A3 adenosine receptor ligand for use in achieving fat reduction effects

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] This disclosure relates to the medical applications of A3AR ligand.

[0002] Background technical literature The following is a list of references that are considered relevant as background information for 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] The recognition of the above references in this specification does not imply that those references are in any way related to the patentability of the subject matter disclosed herein. [Background technology]

[0004] Obesity is a complex disorder characterized by an excessive amount of body fat, and in Western societies, it is considered a major health problem, sometimes even life-threatening.

[0005] Obesity results from an imbalance between food intake, basal metabolism, and energy expenditure. At the individual level, multiple endogenous or environmental factors can contribute to obesity. However, in most cases, a combination of excessive calorie intake and the availability of energy-dense meals is considered the primary cause of obesity.

[0006] While weight loss can improve or prevent health problems associated with obesity, it may be necessary to combine weight loss treatments with medication.

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

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

[0009] International Patent Application Publication No. 17 / 090036 describes the use of an A3AR ligand, specifically Cl-IB-MECA, to reduce ectopic fat accumulation, particularly fat accumulation in fatty liver, and specifically for the treatment of non-alcoholic fatty liver disease (NAFLD). [Overview of the project]

[0010] In a first embodiment thereof, this disclosure provides an A3 adenosine receptor (A3AR) ligand for use in any one of the following: - To reduce the target's weight, - To reduce the target body fat mass, - To treat the obesity of the target, - To reduce the level of target fat cells, and - To suppress the proliferation of target fat cells.

[0011] In a second embodiment, this 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. - To reduce the target's weight, - To reduce the target body fat mass, - To treat the obesity of the target, - To reduce the level of target fat cells, and - To suppress the proliferation of target fat cells.

[0012] Furthermore, the present disclosure provides a method of treatment comprising administering to a subject in need an amount of A3AR ligand effective in achieving at least one of the following fat-reducing effects: - To reduce the target's weight, - To reduce the target body fat mass, - To treat the obesity of the target, - To reduce the level of target fat cells, - To suppress the proliferation of target fat cells.

[0013] In some cases, the A3AR ligand is an A3AR agonist, preferably a 2-chloro-N 6 -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA, also known as CF102 herein). [Brief explanation of the drawing]

[0014] Embodiments will be described with reference to the accompanying drawings, merely as non-limiting examples, in order to better understand the subject matter disclosed herein and to illustrate how it can be put into practice. [Figure 1] Figure 1 is a bar graph showing the dose-dependent inhibition of adipocyte proliferation using two different doses of Cl-IB-MECA(CF102), relative to the control group (RPMI+DMSO). [Figure 2] Figures 2A and 2B are microscopic images showing the accumulation of lipid droplets produced by 3T3-L1 adipocytes. Figure 2A shows the accumulation after treatment with the vehicle alone, and Figure 2B shows the level of accumulation after treatment of the cells with 5 nM Cl-IB-MECA. [Figure 3] Figure 3 is a bar graph showing the optical concentration of lipid droplet accumulation generated by 3T3-L1 adipocytes after treatment with 5 nM Cl-IB-MECA, compared to the control group (DMEM high glucose + 10% FBS). [Figure 4A]Figures 4A-4B show the changes in body weight of mice fed a normal diet during the study period. CF102 or vehicle-based administration was started at week 12 (Figure 4A). Alternatively, the changes in body weight of mice fed a high-fat diet (HFD) throughout the study period are also shown. [Figure 4B] Figures 4A-4B show the changes in body weight of mice fed a normal diet during the study period. Administration of CF102 or vehicle was started at week 12 (Figure 4B). [Figure 5] Figure 5 is a graph showing the effect of CF602, an allosteric modulator of A3AR, on body weight in an experimental rat model of diabetes induced by STZ. [Modes for carrying out the invention]

[0015] This disclosure relates to 2-chloro-N, an A3 adenosine receptor (A3AR) agonist that exhibits high affinity and selectivity for A3ARs. 6 This is based on the finding that -(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA, also known herein as CF102) inhibited adipocyte proliferation 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 reduction in the accumulation of lipids produced in 3T3-L1 adipocytes (Figures 2A-2B and 3).

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

[0018] Furthermore, surprisingly, weight loss was also observed in animal models of diabetes, a known complication of obesity, when treated with the A3AR allosteric modulator N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinoline-4-amine (sometimes abbreviated as LUF6000 or CF602).

