A3 adenosine receptor ligand for use for achieving fat loss effect

Abstract

To provide an A3 adenosine receptor (A3AR) ligand for use in at least one of: reducing weight of a subject; reducing body fat mass in the subject; treating obesity in the subject; lowering a level of adipocytes in the subject; and inhibiting proliferation of adipocytes in the subject.SOLUTION: Provided is a pharmaceutical composition for reducing weight of a subject and for inhibiting proliferation of adipocytes in the subject, the pharmaceutical composition comprising (i) a pharmaceutically acceptable carrier and (ii) an A3AR ligand as an active ingredient, wherein the A3AR ligand is an A3AR agonist selected from 2-chloro-N6-(3-iodobenzyl)-adenosine-5'-N-methyluronamide; or an A3AR allosteric modulator selected from N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine.SELECTED DRAWING: None

Description

Technical Field

[0001] The present disclosure relates to the medical use of A3AR ligands.

[0002] Background Technical Documents The following references are listed as being considered relevant as background to the subject matter disclosed herein. - International Patent Application Publication No. 04 / 007519 - International Patent Application Publication No. 2013 / 111132 - International Patent Application Publication No. 17 / 090036

[0003] Acknowledging the above references in this specification does not mean that they are in any way relevant to the patentability of the subject matter disclosed herein.

Background Art

[0004] Obesity is a complex disease associated with an excessive amount of body fat and is considered a major health problem, sometimes life-threatening, in Western societies.

[0005] Obesity is caused by an imbalance between food intake and basal metabolism and energy consumption. At the individual level, multiple endogenous or environmental causes can lead to obesity. However, in most cases, the combination of excessive calorie intake and the availability of energy-dense meals is considered the main cause of obesity.

[0006] Weight loss can improve or prevent health problems associated with obesity, but in some cases, it may be necessary to combine weight loss treatments with drug therapy.

[0007] International Patent Application Publication No. 04 / 007519 describes compounds that are partial or full A1 adenosine receptor agonists, and their use in treating mammals in various disease states such as diabetic diseases and obesity and in altering adipocyte function.

[0008] International Patent Application Publication No. 2013 / 111132 describes the use of 2-chloro-N 6 -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA, CF102) Cl-IB-MECA for the treatment of hepatocellular carcinoma (HCC) and for maintaining liver function in subjects with chronic liver disease.

[0009] International Patent Application Publication No. 17 / 090036 describes the use of an A3AR ligand, specifically Cl-IB-MECA, for reducing ectopic fat accumulation, particularly fat accumulation in fatty liver, specifically for treating non-alcoholic fatty liver disease (NAFLD). SUMMARY OF THE INVENTION

[0010] In a first aspect thereof, the present disclosure provides an A3 adenosine receptor (A3AR) ligand for use in any one of the following: - reducing the body weight of a subject, - reducing the body fat mass of a subject, - treating obesity of a subject, - reducing the level of adipocytes of a subject, and - suppressing the proliferation of adipocytes of a subject.

[0011] In a second aspect, the present disclosure provides a pharmaceutical composition comprising an A3AR ligand as an active ingredient in an amount effective to achieve at least one of the following fat-reducing effects: - reducing the body weight of a subject, - reducing the body fat mass of a subject, - treating obesity of a subject, - reducing the level of adipocytes of a subject, and - suppressing the proliferation of adipocytes of a subject.

[0012] Further provided by the present disclosure is a method of treatment comprising administering to a subject in need an amount of an A3AR ligand effective to achieve at least one of the fat-reducing effects selected from the following. - reducing the weight of the subject, - reducing the amount of body fat of the subject, - treating obesity of the subject, - reducing the level of adipocytes of the subject, - suppressing the proliferation of adipocytes of the subject.

[0013] In some examples, the A3AR ligand is an A3AR agonist, preferably 2-chloro-N 6 -(3-iodobenzyl)-adenosine-5’-N-methyluronamide (Cl-IB-MECA, also referred to herein as CF102).

Brief Description of the Drawings

[0014] To better understand the subject matter disclosed herein and to illustrate how it may be practiced, embodiments will be described by way of example only, with reference to the accompanying drawings.

Figure 1

Figure 2

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Figure 4A

Figure 4B

Figure 5

DETAILED DESCRIPTION OF THE INVENTION

[0015] The present disclosure relates to 2-chloro-N 6 -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA, also referred to herein as CF102), an agonist of the A3 adenosine receptor (A3AR) having high affinity and selectivity for A3AR, which is based on the finding that it inhibits the proliferation of adipocytes in an in vitro adipocyte proliferation model.

[0016] This unexpected inhibitory effect was demonstrated, in particular, by a decrease in the level of adipocytes (Figure 1) and a decrease in the accumulation of lipids produced in 3T3-L1 adipocytes (Figures 2A-2B and 3).

[0017] In addition, the unexpected effect of Cl-IB-MECA was demonstrated by a statistically significant decrease in the body weight of high-fat diet mice.

[0018] Furthermore, unexpectedly, the decrease in body weight was also shown in an animal model of diabetes, a complication of obesity, when treated with the allosteric modulator of A3AR, N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine (sometimes abbreviated as LUF6000 or CF602).

[0019] Based on these findings, the inventors have concluded that a ligand having high affinity and selectivity for A3AR is an effective tool for achieving a therapeutically beneficial effect, whether it is an A3AR-selective agonist or an A3AR-selective allosteric modulator, in any one or combination of the following: - reducing the weight of a subject, - reducing the body fat mass of a subject, - treating obesity in a subject, - decreasing the level of adipocytes in a subject, and - inhibiting the proliferation of adipocytes in a subject.

[0020] Without being bound by theory, it is believed that the above effects are directly or indirectly linked by the inhibitory effect of the ligand on adipocyte proliferation, and are collectively referred to as the "fat-reducing effect".

[0021] In the context of the present disclosure, each of the above effects or a combination of two or more such effects is considered as a separate embodiment.

[0022] Accordingly, the present disclosure provides an A3AR ligand for use in achieving any of the above fat-reducing effects, as well as a pharmaceutical composition comprising an A3AR ligand for use in achieving any of the above fat-reducing effects and a method of treating a subject in need of any one of the above fat-reducing effects, which treatment also includes preventing an increase in fat, as further described below.

[0023] In one example, the A3AR ligand is for use in treating a subject suffering from obesity.

[0024] In another example, the A3AR ligand is for use in inducing or promoting weight loss in a subject.

[0025] In yet another example, the A3AR ligand is for use in decreasing the level of adipocytes in a subject.

[0026] In yet another further example, the A3AR ligand is for suppressing the proliferation of adipocytes.

[0027] In the context of the present disclosure, when referring to the treatment of weight loss or obesity, it should be understood to equivalently refer to reducing tissues composed of fat, such as adipose tissue intended for storing fat, i.e., subcutaneous and / or peripheral fat, and excluding ectopic fat such as ectopic fat in liver tissue. In some examples, the present disclosure excludes the reduction of ectopic adipose tissue, particularly adipose tissue in the liver.

