WO2014153424A1 - Reducing diabetes in patients receiving hmg-coa reductase inhibitors (statins) - Google Patents

Abstract

The invention provides compositions, methods and uses for reducing insulin resistance or risk of diabetes in a subject taking a statin. The invention also provides compositions, methods and uses for reducing or decreasing increases in blood glucose caused by a statin in a subject taking a statin. The invention further provides compositions, methods and uses for reducing insulin resistance or risk of diabetes in a subject taking statins, and for treating diabetes in a subject taking statins. The invention moreover provides compositions, including an adenosine receptor antagonist, and/or an A2B receptor antagonist, and/or a CD73 antagonist and a statin.

Description

REDUCING DIABETES IN PATIENTS RECEIVING HMG-COA REDUCTASE

INHIBITORS (STATINS)

RELATED APPLICATION INFORMATION

[0001] This application claims priority to application serial no. 61/803, 188, filed March 19, 2013, which application is expressly incorporated herein by reference in its entirety.

GOVERNMENT SUPPORT

This invention received government support from the National Institutes of Health R01 HL37942. The government has certain rights in the invention.

INTRODUCTION

[0003] The conversion of 3-Hydroxy-3-methylglutaryl-CoA (HMG-CoA) to mevalonic acid by HMG-CoA reductase is required for cholesterol biosynthesis. HMG-CoA reductase inhibitors, also known as statins have been enormously successful drugs that are widely used in the United States and throughout the world to treat atherosclerosis and related cardiovascular diseases. The statins include atorvastatin (Lipitor™), which is the best-selling pharmaceutical in history, fluvastatin (Lescol™), lovastatin (Mevacor™), rosuvastatin (Crestor™) and simvastatin (Zocor™). Statins are primarily used for the treatment or prevention of cardiovascular diseases, principally atherosclerosis. Statins cause few side effects; they can produce muscle damage, but this side effect is rare.

[0004] However, in clinical studies, statins have been shown to produce a small but significant increase in the risk of developing diabetes (Sattar et al., 2010). The degree of risk is related to drug dose and patient age (Preiss et al., 2011). Compared to earlier studies, the increased risk of diabetes with statin use was found to be greater in a recent study of postmenopausal women (Culver et al., 2012). In another recent study, statin intake was associated with decreased insulin sensitivity during surgery (Sato et al., 2012). The mechanism by which statins increase diabetes risk is unknown. It has been argued that the beneficial effects of lowering lipids with statin use outweigh the adverse effect of increasing the risk of diabetes. Nevertheless the diabetes side effect of statin use does represent an undesirable complication. It is also significant that many of the patients who are most at risk for atherosclerosis, also have a high risk of developing diabetes, i.e. those with a high body mass index (BMI) and the elderly. SUMMARY

[0005] The present inventors have surprisingly discovered that insulin resistance or risk of diabetes in a subject taking statins can be reduced by either by blocking statin-induced adenosine production by inhibiting a 5 '-nucleosidase, or by blocking an adenosine receptor with a receptor antagonist. Without being limited to any particular theory, increased adenosine production can occur as a result of CD73 activation, which could stimulate A_2B receptors to increase blood levels of adenosine. Accordingly, one embodiment of the invention is to use antagonists of CD73 to reduce, decrease or inhibit statin-induced adenosine production. A second embodiment of the invention is to use antagonists of adenosine A_2B receptors to reduce, decrease or inhibit adenosine A_2B receptor signalling. Both embodiments reduce, decrease, inhibit or suppress statin-induced increases in glucose, insulin resistance and the statin-induced increased risk of diabetes. Such embodiments optionally also do not result in substantially decreasing or inhibiting the beneficial lipid-lowering effects, cholesterol lowering activity and/or cardioprotective effects of statins.

[0006] In accordance with the invention, provided are methods of reducing insulin resistance and methods of reducing risk of diabetes in a subject taking a statin. In one embodiment, a method includes administration of an adenosine receptor antagonist to the subject to reduce insulin resistance and/or reduce risk of diabetes. In another embodiment, a method includes administration of a CD73 antagonist (e.g., 5 '-nucleosidase inhibitor) to the subject to reduce insulin resistance and/or reduce risk of diabetes.

[0007] In accordance with the invention, also provided are methods of reducing or decreasing increases in blood glucose caused by a statin in a subject taking a statin. In one embodiment, a method includes administering an amount of an adenosine receptor antagonist to the subject to reduce or decrease increases in blood glucose caused by the statin. In another embodiment, a method includes administration of a CD73 antagonist (e.g., 5 '-nucleosidase inhibitor) to the subject to reduce or decrease increases in blood glucose caused by the statin.

[0008] The invention additionally provides methods of treating diabetes in a subject taking a statin. In one embodiment, a method includes administration of an adenosine receptor antagonist to the subject. In another embodiment, a method includes administration of a CD73 antagonist (e.g., 5 '-nucleosidase inhibitor) to the subject.

[0009] The invention further provides methods of preventing diabetes in a subject taking a statin. In one embodiment, a method includes administration of an adenosine receptor antagonist to the subject. In another embodiment, a method includes administration of a CD73 antagonist (e.g., 5 '-nucleosidase inhibitor) to the subject. [0010] Methods of the invention include decreasing, reducing, inhibiting, suppressing, limiting or controlling in the subject elevated blood glucose levels. In additional aspects, methods of the invention increase, stimulate, enhance, promote, induce or activate in the subject insulin sensitivity.

[0011] In further aspects, a method does not substantially decrease statin activity. In additional particular aspects, activity of a statin includes but is not limited to lipid-lowering effects or cholesterol lowering activity of a statin, or cardioprotective effects of a statin.

Accordingly, in certain embodiments, methods of the invention do not substantially impair (e.g., by more than 50%) lipid-lowering effects and/or cholesterol lowering activity and/or cardioprotective effects of a statin

[0012] Statins include but are not limited to atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rousuvastatin and simvastatin. Accordingly a subject taking a statin may be taking one or more of the foregoing statins, and/or a method of the invention does not substantially decrease activity of a statin such as one or more of the foregoing statins.

[0013] In accordance with the invention, further provided are compositions that include an adenosine receptor antagonist. In one embodiment, a composition includes an adenosine receptor antagonist and a statin. Examples of statins include but are not limited to atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rousuvastatin and simvastatin. In certain aspects, the composition may further include a CD73 antagonist (e.g., of 5 '-nucleotidase activity).

[0014] In accordance with the invention, further provided are compositions that include a CD73 antagonist (e.g., of 5 '-nucleotidase activity). In one embodiment, a composition includes a CD73 antagonist (e.g., of 5 '-nucleotidase activity) and a statin. Examples of statins include but are not limited to atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rousuvastatin and simvastatin. In certain aspects, the composition may further include an adenosine receptor antagonist.