[0019] Based on these findings, the inventors have concluded that ligands with high affinity and selectivity for A3AR, whether A3AR-selective agonists or A3AR-selective allosteric modulators, are effective tools for achieving therapeutically beneficial effects in any one or combination of the following ways. - To reduce the target's weight, - To reduce the target body fat mass, - To treat the obesity of the target, - To reduce the level of target fat cells, and - To suppress the proliferation of target fat cells.

[0020] While not bound by theory, the above effects are thought to be directly or indirectly linked by the inhibitory effect of ligands on the proliferation of fat cells, and are collectively referred to as the "fat reduction effect."

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

[0022] Accordingly, this disclosure provides an A3AR ligand for use in achieving any of the above-described fat-reducing effects, a pharmaceutical composition comprising the A3AR ligand for use in achieving any of the above-described fat-reducing effects, and a method for treating a subject requiring any one of the above-described fat-reducing effects, the treatment also encompassing the prevention of fat increase, as further described below.

[0023] In one example, A3AR ligands are intended for use in treating subjects suffering from obesity.

[0024] In another example, A3AR ligands are intended for use in inducing or promoting weight loss in subjects.

[0025] In yet another example, A3AR ligands are intended to be used to reduce the levels of target adipocytes.

[0026] In yet another example, A3AR ligands are used to inhibit the proliferation of adipocytes.

[0027] In the context of this disclosure, when referring to weight loss or treatment of obesity, it should be understood that this is equivalent to reducing fat-containing tissue, such as adipose tissue, that is, subcutaneous and / or peripheral fat, and eliminating ectopic fat, such as ectopic fat in liver tissue. In some examples, this disclosure excludes the reduction of ectopic adipose tissue, particularly adipose tissue in the liver.

[0028] In one preferred example, A3AR ligands are used to treat subjects with excess body fat, particularly those suffering from obesity.

[0029] In some cases, a decrease in body fat mass is indicated by a decrease in peripheral fat, particularly adipose tissue.

[0030] In several other cases, a decrease in body fat mass is indicated by a decrease in lipid production from 3T3-L1 adipocytes.

[0031] Excess body fat can be determined by body mass index (BMI). Individuals with a BMI above 25 are considered to have excess body fat, and those with a BMI above 30 are considered obese.

[0032] The effect of the treatment, for example, a reduction in the quantifiable quantities described herein, such as body weight, body fat mass, and / or the level of adipocytes, can be determined by the values ​​obtained at two different time points. In some cases, the first time point is before treatment and the second time point is during treatment.

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

[0034] When referring to a decrease in one of the measured values, it should be understood that the decrease is considered significant by the physician. This decrease could be 1%, 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or sometimes more than 70% compared to a given reference point. Note that there may be multiple points in time between the reference point and the evaluation point where the value is measured.

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

[0036] In some cases, the evaluation time is a predetermined point in time, which is the end of treatment. In other cases, the evaluation time is a point in treatment after the reference time.

[0037] It should be noted that between those two points in time, the subjects received A3AR ligand. However, treatment with A3AR ligand does not need to be long-term, for example, for the entire duration; subjects may receive it periodically.

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

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

[0040] The A3AR ligands described herein are molecules that exert their primary effects through the activation of A3AR, regardless of whether their activation is via a binding site or an allosteric binding site.

[0041] A3AR ligands have high affinity and selectivity for A3AR. This means that at the dosage in 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 more beneficial effects than activation of other adenosine receptors, such as the A1 adenosine receptor, which is known to have cardiovascular side effects not present when using A3AR ligands. In fact, studies have shown that A3AR ligands have protective effects, particularly neuroprotective, chemoprotective, cardioprotective, and hepatoprotective effects.

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

[0044] In another example, "A3 adenosine receptor ligand" is an allosteric modulator (also known as an allosteric effector) of A3AR.

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

[0046] In the context of this disclosure, its affinity for A3AR is compared to any other adenosine receptor (i.e., A1, A3AR). 2a and A 2b A molecule is considered an A3AR agonist (i.e., a molecule that exerts its primary effect through the binding and activation of A3AR) if its affinity for A3AR is at least three times greater (i.e., its Ki to A3AR is at least three times lower), preferably 10 times, more preferably 20 times, and most preferably at least 50 times greater.

[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 radioactive 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 is sometimes measured through an increase or decrease in cAMP levels. When 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, activation of those receptors may occur following such administration in addition to activation of the A3AR. Thus, 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 that is 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 that is 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 several A3AR agonists used in accordance with this disclosure and the methods for preparing them are described in detail, in particular, in U.S. Patents 5,688,774, 5,773,423, 5,573,772, 5,443,836, 6,048,865, International Publications 95 / 02604, 99 / 20284, 99 / 06053, 97 / 27173, and 01 / 19360. All of these documents are incorporated herein by reference.