[0028] In one preferred example, the A3AR ligand is used to treat subjects with excessive body fat, particularly subjects suffering from obesity.

[0029] In some examples, the reduction in body fat mass is indicated by a reduction in peripheral fat, particularly a reduction in adipose tissue.

[0030] In some other examples, the reduction in body fat mass is indicated by a reduction in lipid production in 3T3-L1 adipocytes.

[0031] Excessive body fat can be determined by the body mass index (BMI). A subject with a BMI exceeding 25 is considered to have excessive body fat, and a subject with a BMI exceeding 30 is considered to be suffering from obesity.

[0032] The effect of the treatment, e.g., the reduction in the measured quantities according to the present disclosure, such as body weight, body fat mass and / or the level of adipocytes, can be determined by their values obtained at two different time points. In some examples, the first time point is before treatment and the second time point is during treatment.

[0033] In some examples, the two time points are during treatment. The time difference between the first time point and the second time point may be, for example, 1 day, 1 week, 1 month, or even up to 1 year, during which the subject receives administration of the A3AR ligand according to any of the embodiments of the present disclosure.

[0034] When referring to a decrease in one value of a measurement, it should be understood that the decrease in the value is to the extent considered significant by a physician. That extent can be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or sometimes more than 70% compared to a predetermined reference time point. It should be noted that there may be multiple time points at which the value is measured between the reference time point and the evaluation time point.

[0035] In some examples, the reference time point is before treatment begins. In some other examples, the reference time point is a time point during treatment.

[0036] In some examples, the evaluation time point is a determined time point that is the end of treatment. In some other examples, the evaluation time point is a time point during treatment after the reference time point.

[0037] It should be noted that between those two time points, the subject is receiving administration of an A3AR ligand. However, treatment with an A3AR ligand does not need to be long-term, for example, throughout the entire period, and the subject may receive administration periodically.

[0038] In one example, treatment with an A3AR ligand is long-term treatment, that is, throughout the entire treatment period.

[0039] In the context of the present disclosure, "A3 adenosine receptor ligand" or "A3AR ligand" means any compound that affects the activity of the A3 adenosine receptor, including full or partial activation of the A3 adenosine receptor, either directly (e.g., through a receptor binding site) or indirectly (e.g., through an allosteric binding site).

[0040] An A3AR ligand according to the present disclosure is a molecule that exerts its main action through activation of A3AR, regardless of whether its activation is through a binding site or an allosteric binding site.

[0041] A3AR ligands have high affinity and selectivity for A3AR. This means that at the dosage at which the ligand is administered, it essentially affects only A3AR.

[0042] The specificity and high affinity of ligands for the A3 adenosine receptor result in beneficial effects over the activation of other adenosine receptors, such as the A1 adenosine receptor, which are known to have cardiovascular side effects, especially when using A3AR ligands. In fact, studies have shown that A3AR ligands have protective effects, especially neuroprotective, chemoprotective, cardioprotective, and hepatoprotective effects.

[0043] In one example, an "A3 adenosine receptor ligand" is an A3AR agonist.

[0044] In another example, an "A3 adenosine receptor ligand" is an allosteric modulator of A3AR (which can also be called an allosteric effector).

[0045] When referring to an "A3 adenosine receptor agonist" or an "A3AR agonist", it should be understood to mean any ligand that specifically binds to the A3 adenosine receptor and can thereby fully or partially activate the A3 adenosine receptor.

[0046] In the context of the present disclosure, when its affinity for A3AR is at least 3 times greater (i.e., its Ki for A3AR is at least 3 times lower) than its affinity for any other adenosine receptor (i.e., A1, A 2a and A 2b ), preferably 10 times, desirably 20 times, and most preferably at least 50 times greater, the molecule will be considered an A3AR agonist (i.e., a molecule that exerts its main action through binding to and activation of A3AR).

[0047] The affinity of an A3AR agonist for the human A3AR and its relative affinity for other human adenosine receptors can be determined by a number of assays, such as binding assays. Examples of binding assays include preparing a membrane containing the receptor and measuring the ability of the A3AR agonist to displace the bound radiolabeled agonist, using cells presenting each human adenosine receptor and measuring in a functional assay the ability of the A3AR agonist to activate or inactivate downstream signaling events such as the effect on adenylate cyclase, which may be measured through an increase or decrease in cAMP levels in some cases. The dose of the A3AR agonist is increased such that its blood concentration reaches a level approaching the Ki of the A1, A 2a and A 2b adenosine receptors. Following such administration, activation of those receptors may occur in addition to activation of the A3AR. Therefore, it is preferred that the A3AR agonist be administered at a blood concentration such that essentially only the A3AR is activated.

[0048] In one example, the A3AR agonist has a binding affinity (K i ) for the human A3AR of less than 100 nM, typically less than 50 nM, preferably less than 20 nM, more preferably less than 10 nM, and ideally less than 5 nM. Particularly preferred are A3AR agonists with a K i for the human A3R of less than 2 nM, desirably less than 1 nM.

[0049] In the context of the present disclosure, it should be understood that some A3AR agonists can interact with and activate other adenosine receptors, but have a lower affinity (i.e., a higher Ki).

[0050] In some examples, the A3AR agonist is a molecule having a purine backbone. Purine-containing compounds can be determined as A3AR agonists based on acceptable structure-function activity assays.

[0051] The characteristics of some A3AR agonists used in accordance with the present disclosure and methods for their preparation are described in detail, in particular, in U.S. Patent No. 5,688,774, U.S. Patent No. 5,773,423, U.S. Patent No. 5,573,772, U.S. Patent No. 5,443,836, U.S. Patent No. 6,048,865, International Publication No. 95 / 02604, International Publication No. 99 / 20284, International Publication No. 99 / 06053, International Publication No. 97 / 27173, and International Publication No. 01 / 19360. All of these documents are incorporated herein by reference.

[0052] According to some examples of the present disclosure, the A3AR agonist is a purine derivative falling within the scope of the following general formula (I).

