INTRANASAL FORMULATIONS

RELATED APPLICATIONS

[0001] This application claims the benefit of priority to U.S. Provisional Application No. 61/328,894, filed April 28, 2010, the entire contents of which is incorporated by reference herein in its entirety.

FIELD

[0002] This application relates to methods and compositions of treating a subject with a bone deficit disorder.

BACKGROUND OF THE DISCLOSURE

[0003] Osteoporosis is a disease characterized by low bone mass, microarchitectural deterioration of bone and an increase in bone fragility and risk of fracture. It is a common disease with enormous clinical, societal and economic consequences. It has been estimated that approximately 10 million Americans >50 years old have osteoporosis and that 1.5 million fragility fractures occur each year. Another 34 million Americans have low bone mass and are at risk of developing osteoporosis. The most common sites of osteoporotic fractures are the wrist, hip, and spine, but fractures can occur in any bone. The risk of osteoporotic fractures increases with age, in large part due to the process of age-related bone loss. Osteoporotic fractures occur disproportionately in women, who start with lower bone mass than men, lose bone more quickly during the post-menopausal period and have a longer life expectancy. The occurrence of one osteoporotic fracture greatly increases the risk of a subsequent fracture of the same or different type.

[0004] With increased life expectancy around the globe the number of elderly individuals is rising in every region. These demographic changes are expected to greatly increase the number of osteoporotic fractures and the burden imposed by these events. For example the number of hip fractures worldwide is expected to increase from 1.66 million in 1990 to 6.26 million in 2050.

[0005] The skeleton is continuously being remodeled to repair microfractures that occur in normal use. This process is carried out at localized sites by the sequential processes of bone resorption (excavation of damaged bone) and bone formation (replacement of the excavated bone by new bone). At each site of bone remodeling the amount of bone formation is less than the amount of bone resorption leading to net bone loss. When magnified by time and the rate of bone remodeling this net bone loss leads to strikingly low bone mass. After a certain point the loss of bone strength increases disproportionately to the incremental change in bone mass as the microarchitectural elements of bone that contribute to bone strength are lost. At this point the risk of osteoporotic fracture is greatly increased.

[0006] The goal of anti-osteoporosis therapeutics is to reduce individual risk of fracture by increasing bone strength. Two main strategies are employed: inhibition of bone resorption to restore the balance in the bone remodeling cycle thereby eliminate long term bone loss, and stimulation of bone formation to add new, mechanically sound bone thus directly increasing bone strength. Most drugs in use today are antiresorptives that reduce bone loss. These antiresorptive drugs include estrogen, selective estrogen receptor modulators, calcitonin and bisphosphonates. These drugs are effective in creating a modest amount of new bone and have been shown to reduce fracture rates by 20-60% depending on the agent and type of fracture. Treatment is usually required for many years but frequently patients are not tolerant of prolonged therapy. In general, restoration of microarchitectural structure does not occur with antiresorptive therapy, and with prolonged therapy an excess of microdamage may accumulate due to suppression of bone remodeling.

[0007] The issues of modest efficacy and poor compliance with antiresorptive therapies fuelled the search for anabolic agents that stimulate bone growth and can actively restore bone microarchitecture and bone strength in patients with established osteoporosis who remain at high risk of fracture. To date the only class of approved osteoporosis therapies with an anabolic mechanism of action is PTH and its analogs. PTH produces larger increases in bone mass (particularly in the spine) than those seen with antiresorptive therapies.

[0008] Parathyroid hormone is an 84-amino acid peptide produced by the parathyroid glands. Parathyroid hormone maintains blood calcium concentration within the normal range by promoting calcium reabsorption in the kidney, stimulates the production of the active form of vitamin D that promotes calcium uptake in the gut and stimulates osteoclasts to release calcium from bone to rapidly restore blood calcium concentration to a normal level.

However, when exogenous PTH is given intermittently and at low doses, it is a potent anabolic agent that stimulates the formation of mechanically normal new bone and reduces fractures.

[0009] The only approved bone anabolic agent in the U.S. and some other countries is Forteo®/Forsteo® [recombinant human PTH-(l-34); rhPTH-(l-34); teriparatide]. PREOS® [PTH-(l-84)] is also under development for the treatment of osteoporosis and is approved in Europe (Preotact®) for this indication. Daily SC injections of teriparatide 20 μg reduced the risk of vertebral fractures in a high risk population of post-menopausal women with preexisting vertebral fractures by 65% and non- vertebral fractures by 35% after a mean treatment duration of 19 months.

[0010] ZT-034 is chemical synthesized teriparatide and has the exact chemical sequence as teriparatide (rDNA origin), which is the active pharmaceutical ingredient in Forteo®. However, like many high molecular weight peptide and protein therapeutics, Forteo® use is constrained by several factors, including the requirement for daily, self-administered subcutaneous injection.

[0011] Alternative routes of administration that avoid the need for daily injections would be an advantage in terms of patient acceptance and adherence to therapy. An alternative route of delivery, which would be needle-free, painless, and convenient, is desirable. The nasal mucosa presents a large, accessible, and highly vascular surface area for drug absorption, with the added benefit of elimination of first-pass metabolism by the liver and the gastrointestinal tract. However, peptides are large molecules which are poorly absorbed across mucosal membranes. When given by nasal spray PTH (1-34) has been shown to exhibit poor absorption from the nasal cavity, and doses as high as 1000 μg of PTH (1-34) have been required to achieve plasma concentrations similar to SC Forteo® 20 μg.

(Matsumoto et al, Osteoporos Int (2006) 17: 1532-1538). Therefore permeation enhancing agents are required to provide adequate bioavailability. Various compounds have been used as permeation enhancers, but are variably efficacious and may cause toxicity, principally in the form of local irritation.

[0012] The alkylglycoside class of permeation enhancers has shown the ability to enhance transmucosal delivery of a number of peptide agents. DDM (n-dodecyl-P-D-maltoside) is a 12-carbon chain alkylglycoside with mucosal permeation enhancing properties. DDM is a synthetic molecule which has demonstrated the ability to rapidly and reversibly enhance mucosal absorption of peptides via the intranasal route by increasing paracellular and transcellular transport.

[0013] The foregoing description of related art is not intended in any way as an admission that any of the documents described therein, including pending United States patent applications, are prior art to embodiments of the present disclosure. Moreover, the description herein of any disadvantages associated with the described products, methods, and/or apparatus, is not intended to limit the disclosed embodiments. Indeed, embodiments of the present disclosure may include certain features of the described products, methods, and/or apparatus without suffering from their described disadvantages.

SUMMARY OF THE DISCLOSURE

[0014] According to some embodiments, pharmaceutical compositions are provided comprising one or more PTH-related peptidic analogues (e.g., PTH (1-34)) and at least one transmucosal absorption enhancer. In some embodiments, the at least one transmucosal absorption enhancer includes at least one alkylglycoside. In some embodiments, the pharmaceutical compositions are suitable for intranasal administration. In some

embodiments, the pharmaceutical composition is in the form of an aqueous liquid, nose drops, a spray, or aerosol.

[0015] According to some embodiments, an aqueous pharmaceutical composition is provided comprising one or more PTH-related peptidic analogues (e.g., PTH (1-34)), dodecyl maltoside, EDTA and buffer, wherein the concentration of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) is between 0.5 to 2.3 mg/ml, wherein the concentration of dodecyl maltoside is between 0.15% to 0.25%, and wherein the pH of the aqueous pharmaceutical composition is between 4 to 5.5. In some embodiments, the concentration of dodecyl maltoside is between 0.15% to 0.2%>. In some embodiments, the aqueous pharmaceutical composition is in the form of a liquid, nose drops, intranasal spray, or intranasal aerosol. In some embodiments, the composition is lyophilized and reconstituted prior to use. In some embodiments, the concentration of dodecyl maltoside is about 0.18%. In some embodiments, the buffer is acetate, lactate, or citrate. In some embodiments, the composition further contains a preservative selected form the group consisting of

chlorobutanol, methyl paraben, propyl paraben, butyl paraben, benzalkonium chloride, benzethonium chloride, sodium benzoate, and sorbic acid. In some embodiments, the composition further contains of one or more polyols selected from the group consisting of sucrose, mannitol, sorbitol, trehalose. In some embodiments, the concentration of EDTA is at about 0.05% to 0.2% (e.g., 0.1%. or 0.15%).

[0016] According to some embodiments, methods are provided for treating a bone deficit disorder comprising intranasally administering to a subject in need thereof a therapeutically effective amount of an aqueous pharmaceutical composition, the composition comprising one or more PTH-related peptidic analogues (e.g., PTH (1-34)), dodecyl maltoside, EDTA and buffer, wherein the concentration of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) is between 0.5 to 2.3 mg/ml, wherein the concentration of dodecyl maltoside is between 0.15% to 0.25%>, and wherein the pH of the aqueous pharmaceutical composition is between 4 to 5.5. In some embodiments, the administration achieves a maximum serum concentration, post- dosing (C _max ) of at least 10 pg/mL. In some embodiments, the e composition following intranasal administration to the subject yields a time to maximal plasma concentration (t _max ) of of one or more PTH-related peptidic analogues (e.g., PTH (1 - 34)) in blood plasma of the subject between about 0.1 to 1.0 hours. In some embodiments, the composition, following intranasal administration to the subject, yields a time to maximal plasma concentration (t _max ) of of one or more PTH-related peptidic analogues (e.g., PTH (1 - 34)) in blood plasma of the subject between about 0.1 to 0.3 hours. In some embodiments, the intranasal administration comprises delivering an aerosol having an average mean particle size (Dv,50) between about 10 micron and 100 microns. In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 40 to 160 μg PTH (1-34). In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 80 to 120 μg PTH (1-34). In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 120 to 160 μg PTH (1-34). In some embodiments, the composition is lyophilized and reconstituted prior to use. In some embodiments, the intranasal administration comprises delivering to the subject about 50 and 200 μΐ of the aqueous pharmaceutical composition per actuation. In some embodiments, the pharmaceutical composition is in the form of an aqueous liquid, nose drops, a spray, or an aerosol. [0017] According to some embodiments, methods are provided for administering PTH (1- 34) to a subject in need thereof, comprising intranasally administering to the subject a therapeutically effective amount of an aqueous pharmaceutical composition, wherein the composition comprises one or more PTH-related peptidic analogues (e.g., PTH (1-34)), dodecyl maltoside, EDTA and buffer, wherein the concentration of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) is between 0.5 to 2.3 mg/ml, wherein the concentration of dodecyl maltoside is between 0.15% to 0.25%, and wherein the pH of the aqueous pharmaceutical composition is between 4 to 5.5. The method of claim 21 , wherein administration achieves a maximum serum concentration, post- dosing (C _max ) of at least 10 pg/mL. The method of claim 10 wherein the composition following intranasal administration to the subject yields a time to maximal plasma concentration (t _max ) of of one or more PTH- related peptidic analogues (e.g., PTH (1-34)) in blood plasma of the subject between about 0.1 to 1.0 hours. The method of claim 21 , wherein, the composition, following intranasal administration to the subject, yields a time to maximal plasma concentration (t _max ) of of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in blood plasma of the subject between about 0.1 to 0.3 hours. In some embodiments, the intranasal administration comprises delivering an aerosol having an average mean particle size (Dv,50) between about 10 micron and 100 microns. In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 40 to 160 μg PTH (1-34). In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 80 to 120 μg PTH (1-34). In some embodiments, the intranasal administration comprises delivering to the subject a dose of about 120 to 160 μg PTH (1-34). In some embodiments, the composition is lyophilized and reconstituted prior to use. In some embodiments, the intranasal

administration comprises delivering to the subject about 50 and 200 μΐ of the aqueous pharmaceutical composition per actuation. In some embodiments, the pharmaceutical composition is in the form of an aqueous liquid, nose drops, a spray, or an aerosol.

