WO2016057656A1 - Ppar-delta agonists for use for treating mitochondrial, vascular, muscular, and demyelinating diseases - Google Patents

Abstract

Provided herein are methods for increasing PPAR5 activity and methods for treating PPARδ related diseases (e.g., mitochondrial diseases, muscular diseases, vascular diseases, demyelinating diseases, and metabolic diseases).

Description

PPAR-DELTA AGONISTS FOR USE FOR TREATING MITOCHONDRIAL, VASCULAR, MUSCULAR, AND

DEMYELINATING DISEASES

CROSS-REFERENCE TO RELATED APPLICATIONS This application claims benefit of priority to U.S. Provisional Patent Application No.

62/061,446, filed October 8, 2014. The content of this application is incorporated herein by reference in its entirety.

FIELD

This application concerns methods of treating mitochondrial, vascular, muscular, and demyelinating diseases, and other related conditions, with PPAR delta (PPAR5) agonists.

ACKNOWLEDGMENT OF GOVERNMENT SUPPORT

This invention was made with government support under DK057978-32 awarded by the National Institutes of Health. The United States government has certain rights in the invention.

BACKGROUND

Mitochondrial diseases can be very debilitating and can substantially reduce the quality of life of the patient. Accordingly, there is a great need in the art for novel methods of effectively and reliably mitochondrial diseases. The present invention addresses these and other such needs.

SUMMARY Provided herein, inter alia, are methods of using PPAR5 agonists for treating PPAR5- related diseases {e.g., mitochondrial diseases, muscular diseases, demyelinating diseases, and vascular diseases). In particular, disclosed herein are methods modulating the activity of PPAR5 for the treatment of diseases, developmental delays, and symptoms related to mitochondrial dysfunction (see, e.g., Examples 1-7). For example, the disclosed compounds and compositions are useful in the treatment of mitochondrial diseases, such as Alpers's Disease, CPEO-Chronic progressive external ophthalmoplegia, Kearns-Sayra Syndrome (KSS), Leber Hereditary Optic Neuropathy (LHON), MELAS -Mitochondrial myopathy, encephalomyopathy, lactic acidosis, and stroke-like episodes, MERRF-Myoclonic epilepsy and ragged-red fiber disease, NARP-neurogenic muscle weakness, ataxia, and retinitis pigmentosa, and Pearson Syndrome.

In one embodiment, provided herein is a method of treating a PPAR5-related disease or condition in a subject, comprising administering to the subject a therapeutically effective amount of a PPAR5 agonist, wherein the PPAR5-related disease or condition is a vascular disease, a muscular disease, or a demyelinating disease.

In another embodiment, provided herein is a method of treating a PPAR5-related disease or condition in a subject, comprising administering to the subject a therapeutically effective amount of a PPAR5 agonist, wherein the PPAR5-related disease or condition is a muscle structure disorder, a neuronal activation disorder, a muscle fatigue disorder, a muscle mass disorder, a mitochondrial disease, a beta oxidation disease, a vascular disease, an ocular vascular disease, or a muscular eye disease.

In yet another embodiment, provided herein is a method of increasing or maintaining muscle mass or muscle tone in a subject, comprising administering to the subject a therapeutically effective amount of a PPAR5 agonist.

Also provided herein is the use of one or more PPAR5 agonists, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising one or more PPAR5 agonists, for the preparation of a medicament for the treatment of a PPAR5-related disease or condition, the PPAR5-related disease or condition is a vascular disease, a muscular disease, or a demyelinating disease. Alternatively, the use of the PPAR5 agonists is intended for the treatment of a subject with PPAR5-related disease or condition selected from a muscle structure disorder, a neuronal activation disorder, a muscle fatigue disorder, a muscle mass disorder, a mitochondrial disease, a beta oxidation disease, a vascular disease, an ocular vascular disease, or a muscular eye disease. Alternatively, the use of the PPAR5 agonists is intended for increasing or maintaining muscle mass or muscle tone in a subject.

In another embodiment, provided herein one or more PPAR5 agonists, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising one or more PPAR5 agonists, for use in a method for treating a PPAR5-related disease or condition selected from a muscle structure disorder, a neuronal activation disorder, a muscle fatigue disorder, a muscle mass disorder, a mitochondrial disease, a beta oxidation disease, a vascular disease, an ocular vascular disease, or a muscular eye disease. Alternatively, the PPAR5 agonists are for use in a method to increase or maintain muscle mass or muscle tone in a subject.

BRIEF DESCRIPTION OF THE DRAWINGS

Figures 1A and IB are bar graphs showing recovery of damaged muscle fibers after injury.

Figures 1C-1F show VP16-PPAR5 transgenic animals exhibit accelerated muscle regeneration after acute injury. All error bars are SEM. Figure 1C provides two images of transverse sections of TA of WT and TG animals, with damaged fibers stained by Evans Blue dye 5 days after the injury. Figure ID provides the proportion of stained area over the total cross-sectional area (CSA) of TA (n=5; **P<0.01). Figure IE provides quantification of Evans Blue stain at 12 hours after injury (n=3). Figure IF provides quantification of Evans Blue stain at 36 hours after injury (n=3).