[0053] TIFF2025098147000001.tif36170

[0054] In the formula, -R 11 represents alkyl, hydroxyalkyl, carboxyalkyl or cyanoalkyl, or a group of the following general formula (II),

[0055] TIFF2025098147000002.tif28170

[0056] In the formula, -Y represents oxygen, sulfur or CH2, -X 11 is H, alkyl, R e R f NC(=O)- or HOR g - represents, in the formula, -R e and R f may be the same or different and are selected from the group consisting of hydrogen, alkyl, amino, haloalkyl, aminoalkyl, BOC-aminoalkyl, and cycloalkyl, or are joined together to form a heterocyclic ring containing 2 to 5 carbon atoms, -R g is selected from the group consisting of alkyl, amino, haloalkyl, aminoalkyl, BOC-aminoalkyl, and cycloalkyl, -X 12 is H, hydroxyl, alkylamino, alkylamide or hydroxyalkyl, -X 13 and X 14 are each independently hydrogen, hydroxyl, amino, amide, azide, halo, alkyl, alkoxy, carboxy, nitrilo, nitro, trifluoro, aryl, alkylaryl, thio, thioester, thioether, -OCOPh, -OC(=S)OPh, or both X 13 and X 14 are oxygen atoms that are linked to >C=S to form a 5-membered ring, or X 12 and X 13 form a ring of formula (III),

[0057] TIFF2025098147000003.tif31170

[0058] wherein R' and R'' independently represent an alkyl group, -R 12 is selected from the group consisting of hydrogen, halo, alkyl ether, amino, hydrazide, alkylamino, alkoxy, thioalkoxy, pyridylthio, alkenyl, alkynyl, thio, and alkylthio, -R 13 is a group of the formula -NR 15 R 16 wherein -R 15 is a hydrogen atom or a group selected from alkyl, substituted alkyl or aryl-NH-C(Z)-, Z is O, S, or NR a and R e has the above meanings, where when R 15 is hydrogen, -R 16is R- and S-1-phenylethyl, benzyl, phenylethyl or anilide group substituted with a substituent selected from the group consisting of alkyl, amino, halo, haloalkyl, nitro, hydroxyl, acetamide, alkoxy, and sulfonic acid or its salt at one or more positions, benzo[1,3]dioxol-5-ylmethyl, furfuryl, L-propylalanyl-aminobenzyl, β-alanyl-aminobenzyl, T-BOC-β-alanyl-aminobenzyl, phenylamino, carbamoyl, phenoxy or cycloalkyl, or R 16 is a group of the following formula (IV),

[0059] TIFF2025098147000004.tif32170

[0060] or R 15 when is alkyl or aryl-NH-C(Z)-, R 16 is selected from the group consisting of heteroaryl-NR a -C(Z)-, heteroaryl-C(Z)-, alkyl-NR a -C(Z)-, alkyl-C(Z)-, aryl-NR-C(Z)- and aryl-C(Z)-, and Z represents oxygen, sulfur or amine.

[0061] Exemplary A3AR agonists (disclosed in columns 4, line 67 - column 6, line 16; column 5, lines 40 - 45; column 6, lines 21 - 42; column 7, lines 1 - 11; column 7, lines 34 - 36; and column 7, lines 60 - 61 of U.S. Patent No. 5,688,774): N 6 -(3-iodobenzyl)-9-methyladenine, N 6 -(3-iodobenzyl)-9-hydroxyethyladenine, R-N 6 -(3-iodobenzyl)-9-(2,3-dihydroxypropyl)adenine, S-N 6 -(3-iodobenzyl)-9-(2,3-dihydroxypropyl)adenine, N 6-(3-Iodobenzyladenin-9-yl)acetic acid, N 6 -(3-Iodobenzyl)-9-(3-cyanopropyl)adenine, 2-Chloro-N 6 -(3-Iodobenzyl)-9-methyladenine, 2-Amino-N 6 -(3-Iodobenzyl)-9-methyladenine, 2-Hydrazide-N 6 -(3-Iodobenzyl)-9-methyladenine, N 6 -(3-Iodobenzyl)-2-methylamino-9-methyladenine; 2-Dimethylamino-N 6 -(3-Iodobenzyl)-9-methyladenine, N 6 -(3-Iodobenzyl)-9-methyl-2-propylaminoadenine, 2-Hexylamino-N 6 -(3-Iodobenzyl)-9-methyladenine, N 6 -(3-Iodobenzyl)-2-methoxy-9-methyladenine, N 6 -(3-Iodobenzyl)-9-methyl-2-methylthioadenine, N 6 -(3-Iodobenzyl)-9-methyl-2-(4-pyridylthio)adenine, (1S,2R,3S,4R)-4-(6-Amino-2-phenylethylamino-9H-purin-9-yl)cyclopentane-1,2,3-triol, (1S,2R,3S,4R)-4-(6-Amino-2-chloro-9H-purin-9-yl)cyclopentane-1,2,3-triol, (±)-9-[2α,3α-Dihydroxy-4β-(N-methylcarbamoyl)cyclopent-1β-yl)]-N 6 -(3-Iodobenzyl)-adenine, 2-Chloro-9-(2’-amino-2’,3’-dideoxy-β-D-5’-methyl-arabinofuranamide)-N 6-(3-Iodobenzyl)adenine, 2-Chloro-9-(2’,3’-dideoxy-2’-fluoro-β-D-5’-methyl-arabinofuranosyl)-N 6 -(3-Iodobenzyl)adenine, 9-(2-Acetyl-3-deoxy-β-D-5-methyl-ribofuranosyl)-2-chloro-N 6 (3-Iodobenzyl)adenine, 2-Chloro-9-(3-deoxy-2-methanesulfonyl-β-D-5-methyl-ribofuranosyl)-N 6 -(3-Iodobenzyl)adenine, 2-Chloro-9-(3-deoxy-β-D-5-methyl-ribofuranosyl)-N 6 -(3-Iodobenzyl)adenine, 2-Chloro-9-(3,5-1,1,3,3-tetraisopropyldisiloxanyl-β-D-5-ribofuranosyl)-N 6 -(3-Iodobenzyl)adenine, 2-Chloro-9-(2’,3’-O-thiocarbonyl-β-D-5-methyl-ribofuranosyl)-N 6 -(3-Iodobenzyl)adenine, 9-(2-Phenoxythiocarbonyl-3-deoxy-β-D-5-methyl-ribofuranosyl)-2-chloro-N 6 -(3-Iodobenzyl)adenine, 1-(6-Benzylamino-9H-purin-9-yl)-1-deoxy-N,4-dimethyl-β-D-ribofuranosiduronamide, 2-Chloro-9-(2,3-dideoxy-β-D-5-methyl-ribofuranosyl)-N 6 Benzyladenine, 2-Chloro-9-(2’-azido-2’,3’-dideoxy-β-D-5’-methyl-arabino-furanosyl)-N 6 -Benzyladenine, 2-Chloro-9-(β-D-erythrofuranoside)-N 6 -(3-Iodobenzyl)adenine, N6 -(benzo dioxane methyl) adenosine, 1-(6-furfurylamino-9H-purin-9-yl)-1-deoxy-N-methyl-β-D-ribofuranosyluronamide, N 6 -[3-(L-prolylamino) benzyl] adenosine-5’-N-methyluronamide, N 6 -[3-(β-alanyl amino) benzyl] adenosine-5’-N-methyluronamide, N 6 -[3-(N-T-Boc-β-alanyl amino) benzyl] adenosine-5’-N-methyluronamide 6-(N’-phenylhydrazinyl) purine-9-β-ribofuranoside-5’-N-methyluronamide, 6-(O-phenylhydroxylamino) purine-9-β-ribofuranoside-5’-N-methyluronamide, 9-(β-D-2’,3’-dideoxyerythrofuranosyl)-N 6 -[(3-β-alanyl amino) benzyl] adenosine, 9-(β-D-erythrofuranoside)-2-methylamino-N 6 -(3-iodobenzyl) adenine, 2-chloro-N-(3-iodobenzyl)-9-(2-tetrahydrofuryl)-9H-purin-6-amine, 2-chloro-(2’-deoxy-6’-thio-L-arabinofuranosyl) adenine, and 2-chloro-(6’-thio-L-arabinofuranosyl) adenine.