[0018] In some embodiments, the concentration of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) is between 0.8 to 2.3 mg/ml, 1 to 2.3 mg/ml, 1.2 to 2.3 mg/ml, 1.4 to 2.3 mg/ml, 1.5 to 2.3 mg/ml, 1.6 to 2.3 mg/ml, 0.5 to 2.1 mg/ml, 0.5 to 2.0 mg/ml, 0.5 to 1.9 mg/ml, 0.5 to 1.8 mg/ml, 0.5 to 1.6 mg/ml, 0.5 to 1.4 mg/ml, 0.5 to 1.2 mg/ml, 0.5 to 1 mg/ml, 0.5 to 0.8 mg/ml, 0.8 to 2 mg/ml, 1 to 2 mg/ml, 1.2 to 2 mg/ml, 1.5 to 2 mg/ml, 0.8 to 1.8 mg/ml, 1 to 1.8 mg/ml, 1.2 to 1.8 mg/ml, or 1.5 to 1.8 mg/ml.

[0019] According to some embodiments, compositions are provided comprising one or more PTH-related peptidic analogues (e.g., PTH (1-34)), at least one alkylglycoside, EDTA and buffer. In some embodiments, the at least one alkylglycoside is selected from the group consisting of dodecyl maltoside, tridecyl maltoside, sucrose mono-dodecanoate, sucrose mono-tridecanoate, and sucrose mono-tetradecanoate. In some embodiments, the at least one alkylglycoside is a nontoxic, nonionic alkyl glycoside having a hydrophobic alkyl joined by a linkage to a hydrophilic saccharide, wherein the alkyl has from 9 to 24 carbons. In some embodiments, the glycoside is selected from the group consisting of maltose, sucrose, trehalose and glucose. In some embodiments, the linkage is selected from the group consisting of a glycosidic linkage, a thioglycosidic linkage, an amide linkage, a ureide linkage and an ester linkage.

[0020] According to some embodiments, methods are provided for treating a bone deficit disorder comprising intranasally administering to a subject in need thereof a therapeutically effective amount of pharmaceutical compositions according to the present embodiments. In some embodiments, administration of the compositions achieves a maximum serum concentration, post- dosing (C _max ) of at least 10 pg/mL.

[0021] According to some embodiments, methods are provided for administering one or more PTH-related peptidic analogues (e.g., PTH (1-34)) to a subject in need thereof, comprising intranasally administering to the subject a therapeutically effective amount of a pharmaceutical composition according to the present embodiments.

[0022] Each of the embodiments described herein are equally interchangeable with all other embodiments described herein.

[0023] All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In the case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only not intended to be limiting. Other features and advantages of the disclosure will be apparent from the following detailed description and claims.

BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1. PTH (1-34) concentration in plasma after administration. Data presented is mean data of all the subjects for each dose.

[0025] Figure

injection.

[0026] FFiigguurree 3. Mean AUC of PTH (1-34) following single dose nasal spray or Forteo® injection.

[0027] Figure

[0028] Figure

[0029] Figure

data).

[0030] Figure

the presence or absence of DDM.

[0031] Figure 8. ZT-031 PK profile in rats following ΓΝ administration with 0.18% DDM or SC administration without DDM.

[0032] Figure 9. Pharmacokinetic profile of ZT-034 in rats dosed by intranasal administration with 0.18% DDM or by subcutaneous administration.

DETAILED DESCRIPTION

[0033] The present disclosure provides pharmaceutical compositions and formulations containing suitable PTH-related peptidic analogues for use in methods directed to treating subjects suffering from various bone degenerative or bone deficit disorders (e.g., osteoporosis and related conditions). The PTH-related peptidic analogues described herein induce bone formation in both trabecular and cortical bones, thereby increasing bone mineral density and restoring bones.

[0034] According to some embodiments, there is provided an aqueous therapeutic composition for intranasal delivery of one or more PTH-related peptidic analogues comprising of one or more PTH-related peptidic analogues (e.g., PTH(l -34)) and a transmucosal absorption enhancer. In some embodiments, the administration achieves a maximum serum concentration, post- dosing (C _max ) of the one or more PTH-related peptidic analogues of at least 10 pg/mL.

[0035] According to some embodiments, there is provided an aqueous therapeutic composition for intranasal delivery of one or more PTH-related peptidic analogues comprising of one or more PTH-related peptidic analogues (e.g., PTH(l -34)), at least one alkyl glycoside, EDTA and buffer. In some embodiments, the administration achieves a maximum serum concentration, post- dosing (C _max ) of at least 10 pg/mL.

[0036] In some embodiments, the aqueous therapeutic compositions for intranasal delivery of the present disclosure have a pH of about 4 to about 7. In some embodiments, the compositions have a pH of about 4.5 and to about 5.5.

[0037] The composition of the present embodiments may include EDTA, wherein the concentration of from about 0.001% to about 0.5% w/v (e.g., 0.05%, 0.1%, 0.2%, 0.3%, 0.4%).

[0038] In some embodiments, the transmucosal absorption enhancer comprises one or more alkyl glycosides. In some embodiments, the one or more alkyl glycosides in the composition is from about 0.01% to 2% w/v. In some embodiments, the one or more alkyl glycosides in the composition from about 0.05%> to about 0.2%> w/v. In some embodiments, the one or more alkyl glycosides in the composition from about 0.15% to about 0.2% w/v. In some embodiments, the one or more alkyl glycosides in the composition from about 0.1% to about 0.2% w/v. In some embodiments, the one or more alkyl glycosides in the composition from about 0.1 % to about 0.3%> w/v. In some embodiments, the one or more alkyl glycosides in the composition of about 0.18%.

[0039] In some embodiments, the dodecyl maltoside in the composition is from about 0.01% to 2% w/v. In some embodiments, the dodecyl maltoside in the composition from about 0.05%) to about 0.2%> w/v. In some embodiments, the dodecyl maltoside in the composition from about 0.15% to about 0.2%> w/v. In some embodiments, the dodecyl maltoside in the composition from about 0.1% to about 0.2% w/v. In some embodiments, the dodecyl maltoside in the composition from about 0.1% to about 0.3%> w/v. In some embodiments, the dodecyl maltoside in the composition of about 0.18%.

[0040] The compositions of the present embodiments may include a buffer. The buffer agent may be any known buffer agent known in the art suitable for use in aqueous pharmaceutical preparations. Examples of suitable buffers include acetate, lactate, or citrate.

[0041] In some embodiments, the compositions are self-preserving.

[0042] The compositions of the present embodiments may include a preservative selected form the group consisting of chlorobutanol, methyl paraben, propyl paraben, butyl paraben, benzalkonium chloride, benzethonium chloride, sodium benzoate, and sorbic acid. In some embodiments, the compositions may comprise benzalkonium chloride.

[0043] The compositions of the present embodiments may include a one or more polyols.

The one or more polyols may be selected from the group consisting of sucrose, mannitol, sorbitol, trehalose. In some embodiments, the polyol is trehalose.

[0044] The compositions of the present embodiments may be formulated as liquid droplets. In some embodiments, less than 10% of the droplets are less than 10 microns in diameter.

[0045] In some embodiments, the administration of the compositions of the present embodiments to a subject in need thereof yields enhanced mucosal delivery of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) comprising: a) a peak concentration (C _max ) of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in a blood plasma of said subject that is about 10% or greater (e.g., 10% to 200%, 10% to 100%, 15% to 200%, 15% to 100%, 20% to 200%, 30% to 100%, 50% to 200%, or 60% to 100%) as compared to a peak concentration of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) following subcutaneous injection of an equivalent concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1-34)) to the subject; b) an area under concentration curve (AUC) of PTH (1 -34)) in the blood plasma of the subject that is 10% or greater (e.g., 10% to 200%, 15% to 100%, 20% to 200%, 30% to 100%, 40% to 200%, or 50% to 100%) compared to an AUC of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in blood plasma following subcutaneous injection of an equivalent concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1-34)) to said subject; or c) a time to maximal concentration (T _max ) of one or more PTH-related peptidic analogues (e.g., PTH (1- 34)) in the blood plasma of the subject between about 0.1 to 2.0 hours (e.g., between about 0.1 to 2.0 hours). Between "about 0.1 to 2.0 hours" as used herein includes, between about 0.1 hour to 1 hour, between about 0.2 hours to 1 hour, between about 0.3 hours to 1 hour, between about 0.4 hours to 1 hour, between about 0.5 hours to 1 hour, between about 0.6 hours to 1 hour, between about 0.7 hours to 1 hour, between about 0.8 hours to 1 hour, between about 0.9 hours to 1 hour, between about 0.1 hours to 0.5 hours, between about 0.2 hours to 0.5 hours, between about 0.3 hours to 0.5 hours, and between about 0.4 hours to 0.5 hours.

[0046] In some embodiments, the intranasal administration of the compositions of the present embodiments to a subject in need thereof yields enhanced mucosal delivery of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) comprising a peak concentration (C _max ) of PTH(l-34) in blood plasma of said subject that is 10% or greater (e.g., 10% to 200%, 15% to 100%, 20% to 200%, 30% to 100%, 40% to 200%, or 50% to 100%) as compared to a peak concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1-34)) following subcutaneous injection of an equivalent concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1-34)) to the subject.

[0047] In some embodiments, the intranasal administration of the compositions of the present embodiments to a subject in need thereof yields enhanced mucosal delivery of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) comprising a peak concentration (C _max ) of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in blood plasma of said subject that is 50% or greater (e.g., 50% to 200%, 50% to 500%, 70% to 500%, 70% to 500%), 90%) to 500%), or 90%> to 500%>) as compared to a peak concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1-34)) following subcutaneous injection of an equivalent concentration of the one or more PTH-related peptidic analogues (e.g., PTH (1- 34)) to the subject.

[0048] In some embodiments, the intranasal administration of the compositions of the present embodiments to a subject in need thereof yields a time to maximal plasma concentration (T _max ) of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in blood plasma of said subject between about 0.1 to 2.0 hours (e.g., between about 0.1 to 2.0 hours).

[0049] In some embodiments, the intranasal administration of the compositions of the present embodiments to a subject in need thereof yields a time to maximal plasma concentration (T _max ) of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in blood plasma of said subject between about 0.2 to 0.6 hours.

[0050] The compounds of the present embodiments may be administered in a formulation selected from the group consisting of a drop, a spray, or an aerosol. In some embodiments, the composition may be administered as an intranasal spray. When administered as a nasal spray, the administered dosage of the composition may comprise a total volume of about 0.05 mL to about 0.4 mL (e.g., 0.05 mL to about 0.2 mL such as 0.05 ml, 0.07 ml, 0.1 ml, 0.12 ml, 0.15 ml, 0.18 ml, or 0.2 ml) per administered dose. When administered intranasally, the administered dosage of the composition may comprise a total volume of about 0.05 to 0.1 mL per administered dose. The total administered dose may be divided into two separated administrations. For example, the nasal formulation may be administered to each nostril such that a total volume of about 0.05 to 0.15 mL is administered to each nostril (e.g., 2 x 0.07 ml or 2 x 0.05 ml).

[0051] According to some embodiments, methods are provided for delivering one or more PTH-related peptidic analogues (e.g., PTH (1-34)) via intranasal administration. According to some embodiments, methods are provided for delivering one or more PTH-related peptidic analogues (e.g., PTH (1-34)) in a formulation suitable for intranasal administration, said methods comprising delivering an aerosol having an average mean particle size (Dv,50) between about 5 micron and 100 microns (e.g, 10 to 100 microns, 20 to 100 microns, 30 to 100 microns, 40 to 100 microns, 50 to 100 microns, 60 to 100 microns, 70 to 100 microns, 80 to 100 microns, 90 to 100 microns, 10 to 60 microns, 20 to 60 microns, 30 to 60 microns, 40 to 60 microns, 50 to 60 microns, 30 to 50 microns, 40 to 50 microns, 40 to 55 microns, or 45 to 55 microns).

[0052] According to some embodiments, methods are provided for delivering one or more

PTH-related peptidic analogues (e.g., PTH (1-34)) in a formulation suitable for intranasal administration, wherein said intranasal administration comprises delivering an aerosol having an average mean particle size (Dv,50) between about 40 micron and 60 microns.

[0053] According to some embodiments, methods are provided for delivering one or more

PTH-related peptidic analogues (e.g., PTH (1-34)) in a formulation suitable for intranasal administration, wherein said intranasal administration comprises delivering an aerosol having an average mean particle size (Dv,50) between about 10 micron and 100 microns.