[0015] In accordance with the invention, antagonists include but are not limited to antagonists of the adenosine receptor, A_2B receptor. In particular embodiments, antagonists of adenosine receptor and/or A2B receptor include: Compound 38(OSIP-339,39, (N-[2-[[2-phenyl- 6-[4-(3-phenylpropyl)piperazine-l-carbonyl]-7H-pyrrolo[2,3-d]pyrimidin-4- yl]amino]ethyl]acetamide); ATL-801 (N-[5-(l-cyclopropyl-2,6-dioxo-3-propyl-7H-purin-8- yl)pyridin-2-yl]-N-ethylpyridine-3-carboxamide); CVT-6883 (3-ethyl-l-propyl-8-[l-[[3- (trifluoromethyl)phenyl]methyl]pyrazol-4-yl]-7H-purine-2,6-dione); MRS-1706 (N-(4- acetylphenyl)-2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8-yl)phenoxy]acetamide); MRS- 1754 (N- (4-cyanophenyl)-2-[4-(2,6-dioxo-l ,3-dipropyl-7H-purin-8-yl)phenoxy]acetamide); PSB-603 (8- [4-[4-(4-chlorophenyl)piperazin-l-yl]sulfonylphenyl]-l-propyl-3,7-dihydropurine-2,6-dione); PSB -0788 (8- [4- [4- [(4-chlorophenyl)methyl]piperazin- 1 -yl] sulfonylphenyl] - 1 -propyl-3 ,7- dihydropurine-2,6-dione); PSB-1115 (4-(2,6-dioxo-l-propyl-3,7-dihydropurin-8- yl)benzenesulfonic acid); GS 6201 (3-ethyl-l-propyl-8-[l-[[3-

(trifluoromethyl)phenyl]methyl]pyrazol-4-yl]-7H-purine-2,6-dione); CGS 15943 (9-chloro-2- (furan-2-yl)-[l,2,4]triazolo[l,5-c]quinazolin-5-amine); Caffeine Al and A2B antagonist (1,3,7- trimethylpurine-2,6-dione); Theophylline Adenosine receptor antagonist (l,3-dimethyl-7H- purine-2,6-dione); XAC Adenosine receptor antagonist (N-(2-aminoethyl)-2-[4-(2,6-dioxo-l,3- dipropyl-7H-purin-8-yl)phenoxy]acetamide); CGH 2466 dihydrochloride (4-(3,4- dichlorophenyl)-5-pyridin-4-yl-l,3-thiazol-2-amine;dihydrochloride); XCC Adenosine receptor antagonist (2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8-yl)phenoxy]acetic acid); LAS101057 (N- [5-(3-fluoropyridin-4-yl)-6-pyridin-3-ylpyrazin-2-yl]cyclopropanecarboxamide); MRE-2029- F20 N-(l,3-Benzodioxol-5-yl)-2-{ [5-(2,6-dioxo-l,3-dipropyl-2,3,6,7-tetrahydro-lH-purin-8-yl)- 1 -methyl- 1 H-pyrazol-3 -yl]oxy } acetamide) ; ATL-802 (N- [5 -( 1 -cyclopropyl-2,6-dioxo-3 -propyl - 7H-purin-8-yl)pyridin-2-yl]-N-methyl-6-(trifluoromethyl)pyridine-3-carboxamide);

Theophylline (l,3-dimethyl-7H-purine-2,6-dione); Enprofylline (3-propyl-7H-purine-2,6-dione); and LAS38096 (4-(furan-2-yl)-N-pyridin-3-yl-5-pyrimidin-4-ylpyrimidin-2-amine). In further particular embodiments, antagonists of adenosine receptor and/or A2B receptor include an antibody or antibody fragment that binds to the adenosine receptor or the A2B receptor.

[0016] In accordance with the invention, antagonists include but are not limited to antagonists of CD73 (e.g., of 5 '-nucleotidase activity). In particular embodiments, antagonists of CD73 include: an inhibitor or suppressor of CD73 5'-ecto nucleotidase activity; alpha, beta- methylene adenosine-5' -diphosphate (APCP); or an antibody or antibody fragment that binds to CD73. In particular aspects, a CD73 antagonist includes an antibody having the CDRs of the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9): sc-32299) and/or the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9): sc-32299).

DESCRIPTION OF DRAWINGS

[0017] Figure 1. Shows a flowchart of intervention points and strategies for inhibiting or reducing statin induced increases in blood glucose, risk of diabetes, and insulin resistance, based upon the invention. [0018] Figure 2. Shows structures of exemplary non-limiting adenosine A2B receptor antagonists.

[0019] Figures 3A-3C. Shows effects of adenosine signalling on blood glucose levels. A) Mice were injected IP with the selective A2BR agonist, LUF-6210 (aka Bay-60-6583) with or without prior injection for 30 min with the selective A2BR antagonist, ATL-801. B) Mice were injected IP with the non-selective adenosine receptor agonist, LUF-6210 (aka Bay-60-6583) with or without prior injection for 30 min with the selective A2BR antagonist, ATL-801. C) C57BL/6 wild type mice (B6 WT) or mice lacking the A2BR were injected intravenously with AMP. All mice were fasted for 4 hours prior to injection of agonists or AMP at time 0. Blood glucose was measured at the indicated times, N=4.

[0020] Figures 4A-4C. Shows effects of AMP and Crestor™ on glucose mobilization in mice. Thirty-five mice were fasted for 4 hours and anesthetized with isofluorane prior to injection of AMP or phosphate buffered saline (PBS) vehicle. Blood glucose was measured using a one-touch Ultra meter at the indicated times. Data represent the means + SEM, N=5. (A) C57BL/6 mice were injected with the indicated concentrations of AMP or PBS. AMP produced a dose-depended increase in blood glucose. A small increase in blood glucose in response to PBS injection is attributed to catecholamines released from the stress of IV injection. (B,C) Mice were treated orally with an aqueous solution of Crestor™ ^g in 100 μΐ) or water once daily for 8 or 12 days. They were then injected IV with 15 or 30 mg/kg AMP. AMP injection produced a biphasic increase in blood glucose attributed to a rapid stress response followed by a longer response mediated by CD73-dependent adenosine production from AMP. Crestor™ treatment enhanced AMP-mediated glucose mobilization.

[0021] Figure 5. Shows CD73 expression on the surface of live/TCRbeta/CD4+ cells from mouse splenocytes incubated for 16h at 37 degrees, by FACS. Mouse splenocytes were incubated overnight with various concentrations of Atorvastatin (Lipitor™) and noted a dose dependent-increase in the amount of CD73 on the surface of CD4+ T cells. This is consistent with the idea that statins cause a translocation of CD73 to the cell surface.

DETAILED DESCRIPTION

[0022] It was previously not known how statins increase insulin resistance or the risk of diabetes (type 2). Hence there was no treatment for this complication. Since risk factors for cardiovascular disease largely overlap with risk factors for the type 2 diabetes, i.e. high body mass index (BMI) and western diet, the statin-induced increased insulin resistance risk and diabetes risk is a significant concern, particularly in populations at high risk for development of diabetes.

[0023] Disclosed herein are studies indicating the effect of statins to increase insulin resistance and the risk of type 2 diabetes. As disclosed herein, statins activate the enzyme CD73, an ecto-5' -nucleotidase that converts AMP to adenosine in the extracellular space (Meijer et al., 2009; Sanada et al., 2004). Without being limited to any particular theory, it may be that increased adenosine production causes insulin-resistance mediated by activation of adenosine A₂B receptors.

[0024] As disclosed herein, activation of adenosine receptors (e.g., A₂B) enhances plasma glucose in mice, an effect that opposes insulin action. Namely, injection of a selective A₂B agonist, LUF 6210 (aka Bay 60-6583), into mice causes a rapid increase in blood glucose that is absent in mice that lack the A₂BR. Further, as also disclosed herein statin-induced insulin resistance and the statin-induced increased risk of diabetes can be reduced or decreased either by blocking statin-induced adenosine production by 1) inhibiting CD73; 2) blocking the adenosine A₂B receptor with a receptor antagonist, or 3) a combination of A₂B receptor and CD73 antagonists. This pathway and intervention points are summarized in Figure 1.

[0025] In addition, the increased glucose, insulin resistance, glucose intolerance and risk of diabetes as a consequence of statin use can be reduced, decreased, inhibited or suppressed by coadministration of compounds that antagonize CD-73 (such as alpha,beta-methyleneadenosine-5'- diphosphate or antibodies that block the activity of this enzyme), compounds that non- selectively block adenosine receptors and are known to be safe (such as caffeine and

theophylline), compounds that selectively block the A₂B receptor (Fig. 2) or antibodies that block the adenosine receptor. Since statins are the most prescribed drugs in the world, such novel combinations have a very large use potential.

[0026] In certain embodiments, the invention provides an antagonist that can be

administered to a subject or a patient, or administered with a statin to a subject or a patient, to decrease, reduce or inhibit (counteract) undesirable statin activity. In various embodiments, an antagonist will decrease, reduce or inhibit (counteract) an 1) increase in glucose levels; and/or 2) insulin resistance: and/or 3) the risk of developing diabetes (e.g., type 2) caused by a statin. In particular aspects, an antagonist will not substantially inhibit or impair the beneficial lipid- lowering effects or cholesterol lowering activity of a statin.