[0062] Other exemplary A3AR agonists are disclosed in U.S. Patent No. 5,773,423 and are compounds of formula (V).

[0063] TIFF2025098147000005.tif70170

[0064] Wherein, X1 is R a R bis NC(=O), where R a and R b may be the same or different and are selected from the group consisting of hydrogen, C1-C 10 alkyl, amino, C1-C 10 haloalkyl, C1-C 10 aminoalkyl, and C3-C 10 cycloalkyl, R2 is selected from the group consisting of hydrogen, halo, C1-C 10 alkyoxy, amino, C2-C 10 alkenyl, and C2-C 10 alkynyl, R5 is selected from the group consisting of R- and S-1-phenylethyl, unsubstituted benzyl groups, and benzyl groups substituted with substituents selected from the group consisting of C1-C 10 alkyl, amino, halo, C1-C 10 haloalkyl, nitro, hydroxy, acetamido, C1-C 10 alkoxy, and sulfo at one or more positions.

[0065] More specific A3AR agonists include those of the above formula where R a and R b may be the same or different and are selected from the group consisting of hydrogen and C1-C 10 alkyl, especially those where R2 is hydrogen or halo, particularly those where R2 is hydrogen.

[0066] Additional specific A3AR agonists are compounds where, particularly when R5 is unsubstituted benzyl, R a is hydrogen and R2 is hydrogen.

[0067] More specific A3AR agonists are such compounds where R b is C1-C 10 alkyl or C3-C 10 cycloalkyl, specifically C1-C 10 alkyl, more specifically methyl.

[0068] Particularly specific are those where Ra is hydrogen, R b is C1-C 10 alkyl or C3-C 10 cycloalkyl, R5 is R- or S-1-phenylethyl, or benzyl substituted with a substituent selected from the group consisting of halo, amino, acetamido, C1-C 10 haloalkyl, and sulfo, and those A3AR agonists wherein the sulfo derivative is a salt such as a triethylammonium salt.

[0069] Furthermore, those compounds in which R2 is of the formula R d -C=C- C2-C 10 alkenylene, and R d is C1-C8 alkyl are also specifically described in U.S. Patent No. 5,773,423.

[0070] Similarly specific are compounds in which R2 is other than hydrogen, especially those in which R2 is halo, C1-C 10 alkylamino, or C1-C 10 alkylthio, more preferably those in which furthermore R a is hydrogen, R b is C1-C 10 alkyl, and / or R5 is substituted benzyl.

[0071] Further exemplary A3AR agonists disclosed in U.S. Patent No. 5,773,423 are modified xanthine-7-ribosides having formula (VI).

[0072] TIFF2025098147000006.tif61170

[0073] wherein X is O, R6 is R a R b NC(=O), wherein R a and R b may be the same or different and are hydrogen, C1-C 10 alkyl, amino, C1-C 10Haloalkyl, C1-C 10 Aminoalkyl, and C3-C 10 Selected from the group consisting of cycloalkyl, R7 and R8, which may be the same or different, are C1-C 10 Alkyl, R- and S-1-phenylethyl, unsubstituted benzyl group, and a benzyl group substituted with a substituent selected from the group consisting of C1-C 10 Alkyl, amino, halo, C1-C 10 Haloalkyl, nitro, hydroxy, acetamide, C1-C 10 Alkoxy, and a benzyl group substituted with a substituent selected from the group consisting of sulfo, R9 is selected from the group consisting of halo, benzyl, phenyl, and C3-C 10 Cycloalkyl.

[0074] WO 99 / 06053 discloses compounds of Examples 19 to 33 selected from the following. N 6 -(4-Biphenyl-carbonylamino)-adenosine-5'-N-ethyluronamide, N 6 -(2,4-Dichlorobenzyl-carbonylamino)-adenosine-5'-N-ethyluronamide, N 6 -(4-Methoxyphenyl-carbonylamino)-adenosine-5'-N-ethyluronamide, N 6 -(4-Chlorophenyl-carbonylamino)-adenosine-5'-N-ethyluronamide, N 6 -(Phenyl-carbonylamino)-adenosine-5'-N-ethyluronamide, N 6 -(Benzylcarbamoylamino)-adenosine-5'-N-ethyluronamide, N 6 -(4-Sulfonamido-phenylcarbamoyl)-adenosine-5'-N-ethyluronamide, N 6-(4-Acetyl-phenylcarbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -((R)-α-Phenylethylcarbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -((S)-α-Phenylethylcarbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -(5-Methyl-isoxazol-3-yl-carbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -(1,3,4-Thiadiazol-2-yl-carbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -(4-n-Propoxy-phenylcarbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -Bis-(4-nitrophenylcarbamoyl)-adenosine-5'-N-ethyluronamide, and N 6 -Bis-(5-chloro-pyridin-2-yl-carbamoyl)-adenosine-5'-N-ethyluronamide.

[0075] More specifically disclosed A3AR agonists to be used in accordance with the present disclosure include the following. 2-Chloro-N 6 -(3-Iodobenzyl)-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine, 2-Chloro-N 6 -(3-Iodobenzyl)-adenosine-5'-N-methyluronamide, also known as or by the abbreviation Cl-IB-MECA, N 6 -(3-Iodobenzyl)-2-methylamino-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine, N 6-Also known as (3-iodobenzyl)-adenosine-5'-N-methyluronamide, as 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-D-ribofuranuronamide, or by the abbreviation IB-MECA, N 6 -2-(4-aminophenyl)ethyladenosine (APNEA), N 6 -(4-amino-3-iodobenzyl)adenosine-5'-(N-methyluronamide) (AB-MECA).

[0076] Examples of particularly preferred A3AR agonists include 2-chloro-N 6 -(3-iodobenzyl)-2-methylamino-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine, 2-chloro-N 6 -(3-iodobenzyl)-adenosine-5'-N-methyluronamide, also known as or by the abbreviation Cl-IB-MECA.

[0077] In some examples, Cl-IB-MECA is used to achieve a fat-reducing effect.

[0078] In one particular example, Cl-IB-MECA is used for the treatment of obesity.

[0079] When referring to an "allosteric modulator of A3AR" or "A3ARM", it should be understood to refer to the positive modulation, activation, or increase of receptor activity by the binding of an allosteric modulator at an allosteric site of the receptor, which may be different from the binding site of the endogenous ligand or its agonist.

[0080] In one example, "modulation" means the effect of an A3AR ligand on a receptor as shown by at least a 15% increase in the potency of the A3 adenosine receptor by binding of the compound to the allosteric site of the receptor and / or by a decrease in the dissociation rate of adenosine or an A3AR agonist for the orthosteric binding site.