[0054] According to some embodiments, methods are provided for treating a bone deficit disorder in a human comprising intranasally administering an intranasal formulation of one or more PTH-related peptidic analogues (e.g., PTH (1-34)) according to the present embodiments. In some embodiments, the intranasal administration comprises delivering an aerosol comprising an aqueous solution comprising 0.5 to 2.5 mg/mL of one or more PTH- related peptidic analogues (e.g., PTH (1-34)), which includes 0.5 to 2.5 mg/mL, 0.7 to 2.5 mg/mL, 0.9 to 2.5 mg/mL, 1 to 2.5 mg/mL, 1.2 to 2.5 mg/mL, 1.4 to 2.5 mg/mL, 1.6 to 2.5 mg/mL, 1.8 to 2.5 mg/mL, 2 to 2.5 mg/mL, 2.2 to 2.5 mg/mL, 0.5 to 2.5 mg/mL, 0.5 to 2.3 mg/mL, 0.5 to 2.1 mg/mL, 0.5 to 2 mg/mL, 0.5 to 1.8 mg/mL, 0.5 to 1.5 mg/mL, or 0.5 to 1 mg/mL of one or more PTH-related peptidic analogues (e.g., PTH (1 -34)).

[0055] According to some embodiments, compositions are provided comprising one or more PTH-related peptidic analogues (e.g., PTH (1-34)), wherein the composition is formulated for intranasal administration. The intranasal formulations may comprise from about 40 to 160 μg one or more PTH-related peptidic analogues (e.g., PTH (1-34)), which includes from about 40 to 160 μg, from about 60 to 160 μg, from about 80 to 160 μg, from about 90 to 160 μg, from about 100 to 160 μg, from about 105 to 160 μg, from about 1 10 to 160 μg, from about 1 15 to 160 μg, from about 120 to 160 μg, from about 122 to 160 μg, from about 124 to 160 μg, from about 126 to 160 μg, from about 128 to 160 μg, from about 130 to 160 μg, from about 132 to 160 μg, from about 134 to 160 μg, from about 136 to 160 μg, from about 138 to 160 μg, from about 140 to 160 μg, from about 142 to 160 μg, from about 144 to 160 μg, from about 146 to 160 μg, from about 148 to 160 μg, from about 150 to 160 μg, from about 40 to 150 μg, from about 60 to 150 μg, from about 80 to 150 μg, from about 90 to 150 μg, from about 100 to 150 μg, from about 105 to 150 μg, from about 1 10 to 150 μg, from about 1 15 to 150 μ , from about 120 to 150 μg, from about 122 to 150 μg, from about 124 to 150 μg, from about 126 to 150 μg, from about 128 to 150 μg, from about 130 to 150 μg, from about 132 to 150 μg, from about 134 to 150 μg, from about 136 to 150 μg, from about 138 to 150 μg, from about 140 to 150 μg, from about 142 to 150 μg, from about 144 to 150 μg, from about 146 to 150 μg, from about 148 to 150 μg, from about 40 to 145 μg, from about 60 to 145 μg, from about 80 to 145 μg, from about 90 to 145 μg, from about 100 to 145 μg, from about 105 to 145 μg, from about 1 10 to 145 μg, from about 1 15 to 145 μg, from about 120 to 145 μg, from about 122 to 145 μg, from about 124 to 145 μg, from about 126 to 145 μg, from about 128 to 145 μg, from about 130 to 145 μg, from about 132 to 145 μg, from about 134 to 145 μg, from about 136 to 145 μg, from about 138 to 145 μg, from about 140 to 145 μg, from about 142 to 145 μg, from about 144 to 145 μg, from about 40 to 140 μg, from about 60 to 140 μg, from about 80 to 140 μg, from about 90 to 140 μg, from about 100 to 140 μg, from about 105 to 140 μg, from about 1 10 to 140 μg, from about 1 15 to 140 μg, from about 120 to 140 μg, from about 122 to 140 μg, from about 124 to 140 μg, from about 126 to 140 μg, from about 128 to 140 μg, from about 130 to 140 μg, from about 132 to 140 μg, from about 134 to 140 μg, from about 136 to 140 μg, from about 138 to 140 μg, from about 40 to 130 μg, from about 60 to 130 μg, from about 80 to 130 μg, from about 90 to 130 μg, from about 100 to 130 μg, from about 105 to 130 μg, from about 1 10 to 130 μ , from about 1 15 to 130 μg, from about 120 to 130 μg, from about 122 to 130 μg, from about 124 to 130 μg, from about 126 to 130 μg, and from about 128 to 130 μg.

[0056] In some embodiments, intranasal formulations may be lyophilized and

reconstituted prior to use.

[0057] In some embodiments, the intranasal formulations may be formulated as an aerosol. In some embodiments, the intranasal administration comprises delivering an aerosol comprising from about 40 to 160 μg one or more PTH-related peptidic analogues (e.g., PTH (1-34)). In some embodiments, the intranasal administration comprises delivering an aerosol comprising 80 to 120 μg one or more PTH-related peptidic analogues (e.g., PTH (1-34)). In some embodiments, the aerosol may contain between about 50 and 200 μΐ of solution per actuation

[0058] The compositions of the present embodiments (e.g. , intranasal compositions/ formulations), may in the form of a single or unit dose. In some embodiments, the compositions of the present embodiments do not contain preservatives.

PTH-related peptidic analogues

[0059] PTH-related peptidic analogues of the present disclosure include full-length 1-84, or fragments thereof, including PTH 1-34 and PTH 1-31. Additional PTH-related peptidic analogues of can optionally include less than the first 34 amino acids at the N-terminal end. The PTH peptide analogues of the present disclosure which are less than 34 amino acids in length, when compared to full-length PTH peptides or other PTH peptide analogues which are 34 amino acid residues or longer, trigger less than full activation of phospholipase-C, less bone resorption, and less incidences or lower severity of hypercalcemia, while still maintaining increases in bone mineral density (BMD) at a variety of sites within the body. [0060] As active ingredient, the pharmaceutically acceptable composition or solution described herein may incorporate full-length PTH (1-84), 1-31 and 1-34 fragments, and other fragments, or variants of fragments, including substitutions, deletions, or insertions, of human PTH, or of rat, porcine or bovine PTH that have human PTH activity as determined in the ovariectomized rat model of osteoporosis reported by Kimmel et ah, Endocrinology, 1993, 32(4): 1577. Human PTH activity includes the ability of the PTH to increase trabecular and/or cortical bone growth. The PTH-related peptidic analogues of the present disclosure increase AC activity when administered to a PTH receptor containing or expressing cell in culture, such as an osteoblast or osteoclast. The PTH-related peptidic analogues used in the present disclosure are naturally or non-naturally occurring and may optionally incorporate less than the first 34 N-terminal residues of PTH.

[0061] PTH operates through activation of two second messenger systems, G _s -protein activated adenylyl cyclase (AC) and G _q -protein activated phospho lipase C. The latter results in a stimulation of membrane-bound protein kinase C (PKC) activity. PKC activity has been shown to require PTH residues 29 to 32 (Jouishomme et al (1994) J. Bone Mineral Res. 9, (1179-1189). It has been established that the increase in bone growth, i.e. that effect which is useful in the treatment of osteoporosis, is coupled to the ability of the peptide sequence to increase AC activity.

[0062] The native PTH sequence, and its truncated 1-34 form, has been shown to have all of these activities. The hPTH-(l-34) sequence is: Ser Val Ser Glu He Gin Leu Met His Asn Leu Gly Lys His Leu Asn Ser Met Glu Arg Val Glu Trp Leu Arg Lys Lys Leu Gin Asp Val His Asn Phe-OH (SEQ ID NO: 1).

[0063] AC activity has been shown to require the first few N-terminal residues of the molecule. Thus, in accordance with certain embodiments of the disclosure, it is possible to remove those biological activities associated with the PKC activity by deleting a selected terminal portion of the hPTH-(l-34) molecule. In one embodiment, these shortened analogues are desirably in the form of carboxyl terminal amides. One feature of the analogues of the disclosure therefore comprises variants of the human parathyroid analogues PTH(l-25)-NH ₂ , PTH(l-26)-NH ₂ , PTH(l-27)-NH ₂ , PTH(l-28)-NH ₂ , PTH(l-29)-NH ₂ , PTH(l-30)-NH ₂ , and PTH(1-31)-NH ₂ . [0064] According to another feature of the analogues of the present disclosure, lactams of the PTH-related peptidic analogues are formed, for example, by cyclisation involving the coupling of the side-chains of Glu22 and Lys26, or of the side-chains Lys26 and Asp30, in which Lys27 may be replaced by a Leu or by various other hydrophobic residues, and which has either a C-terminal free amide ending, or has a C-terminal free carboxyl ending. Such substitutions include ornithine, citrulline, alpha-aminobutyric acid, or any linear or branched alpha-amino aliphatic acid, having 2-10 carbons in the side chain, any such analogue having a polar or charged group at the terminus of the aliphatic chain. Example of polar or charged groups include amino, carboxyl, acetamido, guanido and ureido. Isoleucine, norleucine, methionine, and ornithine are expected to be the most active.

[0065] The PTH-related peptidic analogues of the present disclosure may thus feature the formation of a lactam, for example, between either residues Glu22 and Lys26, Ly26 and Asp30, or Glu22 and Lys27. The substitution of Leu for the Lys27 results in a more hydrophobic residue on the hydrophobic face of the amphiphilic helix. This resulted in increased adenylyl cyclase stimulating activity in the PTH receptor containing rat

osteosarcoma (ROS) cell line. It will be appreciated by those skilled in the art that other such substitutions would likely result in analogues with the same or increased activities. These hydrophobic substitutions include residues such as methionine or norleucine. The combined effect of substitution and either lactam formation is expected to stabilize the alpha-helix and increase bioactivity, and to protect this region of the molecule from proteolytic degradation. The presence of the amide at the C-terminus is further expected to protect the peptide against exoproteolytic degradation (Leslie, F. M. and Goldstein, A. (1982) Neuropeptides 2, 185- 196).

[0066] In one preferred embodiment of the methods disclosed, herein, the peptide used in the disclosed method is ΡΤΗ(1-31)-ΝΗ ₂ with the following sequence: Ser-Val-Ser-Glu-Ile- Gln-Leu-Met-His-Asn-Leu-Gly-Lys-His-Leu-Asn-Ser-Met-Glu-Arg- Val-Glu-Trp-Leu-Arg- Lys-Xaa-Leu-Gln-Asp-Val-NH ₂ (SEQ ID NO: 2). Xaa is selected from the group consisting of Lys, Leu, He, Nle and Met. In a preferred embodiment, Xaa is Lys (SEQ ID NO: 3). This embodiment is also referred to as Ostabolin™.

[0067] In another preferred embodiment of the disclosure, the peptide used in the disclosed method is cyclo(22-26)PTH-(l-31)-NH ₂ , cyclized in the form of a lactam between Glu ²² and Lys ²⁶ with the following sequence: Ser-Val-Ser-Glu-Ile-Gln-Leu-Met-His-Asn- Leu-Gly-Lys-His-Leu-Asn-Ser-Met-Glu-Arg-Val-Glu-Trp-Leu-Arg- Lys-Xaa-Leu-Gln-Asp- Val-Y (SEQ ID NO: 4), Xaa is selected from the group consisting of Leu, He, Nle and Met and Y is NH ₂ or OH. When Xaa is Leu and Y is NH ₂ (SEQ ID NO: 5), the PTH is also referred to as Ostabolin-C™.

[0068] The PTH-related peptidic analogues to be used according to the present disclosure can thus be cyclized or linear, and can be optionally amidated at the C-terminus. Alternatives in the form of PTH variants incorporate from 1 to 5 amino acid substitutions that improve PTH stability and half-life, such as the replacement of methionine residues at positions 8 and/or 18 with leucine or other hydrophobic amino acids that improves PTH stability against oxidation and the replacement of amino acids in the 25-27 region with trypsin-insensitive amino acids such as histidine or other amino acid that improves PTH stability against protease. Other suitable forms of PTH include PTHrP, PTHrP(l-34), PTHrP(l-36) and analogues of PTH or PTHrP that activate the PTH1 receptor. These forms of PTH are embraced by the term "parathyroid hormone analogues" as used generically herein. The hormones may be obtained by known recombinant or synthetic methods, such as described in U.S. Pat. Nos. 4,086,196; 5,556,940; 5,955,425; 6,541,450; 6,316, 410; and 6,110,892, incorporated herein by reference.

