEEWQRT QCSPRETCVEVASELGKSTNTFFKPPCVNVFRCGGCCNEESLICMNTSTSYISKQL FEISVPLTSVPELVPVKVANHTGCKCLPTAPRHPYSIIRRSIQIPEEDRCSHSKKLCP IDMLWD SNKCKC VLQEENPLAGTEDHSHLQEP ALCGPHMMFDEDRCEC V CKTP CPKDLIQHPKNCSCFECKESLETCCQKHKLFHPDTCSCEDRCPFHTRPCASGKTA

CAKHCRFPKEKRAAQGPHSRKNP

SEQ ID NO: 7 (Bevacizumab, HC) (Artificial Sequence)

EVQLVESGGGLVQPGGSLRLSCAASGYTFTNYGMNWVRQAPGKGLEWVGWIN T YT GEPT Y A ADFKRRF TF SLDT SK S T A YLQMN SLRAED T A V YY C AK YPH Y Y GS S HW YFD VW GQGTL VT V S S AS TKGP S VFPL AP S SK S T S GGT A ALGCL VKD YFPEP V TV S WN S GALT S GVHTFP A VLQ S S GL Y SL S SWT VP S S SLGTQT YICN VNHKP SNT KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVV D V SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVV S VLTVLHQDWLNG KEYKCK V SNK ALP APIEKTISKAKGQPREPQ VYTLPP SREEMTKNQ V SLT CLVKG FYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCS VMHEALHNHYTQKSLSLSPGK

SEQ ID NO: 8 (Bevacizumab, LC) (Artificial Sequence)

DIQMTQSPSSLSASVGDRVTITCSASQDISNYLNWYQQKPGKAPKVLIYFTSSLHS GVP SRF S GS GS GTDF TLTIS SLQPEDF AT Y Y C QQ YS T VP WTF GQGTK VEIKRT V AA PSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD STYSLSSTLTL SK AD YEKHK V Y ACE VTHQGL S SP VTK SFNRGEC

SEQ ID NO: 9 (IBA111, HC) (Artificial Sequence)

EVKLEESGGGLVQPGGSMKLSCVASGFTFSNYWMNWVRQSPEKGLEWVAEITL K SNN Y ATH Y AE S VKGRF TISRDD SK S S VYLQMNNLRPEDTGI Y Y C TRND YDRAW F AYW GQGTL VTV S AAKTT AP S VYPLAP VCGDTTGS S VTLGCLVKGYFPEP VTLT WN SGSL S S GVHTFP A VLQ SDL YTLS S S VT VT S S T WP S Q SIT CN V AHP A S S TK VDK KIEPRGPTIKPCPPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDD PDVQISWFVNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKV NNKDLPAPIERTISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIY VEWTNNGKTELNYKNTEPVLDSDGS YFMY SKLRVEKKNWVERN S YSC S VVHEG LHNHHTTKSFSRTPGK SEQ ID NO: 10 (IBA111, LC) (Artificial Sequence)

DIVMSQSPSSLAVSVGEKVTLSCKSSQSLLYSSNQKNYLAWYQQEPGQSPKLLIY WASTRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYNYPWTFGGGSK LEIKRAD AAPT V S IFPP S SEQLT S GG AS V V CFLNNF YPKDINVKWKIDGSERQN GV LN S WTDQD SKD ST Y SMS STLTLTKDEYERHN S YT CE ATHKT ST SPIVKSFNRNEC

SEQ ID NO: 11 (IBA111, HC) (Artificial Sequence)

EVQLQESGPSLVKPSQTLSLTCSVTGDSFTSDYWNWIRKFPGNRLEYMGYISYRG VTYYNPSLKSRISITRDTSKNQYYLQLNSVTTEDTATYYCARWGTVPLDYWGQG T S VT V S S AKTTAP S VYPLAP VCGDTTGS S VTLGCLVKGYFPEP VTLTWNSGSLS S GVHTFP AVLQSDLYTLS S S VT VTS STWPSQSITCNVAHP AS STKVDKKIEPRGPTI KPCPPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDDPDVQISWF VNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKVNNKDLPAP IERTISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIYVEWTNNGK TELNYKNTEP VLD SDGS YFMY SKLRVEKKNW VERN S Y SC S VVHEGLHNHHTTK SFSRTPGK

SEQ ID NO: 12 (IBA112, LC) (Artificial Sequence)

DVLMTQTPLSLPVSLGDQASISCRSSQSIVHSNGNTYLEWYLQKPGQSPKLLIYKV SNRF SGVPDRF SGSGSGTDFTLKISRVEAEDLGVYY CF QGSHIP YTF GGGTKLEM KRADAAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLN S WTDQD SKD S T Y SM S S TLTLTKDEYERHN S YT CE ATHKT S T SPIVK SFNRNEC