[0081] In one example, the modulation is by an allosteric modulator of A3AR (A3ARAM) which is an imidazoquinoline derivative.

[0082] In one example, the A3ARAM, or imidazoquinoline derivative, has the following general formula (VII).

[0083] TIFF2025098147000007.tif50170

[0084] Wherein, - R1 is optionally, in an aromatic ring, C1 - C 10 alkyl, halo, C1 - C 10 alkanol, hydroxyl, C1 - C 10 acyl, C1 - C 10 alkoxyl, C1 - C 10 - alkoxycarbonyl, C1 - C 10 alkoxylalkyl, C1 - C 10 thioalkoxy, C1 - C 10 alkyl ether, amino, hydrazide, C1 - C 10 alkylamino, pyridylthio, C2 - C 10 alkenyl, C2 - C 10 alkynyl, thio, C1 - C 10 alkylthio, acetamide, sulfonic acid, represents aryl or alkyl substituted with one or more substituents selected from the group consisting of, or the substituents can together form a cycloalkyl or cycloalkenyl condensed to the aryl, the cycloalkyl or cycloalkenyl optionally containing one or more heteroatoms (provided that the aryl is not an unsubstituted phenyl group), -R2 is hydrogen or C1-C 10 alkyl, C2-C 10 alkenyl, C2-C 10 alkynyl, C4-C 10 cycloalkyl, C4-C 10 cycloalkenyl, a 5- to 7-membered heteroaromatic ring, C5-C 15 fused cycloalkyl, bicyclic aromatic ring or heteroaromatic ring, C1-C 10 alkyl ether, amino, hydrazide, C1-C 10 alkylamino, C1-C 10 alkoxy, C1-C 10 -alkoxycarbonyl, C1-C 10 alkanols, C1-C 10 acyl, C1-C 10 thioalkoxy, pyridylthio, thio, and C1-C 10 alkylthio, acetamide, and sulfonic acid, and represents a substituent selected from the group consisting of pharmaceutically acceptable salts thereof.

[0085] According to some embodiments, the R1 substituent of A3ARAM has the following general formula (VIII).

[0086] TIFF2025098147000008.tif34170

[0087] Wherein n is an integer selected from 0 or 1 to 5, preferably n is 0, 1 or 2, -X1 and X2 may be the same or different and are selected from hydrogen halogen, alkyl, alkanol or alkoxy, indanyl, pyrroline, provided that when n is 0, X1 and X2 are not hydrogen.

[0088] In some further examples, R1 of A3ARAM is a substituent having the above formula (VIII), wherein X1 or X2 may be the same or different and is selected from hydrogen, chloro, methoxy, methanol or a substituent having formula (VIIIa) or (VIIIb).

[0089] TIFF2025098147000009.tif40170

[0090] In the formula, Y is selected from N or CH.

[0091] In some further examples, R2 of A3ARAM is H, C 1-10 alkyl, C 4-10 selected from cycloalkyl, and the alkyl chain may be straight-chain or branched, or may form a 4- to 7-membered cycloalkyl ring.

[0092] In one example, R2 of A3ARAM is selected from 5- to 7-membered heteroaromatic rings.

[0093] In some examples, the R2 substituent of A3ARAM is selected from H, n-pentyl, or a 5-membered heteroaromatic ring having the following formula (IX).

[0094] TIFF2025098147000010.tif30170

[0095] In the formula, Z is selected from O, S or NH, preferably O.

[0096] In one example, R2 of A3ARAM includes one or more fused rings so as to form a bicyclic substituent in particular.

[0097] Non-limiting examples of bicyclic compounds that can be used to form substituents in the context of A3ARAM include bicyclo[2.2.1]heptane, bicyclo[4.1.0]heptane, bicyclo[4.1.0]heptane-3-carboxylic acid, bicyclo[3.1.0]hexane-3-carboxylic acid, bicyclo[4.1.0]heptane-2-carboxylic acid, bicyclo[3.1.0]hexane-2-carboxylic acid, and bicyclo[2.2.1]heptane-2-carboxylic acid.

[0098] In some further other examples, R2 of A3ARAM is selected from 2-cyclohexene and 3-cyclohexene.

[0099] Specific imidazoquinoline derivatives that can be used as allosteric modulators of A3AR are listed below. N-(4-Methyl-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(4-Methoxy-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(4-Chloro-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3-Methanol-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-([3,4-c]Indan)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(1H-Indazol-6-yl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(4-Methoxy-benzyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(1H-Indol-6-yl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(Benzyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(Phenylethyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-cycloheptyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-furyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-cyclobutyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2--1H-imidazo[4,5-c]quinolin-4-amine N-(3,4-Dichloro-phenyl)-2-pentyl-1H-imidazo[4,5-c]quinolin-4-amine

[0100] On the one hand, if the imidazoquinoline derivative has any affinity for the orthosteric binding site of the A1 and A 2A , A 2B adenosine receptors, it is reduced, and the affinity for the orthosteric binding site of the A3 adenosine receptor is reduced. On the other hand, since it has a high affinity for the allosteric site of the A3 adenosine receptor, it is regarded as an allosteric modulator (International Patent Application Publication No. 07 / 089507, incorporated herein by reference).

[0101] In the present disclosure, a specifically preferred imidazoquinoline derivative is N-(3,4-dichloro-phenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine (sometimes also referred to by the abbreviations LUF6000 or CF602), which is an allosteric modulator of A3AR.

[0102] In the context of the general formula disclosed herein, the following meanings are considered for various terms.

[0103] The term "alkyl" is used herein to refer to a straight-chain or branched hydrocarbon chain having 1 to 10 carbon atoms, more preferably 1 to 6 carbon atoms, and examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, n-heptyl, octyl, etc.

[0104] Similarly, the terms "alkenyl" and "alkynyl" each mean a straight-chain or branched hydrocarbon chain having 2 to 10, or 3 to 10 carbon atoms, more preferably 2 to 6, or 3 to 6 carbon atoms, and the alkenyl or alkynyl has at least one unsaturated bond.

[0105] The alkyl, alkenyl or alkynyl substituent may be substituted with a heteroatom-containing group. Thus, although not explicitly stated, any of the alkyl modifications defined above and below herein, such as alkylthio, alkoxy, alkanol, alkylamine, etc., should be understood to also include the corresponding alkenyl or alkynyl modifications, such as alkenylthio, alkenyloxy, alkenol, alkenylamine, or alkynylthio, alkynyloxy, alkynol, alkynylamine, respectively.

[0106] The term "aryl" means an unsaturated aromatic carbocyclic group consisting of 5 to 14 carbon atoms having a single ring (e.g., phenyl) or multiple fused rings (e.g., naphthyl or anthryl). Preferred aryls include phenyl, indanyl, benzimidazole.

[0107] The term "alkaryl" preferably refers to an -alkylene-aryl group having 1 to 10 carbon atoms in the alkylene moiety and 6 to 14 carbon atoms in the aryl moiety. Examples of such alkaryl groups include benzyl, phenethyl, etc.