[0069] Specific embodiments of PTH peptide analogues of described herein include the following: PTH-(1-31)NH ₂ , Ostabolin™; PTH-(1-30)NH ₂ ; PTH-(1-29)NH ₂ ; PTH-(1- 28)NH ₂ ; Leu ²⁷ PTH-(l-31)NH ₂ ; Leu ²⁷ PTH-(l-30)NH ₂ ; Leu ²⁷ PTH-(l-29)NH ₂ ; Leu ²⁷ cyclo(22- 26)PTH-(1-31)NH ₂ Ostabolin-C™; Leu ²⁷ cyclo(22-26)PTH-(l-34)NH ₂ ; Leu ²⁷ cyclo(Lys26- Asp30)PTH-(l-34)NH ₂ ; Cyclo(Lys27-Asp30)PTH-(l-34)NH ₂ ; Leu ²⁷ cyclo(22-26)PTH-(l- 31)NH ₂ ; Ala ²⁷ or Nle ²⁷ or Tyr ²⁷ or He ²⁷ cyclo(22-26)PTH-(l-31)NH ₂ ; Leu ²⁷ cyclo(22- 26)PTH-( 1 -32)NH ₂ ; Leu ²⁷ cyclo(22-26)PTH-( 1 -31 )OH; Leu ²⁷ cyclo(26-30)PTH-( 1 -31 )NH ₂ ; Cys ²² Cys ²⁶ Leu ²⁷ cyclo(22-26)PTH-(l-31)NH ₂ ; Cys ²² Cys ²⁶ Leu ²⁷ cyclo(26-30)PTH-(l-31)NH ₂ ; Cyclo(27-30)PTH-(1-31)NH ₂ ; Leu ²⁷ cyclo(22-26)PTH-(1-30)NH ₂ ; Cyclo(22-26)PTH-(l- 31)NH ₂ ; Cyclo(22-26)PTH-(1-30)NH ₂ ; Leu ²⁷ cyclo(22-26)PTH-(l-29)NH ₂ ; Leu ²⁷ cyclo(22- 26)PTH-( 1 -28)NH ₂ ; Glu ¹⁷ ,Leu ²⁷ cyclo( 13-17)(22-26)PTH-(l -28)NH ₂ ; and

Glu ¹⁷ ,Leu ²⁷ cyclo(13-17)(22-26)PTH-(l-31)NH ₂ . [0070] Some embodiments of PTH peptide analogues include those that when

administered result in reduced phospholipase-C activity, reduced bone resorption, and reduced hypercalcemia levels. As defined in the Definitions section herein, "reduced phospholipase-C activity" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less than full activation of phospholipase-C, as compared to the full-length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length. "Reduced bone resorption" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less bone resorption, as compared to the full-length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length. "Reduced hypercalcemia levels" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less incidences of hypercalcemia, or lower severity of hypercalcemia, as compared to the full- length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length.

[0071] PTH-related peptidic analogues include PTH 1-34 or Teriparatide. In some embodiments, the PTH-related peptidic analogues administered in the formulations and

27 22 26

methods described herein include [Leu ]cyclo[Glu -Lys ]-PTH-(l-31)-NH ₂ , such as

27 advanced by Zelos Therapeutics, Inc. under the tradename Ostabolin-C™ and [Leu ] PTH- (1-31)-NH ₂ . In another embodiment of the disclosure, [Leu ²⁷ ]cyclo[Glu ²² -Lys ²⁶ ]-PTH-(l-30)- NH ₂ is used in the methods described herein. In another embodiment, the hormone can be the linear analogue PTH(l-31), which can have a free carboxyl ending, or be amidated, at the C- terminus. In yet another embodiment, the hormone can be PTH(l-30) , which can have a free

27

carboxyl ending, or be amidated, at the C-terminus; or [Leu ]-PTH(l-30)- NH ₂ . Suitable stabilized solutions of these and other PTH-related peptidic analogues that can be employed in the present methods are described in U.S. Patent Nos. 5,556,940; 5,955,425; 6,541,450; 6,316, 410; and 6,110,892 incorporated herein by reference.

Transmucosal Absorption Enhancer

[0072] Intravail® (Aegis Therapeutics, San Diego, CA) is a patented transmucosal absorption enhancer that comprises a broad class of chemically synthesizable transmucosal absorption enhancement agents that allow non-invasive systemic delivery of potent peptide, protein, nucleotide-related, and other small and large molecule drugs that were previously only deliverable by injection. (See US Patent No. 5,661,130; US Pat No. 7,425,542; European Patent No. EP1789075; PCT Publication No. WO95/00151; U.S. Publication No.

2006/0046969; U.S. Publication No. 2006/0046962; U.S. Publication No. 2006/0045869; U.S. Publication No. 2006/0045868; U.S. Publication No. 2008/0268032; U.S. Publication No.

2008/0194461; U.S. Publication No. 2008/0200418; U.S. Publication No. 2008/0299079; PCT Publication No. WO 2009/029543; and U.S. Publication No. 2009/0047347, each of which are incorporated herein by reference in their entireties).

[0073] Intravail® absorption enhancement agents are mild and non-irritating to mucosal membranes. Moreover, these agents are safe, odorless, tasteless, non-toxic, non-irritating, non-denaturing, and non-mutagenic, chemically synthesized molecules that metabolize to C0 ₂ and H ₂ 0. In fact, these molecules are closely related to mild surfactants, which are widely used in personal care and food products in significantly higher concentrations than those used in Intravail® formulations and are recognized as GRAS (Generally Regarded As Safe) substances for many applications. The use of Intravail® absorption enhancement agents exhibits a high degree of bioavailability, which is comparable to subcutaneous injection, via intranasal, buccal, intestinal, and other mucosal membrane administration routes. Thus, these agents can be used to deliver potent peptide, protein, and large molecule drugs that typically have only been delivered parenterally (e.g. by injection).

[0074] In other embodiments, the compositions of the present embodiments comprise at least one PTH-related peptidic analogue of the present embodiments and at least one alkylglycoside.

[0075] In some embodiments, pharmaceutical compositions are provided comprising at least one PTH-related peptidic analogue of the present embodiments and a suitable nontoxic, nonionic alkylglycoside having a hydrophobic alkyl joined by a linkage to a hydrophilic saccharide. In some embodiments, the alkyl has from 9 to 24 carbons. In some embodiments, the alkyl has from 9 to 14 carbon atoms. In some embodiments, the saccharide is selected from the group consisting of maltose, sucrose and glucose. In some embodiments, the alkylglycoside further has a hydrophile-lipophile balance number in the range of about 10 to 20. In some embodiments, the linkage is selected from the group consisting of a glycosidic linkage, a thioglycosidic linkage, an amide linkage, a ureide linkage and an ester linkage. In some embodiments, the alkylglycoside has a concentration in the range of about 0.01% to 1.0%.

[0076] In some embodiments, pharmaceutical compositions are provided comprising at least one PTH-related peptidic analogue of the present embodiments; a buffering agent; and an alkylglycoside; wherein the alkylglycoside is selected from the group consisting of dodecyl maltoside, tridecyl maltoside, sucrose mono-dodecanoate, sucrose mono- tridecanoate, and sucrose mono-tetradecanoate. In some embodiments, the alkylglycoside has a critical micelle concentration (CMC) of less than about 1 mM (e.g., 0.1 to 1 mM).

[0077] In some embodiments, pharmaceutical compositions are provided comprising at least one PTH-related peptidic analogue of the present embodiments; a buffering agent; and an alkylglycoside, wherein the alkylglycoside is selected from the group consisting of dodecyl maltoside, tridecyl maltoside, sucrose mono-dodecanoate, sucrose mono- tridecanoate, and sucrose mono-tetradecanoate.

[0078] In some embodiments, there are provided formulations comprising at least one PTH related peptide, whether at high or low concentration, and at least one alkylglycoside and/or saccharide alkyl ester surfactant, hereinafter termed "alkylglycosides". As used herein, "alkylglycoside" refers to any sugar joined by a linkage to any hydrophobic alkyl, as is known in the art. The linkage between the hydrophobic alkyl chain and the hydrophilic saccharide can include, among other possibilities, a glycosidic, ester, thioglycosidic, thioester, ether, amide or ureide bond or linkage. Examples of which are described herein. The terms alkylglycoside and alkylsaccharide may be used interchangeably herein.

[0079] In some embodiments, the alkylglycosides of the described herein include, but are not limited to, dodecyl maltoside, tridecyl maltoside, tetradecyl maltoside, sucrose mono- dodecanoate, sucrose mono-tridecanoate, and sucrose mono-tetradecanoate.

[0080] As used herein, "alkylglycoside" refers to any sugar joined by a linkage to any hydrophobic alkyl, as is known in the art. The hydrophobic alkyl can be chosen of any desired size, depending on the hydrophobicity desired and the hydrophilicity of the saccharide moiety. In one aspect, the range of alkyl chains is from 9 to 24 carbon atoms; and further the range is from 10 to 14 carbon atoms.

[0081] As used herein, "Critical Micelle Concentration" or "CMC" is the concentration of an amphiphilic component (alkylglycoside) in solution at which the formation of micelles (spherical micelles, round rods, lamellar structures etc.) in the solution is initiated. In some embodiments, the alkylglycoside has a critical micelle concentration (CMC) of less than about 1 mM (e.g., 0.1 to ImM) in pure water.

[0082] As used herein, "saccharide" is inclusive of monosaccharides, oligosaccharides or polysaccharides in straight chain or ring forms. Oligosaccharides are saccharides having two or more monosaccharide residues.

[0083] As used herein, "sucrose esters" are sucrose esters of fatty acids. Sucrose esters can take many forms because of the eight hydroxyl groups in sucrose available for reaction and the many fatty acid groups, from acetate on up to larger, more bulky fats that can be reacted with sucrose. This flexibility means that many products and functionalities can be tailored, based on the fatty acid moiety used. Sucrose esters have food and non-food uses, especially as surfactants and emulsifiers, with growing applications in pharmaceuticals, cosmetics, detergents and food additives. They are biodegradable, non-toxic and mild to the skin.

[0084] As used herein, a "suitable" alkylglycoside means one that fulfills the limiting characteristics of the methods described, herein, i.e., that the alkylglycoside be nontoxic and nonionic, and that it reduces the immunogenicity or aggregation of a PTH-related peptidic analogue when it is administered via the ocular, nasal, nasolacrimal, sublingual, buccal, inhalation routes or by injection routes such as the subcutaneous, intramuscular, or intravenous routes. Suitable compounds can be determined using the methods set forth in the examples.

[0085] While there are potentially many thousands of alkylglycosides which are synthetically accessible, the alkylglycosides dodecyl, tridecyl and tetradecyl maltoside and sucrose dodecanoate, tridecanoate, and tetradecanoate are particularly useful since they possess desirably low CMC's. Hence, the above examples are illustrative, but the list is not limited to that described herein. Derivatives of the above compounds which fit the criteria of the claims should also be considered when choosing a glycoside.

[0086] Some preferred glycosides include maltose, sucrose, and glucose linked by glycosidic or ester linkage to an alkyl chain of 9, 10, 12, 13 or 14 carbon atoms, e.g., nonyl-, decyl-, dodecyl- and tetradecyl sucroside, glucoside, and maltoside. These compositions are nontoxic, since they are degraded to an alcohol or fatty acid and an oligosaccharide, and amphipathic. [0087] In some embodiments, the compositions comprising at least one PTH-related peptidic analogue may be prepared by admixing the peptide with a surfactant comprising of at least one alkylglycoside and/or sucrose ester, wherein the alkyl has from 10 to 14 carbon atoms.

[0088] In some embodiments, the compositions comprising at least one PTH-related peptidic analogue may be prepared by admixing a PTH-related peptidic analogue of the present embodiments, a stabilizing agent and a buffering agent, wherein the stabilizing agent is at least one alkylglycoside surfactant.