[0052] According to some examples of this disclosure, A3AR agonists are purine derivatives that fall within the range of the following general formula (I).

[0053] TIFF0007894175000001.tif36170

[0054] During the ceremony, -R 11 This represents an alkyl, hydroxyalkyl, carboxyalkyl, or cyanoalkyl group, or a group of the following general formula (II):

[0055] TIFF0007894175000002.tif28170

[0056] During the ceremony, -Y represents oxygen, sulfur, or CH2. -X 11 H, alkyl, R e R f NC(=O)- or HOR g - represents, and in the formula, -R e and R f These may be the same or different, selected from the group consisting of hydrogen, alkyl, amino, haloalkyl, aminoalkyl, BOC-aminoalkyl, and cycloalkyl, or bonded together to form a heterocycle containing 2 to 5 carbon atoms. -R g This 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 Each of these independently represents hydrogen, hydroxyl, amino, amide, azide, halo, alkyl, alkoxy, carboxy, nitrilo, nitro, trifluoro, aryl, alkalil, thio, thioester, thioether, -OCOPh, -OC(=S)OPh, or X 13 and X 14 Both are oxygen atoms that link to >C=S to form a 5-membered ring, or X 12 and X 13 This forms the ring of equation (III),

[0057] TIFF0007894175000003.tif31170

[0058] In the formula, R' and R'' independently represent alkyl groups. -R 12 This 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, formula -NR 15 R 16 It is the basis of, and in the formula, -R 15 is a hydrogen atom or a group selected from alkyl, substituted alkyl, or aryl-NH-C(Z)-, where Z is O, S, or NR a And R e The above has the meaning, and therein R 15 If it is hydrogen, -R 16R- and S-1-phenylethyl, benzyl, phenylethyl or anilide groups substituted with substituents selected from the group consisting of alkyl, amino, halo, haloalkyl, nitro, hydroxyl, acetamide, alkoxy, and sulfonic acid or salts thereof at one or more positions, benzodioxanemethyl, fururyl, L-propylalanyl-aminobenzyl, β-alanylaminobenzyl, T-BOC-β-alanylaminobenzyl, phenylamino, carbamoyl, phenoxy or cycloalkyl, or R 16 is the basis of equation (IV),

[0059] TIFF0007894175000004.tif32170

[0060] or R 15 If R is alkyl or aryl-NH-C(Z)-, 16 is heteroaryl-NR a -C(Z)-, heteroaryl-C(Z)-, alkalil-NR a The group consists of -C(Z)-, alkal-C(Z)-, aryl-NR-C(Z)-, and aryl-C(Z)-, where Z represents oxygen, sulfur, or amine.