[0108] The term "substituted aryl" refers to an aromatic moiety substituted with 1 to 3 substituents defined above. As will be understood by those skilled in the art, various substituents are possible. Nevertheless, some preferred substituents include, but are not limited to, halogen, (substituted) amino, nitro, cyano, alkyl, alkoxy, acyloxy or alkanol, sulfonyl, sulfinyl.

[0109] The term "halo" or "halogen" refers to fluoro, chloro, bromo and iodo, preferably chloro.

[0110] The term "acyl" refers to an H-C(O)-group and an alkyl-C(O)-group.

[0111] The term "alkan ol" refers to a -COH group and an alk-OH group, where "alk" means an alkylene, alkenylene or alkynylene chain.

[0112] The term "alkoxy" is used herein to mean -O-alkyl and includes, but is not limited to, for example, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, etc.

[0113] The term "alkylthio" is used herein to mean -S-alkyl and includes, but is not limited to, for example, methylthio, ethylthio, n-propylthio, isopropylthio, n-butylthio, etc.

[0114] The term "alkoxyalkyl" is used herein to mean -alkyl-O-alkyl and includes, but is not limited to, for example, methoxymethyl, ethoxymethyl, n-propoxymethyl, isopropoxymethyl, n-butoxymethyl, isobutoxymethyl, t-butoxymethyl, etc.

[0115] The term "cycloalkyl" is used herein to mean a cyclic hydrocarbon radical and includes, but is not limited to, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, etc.

[0116] The term "alkoxycarbonyl" is used herein to mean -C(O)O-alkyl and includes, but is not limited to, for example, methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, etc.

[0117] The term "fused cycloalkyl" is used herein to mean any compound or substituent containing at least two aliphatic rings that are fused at only one atom (to form a spiro ring moiety), at two mutually bonded atoms, or across a series of atoms (bridgeheads). The fused rings can include any bicyclic, tricyclic, and polycyclic moieties. In some embodiments of the present disclosure, bicyclic substituents are preferred.

[0118] The present disclosure also utilizes physiologically acceptable salts of A3AR-selective ligands such as the above compounds. "Physiologically acceptable salts" refer to any non-toxic alkali metal, alkaline earth metal, ammonium salts commonly used in the pharmaceutical industry, including sodium, potassium, lithium, calcium, magnesium, barium ammonium, and protamine zinc salts, etc., and are prepared by methods known in the art. The term also includes non-toxic acid addition salts generally prepared by reacting the ligand with a suitable organic or inorganic acid. The acid addition salts retain the biological effects and qualitative properties of the free base and are non-toxic or otherwise not undesirable. Examples include, inter alia, acids derived from mineral acids, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, metaphosphoric acid, etc. Organic acids include, inter alia, tartaric acid, acetic acid, propionic acid, citric acid, malic acid, malonic acid, lactic acid, fumaric acid, benzoic acid, cinnamic acid, mandelic acid, glycolic acid, gluconic acid, pyruvic acid, succinic acid, salicylic acid, arylsulfonic acids, such as p-toluenesulfonic acid, etc.

[0119] The A3AR ligand can be administered as a single dose (a one-time dosing) or as a continuous treatment over several days, weeks, or even several months.

[0120] In one example, the A3AR ligand is used for long-term treatment (treatment over a long period and / or treatment of chronic conditions).

[0121] In the context of the present disclosure, it should be understood that long-term treatment encompasses a treatment duration that continues for at least several days, weeks, or months until sufficient fat loss is demonstrated by parameters well known to any physician.

[0122] Furthermore, in the context of some examples of the present disclosure, long-term treatment includes long-term treatment, for example, long-term daily administration, sometimes even without an assumed endpoint for the treatment. In some examples, long-term treatment includes daily administration of an A3AR ligand for at least one week, sometimes for one month, and sometimes for at least 2, 3, 4, 5, 6, or even 12 months of daily administration of the ligand.

[0123] When referring to "treatment" with an A3AR ligand, it should be understood to refer to any desired pharmacological and physiological effects that result in a medically significant improvement in the health status of the subject, as determined by parameters known in the obesity art. For example, improvement can be determined by at least a 1%, and sometimes a 5%, decrease in the level of the subject's total body mass or the level of body fat mass.

[0124] In some examples, the treatment is for subjects who are defined as suffering from a condition associated with excessive peripheral fat accumulation, such as excessive adipose tissue.

[0125] In some examples, the treatment is actually a prophylactic treatment for subjects with a constitution or tendency to gain weight easily. Sometimes treatment with an A3AR ligand can follow or be combined with another treatment known to cause an increase in body mass (e.g., as a side effect).

[0126] The A3AR ligand can be administered daily or at intervals of one day or more between administrations. In one embodiment, the A3AR ligand is used daily for long-term treatment.

[0127] A3AR ligands can be administered systemically or locally. For this purpose, A3AR ligands are combined with a pharmaceutically acceptable carrier to form a pharmaceutical composition suitable for a particular method of administration and containing an effective amount of the A3AR ligand.

[0128] The term "pharmaceutically acceptable carrier" means any one of the inert and non-toxic materials that do not react with the A3AR ligand and can be added to the ligand to facilitate its delivery to the subject.

[0129] In one embodiment, the carrier is acceptable for the preparation of unit dosage forms for oral administration.

[0130] Oral formulations can be in the form of tablets, capsules, syrups, emulsions, aromatic powders, and various other forms. The carrier is sometimes selected based on the desired form of the formulation. The carrier can also have effects such as improving the delivery or penetration of the active ingredient to the target tissue, improving the stability of the drug, slowing the clearance rate, imparting sustained release, and reducing undesirable side effects. The carrier can also be a substance that stabilizes the formulation (such as a preservative), something for imparting an edible flavor to the formulation, etc. The carrier can be any of the conventionally used ones, limited only by chemical and physical considerations such as solubility and lack of reactivity with the A3AR ligand and the route of administration. Examples of carriers include additives, coloring agents, diluents, buffers, disintegrants, wetting agents, preservatives, flavoring agents, pharmacologically compatible carriers, etc. Furthermore, the carrier can be an adjuvant, which, by definition, is a substance that affects the action of the active ingredient in a predictable manner.

[0131] Typical examples of carriers suitable for oral administration include the following. (a) Suitable liquids such as Cremophor RH40, or suspensions or emulsions in methylcellulose (e.g., Methocel A4M Premium), (b) capsules / tablets (e.g., normal hard shell or soft shell gelatin type containing surfactants, lubricants, and inert fillers), tablets, lozenges (where the active substance is in a flavoring such as sucrose and acacia or tragacanth, or the active substance is in an inert base such as gelatin and glycerin), troches, each containing a predetermined amount of tragacanth as a solid or granule, (c) powders, (d) solutions, typically when combined with solubilizing agents, (e) liposome formulations, etc.