Plasma Concentration - AUC

[0089] The dose may also be selected to provide an effective plasma concentration of PTH-related peptidic analogues or other osteoporosis therapeutic. Examples of an effective maximum plasma concentration of the analogue(s) concentration may range from about 10 pg/mL to about 400 pg/mL, preferably from about 20 pg/mL to about 300 pg/mL; from about 50 pg/mL to about 280 pg/mL; from about 80 pg/mL to about 250 pg/mL; and from about 100 pg/mL to about 150 pg/mL. Other suitable dosage ranges for maximum plasma concentration of PTH-related peptidic analogues include 20-40 pg/mL, 40-60 pg/mL, 60-80 pg/mL, 80-100 pg/mL, 100-120 pg/mL, 120-140 pg/mL, 140-160 pg/mL, 160-180 pg/mL, 180-200 pg/mL, 200-230 pg/mL, 230-260 pg/mL, 260-300 pg/mL, 300-350 pg/mL, and 350- 400 pg/mL.

[0090] In another specific embodiment of the disclosure, the one or more PTH-related peptidic analogues are administered in an effective amount that results in the value for area under the curve (herein referred to as "AUC") in the plasma analogue(s) concentration versus time curve in the range of 5 pg-h/mL - 400 pg-h/mL. More preferably, the range of AUC is between 10 pg-h/mL - 350 pg-h/mL. More preferably, AUC is in the range of 20 pg-h/mL - 300 pg-h/mL. Alternatively, AUC is in the range of 50 pg-h/mL - 250 pg-h/mL; 70 pg-h/mL - 200 pg-h/mL; 90 pg-h/mL - 150 pg-h/mL;or 95 pg-h/mL - 125 pg-h/mL. Other suitable ranges for AUC include 5 pg-h/mL - 20 pg-h/mL, 20 pg-h/mL - 50 pg-h/mL, 50 pg-h/mL - 70 pg-h/mL, 70 pg-h/mL - 90 pg-h/mL, 90 pg-h/mL - 100 pg-h/mL, 100 pg-h/mL - 110 pg-h/mL, 110 pg-h/mL - 120 pg-h/mL, 120 pg-h/mL - 130 pg-h/mL, 130 pg-h/mL - 150 pg-h/mL, 150 pg-h/mL - 175 pg-h/mL, 175 pg-h/mL - 200 pg-h/mL, 200 pg-h/mL -225 pg-h/mL, 225 pg-h/mL - 250 pg-h/mL, 250 pg-h/mL -275 pg-h/mL, 275 pg-h/mL - 300 pg-h/mL, 300 -350 pg-h/mL, and 350 pg-h/mL - 400 pg-h/mL.

[0091] Accordingly, in one aspect, the disclosure provides a pharmaceutical formulation comprising a therapeutically effective amount of a PTH-related peptidic analogue as the active ingredient in a daily dosage range of 80μg to 160 μg or a every other day dosage range of 80 μg to 160 μg, wherein the PTH-related peptidic analogue has reduced phospholipase-C activity and maintains adenylate cyclase activity, in admixture with a pharmaceutically acceptable excipient, diluent, or carrier, or combinations thereof.

Formulations

[0092] A stabilized solution of a parathyroid hormone can include a stabilizing agent, a buffering agent, a preservative, an antibacterial agent and the like. The stabilizing agent incorporated into the solution or composition includes alcohols, ethanol or a polyol which includes a saccharide, preferably a monosaccharide or disaccharide, e.g. , glucose, trehalose, raffinose, or sucrose; a sugar alcohol such as, for example, mannitol, sorbitol or inositol, and a polyhydric alcohol such as glycerin or propylene glycol or mixtures thereof.

[0093] The buffering agent employed in the solution or composition of the described herein may be any acid or salt combination which is pharmaceutically acceptable. Useful buffering systems are, for example, acetate, lactate, or citrate sources. The concentration of buffer may be in the range of about 2 mM to about 500 mM, preferably about 2 mM to 100 mM. Any pharmaceutically acceptable buffer may be used in the present formulations/ compositions. Preferably, the buffer is present in a concentration of from about 1 mM to about 100 mM (e.g., about 5 mM, about 10 mM, about 15 mM, about 20 mM, about 25 mM, about 30 mM, about 35 mM, about 40 mM, about 50 mM, about 55 mM, about 60 mM, about 65 mM, about 70 mM, about 75 mM, about 80 mM, about 85 mM, about 90 mM, or about 95 mM).

[0094] The buffer to be included in the formulations may include, but is not limited to, the following: acetate (e.g., sodium acetate), sodium carbonate, citrate (e.g., sodium citrate), tartrate, lactate, glycylglycine, histidine, glycine, lysine, arginine, sodium dihydrogen phosphate, disodium hydrogen phosphate, sodium phosphate, and tris(hydroxymethyl)- aminomethan, or mixtures thereof. Each one of these specific buffers constitutes an alternative embodiment. [0095] The buffers are added to maintain the pH of the formulation to between about 3 to about 8 (e.g., about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 6.8, about 7, about 7.2, about 7.3, about 7.4, about 7.5, about 7.6, about 7.7, about 7.8, about 7.9, or about 8.0). According to some embodiments, the pH of the formulation may be between about 3 to about 6, between about 3 to about 5.5, between about 3 to about 5.2, between about 3 to about 4.5, between about 3 to about 4, between about 4 to about 5.5, between about 4.5 to about 5.5, between about 4 to about 6, between about 4 to about 7, between about 3 to about 7, between about 4 to about 7.5, between about 5 to about 7.5, between about 5 to about 6.5, between about 5 to about 8, between about 6 to about 8, between about 6.5 to about 7.5, or between about 7.5 to about 8.0).

[0096] Where a pharmaceutically acceptable preservative is to be included in the formulations described herein, the preservative is selected from the group consisting of phenol, m-cresol, benzalkonium chloride, chloroethanol, methyl p-hydroxybenzoate, propyl p-hydroxybenzoate, parabens, 2-phenoxyethanol, butyl p-hydroxybenzoate, 2-phenylethanol, benzyl alcohol, chlorobutanol, and thiomerosal, or mixtures thereof. Each one of these specific preservatives constitutes an alternative embodiment.

Dosing Regimen

[0097] Administration as described herein may consist of one or more cycles; during these cycles one or more periods of osteoclastic and osteoblastic activity will occur, as well as one or more periods when there is neither osteoclastic nor osteoblastic activity. Alternatively, administration may be conducted in an uninterrupted regimen; such a regimen may be a long term regimen, e.g., a permanent regimen.

[0098] It will be understood that the dosages of compositions and the duration of administration as described herein will vary depending on the requirements of the particular subject. The precise dosage regime will be determined by the attending physician or veterinary surgeon who will, inter alia, consider factors such as body weight, age and symptoms (if any). The compositions may, if desired, incorporate one or more further active ingredients.

[0099] During the dosing regimen, the hormone can be administered regularly (e.g., once or more each day or week), intermittently (e.g., irregularly during a day or week), or cyclically (e.g., regularly for a period of days or weeks followed by a period without administration). Regular administration can include once daily, once every two days, once every three days, once every four days, once every five days, once every six days, or once every seven days (once/week). Preferably PTH is administered once daily for 1-7 days for a period ranging from 3 months for up to 3 years in osteoporotic patients. In additional embodiments, PTH is administered for no less than 8 days. The present disclosure also encompasses embodiments wherein PTH is administered on an every other day or bi-weekly basis.

Kits

[00100] The present disclosure also encompasses a kit including the present pharmaceutical compositions and to be used with the methods described herein. The kit can contain a vial, for example, which contains a formulation as described herein and suitable carriers, either dried or in liquid form. The kit further includes instructions in the form of a label on the vial and/or in the form of an insert included in a box in which the vial is packaged, for the use and administration of the compounds. The instructions can also be printed on the box in which the vial is packaged. The instructions contain information such as sufficient dosage and administration information so as to allow a worker in the field to administer the drug. It is anticipated that a worker in the field encompasses any doctor, nurse, or technician who might administer the drug, or a patient who might self-administer the pharmaceutical composition.

Bone Disorders and Diseases

[00101] Bone Deficits

[00102] In one aspect, the subject in need has a bone deficit, which means that they will have less bone than desirable or that the bone will be less dense or strong than desired. A bone deficit may be localized, such as that caused by a bone fracture or systemic, such as that caused by osteoporosis. Bone deficits may result from a bone remodelling disorder whereby the balance between bone formation and bone resorption is shifted, resulting in a bone deficit. Examples of such bone remodelling disorders include, for example, osteoporosis, Paget's disease, renal osteodystrophy, renal rickets, osteoarthritis, rheumatoid arthritis,

achondroplasia, osteochondrytis, hyperparathyroidism, osteogenesis imperfecta, congenital hypophosphatasia, fribromatous lesions, fibrous displasia, multiple myeloma, abnormal bone turnover, osteolytic bone disease and periodontal disease. Bone remodelling disorders includes metabolic bone diseases which are characterized by disturbances in the organic matrix, bone mineralization, bone remodelling, endocrine, nutritional and other factors which regulate skeletal and mineral homeostasis. Such disorders may be hereditary or acquired and generally are systemic, affecting the entire skeletal system.

[00103] Thus, in one aspect the human subject may have a bone remodelling disorder. Bone remodelling as used herein refers to the process whereby old bone is being removed and new bone is being formed by a continuous turnover of bone matrix and mineral that involves bone resorption by osteoclasts and bone formation by osteoblasts.

[00104] Bone Trauma

[00105] The methods described herein provide benefit to a subject that may suffer or have suffered trauma to one or more bones. The method can benefit mammalian subjects, such as humans, horses, dogs, and cats, in particular, humans. Bone trauma can be a problem for racing horses and dogs, and also for household pets. A human can suffer any of a variety of bone traumas due, for example, to accident, medical intervention, disease, or disorder. In the young, bone trauma is likely due to fracture, medical intervention to repair a fracture, or the repair of joints or connective tissue damaged, for example, through athletics. Other types of bone trauma, such as those from osteoporosis, degenerative bone disease (such as arthritis or osteoarthritis), hip replacement, or secondary conditions associated with therapy for other systemic conditions (e.g., glucocorticoid osteoporosis, burns or organ transplantation) are found most often in older people.

[00106] Osteoporosis can lead, for example, to vertebral and/or non-vertebral fractures. Vertebral fractures are those involving the spinal column and non-vertebral fractures refers to any fracture not involving the spinal column. Non- vertebral fractures are more common than fractures of the vertebrae— an estimated 850,000 non-vertebral compared with 700,000 vertebral fractures occur annually in the United States. Non-vertebral fractures include more than 300,000 hip and 250,000 wrist fractures, in addition to 300,000 fractures at other non- vertebral sites. Other examples of non- vertebral fractures include a hip fracture, a fracture of a distal forearm, a fracture of a proximal humerus, a fracture of a wrist, a fracture of a radius, a fracture of an ankle, a fracture of an humerus, a fracture of a rib, a fracture of a foot, a fracture of a pelvis, or a combination of these.

[00107] The methods described herein can be used to decrease the risk of such fractures or for treating such fractures. The risk of fracture is diminished and the healing of a fracture is aided by increasing the strength and/or stiffness of bone, for example, in the hip, the spine or both. A typical woman at risk for osteoporosis is a postmenopausal woman or a

premenopausal, hypogonadal woman. A preferred subject is a postmenopausal woman, and is independent of concurrent hormone replacement therapy (HRT), estrogen or equivalent therapy, or antiresorptive therapy. The methods described herein can benefit a subject at any stage of osteoporosis, but especially in the early and advanced stages.

[00108] The present disclosure provides a method, in particular, effective to prevent or reduce the incidence of fractures in a subject with or at risk of progressing to osteoporosis. For example, the compositions and methods described herein can reduce the incidence of vertebral and/or non-vertebral fracture, reduce the severity of vertebral fracture, reduce the incidence of multiple vertebral fracture, improve bone quality, and the like. In another embodiment, the methods described herein can benefit patients with low bone mass or prior fracture who are at risk for future multiple skeletal fractures, such as patients in which spinal osteoporosis may be progressing rapidly.