[0027] An "antagonist" is capable of decreasing, reducing, suppressing or inhibiting one or more functions or activities of a target molecule, such as signal transduction or enzyme activity of an adenosine receptor (e.g., A2B receptor), or CD73 (an ecto-5 '-nucleotidase that converts AMP to adenosine). Antagonists may interfere with the function or activity of an adenosine receptor (e.g., A2B receptor), or CD73. The antagonist may but need not completely block the activity or function an adenosine receptor (e.g., A2B receptor), or CD73.

[0028] Also provided are methods of reducing or decreasing insulin resistance and risk of diabetes (e.g., type 2) and methods of treating diabetes (e.g., type 2) in subjects taking statins. In one embodiment, a method includes administration of an adenosine receptor antagonist to the subject or administration of a 5 ' -nucleotidase (CD73) antagonist to the subject in an amount to reduce or decrease insulin resistance, reduce or decrease risk of diabetes (e.g., type 2) and/or treat diabetes (e.g., type 2).

[0029] The term "hyperglycemic" or "hyperglycemia," when used in reference to a condition means a transient or chronic abnormally high level of glucose present in the blood of a subject. The condition can be caused by a delay in glucose metabolism or absorption such that the subject exhibits glucose intolerance or a state of elevated glucose not typically found in normal subjects (e.g. , in glucose-intolerant subdiabetic subjects at risk of developing diabetes, or in diabetic subjects).

[0030] Fasting plasma glucose (FPG) levels for normoglycemia are less than about 1 10 mg/dl, for impaired glucose metabolism, between about 1 10 and 126 mg/dl, and for diabetics greater than about 126 mg/dl. Accordingly, invention methods and uses include, decreasing, reducing or suppressing statin induced increases of glucose such that glucose levels approach (e.g., between about 110 and 126 mg/dl) or are within normalglycemia, e.g., less than about 110 mg/dl.

[0031] Thus, in various embodiments, methods and uses of decreasing glucose, improving glucose tolerance, decreasing risk of insulin resistance, and treating a hyperglycemic condition (e.g. , diabetes) are provided. Additionally provided are methods and uses for treating a physiological disorder associated with or resulting from a hyperglycemic condition. Such disorders include, for example, diabetic neuropathy (autonomic), nephropathy (kidney damage), skin infections and other cutaneous disorders, slow or delayed healing of injuries or wounds (e.g. , that lead to diabetic carbuncles), eye damage (retinopathy, cataracts) which can lead to blindness, diabetic foot and accelerated periodontitis. Such disorders also include increased risk of developing coronary heart disease, stroke, peripheral vascular disease, dyslipidemia, hypertension and obesity.

[0032] In accordance with certain embodiments of the invention, adenosine receptor (e.g., A2B receptor) antagonists may be used to reduce, decrease, inhibit or suppress statin-induced increases in glucose, insulin resistance, and/or risk of diabetes. Non-limiting structures of adenosine receptor (e.g., A2B receptor) antagonists are described in US Patent Nos.: 8,324,224, 8,188,099, 7,947,692, 7,504,421, 6,117,878, and US patent application Nos., 20100273757, 20070270433, 20070225335. In additional embodiments, clinically approved non-selective adenosine receptor antagonists, such as theophylline and caffeine, also could be used for this purpose. Some example antagonist structures are shown in Figure 2.

[0033] In accordance with certain embodiments of the invention, CD73 antagonists may be used to reduce, decrease, inhibit or suppress statin-induced increases in glucose, insulin resistance, and/or risk of diabetes. Non- limiting examples of CD73 antagonists (e.g., an inhibitor or suppressor of CD73 5'-ecto nucleotidase activity) include: alpha, beta- methylene adenosine- 5' -diphosphate (APCP).

[0034] Additional examples of antagonists include antibodies and fragments of antibodies that bind to a target. In particular embodiments, antibodies and fragments of antibodies bind to adenosine receptor (e.g., A2B receptor) and/or CD73.

[0035] The term "antibody" refers to a protein that binds to another molecule (antigen) via heavy and light chain variable domains, denoted VH and VL, respectively. "Antibody" refers to any polyclonal or monoclonal immunoglobulin molecule, or mixtures thereof, such as IgM, IgG, IgA, IgE, IgD. Antibodies belong to any antibody class or subclass. Exemplary subclasses for IgG are IgGi, IgG₂, IgG₃ and IgG₄.

[0036] Antibodies include polyclonal and monoclonal antibodies and fragments thereof. The term "monoclonal," when used in reference to an antibody, refers to an antibody that is based upon, obtained from or derived from a single clone, including any eukaryotic, prokaryotic, or phage clone. A "monoclonal" antibody is therefore defined herein structurally, and not the method by which it is produced.

[0037] Antibodies include antibodies having kappa or lambda light chain sequences, either full length as in naturally occurring antibodies, mixtures thereof (i.e., fusions of kappa and lambda chain sequences), and subsequences/fragments thereof. Naturally occurring antibody molecules contain two kappa and two lambda light chains. The primary difference between kappa and lambda light chains is in the sequences of the constant region.

[0038] An antibody that includes or consists of a Heavy (H) chain and/or Light (L) chain or fragment of a Heavy (H) chain or Light (L) chain can include a single H or L chain or a single H or L chain fragment, or a plurality (2, 3, 4 or more) of Heavy (H) chains and/or Light (L) chains, or a plurality of fragments of Heavy (H) chains and/or Light (L) chains. An antibody or fragment thereof that includes a Heavy (H) chain and/or Light (L) chain of an antibody or fragment can but is not required to include 2 Heavy (H) chains and 2 Light (L) chains. An antibody or fragment thereof may be an oligomeric (higher order or valent) forms, such as a trimer, tetramer, pentamer, hexamer, hep tamer, and so forth, with other antibodies, fragments thereof, Heavy (H) chain, Light (L) chain, or polypeptides distinct from an antibody Heavy (H) or Light (L) chain.

[0039] The invention provides compositions including an adenosine receptor antagonist (e.g., A2B receptor antagonist), and/or a CD73 antagonist. Such compositions include sterile compositions, for example, for methods and uses of treatment of a subject as set forth herein.

[0040] The invention further provides pharmaceutical compositions. A pharmaceutical composition refers to "pharmaceutically acceptable" and "physiologically acceptable" carriers, diluents or excipients. As used herein, the term "pharmaceutically acceptable" and

"physiologically acceptable," when referring to carriers, diluents or excipients includes solvents (aqueous or non-aqueous), detergents, solutions, emulsions, dispersion media, coatings, isotonic and absorption promoting or delaying agents, compatible with pharmaceutical administration and with the other components of the formulation. Such formulations can be contained in a tablet (coated or uncoated), capsule (hard or soft), microbead, emulsion, powder, granule, crystal, suspension, syrup or elixir.

[0041] Pharmaceutical compositions can be formulated to be compatible with a particular route of administration or use. Compositions for parenteral, intradermal, or subcutaneous administration can include a sterile diluent, such as water, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents. The preparation may contain one or more preservatives to prevent microorganism growth (e.g., antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as EDTA; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose).

[0042] Pharmaceutical compositions for injection include sterile (e.g., aqueous) solutions (where water soluble) or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersion. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor EL™ (BASF, Parsippany, NJ) or phosphate buffered saline (PBS). The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (e.g., glycerol, propylene glycol, and polyetheylene glycol), and suitable mixtures thereof. Fluidity can be maintained, for example, by the use of a coating such as lecithin, or by the use of surfactants. Antibacterial and antifungal agents include, for example, parabens, chlorobutanol, phenol, ascorbic acid and thimerosal. Including an agent that delays absorption, for example, aluminium monostearate and gelatin can prolong absorption of injectable compositions. Polysorbate 20 and polysorbate 80 can be added into the formulation mixture, for example, up to 1%. Other non- limiting additives include histidine HC1, α,α- treahlose dehydrate.