[0061] Exemplary A3AR agonists (disclosed in columns 4, 67-6, 16; 5, 40-45; 6, 21-42; 7, 1-11; 7, 34-36; and 7, 60-61 of U.S. Patent No. 5,688,774): N 6 -(3-iodobenzyl)-9-methyladenine, N 6 -(3-iodobenzyl)-9-hydroxyethyladenine, RN 6 -(3-iodobenzyl)-9-(2,3-dihydroxypropyl)adenine, SN 6 -(3-iodobenzyl)-9-(2,3-dihydroxypropyl)adenine, N 6-(3-iodobenzyladenine-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-methylthioadene, N 6 -(3-iodobenzyl)-9-methyl-2-(4-pyridylthio)adenine, (1S, 2R, 3S, 4R)-4-(6-amino-2-phenylethylamino-9H-purine-9-yl)cyclopentan-1,2,3-triol, (1S, 2R, 3S, 4R)-4-(6-amino-2-chloro-9H-purine-9-yl)cyclopentan-1,2,3-triol, (±)-9-[2α,3α-dihydroxy-4β-(N-methylcarbamoyl)cyclopenta-1β-yl)]-N 6 -(3-iodobenzyl)-adenine, 2-Chloro-9-(2'-amino-2',3'-dideoxy-β-D-5'-methyl-arabino-fronamide)-N 6-(3-iodobenzyl)adenine, 2-Chloro-9-(2',3'-dideoxy-2'-fluoro-β-D-5'-methyl-arabinoflonamide)-N 6 -(3-iodobenzyl)adenine, 9-(2-acetyl-3-deoxy-β-D-5-methyl-ribofronamide)-2-chloro-N 6 (3-iodobenzyl)adenine, 2-Chloro-9-(3-deoxy-2-methanesulfonyl-β-D-5-methyl-ribofronamide)-N 6 -(3-iodobenzyl)adenine, 2-Chloro-9-(3-deoxy-β-D-5-methyl-ribofronamide)-N 6 -(3-iodobenzyl)adenine, 2-Chloro-9-(3,5-1,1,3,3-tetraisopropyldisiloxyl-β-D-5-ribofuranosyl)-N 6 -(3-iodobenzyl)adenine, 2-Chloro-9-(2',3'-O-thiocarbonyl-β-D-5-methyl-ribofronamide)-N 6 -(3-iodobenzyl)adenine, 9-(2-phenoxythiocarbonyl-3-deoxy-β-D-5-methyl-ribofronamide)-2-chloro-N 6 -(3-iodobenzyl)adenine, 1-(6-benzylamino-9H-purine-9-yl)-1-deoxy-N,4-dimethyl-β-D-ribofuranosiduronamide 2-Chloro-9-(2,3-dideoxy-β-D-5-methyl-ribofronamide)-N 6 benzyladenine, 2-Chloro-9-(2'-azido-2',3'-dideoxy-β-D-5'-methyl-arabino-fronamide)-N 6 -benzyladenine, 2-Chloro-9-(β-D-erythrofuranoside)-N 6 -(3-iodobenzyl)adenine, N6 -(benzodioxanemethyl)adenosine, 1-(6-furfurylamino-9H-purine-9-yl)-1-deoxy-N-methyl-β-D-ribofuranosiduramide, N 6 -[3-(L-prolylamino)benzyl]adenosine-5'-N-methyluronamide, N 6 -[3-(β-alanylamino)benzyl]adenosine-5'-N-methyluronamide, N 6 -[3-(NT-Boc-β-alanylamino)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-alanylamino)benzyl]adenosine, 9-(β-D-erythrofuranoside)-2-methylamino-N 6 -(3-iodobenzyl)adenine, 2-Chloro-N-(3-iodobenzyl)-9-(2-tetrahydrofuryl)-9H-purine-6-amine, 2-Chloro-(2'-deoxy-6'-thio-L-arabinosyl)adenine, and 2-Chloro-(6'-thio-L-arabinosyl)adenine.

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

[0063] TIFF0007894175000005.tif70170

[0064] During the ceremony, 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, acetamide, 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, acetamide, C1-C 10 haloalkyl, and sulfo, and the sulfo derivative is a salt such as a triethylammonium salt, and they are those A3AR agonists.

[0069] Furthermore, 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, particularly compounds in which R2 is halo, C1-C 10 alkylamino, or C1-C 10 alkylthio, more preferably, further 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] TIFF0007894175000006.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 groups, R7 and R8 may be the same or different, C1-C 10 Alkyl, R- and S-1-phenylethyl, unsubstituted benzyl group, and C1-C at one or more positions. 10 Alkyl, amino, halo, C1-C 10 Haloalkyl, nitro, hydroxy, acetamide, C1-C 10 Selected from the group consisting of benzyl groups substituted with substituents selected from the group consisting of alkoxys and sulfo, R9 is a compound of halo, benzyl, phenyl, and C3-C 10 Selected from the group consisting of cycloalkyl groups.

[0074] International Publication No. 99 / 06053 discloses 19 to 33 compounds selected from the following examples. 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-sulfonamide-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-isoxazole-3-yl-carbamoyl)-adenosine-5'-N-ethyluronamide, N 6 -(1,3,4-thiadiazole-2-ylcarbamoyl)-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-chloropyridine-2-ylcarbamoyl)-adenosine-5'-N-ethyluronamide.

[0075] More specifically, the A3AR agonists to be used in accordance with this disclosure include the following: 2-Chloro-N 6 -(3-iodobenzyl)-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine,2-chloro-N 6 Also known as -(3-iodobenzyl)-adenosine-5'-N-methyluronamide, or by the abbreviation Cl-IB-MECA, N 6 -(3-iodobenzyl)-2-methylamino-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine, N 6It is also known as -(3-iodobenzyl)-adenosine-5'-N-methyluronamide, 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purine-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] A particularly preferred example of an A3AR agonist is 2-chloro-N 6 -(3-iodobenzyl)-2-methylamino-9-[5-(methylamide)-β-D-ribofuranosyl]-adenine,2-chloro-N 6 It is also known as -(3-iodobenzyl)-adenosine-5'-N-methyluronamide, or by the abbreviation Cl-IB-MECA.