[00109] Other subjects can also be at risk of or suffer bone trauma and can benefit from the methods described herein. For example, a wide variety of subjects at risk of one or more of the fractures identified above, can anticipate surgery resulting in bone trauma, or may undergo an orthopedic procedure that manipulates a bone at a skeletal site of abnormally low bone mass or poor bone structure, or deficient in mineral. For example, recovery of function after a surgery such as a joint replacement (e.g. knee or hip) or spine bracing, or spinal fusion indications or other procedures that immobilize a bone or skeleton can improve due to the methods described herein. The methods described herein can also aid recovery from orthopedic procedures that manipulate a bone at a site of abnormally low bone mass or poor bone structure, which procedures include surgical division of bone, including osteotomies, joint replacement where loss of bone structure requires restructuring with acetabulum shelf creation and prevention of prosthesis drift, for example. Other suitable subjects for practice of the methods and compositions described herein include those suffering from

hypoparathyroidism or kyphosis, who can undergo trauma related to, or caused by, hypoparathyroidism or progression of kyphosis.

[00110] Bone Toughness and Stiffness [00111] The methods described herein reduce the risk of trauma or aids recovery from trauma by increasing bone toughness, stiffness or both. Generally toughness or stiffness of bone results from mass and strength of cortical and trabecular (cancellous) bone. The methods described herein can provide levels of bone toughness, stiffness, mass, and/or strength within or above the range of the normal population. Preferably the methods and composition described herein provide increased levels relative to the levels resulting from trauma or giving rise to risk of trauma. Increasing toughness, stiffness, or both decreases risk or probability of fracture compared to an untreated control population.

[00112] Certain characteristics of bone when increased provide increased bone toughness and/or stiffness. Such characteristics include bone mineral density (BMD), bone mineral content (BMC), activation frequency or bone formation rate, trabecular number, trabecular thickness, trabecular and other connectivity, periosteal and endocortical bone formation, cortical porosity, cross sectional bone area and bone mass, resistance to loading, and/or work to failure. An increase in one or more of these characteristics is a preferred outcome of the methods described herein.

[00113] The methods described herein are effective for increasing the toughness and/or stiffness of any of several bones. For example, the present method can increase the toughness and/or stiffness of bones including a hip bone, such as an ilium, a leg bone, such as a femur, a bone from the spine, such as a vertebra, or a bone from an arm, such as a distal forearm bone or a proximal humerus. This increase in toughness and/or stiffness can be found throughout the bone, or localized to certain portions of the bone. For example, toughness and/or stiffness of a femur can be increased by increasing the toughness and/or stiffness of a femur neck or a femur trochanter. Toughness and/or stiffness of a hip can be increased by increasing the toughness and/or stiffness of an iliac crest or iliac spine.

Toughness and/or stiffness of a vertebra can be increased by increasing the toughness and/or stiffness of a pedicle, lamina, or body. Advantageously, the effect is on vertebra in certain portions of the spine, such as cervical, thoracic, lumbar, sacral, and/or coccygeal vertebrae. Preferably the effect is on one or more mid-thoracic and/or upper lumbar vertebrae.

[00114] The increase in toughness and/or stiffness can be found in each of the types of bone, or predominantly in one type of the bone. Types of bone include spongy (cancellous, trabecular, or lamellar) bone and compact (cortical or dense) bone and the fracture callus. The methods described herein preferably increase toughness and/or stiffness through its effects on cancellous and cortical bone, or on cortical bone alone. Trabecular bone, bone to which connective tissue is attached can also be toughened and/or stiffened by the present method. For example, it is advantageous to provide additional toughness at a site of attachment for a ligament, a tendon, and/or a muscle.

[00115] In another aspect of the methods and compositions described herein, increasing toughness or stiffness can reduce incidence of fracture. In this aspect, increasing toughness or stiffness can include reducing incidence of vertebral fracture, reducing incidence of severe fracture, reducing incidence of moderate fracture, reducing incidence of non-vertebral fracture, reducing incidence of multiple fracture, or a combination thereof.

[00116] The methods described herein may also be used to enhance bone formation in conditions where a bone deficit is caused by factors other than bone remodelling disorders. Such bone deficits include fractures, bone trauma, conditions associated with post-traumatic bone surgery (e.g., bone grafts or bone fusions), post-prosthetic joint surgery, post plastic bone surgery, post dental surgery, bone chemotherapy, and bone radiotherapy. Fractures include all types of microscopic and macroscopic fractures. Examples of fractures and/or injuries include avulsion fracture, comminuted fracture, non-union fracture, transverse fracture, oblique fracture, spiral fracture, segmental fracture, a segmental gap, displaced fracture, impacted fracture, greenstick fracture, torus fracture, fatigue fracture, intra-articular fracture (epiphyseal fracture), closed fracture (simple fracture), open fracture (compound fracture), a bone void, and occult fracture in any bones of the subject.

[00117] As previously mentioned, a wide variety of bone diseases may be treated in accordance with the present disclosure, for example all those bone diseases connected with the bone-remodelling cycle. Examples of such diseases include all forms of osteoporosis, osteomalacia and rickets. Osteoporosis, especially of the post-menopausal, male, post- transplant, and steroid-induced types, is of particular note. In addition, PTH peptide analogues find use as bone promotion agents, and as anabolic bone agents. Such uses form another aspect of the present disclosure.

Definitions

[00118] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the methods and compositions described herein, suitable methods and materials are described below.

[00119] For the purposes of promoting an understanding of the embodiments described herein, reference will be made to preferred embodiments and specific language will be used to describe the same. The terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present discloure. As used throughout this disclosure, the singular forms "a," "an," and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, a reference to "a

composition" includes a plurality of such compositions, as well as a single composition, and a reference to "a therapeutic agent" is a reference to one or more therapeutic and/or

pharmaceutical agents and equivalents thereof known to those skilled in the art, and so forth.

[00120] Throughout this application, the term "about" is used to indicate that a value includes the standard deviation of error for the device or method being employed to determine the value.

[00121] Reference to numeric ranges throughout this specification encompasses all numbers falling within the disclosed ranges. Thus, for example, the recitation of the range of about 1% to about 5% includes 1%, 2%, 3%, 4%, and 5%, as well as, for example, 2.3%, 3.9%, 4.5%, etc.

[00122] The use of the term "or" in the claims is used to mean "and/or" unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and "and/or."

[00123] As used in this specification and claim(s), the words "comprising" (and any form of comprising, such as "comprise" and "comprises"), "having" (and any form of having, such as "have" and "has"), "including" (and any form of including, such as "includes" and "include") or "containing" (and any form of containing, such as "contains" and "contain") are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[00124] As used herein, the "PTH peptide analogues" of the as described herein are preferably, but not exclusively, non-naturally occurring and may be obtained either recombinantly or by peptide synthesis. The PTH analogues described herein include full length PTH (1-84), 1-31 and 1-34 fragments, and other fragments or variants of fragments of human, rat, porcine, or bovine PTH that have human PTH activity as determined in the ovarectomized rat model of osteoporosis (Kimmel et al, Endocrinology, 1993, 32(4): 1577). Human PTH activity includes the ability of the PTH to increase trabecular and/or cortical bone growth. The PTH analogues described herein increase AC activity when administered to a PTH receptor containing or expressing cell in culture, such as an osteoblast or an osteoclast. The PTH analogues described herein have certain additional functional activities, as defined below.

[00125] As used herein, a PTH peptide analogue that has a "reduced phospholipase-C activity" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less than full activation of phospholipase-C, as compared to the full- length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length.

[00126] As used herein, a PTH peptide analogue that leads to "reduced bone resorption" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less bone resorption, as compared to the full-length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length.

[00127] As used herein, a PTH peptide analogue that leads to "reduced hypercalcemia levels" refers to a PTH peptide analogue that has been truncated or modified in some manner so as to trigger less incidences of hypercalcemia, or lower severity of hypercalcemia, as compared to the full-length PTH peptide or other PTH peptide analogues which are at least 34 amino acid residues in length.

[00128] As used herein, "treating" or "treatment of a condition or subject refers to taking steps to obtain beneficial or desired results, including clinical results. For purposes of this disclosure, beneficial or desired clinical results include, but are not limited to, alleviation or amelioration of one or more disease, symptom, or condition related to bone deficiency.

Generally, such bone deficit disease, symptoms, and conditions are treated by inducing bone formation as measured by an increase in bone mineral density ("BMD"). For example, symptoms of osteoporosis include back pain, loss of height and stooped posture, a curved backbone (dowager's hump), or fractures that may occur with a minor injury (especially of the hip, spine, or wrist). Symptoms of Paget's disease most commonly include bone pain. Other symptoms can include: headaches and hearing loss, neck pain, pressure on nerves, increased head size or bending of spine, hip pain, damage to cartilage of joints (which may lead to arthritis), and Barrel-shaped chest. As used herein, "reduction" of a symptom or symptoms (and grammatical equivalents of this phrase) means decreasing of the severity or frequency of the symptom(s), or elimination of the symptom(s).

[00129] As used herein, "administering" or "administration of a drug or pharmaceutical composition or formulation described herein to a subject (and grammatical equivalents of this phrase) includes both direct administration, including self-administration, and indirect administration, including the act of prescribing a drug. For example, as used herein, a physician who instructs a patient to self-administer a drug and/or provides a patient with a prescription for a drug is administering the drug to the patient.

[00130] A variety of administration routes can be used in accordance with the compositions and methods described herein. An effective amount of the peptide described herein can be administered parenterally, orally, by inhalation, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir.

[00131] As used herein, a "therapeutically effective amount" of a drug or pharmaceutical composition or formulation, or agent, described herein is an amount of a drug or agent that, when administered to a subject with a disease or condition, will have the intended therapeutic effect, e.g. , alleviation, amelioration, palliation or elimination of one or more manifestations of the disease or condition in the subject. The full therapeutic effect does not necessarily occur by administration of one dose and may occur only after administration of a series of doses. Thus, a therapeutically effective amount may be administered in one or more administrations.

[00132] As used herein, a "prophylactically effective amount" of a drug or pharmaceutical composition or formulation, or agent, described herein is an amount of a drug or agent that, when administered to a subject, will have the intended prophylactic effect, e.g., preventing or delaying the onset (or reoccurrence) of disease or symptoms, or reducing the likelihood of the onset (or reoccurrence) of disease or symptoms. The full prophylactic effect does not necessarily occur by administration of one dose and may occur only after administration of a series of doses. Thus, a prophylactically effective amount may be administered in one or more administrations.

[00133] Administration of a bone enhancing agent "in combination with" a drug or pharmaceutical composition or formulation described herein includes parallel administration (i.e., administration of both the drug and the agents to the subject over a period-of time, coadministration (in which both the drug and agents are administered at approximately the same time, e.g., within about a few minutes to a few hours of one another), and co-formulation (in which both the drug and agents are combined or compounded into a single dosage form suitable for oral or parenteral administration).

[00134] A "subject" is a mammal, preferably a human, but can also be an animal in need of veterinary treatment, e.g., companion animals (e.g., dogs, cats, and the like), farm animals (e.g., cows, sheep, pigs, horses, and the like) and laboratory animals (e.g., rats, mice, guinea pigs, and the like).

[00135] Additional active ingredients can be included in the present compositions. Choices are not limited, but may be chosen for a desired combined therapeutic effect. For example, active ingredients that may be added for a complementary therapeutic effect include, but are not limited to, vitamin D and analogs, estrogen, calcitonin, bisphosphonates, and mixtures thereof. A particularly desirable choice is calcitonin.

Examples

Example 1: Clinical Evaluation of a Nasal Spray Formulation of

Teriparatide Demonstrates Rapid Absorption and Similar Systemic

Exposure to Marketed Subcutaneous Teriparatide.