[0043] Additional pharmaceutical formulations and delivery systems are known to the skilled artisan and are applicable in the methods of the invention (see, e.g., Remington's Pharmaceutical Sciences (1990) 18th ed., Mack Publishing Co., Easton, PA; The Merck Index (1996) 12th ed., Merck Publishing Group, Whitehouse, NJ; Pharmaceutical Principles of Solid Dosage Forms, Technonic Publishing Co., Inc., Lancaster, Pa., (1993); and Poznansky, et al., Drug Delivery Systems, R. L. Juliano, ed., Oxford, N.Y. (1980), pp. 253-315).

[0044] The term "isolated," when used as a modifier of a composition, means that the composition is made by the hand of man or is separated, completely or at least in part, from their naturally occurring in vivo environment. Generally, an isolated composition is substantially free of one or more materials with which it normally associates with in nature. The term "isolated" does not exclude alternative physical forms, such as multimers/oligomers, variants,

modifications or derivatized forms, or forms produced by the hand of man. The term "isolated" also does not exclude forms (e.g., pharmaceutical formulations and combination compositions) in which there are combinations, any one of which is produced by the hand of man.

[0045] The term "bind," or "binding," when used in reference to an antagonist means that the antagonist binds to all or a part of a target, such as an adenosine receptor or A2B receptor, or an CD73. The binding of an antagonist to a target may be specific or non-specific. Specific binding refers to binding to a target protein. Binding to a non-target protein that does not significantly interfere with binding to a target protein is also considered specific.

[0046] The term "contacting" means direct or indirect interaction between two or more entities. A particular example of direct interaction is binding, e.g., an antagonist binds to adenosine receptor or A2B receptor (an adenosine receptor or A2B receptor antagonist), or an antagonist binds to CD73 (CD73 antagonist). Contacting as used herein includes in in vitro, ex vivo, and in vivo. Contacting in vivo can be referred to as administering, administration, to administer, delivering or delivery to a subject or patient.

[0047] In methods and uses of the invention, an adenosine receptor antagonist (e.g., A2B receptor antagonist), CD73 antagonist, compositions thereof, including combinations (e.g., with other drugs or agents such as statins), and pharmaceutical formulations as set forth herein can be administered prior to, substantially contemporaneously with or following administration of a statin. Thus, methods and uses of the invention may be practiced prior to (i.e. prophylaxis), concurrently with or after a subject takes a statin. Thus in certain embodiments, the present invention provides administering an adenosine receptor antagonist (e.g., A2B receptor antagonist), CD73 antagonist, a composition thereof, including combinations, and

pharmaceutical formulations set forth herein prior to, concurrently with or immediately following administration of a statin, or development of an adverse symptom caused by a statin.

[0048] The invention provides combination compositions, methods and uses, such as an adenosine receptor antagonist (e.g., A2B receptor antagonist), CD73 antagonist, compositions thereof, including combinations, methods and uses and a second drug or agent (e.g., such as a statin). An adenosine receptor antagonist (e.g., A2B receptor antagonist), CD73 antagonist, or composition thereof can be formulated and/or administered in combination with a second agent, drug or treatment. Accordingly, an adenosine receptor antagonist (e.g., A2B receptor antagonist), CD73 antagonist, composition thereof, combination, methods and uses as set forth herein can be formulated as a combination (e.g., with other drugs or agents such as statins), and/or administered in combination to a subject.

[0049] Methods and uses of the invention include administration of a sufficient or effective amount to a subject. An "effective amount" or "sufficient amount" refers to an amount that provides, in single or multiple doses, alone or in combination, with one or more other compositions (therapeutic agents such as a drug), treatments, protocols, or therapeutic regimens agents, a detectable response of any duration of time (long or short term), an expected or desired outcome in or a benefit to a subject of any measurable or detectable degree or for any duration of time (e.g., for minutes, hours, days, months, years, or cured).

[0050] The "effective amount" or "sufficient amount" for treatment (e.g., to ameliorate or to provide a therapeutic benefit or improvement) typically provides a response in a subject, for example, a reduction or decrease or inhibition of an adverse effect of a statin, such as an increase in blood glucose, to a measurable extent. Stabilizing, limiting or controlling an adverse effect of a statin is also considered a satisfactory outcome.

[0051] As is typical for treatment methods and uses, some subjects will exhibit a greater response, or less or no response to a given treatment method or use. An effective amount or a sufficient amount therefore need not be effective in each and every subject treated,

prophylactically or therapeutically, nor a majority of treated subjects in a given group or population. An effective amount or a sufficient amount means effectiveness or sufficiency in a particular subject, not a group or the general population. Appropriate amounts will depend upon, among other things, the individual subject (e.g., the bioavailability within the subject, gender, age, etc.). [0052] Accordingly, an effective amount or a sufficient amount can but need not be provided in a single administration, may require multiple administrations, and, can but need not be, administered alone or in combination with another composition (e.g., agent), treatment, protocol or therapeutic regimen. In addition, an effective amount or a sufficient amount need not be effective or sufficient if given in single or multiple doses without a second composition (e.g., another drug or agent), treatment, protocol or therapeutic regimen, since additional doses, amounts or duration above and beyond such doses, or additional compositions (e.g., drugs or agents), treatments, protocols or therapeutic regimens may be included in order to be considered effective or sufficient in a given subject.

[0053] The term "ameliorate" refers to a detectable or measurable improvement in a subject's condition or a response indicative of a beneficial effect. A detectable or measurable improvement would include a decrease, reduction, inhibition, suppression, in the amount or the duration of increased glucose caused by a statin in a subject. A detectable or measurable improvement would also include a limit or control on increased glucose caused by a statin in a subject. Since increased glucose can lead to insulin resistance and diabetes, such a decrease, reduction, inhibition, suppression, limit or control in the amount or the duration of increased glucose caused by a statin in a subject also is considered a beneficial effect. Such effects are considered a successful treatment outcome as they lead to a "therapeutic effect," or "benefit" in a subject.

[0054] A therapeutic benefit or improvement therefore need not be complete ablation of any one, most or all adverse symptoms caused by a statin in a subject. Thus, a satisfactory endpoint may be achieved when there is an incremental response, or a partial response to decrease, reduce, inhibit, suppress, limit or control an adverse symptom caused by a statin in a subject.

[0055] Effectiveness of a method or use, such as a treatment that provides a therapeutic benefit or improvement, such as an undesirable or aberrant side effect of a statin, can be ascertained by various methods. Such methods include, for example, a reduction of blood glucose levels, and inhibition of increases in glucose caused by a statin, reduction of risk of diabetes, reduction in insulin resistance.

[0056] The term "subject" or "patient" refers to animals, typically mammalian animals, such as humans, non-human primates (e.g., apes, gibbons, chimpanzees, orangutans, macaques), domestic animals (e.g., dogs and cats), farm animals (e.g., horses, cows, goats, sheep, pigs) and experimental animals (e.g., mouse, rat, rabbit, guinea pig). Subjects include animal disease models, for example, animal models of statin-induced increases of glucose, insulin resistance or diabetes, aberrant glucose homeostasis, diabetes, insulin resistance or intolerance, atherosclerosis or obesity.

[0057] Subjects appropriate for treatment include those taking a statin. Subjects appropriate for treatment include those that are candidates for statin treatment but may have other medical conditions where statin treatment could worsen or exacerbate the condition. For example a subject with diabetes, insulin resistance or aberrant or abnormal glucose homeostasis, may not be an ideal candidate for statin treatment in view of the risk that a statin will worsen or exacerbate diabetes, insulin resistance or aberrant or abnormal glucose homeostasis. Thus, by way of example a diabetic subject may not be taking a statin but can be treated in accordance with the invention method and then be able to take a statin with a reduced risk that the statin will worsen or exacerbate their diabetes. Accordingly, subjects also include those that would benefit from a statin or are in need of treatment with a statin, but may not be currently taking a statin.