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

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

[0079] When referring to "A3AR allosteric modulators" or "A3ARMs," it should be understood that this refers to the positive regulation, activation, or increase of receptor activity resulting from the binding of an allosteric modulator to the receptor's allosteric site, which may differ from the binding site of the endogenous ligand or its agonist.

[0080] In one example, “modulation” refers to the effect of an A3AR ligand on the receptor, which is manifested by an increase of at least 15% in the potency of the A3 adenosine receptor due to the compound binding to the allosteric site of the receptor, and / or by a decrease in the dissociation rate of adenosine or an A3AR agonist to the orthosteric binding site.

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

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

[0083] TIFF0007894175000007.tif50170

[0084] During the ceremony, -R1 is optionally C1-C in the aromatic ring. 10 Alkyl, Halo, C1-C 10 Alkanol, Hydroxyl, C1-C 10 Ashiru, C1-C 10 Alkoxyl, C1-C 10 -alkoxycarbony, C1-C 10 Alkoxyalkyl, C1-C 10 Thioalkoxy, C1-C 10 Alkyl ethers, amino acids, hydrazides, C1-C 10 Alkylamino, pyridylthio, C2-C 10 Alkenyl, C2-C 10 Alkinyl, Thio, C1-C 10 Represents an aryl or alkaryl substituted with one or more substituents selected from the group consisting of alkylthio, acetamide, and sulfonic acid, or the substituents may together form a cycloalkyl or cycloalkenyl condensed with the aryl, the cycloalkyl or cycloalkenyl optionally containing one or more heteroatoms (provided the aryl is not an unsubstituted phenyl group), -R2 is hydrogen or C1-C 10 Alkyl, C2-C 10 Alkenyl, C2-C 10 Alkinyl, C4-C 10 Cycloalkyl, C4-C 10 Cycloalkenyl, a heterocyclic aromatic ring with 5-7 members, C5-C 15 Condensed cycloalkyl, bicyclic aromatic ring or heteroaromatic ring, C1-C 10 Alkyl ethers, amino acids, hydrazides, C1-C 10 Alkylamino, C1-C 10 Alkyl, C1-C 10 -alkoxycarbon, C1-C 10 Alkanol, C1-C 10 Ashiru, C1-C 10 Thioalkoxy, pyridylthio, thio, and C1-C 10 Alkylthio, acetamide, and sulfonic acid, This represents substituents selected from the group consisting of and pharmaceutically acceptable salts thereof.

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

[0086] TIFF0007894175000008.tif34170

[0087] In the formula, 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 halogens, alkyls, alkanols or alkoxys, indanyls, and pyrrolines, 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), where X1 or X2 may be the same or different, and is selected from hydrogen, chloro, methoxy, methanol, or substituents having the formula (VIIIa) or (VIIIb).

[0089] TIFF0007894175000009.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 cycloalkyls, the alkyl chain may be linear, branched, or form a 4-7 membered cycloalkyl ring.

[0092] In one example, R2 of A3ARAM is selected from a 5- to 7-membered heterocyclic aromatic ring.

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

[0094] TIFF0007894175000010.tif30170

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

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

[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] Furthermore, in several other examples, R2 of A3ARAM is selected from 2-cyclohexene and 3-cyclohexene.

[0099] The following is a list of specific imidazoquinoline derivatives that can be used as allosteric modulators for A3AR. N-(4-methylphenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(4-methoxyphenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(4-chlorophenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3-methanol-phenyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-([3,4-c]indan)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(1H-indazole-6-yl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(4-methoxybenzyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(1H-indole-6-yl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(benzyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(phenylethyl)-2-cyclopentyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-cycloheptyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-furyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-cyclobutyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2--1H-imidazo[4,5-c]quinoline-4-amine N-(3,4-dichlorophenyl)-2-pentyl-1H-imidazo[4,5-c]quinoline-4-amine

[0100] The above imidazoquinoline derivatives are, on the one hand, A1 and A 2A , A 2B The affinity for the orthosteric binding site of the adenosine receptor is reduced, if any, and the affinity for the orthosteric binding site of the A3 adenosine receptor is reduced, while on the other hand, the affinity for the allosteric site of the A3 adenosine receptor is high, and therefore it is considered an allosteric modulator (International Patent Application Publication No. 07 / 089507, incorporated herein by reference).

[0101] In this disclosure, a specifically preferred imidazoquinoline derivative is N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinoline-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 formulas disclosed herein, the following meanings are considered for various terms:

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

[0104] Similarly, the terms “alkenyl” and “alkynyl” refer to a straight 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, respectively, where an alkenyl or alkynyl has at least one unsaturated bond.