[0132] The A3AR ligand is used in an effective amount to achieve at least one fat-reducing effect. The "effective amount" can be readily determined in the present disclosure by administering various amounts of the A3AR ligand to a plurality of test subjects and then plotting the response (e.g., a combination of several beneficial effects) as a function of the amount. Sometimes, the amount used can be influenced by various factors such as the method of administration, the age, weight, body surface area, gender, health status, and genetic factors of the subject, and other administered drugs.

[0133] The effective amount of the A3AR ligand can be defined by the unit dosage form. The term "unit dosage form" refers to physically discrete units suitable for unit administration to human subjects and other mammals, and each unit contains a predetermined amount of the active substance calculated to exert the desired therapeutic effect, together with suitable pharmaceutical excipients.

[0134] When the A3AR ligand is an A3AR agonist, the effective amount can be, for example, an amount of at least about 10 mg / day, e.g., at least about 10 mg for once-daily treatment regimens, at least about 5 mg for twice-daily treatment, at least about 3.3 mg for three-times-daily treatment, etc.

[0135] A dosage of at least about 10 mg / day can be at least about 15 mg / day, at least about 20 mg / day, at least about 25 mg / day. In some embodiments, the dosage is 25±5 mg / day.

[0136] The total amount of the A3AR ligand administered to a patient in a day, regardless of the number of administrations, is referred to herein as the "daily therapeutic dosage".

[0137] Thus, in one example, the A3AR ligand is formulated into a unit dosage form for administering a daily therapeutic dosage of at least 10 mg / day. If the dosage form is intended to be administered to a subject in a treatment regimen that includes n administrations per day, the unit dosage form can include 1 / n of the daily therapeutic dosage (e.g., if the intended daily therapeutic dosage is 20 mg and the treatment regimen is twice a day, each unit dosage form will have a dosage of 10 mg, and if the intended daily therapeutic dosage is 25 mg and the treatment regimen is twice a day, each unit dosage form will have a dosage of 12.5 mg).

[0138] In some examples, the A3AR ligand is administered in combination with an anti-obesity treatment. In the context of the present disclosure, anti-obesity treatment can include any treatment known in the art, including diet plans, special diets, physical activity plans, treatment with drugs, bariatric surgery, weight loss devices.

[0139] In one example, the A3AR ligand is used in combination with a commercially available, medically acceptable weight loss drug. Examples of commercially available FDA-approved drugs include orlistat (Alli, Xenical), lorcaserin (Belviq), phentermine and topiramate (Qsymia), bupropion and naltrexone (Contrave), liraglutide (Saxenda, Victoza).

[0140] It should be understood that in a subject diagnosed with obesity or having a body mass index (BMI) of 30 or more, treatment should be considered effective if there is at least a weight loss of at least 3%, sometimes at least 4% or at least 5% of the total body weight compared to the weight before the start of treatment. In some examples, an effective treatment is one with a weight loss of at least 3% - 6%, or 5% - 10% of the total body weight before the start of treatment.

[0141] As used herein, the forms "a", "an", and "the" include both the singular and the plural unless the context clearly indicates otherwise. For example, the term "A3AR ligand" includes one or more compounds that can directly or indirectly, fully or partially specifically affect the activity of A3AR.

[0142] Furthermore, as used herein, the term "comprising" is intended to mean that the composition includes the recited active substance, i.e., the A3AR ligand, but does not exclude other elements such as physiologically acceptable carriers and excipients and other active substances. The term "consisting essentially of" is used to define a composition that includes the recited elements but excludes other elements that may have an essential importance for any one of the fat-reducing effects. Thus, "consisting of" is meant to exclude other elements in amounts greater than trace amounts. Embodiments defined by each of these transition terms are within the scope of the present invention.

[0143] Furthermore, when referring to the amount or range of the components constituting a composition containing, for example, an A3AR ligand as an active ingredient, all numerical values are approximate values that can vary from the recited value by up to 20% (+) or (-), sometimes up to 10%. It should be understood that although not always explicitly stated, the word "about" precedes all numerical representations.

[0144] Next, the present invention is illustrated in the following description of experiments conducted in accordance with the present invention. It should be understood that these examples are intended to be illustrative in nature and not limiting. Clearly, many modifications and variations of these examples are possible in light of the above teachings. Thus, within the scope of the appended claims, it should be understood that the present invention can be practiced in innumerable possible ways other than the methods specifically described below.

Examples

[0145] Example 1-3 Effect of CF102 on the proliferation of 3T3-L1 adipocytes 3T3-L1 preadipocytes were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). The preadipocytes were maintained in Dulbecco's Modified Eagle Medium (DMEM, HyClone, South Logan, UT, USA) supplemented with 10% newborn calf serum (HyClone) and antibiotics (HyClone) at 37°C and 5% CO2.

[0146] To induce adipocyte differentiation, preadipocytes were seeded in 24-well plates. After reaching confluence, the medium was changed and 10 μg / ml insulin, 1 μM DEX, and 0.5 mM IBMX were added. After 2 days, the medium was changed to a basal medium supplemented with 10% FBS and 10 μg / ml insulin. During the period of 8 - 14 days, the medium was changed every 2 days.

[0147] For the evaluation of cell proliferation, 3 the H-thymidine incorporation assay was used. 3T3-L1 cells (5,000 cells / well) were incubated in 96-well plates with 5 nM or 10 nM CF102 for 48 hours. During the last 24 hours, each well was pulsed with 1 mCi of 3 H-thymidine. The cells were harvested and the amount of 3 H-thymidine incorporation was measured using an LKB liquid scintillation counter (LKB, Piscataway, NJ, USA).

[0148] Results: Figure 1 shows the dose-dependent inhibitory effect of CF102 on the proliferation of 3T3-L1 adipocytes. Adipocytes incubated with 5 nM and 10 nM of CF102 showed decreases of more than 20% and 40%, respectively, compared to the control measurements.

[0149] Example 2 - Effect of CF102 on Lipogenesis in 3T3-L1 Adipocytes Preadipocytes were cultured in a differentiation medium (DMEM high glucose + 10% FBS) containing 10 μg / ml insulin, 1 μM DEX, and 0.5 mM IBMX, and treated with 5 nM CF102 for 48 hours to evaluate the potential of its anti-adipogenic effect.

[0150] Accumulation of lipid droplets was evaluated by staining with Oil Red O. Briefly, cells were washed with PBS, incubated with 3.7% HCHO for 1 hour, and then incubated with Oil Red O solution for 45 minutes. Cells were washed very well to remove excess Oil Red O, visualized under an Olympus microscope with a Leica camera, and photographed. To quantify lipid accumulation, cells were dissolved in isopropanol and the optical density was read at 595 nm using a Microelisa reader from Dynatech Corp. (Chantilly, VA). The degree of accumulation of lipid droplets was proportional to the optical density.

[0151] Accumulation of lipid droplets was visualized under a microscope. Accumulation of lipid droplets is shown in Figures 2A - 2B. Figure 2A shows a microscopic image of the control measurement, and Figure 2B shows a microscopic image of lipid droplets derived from preadipocytes treated with 5 nM CF102.