[0058] Adenosine receptor antagonists, (e.g., A2B receptor antagonists) and CD73 antagonists, and compositions thereof, including combinations (e.g., with other drugs or agents such as statins), methods and uses in accordance with the invention, and pharmaceutical formulations can be packaged in dosage unit form for ease of administration and uniformity of dosage. "Dosage unit form" as used herein refers to physically discrete units suited as unitary dosages treatment; each unit contains a quantity of the composition in association with the carrier, excipient, diluent, or vehicle calculated to produce the desired treatment or therapeutic (e.g., beneficial) effect. The unit dosage forms can be varied according to factors including, but not necessarily limited to, the particular composition employed, the disorder or disease treated, the effect to be achieved, and the subject to be treated.

[0059] Non-limiting exemplary unit doses range from about 25-250, 250-500, 500-1,000, 1,000-2,500, 2,500-5,000, 5,000-25,000, or 5,000-50,000 pg; from about 50-500, 500-5,000, 5,000-25,000 or 25,000-50,000 ng; from about 50-500, 500-5,000, 5,000-25,000 or 25,000- 50,000 μg; from about 25-250, 250-500, 500-1,000, 1,000-2,500, 2,500-5,000, 5,000-25,000, or 5,000-50,000 mg; and from about 1-5, 5-10, 10-25, 25-50, 50-100, 100-250, 250-500, 500- 1,000, 1,000-2,500, or 2,500-5,000 grams.

[0060] Adenosine receptor antagonists (e.g., A2B receptor antagonists) and CD73 antagonists, and compositions thereof, methods and uses in accordance with the invention, including combinations, and pharmaceutical formulations may be contacted or provided in vitro, ex vivo or administered or delivered in vivo to a subject or patient in various doses and amounts, and frequencies. For example, an adenosine receptor antagonist (e.g., A2B receptor antagonists) and/or CD73 antagonist, composition thereof, including combinations (with other drugs or agents such as statins), and pharmaceutical formulations can be can be administered or delivered to provide the intended effect, as a single or as multiple dosages, for example, in an effective or sufficient amount. Exemplary doses range from about 25-250, 250-500, 500-1,000, 1,000- 2,500, 2,500-5,000, 5,000-25,000, or 5,000-50,000 pg/kg; from about 50-500, 500-5,000, 5,000- 25,000 or 25,000-50,000 ng/kg; from about 50-500, 500-5,000, 5,000-25,000 or 25,000-50,000 μg/kg; and from about 25-250, 250-500, 500-1,000, 1,000-2,500, 2,500-5,000, 5,000-25,000, or 5,000-50,000 mg/kg, on consecutive days, alternating days or intermittently.

[0061] Single or multiple (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more times) administrations or doses can be administered on the same or consecutive days, alternating days or intermittently. For example, an adenosine receptor antagonist (e.g. , A2B receptor antagonists) and/or CD73 antagonist, composition thereof, combination or pharmaceutical formulation can be administered one, two, three, four or more times daily, on alternating days, bi-weekly, weekly, monthly, bimonthly, or annually. A fusion polypeptide or composition thereof can be administered for any appropriate duration, for example, for period of 1 hour, or less, e.g., 30 minutes or less, 15 minutes or less, 5 minutes or less, or 1 minute, or less.

[0062] An adenosine receptor antagonist (e.g., A2B receptor antagonist) and/or CD73 antagonist, composition thereof, combination or pharmaceutical formulation can be administered to a subject and methods and uses may be practiced prior to, substantially contemporaneously with, or within about 1-60 minutes, hours (e.g., within 1, 2, 3, 4 , 5, 6, 8, 12, 24 hours), or days (1 , 2, 3, 4, 5, 6, 7, 7-14, 14-21 , 21-28, 28-45, 45-60, 60-90, etc.) of administration or ingestion of a statin, or a symptom or onset of statin induced increases in blood glucose, risk of diabetes, insulin resistance, etc.

[0063] An adenosine receptor antagonist (e.g., A2B receptor antagonists) and/or CD73 antagonist, composition thereof, combination or pharmaceutical formulation thereof can be administered and methods and uses may be practiced via systemic, regional or local

administration, by any route. For example, an adenosine receptor antagonist (e.g., A2B receptor antagonists) and/or CD73 antagonist, composition thereof, combination or pharmaceutical formulation may be administered systemically, regionally or locally, via injection, infusion, orally (e.g., ingestion or inhalation), topically, intravenously, intra-arterially, intramuscularly, intraperitoneally, intradermally, subcutaneously, intracavity, intracranially, transdermally (topical), parenterally, e.g. transmucosally or intrarectally (enema) catheter, or optically.

[0064] Further provided are kits including an adenosine receptor antagonist (e.g., A2B receptor antagonist) and/or CD73 antagonist, composition thereof, combination or

pharmaceutical formulation thereof, packaged into suitable packaging material. A kit optionally includes a label or packaging insert including a description of the components or instructions for use in vitro, in vivo, or ex vivo, of the components therein. Exemplary instructions include instructions for a method, treatment protocol or therapeutic regimen.

[0065] A kit can contain a collection of such components, e.g., two or more adenosine receptor antagonists (e.g., A2B receptor antagonists) and/or CD73 antagonist, composition thereof, combination or pharmaceutical formulation alone, or in combination with another therapeutically useful agent or drug, such as a statin. The term "packaging material" refers to a physical structure housing the components of the kit. The packaging material can maintain the components sterilely, and can be made of material commonly used for such purposes {e.g., paper, corrugated fiber, glass, plastic, foil, ampules, vials, tubes, etc.).

[0066] Kits can include labels or inserts. Labels or inserts include "printed matter," e.g., paper or cardboard, or separate or affixed to a component, a kit or packing material (e.g., a box), or attached to an ampule, tube or vial containing a kit component. Labels or inserts can additionally include a computer readable medium, optical disk such as CD- or DVD- ROM/RAM, DVD, MP3, magnetic tape, or an electrical storage media such as RAM and ROM or hybrids of these such as magnetic/optical storage media, FLASH media or memory type cards.

[0067] Labels or inserts can include identifying information of one or more components therein, dose amounts, clinical pharmacology of the active ingredient(s) including mechanism of action, pharmacokinetics (PK) and pharmacodynamics (PD). Labels or inserts can include information identifying manufacturer information, lot numbers, manufacturer location and date.

[0068] Labels or inserts can include information for which a kit component may be used. Labels or inserts can include instructions for the clinician or for a subject for using one or more of the kit components in a method, treatment protocol or therapeutic regimen. Instructions can include dosage amounts, frequency or duration, and instructions for practicing any of the methods, treatment protocols or therapeutic regimes set forth herein. Kits of the invention therefore can additionally include labels or instructions for practicing any of the methods and uses of the invention described herein.

[0069] Labels or inserts can include information on any benefit that a component may provide, such as a prophylactic or therapeutic benefit. Labels or inserts can include information on potential adverse side effects, such as warnings to the subject or clinician regarding situations where it would not be appropriate to use a particular composition. Adverse side effects could also occur when the subject has, will be or is currently taking one or more other medications that may be incompatible with the composition, or the subject has, will be or is currently undergoing another treatment protocol or therapeutic regimen which would be incompatible with the composition and, therefore, instructions could include information regarding such

incompatibilities.

[0070] As disclosed herein, kits can additionally include other components. Each component of the kit can be enclosed within an individual container and all of the various containers can be within a single package. Invention kits can be designed for cold storage.

[0071] 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 invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, suitable methods and materials are described herein.

[0072] All applications, publications, patents and other references, GenBank citations and ATCC citations cited herein are incorporated by reference in their entirety. In case of conflict, the specification, including definitions, will control.