[0105] Alkyl, alkenyl, or alkynyl substituents may be substituted with heteroatom-containing groups. Therefore, although not explicitly stated, it should be understood that any of the alkyl modifications defined above and below herein, such as alkylthio, alkoxy, akanol, and alkylamine, also include the corresponding alkenyl or alkynyl modifications, such as akenylthio, akenyloxy, alkenol, alkenylamine, or akinylthio, alkynyloxy, alkinol, and alkynylamine, respectively.

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

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

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

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

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

[0111] The term "alkanol" refers to the -COH group and the alk-OH group, and "alk" means alkylene, alkenylene, or alkynylene chain.

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

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

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

[0115] The term "cycloalkyl" is used herein to mean a cyclic hydrocarbon radical, and examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl.

[0116] The term "alkoxycarbonyl" is used herein to mean -C(O)O-alkyl, and examples include, but are not limited to, methoxycarbonyl, ethoxycarbonyl, and propoxycarbonyl.

[0117] The term "condensed cycloalkyl" is used herein to mean any compound or substituent comprising at least two aliphatic rings linked at just one atom (to form a spiro ring moiety), at two mutually bonded atoms, or across a series of atoms (bridgeheads). The condensed ring may include any bicyclic, tricyclic, or polycyclic moiety. In some embodiments of this disclosure, bicyclic substituents are preferred.

[0118] This disclosure also utilizes physiologically acceptable salts of A3AR-selective ligands such as the compounds described above. “Physiologically acceptable salts” refers to any non-toxic alkali metal, alkaline earth metal, or ammonium salt commonly used in the pharmaceutical industry, including sodium, potassium, lithium, calcium, magnesium, barium ammonium, and protamine zinc salts, which are prepared by methods known in the art. The term also encompasses non-toxic acid addition salts, which are commonly prepared by reacting ligands with suitable organic or inorganic acids. Acid addition salts retain the biological effects and qualitative properties of the free base and are non-toxic, or otherwise undesirable. Examples include, among others, acids derived from mineral acids, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and metaphosphoric acid. Examples of organic acids include, in particular, 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, aryl sulfonic acid, such as p-toluenesulfonic acid.

[0119] A3AR ligands can be administered as a single dose (one-time medication) or as part of a continuous treatment over several days, weeks, or even months.

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

[0121] In the context of this disclosure, long-term treatment should be understood to encompass a treatment time range of at least several days, weeks, or months, until sufficient fat reduction is demonstrated by parameters well known to any physician.

[0122] Furthermore, in the context of some examples of this disclosure, long-term treatment includes treatment over a long period, such as long-term daily administration, sometimes even without an expected endpoint for treatment. In some examples, long-term treatment includes daily administration of A3AR ligand for at least one week, sometimes daily administration for one month, and sometimes daily administration of ligand for at least two, three, four, five, six, or even twelve months.

[0123] When referring to "treatment" with A3AR ligands, it should be understood that this refers to any desired pharmacological and physiological effect that results in a medically significant improvement in the subject's health status, as judged by parameters known in the obesity industry (world). For example, improvement can be determined by a reduction of at least 1%, sometimes 5%, in the subject's total body mass level or body fat level.

[0124] In some cases, treatment is defined as being for individuals suffering from a condition associated with excessive peripheral fat accumulation, such as an excess of adipose tissue.

[0125] In some cases, the treatment is actually a preventative measure for individuals with a predisposition or tendency to gain weight easily. Sometimes, treatment with A3AR ligands can follow or be combined with other treatments that are known to cause increased body mass (e.g., as a side effect).

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

[0127] A3AR ligands can be administered systemically or topically. For this purpose, A3AR ligands are combined with pharmaceutically acceptable carriers to form pharmaceutical compositions that are suitable for specific administration methods and contain an effective amount of A3AR ligand.

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

[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. Carriers are sometimes selected based on the desired form of the formulation. Carriers may also have effects such as improving the delivery or penetration of the active ingredient to target tissue, improving drug stability, slowing the clearance rate, providing sustained release, or mitigating undesirable side effects. Carriers may also be substances that stabilize the formulation (e.g., preservatives), or substances that impart an edible flavor to the formulation. Carriers may be any conventionally used ones, limited only by chemical and physical considerations such as solubility and non-reactivity with A3AR ligands, and by the route of administration. Examples of carriers include additives, colorants, diluents, buffers, disintegrants, wetting agents, preservatives, flavorings, and pharmacologically suitable carriers. Furthermore, carriers may be adjuvants, which, by definition, are substances that predictably influence the action of the active ingredient.