Figures 1G-1J illustrate VP16-PPAR5 transgenic animals that exhibit accelerated muscle regeneration after acute injury. All error bars are SEM. *P<0.05; **P<0.01;

***P<0.001; n.s.=not significant. Figure 1G provides H&E stained transverse sections of injured transversus abdominis muscle (TVA) from wildtype (WT) and transgenic (TG) animals. Representative images are from 3, 5 and 7 days after injury. Arrows = regenerating fibers with centralized nuclei. Arrowheads = hollowed remains of basal lamina. Asterisks = uninjured fibers. Figure 1H illustrates the average number of regenerating fibers per field. Figure II illustrates the average CSA of regenerating myofiber (n=5 for day 5; n=l 1 for day 7). Figure 1J illustrates the average CSA of regenerating myofiber, 21 days after injury (n=5).

Figures 2A-E illustrate that PPAR5 activation promotes a temporal shift in gene expression profile of the regenerative process. *P<0.05. All error bars are SEM. Figure 2A providesa GO classification of injury specific upregulated genes in TG (n=3). Figure 2B shows the relative expression of regeneration markers in TG. Figure 2C is a graph of relative expression versus days post injury, illustrating post injury temporal gene expression profiles of inflammatory marker CD68, measured by QPCR (n=5). Figure 2D is a graph of relative expression versus days post injury, illustrating post injury temporal gene expression profiles of a myogenic marker MyoD by Q-PCR (n=5). Figure 2E is a bar graph showing the Myh8 mRNA level 5 days post injury (n>5). Figures 3A-3G illustrate that PPAR5 regulates FGFla to promote micro- vascularization. *P<0.05; **P<0.01. All error bars are SEM. Figure 3A provides immunofluorescence staining for CD31 on transverse sections of uninjured TA from WT and TG animals. Figure 3B provides quantification of CD31 positive capillary number (n=4). Figure 3C illustrates the FGFla mRNA level in TA of WT and TG by QPCR (n=5). Figure 3D provides a Western blot for FGF1. Figure 3E provides immunofluorescence staining for CD31 positive capillaries on transverse sections of TA, 5 days after the injury (n=3). Figure 3F provides quantification for CD31 positive capillaries on transverse sections of TA, 5 days after the injury (n=3). Figure 3G provides luciferase reporter assays of FGFla promoter co- transfected with PPAR5 with or without the ligand, GW501516.

Figures 4A-4E illustrate that the skeletal muscle specific activation of PPAR5 increases the quiescent satellite cell pool. All error bars are SEM. *P<0.05; **P<0.01.

Figure 4A provides digital images of isolated myofibers from lateral gastrocnemius of 8- week-old nestin reporter mice with or without VP16-PPAR5 transgene. Figure 4B is a bar graph showing quantification of GFP+ satellite cells per unit length of myofiber (n=3).

Figure 4C is a bar graph showing the proportion of BrdU positive nuclei at 0.5, 1 and 2 days after injury (n=5). Figure 4D is a bar graph showing VP 16 mRNA levels in whole TA or satellite cells (SC) from WT and TG. Figure 4E is a bar graph showing PPAR5 mRNA levels in whole TA or satellite cells (SC) from WT and TG.

Figures 5A-5E illustrate that acute pharmacological activation of PPAR5 confers regenerative advantage. *P<0.05; **P<0.01; ***P<0.001. All error bars are SEM. Figure 5A is a series of bar graphs showing PPAR5 target gene expression in TA after 9 day treatment with either vehicle or GW501516 (n=6). Figure 5B provides digital images of transverse TA sections showing Evans Blue dye uptake 5 days after the injury. Figure 5C is a bar graph showing the proportions of stained area (n=5) in the images of Figure 5B.

Figure 5D is a bar graph showing the percentage of BrdU positive nuclei 2 days after injury (n=4). Figure 5E is a series of bar graphs showing TNFa and F480 levels 3 days after injury measured by QPCR (n=6).

Figures 6A-6E show VP16-PPAR5 transgenic animals exhibit accelerated muscle regeneration after the acute injury. All error bars are SEM. Figure 6A shows werum creatine kinase levels in wildtpe and VP16-PPAR5 transgenic animals. Figure 6B shows transverse sections of TA of WT and TG animals. Staining of damaged fibers by Evans Blue dye 5 days after the injury. Figure 6C shows proportion of stained area over the total CSA of TA (n=5; **P<0.01). Figures 6D and 6E show quantification of Evans Blue stain at 12 and 36 hours after injury (n=3).

Figure 7A shows transverse sections of TA of WT and TG animals. Staining of damaged fibers by Evans Blue 3 days after the injury.

Figure 7B shows Injury dependent induction of PPAR5 by QPCR (n=5).

Figure 7C shows post injury temporal gene expression profiles of inflammatory markers TNFcc.

Figure 7D shows induction of VEGFa in TA muscle, as measured by Western Blot, in TG animals.