[0073] As used herein, the singular forms "a", "and," and "the" include plural referents unless the context clearly indicates otherwise. Thus, for example, reference to "an adenosine receptor antagonist (e.g., A2B receptor antagonist) and/or CD73 antagonist, etc." includes a plurality of such adenosine receptor antagonists (e.g., A2B receptor antagonists), CD73 antagonists, compositions thereof, combinations and pharmaceutical formulations, and so forth.

[0074] As used herein, numerical values are often presented in a range format throughout this document. The use of a range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention unless the context clearly indicates otherwise. Accordingly, a range expressly includes all possible subranges, all individual numerical values within that range, and all numerical values or numerical ranges including integers within such ranges and fractions of the values or the integers within ranges unless the context clearly indicates otherwise. This construction applies regardless of the breadth of the range and in all contexts throughout this patent document. Thus, for example, reference to a range of 90-100% includes 91-99%, 92-98%, 93-95%, 91-98%, 91-97%, 91-96%, 91-95%, 91-94%, 91-93%, and so forth. Reference to a range of 90-100% also includes 91%, 92%, 93%, 94%, 95%, 95%, 97%, etc., as well as 91.1 %, 91.2%, 91.3%, 91.4%, 91.5%, etc., 92.1%, 92.2%, 92.3%, 92.4%, 92.5%, etc., and so forth. In addition, reference to a range of 1- 5,000 fold includes 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, fold, etc., as well as 1.1, 1.2, 1.3, 1.4, 1.5, fold, etc., 2.1, 2.2, 2.3, 2.4, 2.5, fold, etc., and any numerical range within such a ranges, such as 1-2, 5-10, 10-50, 50-100, 100-500, 100-1000, 500-1000, 1000- 2000, 1000-5000, etc.

[0075] As also used herein a series of ranges are disclosed throughout this document. The use of a series of ranges includes combinations of the upper and lower ranges to provide another range. This construction applies regardless of the breadth of the range and in all contexts throughout this patent document. Thus, for example, reference to a series of ranges such as 5- 10, 10-20, 20-30, 30-40, 40-50, 50-75, 75-100, 100-150, 150-200, etc., includes ranges such as 5-20, 5-30, 5-40, 5-50, 5-75, 5-100, 5-150, 5-200, and 10-30, 10-40, 10-50, 10-75, 10-100, 10- 150, 10-171, and 20-40, 20-50, 20-75, 20-100, 20-150, 20-200, and so forth.

[0076] The invention is generally disclosed herein using affirmative language to describe the numerous embodiments. The invention also specifically includes embodiments in which particular subject matter is excluded, in full or in part, such as substances or materials, method steps and conditions, protocols, procedures, assays or analysis. Thus, even though the invention is generally not expressed herein in terms of what the invention does not include aspects that are not expressly included in the invention are nevertheless disclosed herein.

[0077] A number of embodiments of the invention have been described. Nevertheless, it will be understood that various modifications may be made without departing from the spirit and scope of the invention. Accordingly, the following examples, which include data demonstrating the effects of A2B antagonists on blood glucose, are intended to illustrate but not limit the scope of the invention.

EXAMPLES

Example 1: A2B Antagonist Blocks Increase in Blood Glucose

[0078] In order to test whether adenosine stimulates glucose production by activation of A₂B receptors, the present inventors conducted experiments with recently developed selective A₂BR agonists and antagonists. Fig. 3A shows that injection of mice with a selective A2BR agonist, LUF-6210 (LUF) evokes a rapid increase in blood glucose. The present inventors found that the response is blocked by prior treatment with a selective A₂BR antagonist, ATL-801. Fig. 3B shows that injection of mice with a non-selective adenosine agonist, NECA, stimulates an increase in blood glucose that is mostly blocked by prior injection of ATL801. Without being limited to any particular theory, AMP may be metabolized by CD-73 in vivo to form adenosine and stimulate glucose production. Fig. 3C shows that IV injection of AMP does result in an increase in blood glucose that is completely eliminated in A₂BR ^_ mice. These findings support that the effects of Luf, NECA and AMP/adenosine to enhance glucose production are mediated by activation of the A₂B receptor, and not by simulation of other adenosine receptor subtypes.

[0079] Without being limited to any particular theory, this enzyme may convert AMP to adenosine which activates adenosine A2B receptors to enhance glucose release for tissues and/or to inhibit glucose uptake into tissues. In order to test this hypothesis further the present inventors examined: 1) the effect of AMP injection on glucose mobilization in mice; and 2) the effect of treating mice with rosuvastatin (Crestor™) on AMP-induced glucose mobilization. The results shown in Figure 4 indicate that Crestor™ treatment increases glucose mobilization in response to AMP injection. These findings demonstrate that Crestor™ increases glucose by increasing the activity of CD73.

Example 2: Effects of AMP and Crestor™ on glucose mobilization in mice

[0080] Thirty-five mice were fasted for 4 hours and anesthetized with isofluorane prior to injection of AMP or phosphate buffered saline (PBS) vehicle. Blood glucose was measured using a one-touch Ultra meter at the indicated times. Data represent the means + SEM, N=5. In Figure 4A C57BL/6 mice were injected with the indicated concentrations of AMP or PBS. AMP produced a dose-depended increase in blood glucose. A small increase in blood glucose in response to PBS injection is attributed to catecholamines released from the stress of IV injection. Figures 4B and 4C show mice that were treated orally with an aqueous solution of Crestor™ ^g in 100 μΐ) or water once daily for 8 or 12 days. They were then injected IV with 15 or 30 mg/kg AMP. AMP injection produced a biphasic increase in blood glucose attributed to a rapid stress response followed by a longer response mediated by CD73-dependent adenosine production from AMP. Crestor treatment enhanced AMP-mediated glucose mobilization.

[0081] Methods: Male C57BL/6 or Balb C mice, 8 -10 weeks old, mice treated once daily by oral gavage with Crestor™ (100 μg/mouse) or water once per day for 8 or 12 days. Mice were then fasted for 4 hours before IV injection of AMP. Blood glucose was measured before and after AMP injection as indicated.

Example 3: Effects of Statins on CD73 Expression on Splenocytes

[0082] Splenocytes isolated from mice were incubated overnight with various concentrations of Atorvastatin (Lipitor™) in vitro. A dose-dependent increase in CD73 expression on CD4+ T cells was observed as shown in Figure 5. This demonstrates that statins cause a translocation of CD73 to the cell surface. References

Broady R and Levings MK (2008) Graft- versus-host disease: suppression by statins. Nat Med 14(11): 1155-1156.

Culver AL, Ockene IS, Balasubramanian R, Olendzki BC, Sepavich DM, Wactawski-Wende J, Manson JE, Qiao Y, Liu S, Merriam PA, Rahilly-Tierny C, Thomas F, Berger JS, Ockene JK, Curb JD and Ma Y (2012) Statin use and risk of diabetes mellitus in postmenopausal women in the Women's Health Initiative. Arch Intern Med 172(2): 144-152.

Lappas CM, Liu PC, Linden J, Kang EM and Malech HL (2010) Adenosine A2A receptor activation limits graft-versus-host disease after allogenic hematopoietic stem cell

transplantation. J Leukoc Biol 87(2):345-354.

Ledoux S, Laouari D, Essig M, Runembert I, Trugnan G, Michel JB and Friedlander G (2002) Lovastatin enhances ecto-5 '-nucleotidase activity and cell surface expression in endothelial cells: implication of rho-family GTPases. Circ Res 90(4):420-427.

Matetzky S, Fefer P, Shenkman B, Shechter M, Novikov I, Savion N, Varon D and Hod H (2011) Statins have an early antiplatelet effect in patients with acute myocardial infarction. Platelets 22(2): \m-\ \Q.

Meijer P, Oyen WJ, Dekker D, van den Broek PH, Wouters CW, Boerman OC, Scheffer GJ, Smits P and Rongen GA (2009) Rosuvastatin increases extracellular adenosine formation in humans in vivo: a new perspective on cardiovascular protection. Arterioscler Thromb Vase Biol 29(6):963-968.