[0131] Typical examples of carriers suitable for oral administration include: (a) suspensions or emulsions in suitable liquids such as Cremofor RH40, or in methylcellulose (e.g., Methocel A4M Premium); (b) capsules / prescriptions (e.g., typical hard-shell or soft-shell gelatin types containing surfactants, lubricants, and inert fillers), tablets, lozenges (where the active substance is in sucrose and flavorings such as acacia or in tragacanth, or in an inert base such as gelatin and glycerin), troches, each containing a predetermined amount of tragacanth as solid or granules; (c) powders; (d) solutions, typically in combination with a dissolution accelerator; (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 this disclosure by administering different amounts of the A3AR ligand to multiple 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 may depend on various factors such as the method of administration, the subject’s age, weight, body surface area, sex, health status and genetic factors, and other administered drugs.

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

[0134] When the A3AR ligand is an A3AR agonist, the effective dose may be, for example, at least about 10 mg / day, for example, at least about 10 mg in a once-daily treatment regimen, at least about 5 mg in a twice-daily regimen, and at least about 3.3 mg in a three-times-daily regimen.

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

[0136] In this specification, the total amount of A3AR ligand administered to a patient per day, regardless of the number of doses, is referred to as the "daily therapeutic dose."

[0137] Therefore, in one example, the A3AR ligand is formulated into a unit dosage form to administer a daily therapeutic dose of at least 10 mg / day. If the dosage form is intended to be administered to a subject in a treatment regimen involving n doses per day, the unit dosage form may contain 1 / n of the daily therapeutic dose (for example, if the intended daily therapeutic dose is 20 mg and the treatment regimen is twice daily, each unit dosage form would have a dose of 10 mg; if the intended daily therapeutic dose is 25 mg and the treatment regimen is twice daily, each unit dosage form would have a dose of 12.5 mg).

[0138] In some cases, A3AR ligands are administered in combination with anti-obesity treatments. In the context of this disclosure, anti-obesity treatments may include any treatment known in the art, including dietary plans or special diets, physical activity plans, drug therapy, weight-loss surgery, and weight-loss devices.

[0139] In some cases, A3AR ligands are used in combination with commercially available, medically acceptable weight-loss medications. Examples of commercially available, FDA-approved medications include orlistat (Alli, Xenical), lorcaserin (Belviq), phentermine and topiramate (Qsymia), bupropion and naltrexone (Contrave), and liraglutide (Saxenda, Victoza).

[0140] It should be understood that in individuals diagnosed with obesity or with a body mass index (BMI) of 30 or higher, treatment should be considered effective if there is a weight loss of at least 3%, sometimes at least 4%, or at least 5% of total body weight compared to the weight before treatment. In some cases, effective treatment is defined as a weight loss of at least 3% to 6%, or 5% to 10%, of total body weight compared to the weight before treatment.

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

[0142] Furthermore, as used herein, the term “comprising” is intended to mean that the composition contains the described active substance, namely the A3AR ligand, but does not exclude other elements such as physiologically acceptable carriers and excipients and other active substances. The term “essentially consisting of” is used to define a composition that contains the described elements but excludes other elements that may be of essential importance to any one of the fat-reducing effects. Thus, “consisting of” shall mean excluding 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 quantity or range of components constituting a composition containing, for example, A3AR ligand as an active ingredient, all numerical values ​​are approximations that may vary by up to 20% (+) or (-) from the stated value, and sometimes up to 10%. It should be understood that, although not always explicitly stated, the word "approximately" 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, not limiting. Obviously, many modifications and variations of these examples are possible in light of the above teachings. Therefore, it should be understood that within the scope of the appended claims, the present invention can be carried out in countless possible ways other than those specifically described below. [Examples]

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

[0146] To induce adipocyte differentiation, preadipocytes were seeded in 24-well plates. After 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 basal medium supplemented with 10% FBS and 10 μg / ml insulin. The medium was changed every 2 days for a period of 8 to 14 days.

[0147] For evaluating cell proliferation, 3 An H-thymidine uptake assay was used. 3T3-L1 cells (5,000 cells / well) were incubated in a 96-well plate with 5 nM or 10 nM CF102 for 48 hours. For the last 24 hours, each well was treated with 1 mCi of CF102. 3 The cells were pulsed with H-thymidine. Cells were collected and measured using an LKB liquid scintillation counter (LKB, Piscataway, New Jersey, USA). 3 The amount of H-thymidine taken up was measured.

[0148] result: 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 CF102 showed reductions of over 20% and 40%, respectively, compared to control measurements.