[0152] Results Figure 3 shows the suppression of lipid accumulation in cells pre-exposed to 5 nM CF102. Data are represented as OD values at 595 nm of the vehicle administration group (control) vs. the CF102 administration group (p = 0.01).

[0153] Example 3 - Effect of CF102 on the Body Weight of High-Fat Diet (HFD) Mice vs. Mice Fed a Normal Diet Complications resulting from obesity, such as diabetes and cardiovascular diseases, usually take decades to develop. Therefore, surrogate animal models are important for studying the molecular aspects of obesity and its pathophysiological effects. One of these increasingly prominent models is the diet-induced obesity model in mice. This protocol is designed to evaluate the effect of CF102 in a murine in vivo model of diet-induced obesity.

[0154] Mouse The mice used were male C57BL / 6J, 4 - 6 weeks old (Jackson Laboratory, stock 000664).

[0155] Diet 60 kcal% fat diet (Research Diets, D12492i)

[0156] CF102 Administration Two types of administration, preventive and therapeutic, will be investigated.

[0157] Experimental Design Groups I - II Naïve animals fed a normal diet (n = 20) were used as the control group and defined as the "Lean Diet". After 12 weeks, the mice were divided into two groups (n = 10 per group). The first group was orally administered CF102 (Canceright Biopharma, catalog number A14402 - 10, 100 μg / Kg) daily for 4 weeks, and the second group was administered only the vehicle. Groups III - IV Naïve mice (n = 20) were fed a HFD for 12 weeks. After 12 weeks, the mice were divided into two groups (n = 10 per group). The first group was orally administered CF102 (Canceright Biopharma, catalog number A14402 - 10, 100 μg / Kg) daily for 4 weeks, and the second group was administered only the vehicle.

[0158] Results and Analysis In the lean diet group, CF102 caused weight loss in the animals compared to the control.

[0159] Specifically, Figure 4A shows that naive mice administered with CF102 under a low-fat diet did not show weight loss compared to non-administered naive mice. However, as shown in Figure 4B, mice under HFD showed a significant (p < 0.001) weight loss effect when administered with CF102 (the arrow indicates the start point of administration).

[0160] Example 4 - Effect of the allosteric modulator CF602 of A3AR on the body weight of diabetic rats Diabetes is one of the complications of obesity. Therefore, the effect of an A3AR ligand on the body weight of a diabetes model was also evaluated.

[0161] Method A diabetes model was created by intraperitoneally injecting streptozotocin (STZ) into Sprague-Dawley male rats (8 - 10 weeks old) at a dose of 60 mg / kg in citrate buffer. Only animals with blood glucose levels higher than 250 mg / dL were included in the study. The allosteric A3AR ligand CF602 was orally administered twice a day for 5 days at a dose of 100 μg / kg. The body weight was monitored 5 days later.

[0162] Results Figure 5 shows the change in body weight from the baseline, that is, the value obtained by subtracting the body weight before administration from the body weight at the end of administration (i.e., the difference). The weight loss was a significant (p < 0.03) decrease in body weight in animals administered with CF602.

Claims

1. (a) reducing the body weight of a subject; (b) reducing body fat mass in a subject; (c) treating obesity in a subject, and (d) inhibiting the proliferation of adipocytes in a subject. A for use in achieving at least one of the fat reducing effects selected from ３ Adenosine receptor (A ３ AR) Ligand.

2. A for use according to claim 1, in a dosage form suitable for daily administration to the subject. ３ AR ligand.

3. 3. The method according to claim 2, wherein the dosage form is suitable for administration once or twice a day. ３ AR ligand.

4. A for use according to any one of claims 1 to 3, which is in a dosage form suitable for oral administration. ３ AR ligand.

5. A ３ A for use according to any one of claims 1 to 4 which is an AR agonist ３ AR ligand.

6. The above A ３ The AR agonist is ６ -2-(4-aminophenyl)ethyl adenosine (APNEA), ６ -(4-amino-3-iodobenzyl)adenosine-5'-(N-methyluronamide) (AB-MECA), ６ -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (IB-MECA) and 2-chloro-N ６ A for the use according to claim 5, selected from the group consisting of -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA). ３ AR ligand.

7. The above A ３ 7. The method according to claim 6, wherein the AR agonist is Cl-IB-MECA. ３ AR ligand.

8. A ３ A for use according to any one of claims 1 to 4 which is an allosteric modulator of AR. ３ AR ligand.

9. The above A ３ Allosteric modulators of the AR N-(3,4-dichloro-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinolin-4-amine, N-(3,4-dichloro-phenyl)-2-cycloheptyl-1H-imidazo[4,5-c]quinolin-4-amine, N-(3,4-dichloro-phenyl)-2-cyclobutyl-1H-imidazo[4,5-c]quinolin-4-amine, and N-(3,4-dichloro-phenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine A for the use according to claim 8, selected from the group consisting of ３ AR ligand.

10. A for the use according to any one of claims 1 to 9 in combination with an anti-obesity treatment ３ AR ligand.

11. A pharma- ceutically acceptable carrier and A as an active ingredient ３ AR ligand - reducing the weight of a subject; - reducing body fat mass in a subject; - treating obesity in a subject; - Inhibiting the proliferation of fat cells in a subject A pharmaceutical composition comprising an effective amount of a compound selected from the group consisting of:

12. The A to the subject ３ 12. The pharmaceutical composition of claim 11 in a dosage form suitable for daily administration of an AR ligand.

13. 13. The pharmaceutical composition of claim 12 in a dosage suitable for administration once or twice daily.

14. A pharmaceutical composition according to any one of claims 11 to 13 for the long-term treatment of said subject.

15. The pharmaceutical composition according to any one of claims 11 to 14, which is in a dosage form suitable for oral administration.

16. The above A ３ AR ligand is A ３ The pharmaceutical composition according to any one of claims 11 to 15, which is an AR agonist.

17. The above A ３ The pharmaceutical composition of claim 16, wherein the AR agonist is Cl-IB-MECA.

18. The above A ３ AR ligand is A ３ The pharmaceutical composition according to any one of claims 11 to 17, which is an allosteric modulator of the AR.

19. The above A ３ 19. The pharmaceutical composition of claim 18, wherein the allosteric modulator of AR is N-(3,4-dichloro-phenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinolin-4-amine (CF602).

20. 1. A method for treating a subject, comprising administering to the subject: - reducing the body weight of said subject; - reducing body fat mass in said subject; - treating obesity in said subject, - Inhibiting the proliferation of fat cells in a subject A. ３ The method comprises administering an AR ligand.

21. 21. The method of claim 20 for treating obesity.

22. A kit comprising: (a) A according to any one of claims 11 to 19 ３ A pharmaceutical composition comprising an AR ligand; (b) The following effects: - reducing the body weight of said subject; - reducing body fat mass in said subject; - treating obesity in said subject, - Inhibiting the proliferation of fat cells in a subject and instructions for use of said pharmaceutical composition to achieve at least one of the following: Including the kit.

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