Meijer P, Wouters CW, van den Broek PH, de Rooij M, Scheffer GJ, Smits P and Rongen GA (2010) Upregulation of ecto-5 '-nucleotidase by rosuvastatin increases the vasodilator response to ischemia. Hypertension 56(4):722-727.

Preiss D, Seshasai SR, Welsh P, Murphy SA, Ho JE, Waters DD, DeMicco DA, Barter P, Cannon CP, Sabatine MS, Braunwald E, Kastelein JJ, de Lemos JA, Blazing MA, Pedersen TR, Tikkanen MJ, Sattar N and Ray KK (2011) Risk of incident diabetes with intensive-dose compared with moderate-dose statin therapy: a meta-analysis. Jama 305(24):2556-2564.

Sanada S, Asanuma H, Minamino T, Node K, Takashima S, Okuda H, Shinozaki Y, Ogai A, Fujita M, Hirata A, Kim J, Asano Y, Mori H, Tomoike H, Kitamura S, Hori M and Kitakaze M (2004) Optimal windows of statin use for immediate infarct limitation: 5'-nucleotidase as another downstream molecule of phosphatidylinositol 3-kinase. Circulation 110(15):2143-2149.

Sato H, Carvalho G, Sato T, Hatzakorzian R, Lattermann R, Codere-Maruyama T, Matsukawa T and Schricker T (2012) Statin intake is associated with decreased insulin sensitivity during cardiac surgery. Diabetes Care 35(10):2095-2099.

Sattar N, Preiss D, Murray HM, Welsh P, Buckley BM, de Craen AJ, Seshasai SR, McMurray JJ, Freeman DJ, Jukema JW, Macfarlane PW, Packard CJ, Stott DJ, Westendorp RG, Shepherd J, Davis BR, Pressel SL, Marchioli R, Marfisi RM, Maggioni AP, Tavazzi L, Tognoni G, Kjekshus J, Pedersen TR, Cook TJ, Gotto AM, Clearfield MB, Downs JR, Nakamura H, Ohashi Y, Mizuno K, Ray KK and Ford I (2010) Statins and risk of incident diabetes: a collaborative meta-analysis of randomised statin trials. Lancet 375(9716):735-742.

Varani K, Portaluppi F, Merighi S, Ongini E, Belardinelli L and Borea PA (1999) Caffeine alters A2A adenosine receptors and their function in human platelets. Circulation 99(19):2499-2502.

Claims

WHAT IS CLAIMED:

1. A method of reducing insulin resistance or risk of diabetes in a subject taking a statin, the method comprising administration of an adenosine receptor antagonist to the subject.

2. A method of reducing or decreasing increases in blood glucose caused by a statin in a subject taking a statin, comprising administering an amount of an adenosine receptor antagonist to the subject to reduce or decrease increases in blood glucose caused by the statin.

3. The method of claim 1 or claim 2 wherein the adenosine receptor is an A2B receptor.

4. The method of any one of claims 1 to 3, wherein the adenosine receptor antagonist or A2B receptor antagonist comprises: Compound 38(OSIP-339,39, (N-[2-[[2-phenyl-6-[4- (3-phenylpropyl]piperazine-l-carbonyl]-7H-pyrrolo[2,3-d]pyrimidin-4- yl]amino] ethyl] acetamide}; ATL-801 (N-[5-(l-cyclopropyl-2,6-dioxo-3-propyl-7H- purin-8-yl}pyridin-2-yl]-N-ethylpyridine-3-carboxamide}; CVT-6883 (3-ethyl-l-propyl- 8-[l-[[3-(trifluoromethyl}phenyl]methyl]pyrazol-4-yl]-7H-purine-2,6-dione}; MRS- 1706 (N-(4-acetylphenyl]-2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8- yl]phenoxy]acetamide}; MRS-1754 (N-(4-cyanophenyl]-2-[4-(2,6-dioxo-l,3-dipropyl- 7H-purin-8-yl]phenoxy]acetamide}; PSB-603 (8-[4-[4-(4-chlorophenyl]piperazin-l- yl]sulfonylphenyl]-l-propyl-3,7-dihydropurine-2,6-dione}; PSB-0788 (8-[4-[4-[(4- chlorophenyl]methyl]piperazin-l-yl]sulfonylphenyl]-l-propyl-3,7-dihydropurine-2,6- dione}; PSB-1115 (4-(2,6-dioxo-l-propyl-3,7-dihydropurin-8-yl]benzenesulfonic acid}; GS 6201 (3-ethyl-l-propyl-8-[l-[[3-(trifluoromethyl]phenyl] methyl] pyrazol-4-yl] -7H- purine-2,6-dione}; CGS 15943 (9-chloro-2-(furan-2-yl]-[l,2,4]triazolo[l,5- c]quinazolin-5-amine}; Caffeine Al and A2B antagonist (l,3,7-trimethylpurine-2,6- dione}; Theophylline Adenosine receptor antagonist (l,3-dimethyl-7H-purine-2,6- dione}; XAC Adenosine receptor antagonist (N-(2-aminoethyl]-2-[4-(2,6-dioxo-l,3- dipropyl-7H-purin-8-yl]phenoxy]acetamide}; CGH 2466 dihydrochloride (4-(3,4- dichlorophenyl}-5-pyridin-4-yl-l,3-thiazol-2-amine;dihydrochloride}; XCC Adenosine receptor antagonist (2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8-yl]phenoxy]acetic acid}; LAS101057 (N-[5-(3-fluoropyridin-4-yl]-6-pyridin-3-ylpyrazin-2- yl]cyclopropanecarboxamide}; MRE-2029-F20 N-(l,3-Benzodioxol-5-yl}-2-{[5-(2,6- dioxo-l,3-dipropyl-2,3,6,7-tetrahydro-lH-purin-8-yl]-l-methyl-lH-pyrazol-3- yl]oxy}acetamide}; ATL-802 (N-[5-(l-cyclopropyl-2,6-dioxo-3-propyl-7H-purin-8- yl}pyridin-2-yl]-N-methyl-6-(trifluoromethyl}pyridine-3-carboxamide}; Theophylline (l,3-dimethyl-7H-purine-2,6-dione}; Enprofylline (3-propyl-7H-purine-2,6-dione}; or LAS38096 (4-(furan-2-yl}-N-pyridin-3-yl-5-pyrimidin-4-ylpyrimidin-2-amine}.

5. The method of any one of claims 1 to 3, wherein the adenosine receptor antagonist or A2B receptor antagonist comprises an antibody or antibody fragment that binds to the adenosine receptor or the A2B receptor.

6. The method of any one of claims 1 to 5, wherein the method comprises decreasing, reducing, inhibiting, suppressing, limiting or controlling in the subject elevated blood glucose levels.

7. The method of any one of claims 1 to 6, wherein the method comprises increasing, stimulating, enhancing, promoting, inducing or activating in the subject insulin sensitivity.

8. The method of any one of claims 1 to 7, wherein the method does not substantially decrease statin activity.

9. The method of any one of claims 1 to 8, wherein the method does not substantially decrease the lipid-lowering effect or cholesterol lowering activity of the statin.

10. The method of any one of claims 1 to 9, wherein the method does not substantially decrease the cardioprotective effects of the statin.

11. A method of treating diabetes in a subject taking a statin comprising administration of an adenosine receptor antagonist to the subject.