[0149] Example 2-3: Effect of CF102 on lipid synthesis in T3-L1 adipocytes Preadipocytes were cultured in differentiation medium (DMEM high glucose + 10% FBS) containing 10 μg / ml insulin, 1 μM DEX, and 0.5 mM IBMX, treated with 5 nM CF102 for 48 hours, and their potential anti-lipid activity was evaluated.

[0150] Lipid droplet accumulation was assessed by staining with Oil Red O. Briefly, cells were washed with PBS, incubated in 3.7% HCHO for 1 hour, and then incubated in Oil Red O solution for 45 minutes. Cells were thoroughly washed 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 optical density was read at 595 nm using a Dynatech Corp. Microelisa reader (Chantilly, Virginia). The degree of lipid droplet accumulation was proportional to the optical density.

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

[0152] result Figure 3 shows the suppression of lipid accumulation in cells pre-exposed to 5 nM CF102. The data are expressed as OD values ​​at 595 nm for the vehicle administration group (control) versus the CF102 administration group (p=0.01).

[0153] Example 3 - Effect of CF102 on body weight of mice fed a high-fat diet (HFD) versus mice fed a normal diet Obesity-related complications such as diabetes and cardiovascular disease typically take decades to develop, making surrogate animal models crucial for studying the molecular aspects of obesity and its pathophysiological effects. One such model gaining increasing attention is the mouse diet-induced obesity model. This protocol is designed to evaluate the effects of CF102 in an in vivo mouse model of diet-induced obesity.

[0154] mouse The mouse used was a C57BL / 6J male, 4-6 weeks old (Jackson Laboratory, stock 000664).

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

[0156] CF102 administration We will investigate two types of administration: prophylactic and therapeutic.

[0157] Experimental Design Groups I and II consisted of naive animals (n=20) fed a standard diet, which 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). Group 1 was orally administered CF102 (Canfite BioPharma, catalog number A14402-10, 100 μg / kg) daily for 4 weeks, while Group 2 was administered only the vehicle. Groups III-IV naive mice (n=20) were fed HFD for 12 weeks. After 12 weeks, the mice were divided into two groups (n=10 per group). Group 1 was orally administered CF102 (Canfite BioPharma, catalog number A14402-10, 100 μg / kg) daily for 4 weeks, while Group 2 was administered only the vehicle.

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

[0159] Specifically, Figure 4A shows that naive mice administered CF102 under a low-fat diet did not show weight loss compared to unadministered naive mice. However, as shown in Figure 4B, mice under HFD (High Fat Depletion) showed a significant weight loss effect (p<0.001) when administered CF102 (arrows indicate the starting point of administration).

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

[0161] method A diabetic model was created in male Sprague Dolly rats (8-10 weeks old) by intravenous administration of streptozotocin (STZ) 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 at a dose of 100 μg / kg twice daily for 5 days. Body weight was monitored after 5 days.

[0162] result Figure 5 shows the change in body weight from baseline, i.e., the difference between body weight at the end of administration and body weight before administration. Body weight loss was significant (p<0.03) in animals administered CF602.

Claims

1. A pharmaceutical composition for reducing the body weight of a subject and for inhibiting the proliferation of fat cells of the subject, wherein the pharmaceutical composition comprises (i) a pharmaceutically acceptable carrier and (ii) an active ingredient A ３ Contains AR ligand, A ３ The AR ligand is an A3 AR agonist selected from 2-chloro-N6-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA); or A3 is an AR allosteric modulator selected from N-(3,4-dichlorophenyl)-2-cyclohexyl-1H-imidazo[4,5-c]quinoline-4-amine. A pharmaceutical composition characterized by the following features.

2. In the pharmaceutical composition according to claim 1, the pharmaceutical composition provides the A to the target. ３ A pharmaceutical composition characterized by being a dosage form suitable for daily administration of an AR ligand.

3. A pharmaceutical composition according to claim 2, characterized in that the pharmaceutical composition is in a dosage suitable for administration once or twice a day.

4. A pharmaceutical composition according to any one of claims 1 to 3 for the long-term treatment of the subject mentioned above.

5. A pharmaceutical composition according to any one of claims 1 to 4, characterized in that the pharmaceutical composition is in a dosage form suitable for oral administration.

6. A kit for reducing the weight of a subject and for suppressing the proliferation of fat cells in the subject, (a) The pharmaceutical composition according to any one of claims 1 to 5, (b) Instructions for the use of the pharmaceutical composition for reducing the weight of the subject and for inhibiting the proliferation of fat cells in the subject. A kit characterized by containing the following.

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