[00136] In this study 25 health postmenopausal women were treated with various doses of nasal (NS) ZT-034 between 20 and 160 μg/day for a seven day period. Pharmacokinetic profiles were assessed on Day 1 and Day 7. Patients were dosed 20 μg (1 spray), 40 μg (1 spray), 80 μg (2 sprays), or 160 μg (4 sprays of 40 μg each). PTH (1-34) mean maximum concentrations C _max , and exposure, AUC, increased in approximately dose-proportional manner over the dose range tested following administration of NS ZT-034. Median T _max for PTH (1-34) following intranasal administration of ZT-034 occurred at approximately 0.08 to 0.33 hours across all treatment groups and indicated a consistent absorption of NS ZT-034. The mean T ₂ of PTH (1-34) ranged from 0.19 to 0.65 hours across all treatment groups and demonstrated the rapid elimination of drug from plasma. The bioavailability (F) of NS ZT- 034 was evaluated by comparing the C _max and AUC for the intranasal administration versus subcutaneous administration of Forteo® 20 μg. The average bioavailability of ZT-034 based on AUC for the 20, 40, 80, 120, and 160 μg dose groups was 22%, 66%, 58%, 90%, and 54% on Day 1 and 28%, 47%, 92%, 88%, and 69% on Day 7, respectively. Based on C _max , the average bioavailability was 40%, 150%, 120%, 160%, and 130% on Day 1 and 65%, 110%, 200%, 100%, and 160% on Day 7 for the 20, 40, 80, 120, and 160 μg dose groups, respectively. No changes in bioavailability were evident following repeated administration of ZT-034. The ZT-034 bioavailability was, however, greater with increasing dose.

[00137] These data suggest that doses in the range of 80-160 μg/day are most comparable to Forteo® in terms of AUC. For C _max , bioavailability in the 40-160 μg/day dose groups ranged from approximately 100%-200% and demonstrated a trend towards a higher bioavailability for ZT-034. These data suggest that NS ZT-034 is able to achieve levels of PTH (1-34) that are comparable or slightly higher than subcutaneous Forteo® 20 μg when doses of 80-160 μg are administered. See Figure 6.

[00138] The quantitative composition of intranasal ZT-034 is provided in the table below. These were ready to use liquid formulations and were intended to deliver 50 μΐ volume with an attached intranasal spray device.

[00139] Table 1-1 : Quantitative Composition for Intranasal ZT-034.

*API amount is adjusted for potency. ** EDTA amount is adjusted for dihydrate EDTA.

[00140] Table 1-2: Observed Pump Parameters for Three Different Concentrations of Intranasal (IN) ZT-034

1.6 50.7±2.9 52.06±4.32 24.15±0.80 47.69±2.11 103.57±7.81 1.02±0.10

[00141] Table 1-3: Pharmacokinetic profiles

Example 2: A 6 Period Sequential Dose Study to Evaluate the

Pharmacokinetics and Tolerability of Nasal Spray ZT-034 Compared to

Teriparatide Following Single Dose Administration in Healthy Adults

[00142] Objectives: To compare the pharmacokinetics (PK) of nasal spray (NS) ZT-034 to the PK of subcutaneous (SC) teriparatide 20 μg (Forteo®) following single dose

administration.

[00143] Design: This was an open label, single dose, 6-period sequential dose design.

Subjects received each of the five treatment regimens listed below, in six separate study periods. Doses of study medication were separated by at least 72 hours. Subjects received the Regimens A, B, C, D and E, in escalating order starting in the second period.

[00144] Blood samples were obtained predose, and at 10, 15, 20, 30, 45, 60, 90, 120 and 240 minutes post dose and PTH (1-34) levels evaluated by a validated ELISA assay. Four strengths of lyophilized product were made 1.71, 3.43, 5.14, and 6.86 mg corresponding to 0.57, 1.14, 1.71, and 2.29 mg/mL after reconstitution with 3.0 mL distilled water. These four strengths deliver 40, 80,120, and 160 μg, respectively per 70 μΙ_, spray administered via a nasal spray pump.

[00145] The quantitative composition of intranasal ZT-034 is provided in the following Table 1-1.

Table 1-1 : Quantitative Composition for Intranasal ZT-034

*APi amount Is adjusted for potency.

** Amount adjusted for hydrate and salt content

[00146] The spray pump that was used delivered about 70 μΐ per actuation to deliver the corresponding dose for each strength. Using distilled water, the pump gave an individual shot weight of 70 mg +/- 15% and a median droplet size distribution (D, 0.5) of 40 microns, and gave 2% of droplets that were < 10 microns.

[00147] Administration of teriparatide by nasal spray produces a dose proportional increase in exposure (AUC) and maximum achieved concentration (C _max ) (Figures 1-5 and Table 1-2). Unexpectedly the concentration time profile of nasal spray teriparatide differed from that of injectable teriparatide with more rapid absorption and more rapid clearance from the circulation. C _max trended to higher values with nasal spray than with injection whereas AUC trended lower (Table 1-3). Table 1-2: Pharmacokinetic parameters of teriparatide delivered by nasal spray or subcutaneous injection.

Table 1-3: C _max and AUC of nasal spray teriparatide relative to injectable teriparatide

(geometric LS means).

[00148] Consistent with this PK profile changes in calcium metabolism, including fractional excretion of calcium, serum total calcium and suppression of endogenous PTH levels all demonstrated earlier onset of effect and more rapid return to baseline with nasal spray than with injectable teriparatide (Figures 1-5). This finding suggests that the more rapid clearance of teriparatide from the circulation observed with nasal spray teriparatide is associated with a reduced duration of the pharmacological effects on calcium metabolism normally associated with administration of exogenous teriparatide.

Example 3: New Zealand White Male and Female Rabbits

[00149] New Zealand White male and female rabbits were administered ZT-034 at 10 and 50 μg/kg with either 0.06% or 0.125% DDM (both 10 mM acetate, pH 5), intranasally (IN), once daily into alternate nostrils for 28 days as shown in Table 3-1 below:

[00150] Table 3-1 : IN Study Design in Rabbits

[00151] The dose volume was 50 μί/day via drop instillation for all dose groups.

Toxicokinetic evaluations were made on Days 1 and 28 on the terminal animals. [00152] Based on TK data analysis (Table 3.2), male and female rabbits were exposed to quantifiable concentrations of ZT-034 on Days 1 and 28 after intranasal administrations of 10 and 50 ZT-034 with 0.06% or 0.125% DDM. In general, the toxicokinetics of ZT- 034 appeared to be non-linear with dose in either 0.06 or 0.125% DDM showing a greater than dose-proportional increase. In the study period of 28 days, it appears that there was no accumulation of test article in rabbits of both sexes after repeated administration of ZT-034. On Day 1, systemic exposure to ZT-034 in 0.06 and 0.125% DDM were generally greater than the systemic exposure to ZT-034 in both concentrations of DDM on Day 28. There were no clear consistent patterns for gender differences. The T _max in the range of 5-25 min and the Ti/2 in the range of 9-28 minutes are consistent with rapid absorption across the nasal epithelium and rapid clearance, respectively, following IN delivery of ZT-034 with DDM.

[00153] Table 3-2: IN Toxicokinetic Parameters in the Rabbit on Day 1 and 28 of Dosing.

Example 4: New Zealand White male rabbits

[00154] Twenty- five male New Zealand white rabbits were assigned to five groups (5 rabbits/group). ZT-031 was dosed at 2, 4, 8, 20 μg/kg in the presence or absence of DDM at 0.18% by intranasal administration as a 100 instillation to both nares of each animal (50 μΐ, per nare). An additional group of rabbits was dosed at 20 μg/kg by intranasal

administration in an alternate formulation, Rhinaris® (Rhinaris is a saline moisturizing nasal spray used to relieve dry and irritated nasal passages containing Sodium Chloride (0.2%) and other inactive ingredients: Benzalkonium Chloride Solution, Polyethylene Glycol, Potassium Chloride, Potassium Phosphate Monobasic, Propylene Glycol, Water (Purified), Sodium

Phosphate Dibasic. Blood samples of 1 mL in volume were drawn from the ear vein over a two hour time period at 0, 5, 10, 15, 20, 30, 45, 60, and 120 minutes following intranasal administration of ZT-031 dosing solutions. Each rabbit was bled for a total of 9 times. At 2 hours (120 minutes) post-dose, the last blood sample was collected and each rabbit was observed for an hour and then euthanized with C0 ₂ . Following blood collection, plasma was immediately prepared from each blood sample using lithium/heparin as the anticoagulant and divided into two aliquots and immediately frozen on dry ice. Levels of ZT-031 in plasma samples were determined using an ELISA assay. The individual plasma concentration-time data at each dose level was used to calculate composite profiles to be used in the calculation of pharmacokinetic parameters of ZT-031 using Pharmacokinetic function component in

Excel 2007. Pharmacokinetic parameters are summarized in Table 4-1.

[00155] Dose-proportional increases in drug exposures as determined by both C _max and

AUC last were observed with increases in dose from 2 to 20 μg/kg (with 0.18% DDM). At the 20 μg/kg dose, the C _max was approximately 2.5-fold higher in the presence of DDM than in the absence, 874 and 369 pg/mL respectively. The T _max following IN administration of all doses with 0.18% DDM was 5 minutes, whereas, the T _max following IN administration of ZT- 031 in the absence of DDM was extended to 15 minutes indicating faster absorption in the presence of DDM. The apparent elimination half-life (T ₂ ) in all the IN dose groups ranged from 8-34 minutes independent of increasing doses of ZT-031 or in the presence or absence of DDM. Administration of ZT-031 in the Rhinaris formulation did not give any quantifiable ZT-031 plasma levels.

[00156] A summary of dosing and blood collection times is shown in Table 4-1. See also Figure 7.

[00157] Table 4- 1 : Summary of Dosing and Blood Collection

Group Number Test Dose Level Route of

Administratio

Number of Males Article ^g/kg) n Blood Collection Times 1 5 ZT-031 with Intravail® A3 2

2 5 ZT-031 with Intravail® A3 4

3 5 ZT-031 with Intravail® A3 8

0,5, 10, 15, 20, 30, 45, 60 4 5 ZT-031 with Intravail® A3 20

and 120 minutes 5 5 ZT-031 in Rhinaris® 20

ZT-031 without Intravail®

A3 20 Intranasal

100 intranasal (IN) instillation to a both nare, 50 μΐ per nare

Lithium/heparin was anticoagulant

Table 4-2: Summary of Mean Pharmacokinetic Parameters of ZT-031 Following Intranasal Administration to New Zealand Rabbits (with and without Intravail® A3)

Example 5: ZT-031 and ZT-034 in an in vivo rat model.

[00158] One-hundred female Sprague-Dawley rats were assigned to ten groups (10

rats/group). Doses were administered intranasally as a 20 μΐ _^ instillation to a single nare or subcutaneous ly as a 100 μΙ_, injection between the skin and underlying tissue layers in the scapular region on the back of each animal. Blood samples of 350 μΐ _^ in volume were drawn by orbital bleed over a three hour time period at 0, 5, 10, 15, 25, 40, 60, 120, and 180 minutes following either intranasal or subcutaneous administration of ZT-031 or PTH 1-34 dosing solutions. Each group of ten rats was divided into 2 subgroups and blood was collected from each subgroup at alternating sample times. Each rat was bled a total of 4 or 5 times depending on the subgroup. At 3 hours (180 minutes) the last blood sample was collected and each rat was euthanized with C0 ₂ . Following blood collection, plasma was immediately prepared from each blood sample using K2EDTA as the anticoagulant. Blood levels of compound were measured by ELISA.

[00159] Rats in Groups 1 to 5 received ZT-031. Respectively, groups 1-4 received a 20 instillation of ZT-031 at 10, 75, 125 or 200 μg/kg average body mass/group to a single nare with DDM (buffer). Female rats in group 5 received a 100 subcutaneous administration of ZT-031 at 75 μg/kg average body mass/group without DDM.

[00160] Rats in Groups 6 to 10 received PTH 1 34. Respectively, Groups 6 through 9 received a 20 μΐ, instillation of PTH 1-34 at 10, 75, 125 μg/kg or 200 μg/kg average body mass/group to a single nare with DDM (buffer). Rats in Group 10 received a 100 μί subcutaneous administration of PTH 1 34 at 75 μg/kg average body mass/group without DDM.

[00161] Table 5-1 : Test Article Preparation: calculations based on average weight of 250g/rat.

[00162] Table 5-2: Dose administration (at 250g average body weight).