12. The method of claim 11, wherein the adenosine receptor is an A2B receptor.

13. The method of claims 11 or 12, wherein the adenosine receptor antagonist or A2B

receptor antagonist comprises: Compound 38(OSIP-339,39, (N-[2-[[2-phenyl-6-[4-(3- phenylpropyl]piperazine-l-carbonyl]-7H-pyrrolo[2,3-d]pyrimidin-4- yl]amino] ethyl] acetamide}; ATL-801 (N-[5-(l-cyclopropyl-2,6-dioxo-3-propyl-7H- purin-8-yl}pyridin-2-yl]-N-ethylpyridine-3-carboxamide}; CVT-6883 (3-ethyl-l-propyl- 8-[l-[[3-(trifluoromethyl}phenyl]methyl]pyrazol-4-yl]-7H-purine-2,6-dione}; MRS- 1706 (N-(4-acetylphenyl]-2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8- yl]phenoxy]acetamide}; MRS-1754 (N-(4-cyanophenyl]-2-[4-(2,6-dioxo-l,3-dipropyl- 7H-purin-8-yl]phenoxy]acetamide}; PSB-603 (8-[4-[4-(4-chlorophenyl]piperazin-l- yl]sulfonylphenyl]-l-propyl-3,7-dihydropurine-2,6-dione}; PSB-0788 (8-[4-[4-[(4- chlorophenyl]methyl]piperazin-l-yl]sulfonylphenyl]-l-propyl-3,7-dihydropurine-2,6- dione}; PSB-1115 (4-(2,6-dioxo-l-propyl-3,7-dihydropurin-8-yl]benzenesulfonic acid}; GS 6201 (3-ethyl-l-propyl-8-[l-[[3-(trifluoromethyl]phenyl] methyl] pyrazol-4-yl] -7H- purine-2,6-dione}; CGS 15943 (9-chloro-2-(furan-2-yl]-[l,2,4]triazolo[l,5- c]quinazolin-5-amine}; Caffeine Al and A2B antagonist (l,3,7-trimethylpurine-2,6- dione}; Theophylline Adenosine receptor antagonist (l,3-dimethyl-7H-purine-2,6- dione}; XAC Adenosine receptor antagonist (N-(2-aminoethyl]-2-[4-(2,6-dioxo-l,3- dipropyl-7H-purin-8-yl]phenoxy]acetamide}; CGH 2466 dihydrochloride (4-(3,4- dichlorophenyl}-5-pyridin-4-yl-l,3-thiazol-2-amine;dihydrochloride}; XCC Adenosine receptor antagonist (2-[4-(2,6-dioxo-l,3-dipropyl-7H-purin-8-yl]phenoxy]acetic acid}; LAS101057 (N-[5-(3-fluoropyridin-4-yl]-6-pyridin-3-ylpyrazin-2- yl]cyclopropanecarboxamide}; MRE-2029-F20 N-(l,3-Benzodioxol-5-yl}-2-{[5-(2,6- dioxo-l,3-dipropyl-2,3,6,7-tetrahydro-lH-purin-8-yl]-l-methyl-lH-pyrazol-3- yl]oxy}acetamide}; ATL-802 (N-[5-(l-cyclopropyl-2,6-dioxo-3-propyl-7H-purin-8- yl}pyridin-2-yl]-N-methyl-6-(trifluoromethyl}pyridine-3-carboxamide}; Theophylline (l,3-dimethyl-7H-purine-2,6-dione}; Enprofylline (3-propyl-7H-purine-2,6-dione}; or LAS38096 (4-(furan-2-yl}-N-pyridin-3-yl-5-pyrimidin-4-ylpyrimidin-2-amine}.

14. The method of claim 11 or 12, wherein the adenosine receptor antagonist or A2B

receptor antagonist comprises an antibody or antibody fragment that binds to the adenosine receptor or the A2B receptor.

15. The method of any one of claims 11 to 14, wherein the method comprises decreasing, reducing, inhibiting, suppressing, limiting or controlling in the subject elevated blood glucose levels.

16. The method of any one of claims 11 to 15, wherein the method comprises increasing, stimulating, enhancing, promoting, inducing or activating in the subject insulin sensitivity.

17. The method of any one of claims 11 to 16, wherein the method does not substantially decrease statin activity.

18. The method of any one of claims 11 to 17, wherein the method does not substantially decrease the lipid-lowering effects or cholesterol lowering activity of the statin.

19. The method of any one of claims 11 to 18, wherein the method does not substantially decrease the cardioprotective effects of the statin.

20. A method of reducing insulin resistance or risk of diabetes in a subject taking statins the method comprising administration of a CD73 antagonist to the subject.

21. A method of reducing or decreasing increases in blood glucose caused by a statin in a subject taking a statin, comprising administering an amount of a CD73 antagonist to the subject to reduce or decrease increases in blood glucose caused by the statin.

22. The method of claim 20 or 21, wherein the CD73 antagonist comprises an inhibitor or suppressor of CD73 5'-ecto nucleotidase activity; alpha, beta-methylene adenosine-5'- diphosphate (APCP}; or an antibody or antibody fragment that binds to CD73.

23. The method of any one of claims 20 to 22, wherein the CD73 antagonist comprises an antibody having the CDRs of the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9): sc-32299}.

24. The method of any one of claims 20 to 22, wherein the CD73 antagonist comprises an antibody having the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9}: sc-32299}.

25. The method of any one of claims 20 to 24, wherein the method comprises decreasing, reducing, inhibiting, suppressing, limiting or controlling in the subject elevated blood glucose levels.

26. The method of any one of claims 20 to 25, wherein the method comprises increasing, stimulating, enhancing, promoting, inducing or activating in the subject insulin sensitivity.

27. The method of any one of claims 20 to 26, wherein the method does not substantially decrease statin activity.

28. The method of any one of claims 20 to 27, wherein the method does not substantially decrease the beneficial lipid-lowering effects or cholesterol lowering activity of statins.

29. The method of any one of claims 20 to 28, wherein the method does not substantially decrease the cardioprotective effects of the statin.

30. A method of treating diabetes in a subject taking statins comprising administration of a CD73 antagonist to the subject.

31. The method of claim 30, wherein the CD73 antagonist comprises an inhibitor or

suppressor of CD73 5'-ecto nucleotidase activity; alpha, beta-methylene adenosine-5'- diphosphate (APCP}; or an antibody or antibody fragment that binds to CD73.

32. The method of claim 30 or claim 31, wherein the CD73 antagonist comprises an

antibody having the CDRs of the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9): sc-32299}.

33. The method of claim 30 or claim 31, wherein the CD73 antagonist comprises an

antibody having the heavy and the light chain variable regions of CD73 antibody IE9 (Santa Cruz Biotechnology CD73 Antibody (IE9}: sc-32299}.

34. The method of any one of claims 30 to 33, wherein the method comprises decreasing, reducing, inhibiting, suppressing, limiting or controlling in the subject elevated blood glucose levels.

35. The method of any one of claims 30 to 34, wherein the method comprises increasing, stimulating, enhancing, promoting, inducing or activating in the subject insulin sensitivity.

36. The method of any one of claims 30 to 35, wherein the method does not substantially decrease statin activity.

37. The method of any one of claims 30 to 36, wherein the method does not substantially decrease the beneficial lipid-lowering effects or cholesterol lowering activity of statins.

38. The method of any one of claims 30 to 37, wherein the method does not substantially decrease the cardioprotective effects of the statin.

39. A composition comprising an adenosine receptor antagonist and a statin.

40. The composition of claim 39, wherein the adenosine receptor is an A2B receptor.

41. The method of any of claims 1 to 19, or the composition of claim 39 or 40, wherein the statin is atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rousuvastatin or simvastatin.

42. The method of any of claims 1 to 19, or the composition of claim 39 or 40, further comprising a CD73 antagonist.

43. A composition comprising a CD73 antagonist and a statin.

44. The composition of claim 43, wherein the CD73 antagonist comprises an inhibitor or suppressor of CD73 5'-ecto nucleotidase activity; alpha, beta-methylene adenosine-5'- diphosphate (APCP}; or an antibody or antibody fragment that binds to CD73.

45. The method of any of claims 20 to 38, or the composition of claim 43 or 44, wherein the statin is atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rousuvastatin or simvastatin.

46. The method of any of claims 20 to 38, or the composition of claim 43 or 44, further comprising an adenosine receptor antagonist.