[00163] Rats were placed in an anesthesia induction box in an atmosphere of 2% Isoflurane prior to dosing. Intranasal instillation (20ul/naris) or subcutaneous injection between the skin and underlying tissue layers in the scapular region (100 μΐ/rat) were administered to each anesthetized rat.

[00164] Table 5-3: Summary of Dosing and Blood Collection.

Grasp, Test DoSe Level Route of

Article Adirmiis ratseii Blood- C ^' oltectkfii ijrses i. ZT-031. with la rs^ ^■ ail A3 10 Siirsass l

9 •1© PTH 1-3- ί with Mi'a :vail A3 200 Mr¾Jia¾si

10 10 PTH 1-34

Groups !· -4 <¾isd 6-9 : : 20 u-L iat aa sal ^' (∑_· <?) iastillatioa to -a. ¾ iiiiglc aare

Groups. 5 · · · l¾ ;¾½Beom ·; i.

[00165] Following intranasal (IN) or subcutaneous (SC) administration of ZT-031 and PTH 1-34, T _max ranged from 5 to 15 minutes. Plasma levels declined in a similar manner following ZT-031 and PTH 1-34 IN dosing with apparent elimination half-lives (Ti _/2 ) increasing with increasing doses of ZT-031 (ranging from 5.1 to 35.8 minutes) and PTH 1-34 (ranging from 7.4 to 50.7 minutes) when they were administered with DDM. Following a single SC dose (without DDM), T ₂ was 22.3 minutes for ZT-031 and 31.1 minutes for PTH 1-34.

[00166] Following IN doses of ZT-031, increases in dose from 10 to 200 μg/kg (with Intravail® A3) resulted in greater than dose-proportional increases in AUCinf. Similar increases in AUCinf were observed following IN dosing of PTH 1-34 with DDM except, an increase in dose from 75 to 125 μg/kg , resulted in a less than dose-proportional increase in PTH 1-34 AUCinf.

[00167] Following single IN doses of 10, 75 125 and 200 μg/kg ZT-031 administered with DDM, the relative bioavailability of ZT 031, relative to a single SC dose of 75 μg/kg ZT-031 administered without DDM, was 33.8, 514, 985 and 2091%, respectively. Following single IN doses of 10, 75, 125 and 200 μg/kg PTH 1-34 administered with DDM, the relative bioavailability of PTH 1-34, relative to a single SC dose of 75 μg/kg PTH 1-34 administered without DDM, was 16.4, 204, 133 and 283%, respectively.

[00168] See Figures 8 and 9. [00169] Table 5-4: Summary of Mean Pharmacokinetic Parameters of ZT-031 and PTH 1 34 Following Subcutaneous (without DDM) and Intranasal (with DDM) Administration to Female Sprague Dawley Rats.

Alitor F

Te st Aitfcte t &i Cpg-saitii'mL) (%)

ZT-031 sr 75 S54S 5 8832» 92630 92.940 22.3

lis 10 5 4055 4tS8 1S6 5J 33.8

ZT-03 i with s? 75 43020 5 47ei29 4770SS 477320 25 514

DDM IN 125 72853 10' ί 522443 1522443 525072 35,8 SS5

IN 200 152760 15 5178784· 5 7S7S4 51 SI 659 IS. 2ϋ ϊ

PTHl-34 SC ^• 75 i t s S 5 3S2031 3S2031 3S490 31.1

ΪΜ 10 714 5 S098 S3S0 S397 7.4 16.4

PTH 1-14 ¾¾S IN 75 5 7S¾2¾3? S29.37 785104 34,5. 204

[00170] Abbreviations:

AUC(0-180m) Area under the plasma concentration-time curve from time zero to 180 minutes

post-dose

AUC _last Area under the plasma concentration-time curve from time zero to the time of the last measurable concentration

AUC _inf Area under the plasma concentration-time curve from time zero to infinity r ^max The maximum observed plasma concentration

ELISA Enzyme-Linked Immunosorbent Assay

F Bioavailability

IN intranasal

LLOQ Lower Limit of Quantification

NC Not Calculated

ND Not Determined

NS No Sample

PTH 1-34 Parathyroid Hormone 1-34

SC subcutaneous

SE Standard error

l/2 Terminal half-life

T ¹ max Time to reach C _mj

ULOQ Upper Limit of Quantification

ZT-031 Zelos' Parathyroid Hormone Analog Ostabolin-C Example 6: Relative Bioavialbility of Intranasal Dosing Relative to

Subcutaneous Dosing in Monkeys

[00171] Methods

[00172] Ten cynomolgus monkeys were allocated to the study. On the first administration day, the body weight ranged from 3.1 and 5.0 kg.

[00173] This study was divided into two different phases:

[00174] Phase I

[00175] For Phase I, the animals were allocated to two groups of five females (Group l(low dose): and group 2 (high dose)). Animals received the test items ZT-031 and PTH 1-34 in 10 mM Sodium Acetate buffer, pH 5.0 with 0.1% EDTA and 0.18% DDM for the intranasal administration and in 10 mM Sodium Acetate Buffer, mannitol at pH 4.0 for the subcutaneous injection. For each test item, the animals in Groups 1 and 2 were first dosed by a single intranasal administration and then by subcutaneous injection. The design of the Phase I study is shown in the table below:

Dose Dose* Dose-volume Concentration

Test item Period Route Group

^g/kg) ( ¾) ^L or mL/kg) ^g mL)

IN 1 5 15 43.5 μί** 344.8

Day 1

with 0.18% DDM 2 ^a 50 150 43.5 μί** 3448.0

ZT-031

1 5 15 0.2 mL/kg 25

Day 8 SC

2 ^a 50 150 0.2 mL/kg 250

Day 15 IN 1 5 15 43.5 μί** 344.8

PTH 1-34 with 0.18% DDM ₂ ^a 50 150 43.5 μί** 3448.0

15 0.2 mL/kg 25

Day 22 SC

150 0.2 mL/kg 250

*: dose based on 3 kg monkey; **: adjustment to device volume output.

IN: intranasal route; SC: subcutaneous route.

[00176] At the end of the phase I all animals were moved to phase II.

[00177] Phase II

[00178] After a 1-week washout/recovery period at the end of phase I, the ten female monkeys were placed into three groups of three/four animals per group. The monkeysin each group were dosed the test item ZT-031 formulated in 10 mM Sodium Acetate Buffer, 0.1 %> EDTA at pH 5.0 and at three concentrations (0%, 0.06% and 0.125%) of DDM by the intranasal route as follows: Number of Percentage Dose

Test Dose Dose' Concentration

Period Route Group animal of DDM volume

item g/kg) fog) g/mL)

Animal number (%) foL)

43.5

3 females 0 50 150 3448.0 uL**

43.5

ZT-031 Day 29 IN 3 females 0.06 50 150 3448.0

43.5

4 females 0.125 50 150 3448.0

*: dose based on 3 kg monkey.

**: adjustment to device volume output.

[00179] Blood samples for the determination of plasma levels of the test items were collected from all animals before test item administration of phases I and II (pre-dose sample) and then at 5, 10, 15, 25, 60 and 180 min after each administration.

[00180] Venous blood was collected into tubes containing EDTA. Pharmacokinetic analysis was performed by a non-compartmental method using the validated WinNonLin® software.

[00181] Results

[00182] The relative intranasal bioavailability of ZT-031 compared to subcutaneous injection ranged between 35-40% (AUC) with 0.18% DDM. This was significantly higher than the bioavailability observed when no DDM was present in the dosing solutions (~5%). Increases of DDM concentration up to 0.125% only showed modest improvement of relative bioavailability (5-1 1%). Good systemic exposure of ZT-031 was achieved following IN dosing with dose proportionality among the low and high dose groups (AUC).

[00183] ZT-031 IN PK Parameters in Monkeys

nc: not calculated as not enough point in the terminal phase or as AUC extrapolated hig

20%.

IN: intranasal; SC: subcutaneous. N/A: Not Applicable [00184] Relative Bioavailability (Rel F%) = ((AUC _{inf m} xDose _sc ) / ( AUC _{inf sc} x Dose

IN))X 100

[00185] ZT-031 IN PK Parameters in the Presence or Absence of DDM in Monkeys

nc: not calculated as not enough point in the terminal phase or as AUC _ex trapoiated higher than 20%. IN: intranasal.

[00186] Relative Bioavailability (Rel F%) = ((AUC _{inf m} xDose _sc ) / ( AUC _{inf sc} x Dose

IN))X 100.

[00187] ZT-034

[00188] Mean plasma exposure (AUC _0-t ) values of ZT-034 in the presence of 0.18%

DDM were found to be non-linear showing greater than dose proportionality with increase in dose from 5 to 50 μg/kg (

[00189]

[00190] Table 6-2). The IN bioavailability of ZT-034 at the 5 μg/kg dose relative to SC injection was very high (82%), but only one monkey had quantifiable levels of ZT-034. However, at the high dose (50 μg/kg) the ZT-034 relative bioavailability was approximately 20%.

[00191] The T _max values following IN or SC administration ranged between 10 and 25 minutes for ZT-034. The elimination half-life of approximately 33 min was similar following both IN and SC dosing.

Table 6-2: Pharmacokinetic Parameters Following Intranasal or Subcutaneous Administration of ZT-034 in the Presence of Absence of DDM

ZT-034 189 7513

5. nc 15 nc n/a SC ±73 ±4037

ZT-034 32.9 19422 1436386 910100

50 25 n/a

SC ±3.9 ±14009 ±1311248 ±534809

* PK data from one animal; nc: not calculated; in: intranasal; sc: subcutaneous; n/a: Not Applicable; Relative Bioavailability (Rel F%) = ((AUC _M xDose _sc ) / ( AUC _m - _{f sc} x Dose IN))X 100

[00193] t _½ (half life) following both IN and SC dosing ranged ~ 1 -2 h

[00194] Lowering the concentration of DDM below 0.18% in the intranasal formulation does not effectively increase absorption of the peptide.

Example 7: Intranasal Study Dosing in Rats

[00195] Sprague-Dawley male and female rats were administered ZT-034 at 80 and 200 μg/kg with either 0.06% or 0.125% DDM (both with lOmM acetate buffer, pH 5)

intranasally, once daily into alternate nostrils for at 28 days as shown in Table 7-1 below:

[00196] The dose volume was 20 μί/day for all dose groups and all solutions contained 10 mM acetate pH 5). Animals were dosed and bled on Days 1 and 28 for toxicokinetic analysis.

[00197] Based on toxicokinetic data analysis (Table 7.2), male and female rats were exposed to quantifiable concentrations of ZT-034 on Days 1 and 28 after administrations of 80 and 200 μg/kg/day ZT-034 with 0.06% or 0.125% DDM. In general, the toxicokinetics of ZT-034 appeared to be non-linear with dose in either 0.06 or 0.125% DDM. In the study period of 28 days, it appears that with one exception, some accumulation of test article occurs in rats of both sexes after repeated administration of ZT-034. There were no consistent patterns for gender differences. Systemic exposures to ZT-034 in 0.06%> DDM were generally greater than or approximately equal (within±30%) to the systemic exposure to ZT- 034 in 0.125% DDM. [00198] Plasma exposures up to 30-fold higher than the human therapeutic plasma levels were achieved at both 80 and 200 μg/kg doses as demonstrated by the C _max and AUC values. The T _max in the range of 5-15 min and the T ₂ in the range of 8-17 minutes are consistent with rapid absorption across the nasal epithelium and rapid clearance respectively following IN delivery of ZT-034 with DDM.

[00199] Table 7-2: Toxicokinetic Evaluation of ZT-034 Following IN Administration to Rats in the Presence of DDM Concentrations.

*ϊ? value ¾1 5

[00200] It is understood that modifications which do not substantially affect the activity of the various embodiments described herein are also provided within the definition of the disclosure provided herein. Accordingly, the disclosed examples are intended to illustrate but not limit the present disclosure. While the methods and compositions described herein have been described in detail and with reference to specific embodiments thereof, it will be apparent to one of ordinary skill in the art that various changes and modifications can be made without departing from the spirit and scope thereof. Thus, for example, those skilled in the art will recognize, or be able to ascertain, using no more than routine experimentation, numerous equivalents to the specific substances and procedures described herein. Such equivalents are considered to be within the scope of this disclosure, and are covered by the following claims.