Figure 7E shows quantification of TNFa Western Blot.

DETAILED DESCRIPTION

Peroxisome proliferator- activated receptor delta (PPAR5), also known as peroxisome proliferator- activated receptor beta (PPARP) or as NR1C2 (nuclear receptor subfamily 1, group C, member 2), refers to a nuclear receptor protein that function as a transcription factor regulating the expression of genes. Ligands of PPAR5 can promote myoblast proliferation after injury, such as injury to skeletal muscle. PPAR5 (OMIM 600409) sequences are publically available, for example from GenBank® sequence database (e.g., accession numbers NP_001165289.1 (human, protein) NP_035275 (mouse, protein), NM_001171818 (human, nucleic acid) and NM_011145 (mouse, nucleic acid)).

Herein, the phrase "PPAR5 agonist" refers to substances that increase the activity of PPAR5. Substances can be tested for their PPAR5 agonist activity by contacting the substance with cells expressing PPAR5, detecting their binding with PPAR5 and then detecting signals that serve as the indicator of the activation of PPAR5.

As shown in PCT/2014/033088 (incorporated herein by reference), modulating the activity of PPAR5 is useful in the treatment of diseases, developmental delays, and symptoms related to mitochondrial dysfunction.

Definitions

The term "aliphatic" refers to a saturated or unsaturated linear or branched

hydrocarbon group, and encompasses, for example, alkyl, alkenyl, and alkynyl groups. "Heteroaliphatic" means a saturated or unsaturated linear or branched hydrocarbon group containing one or more heteroatoms, and encompasses, for example, heteroalkyl, heteroalkenyl, and heteroalkynyl groups.

The term "alkyl" used alone or as part of a larger moiety, such as "alkoxy",

"haloalkyl", "cycloalkyl", "heterocycloalkyl", and the like, means saturated aliphatic straight- chain or branched monovalent hydrocarbon radical. Unless otherwise specified, an alkyl group typically has 1 to 4 carbon atoms, i.e., Ci-C₄-alkyl. As used herein, a "Ci-C₄-alkyl" group is means a radical having from 1 to 4 carbon atoms in a linear or branched

arrangement.

The term "alkenyl" used alone or as part of a larger moiety, such as "alkenoxy",

"cycloalkenyl", "heterocycloalkenyl", "haloalkenyl", and the like, means an unsaturated aliphatic straight-chain or branched monovalent hydrocarbon radical having one or more carbon-carbon double bonds.

The term "alkynyl" used alone or as part of a larger moiety, such as "alkynoxy", "haloalkynyl", and the like, means an unsaturated aliphatic straight-chain or branched monovalent hydrocarbon radical having one or more carbon-carbon triple bonds.

"Alkylsulfonyl" refers to a radical— S(0)₂R^xx where R^xx is an alkyl or cycloalkyl group as defined herein. Representative examples include, but are not limited to,

methylsulfonyl, ethylsulfonyl, propylsulfonyl, butylsulfonyl and the like.

"Alkoxy" means an alkyl radical attached through an oxygen linking atom, represented by -O-alkyl. For example, "Ci-C3-alkoxy" includes methoxy, ethoxy, propoxy, and butoxy.

The terms "haloalkyl" and "haloalkoxy" mean alkyl or alkoxy, as the case may be, substituted with one or more halogen atoms.

"Acyl" refers to a moiety of the formula— C(=0)— R^z, where R^z is aliphatic as defined herein.

"Amino" refers to the radical -NH₂.

"Amide" refers to -C(0)-NH-R^w, wherein R^w is hydrogen, alkyl, aryl, alkylaryl or hydrogen.

"Sulfonyl" refers to -S(0)₂-R^x, where R^x is aryl, C(CN)=C-aryl, CH₂CN, alkylaryl, sulfonamide, NH-alkyl, NH-alkylaryl, or NH-aryl.

"Carboxyl" refers to the radical— C(0)OH.

"Carboxyl ester" refers to the radical— C(0)OR^v, wherein R^v is hydrogen, alkyl, aryl, or alkylaryl. The term "carboxyl bioisostere" is a term familiar to medicinal chemists (see for example "The Organic Chemistry of Drug Design and Drug Action", by Richard B.

Silverman, pub. Academic Press, 1992), and refers to a group which has similar acid-base characteristics to those of a carboxyl group. Well known carboxyl bioisosteres include -S0₂NHR or -P(=0)(OH)(OR^q) wherein R^q is, for example, hydrogen methyl or ethyl, -S0₂OH, -P(=0)(OH)(NH₂), -C(=0)NHCN, and groups of the formulae:

The term "halogen" means fluorine or fluoro (F), chlorine or chloro (CI), bromine or bromo (Br), or iodine or iodo (I).

The term "ring" used herein means a cyclic group, which includes cycloalkyl, heterocycloaklyl, aryl, and heteroaryl, each of which can be monocyclic, bicyclic (e.g. , a bridged bicyclic ring), polycyclic (e.g. , tricyclic), or fused.

The term "aryl group" means an aromatic hydrocarbon ring system having six to fourteen carbon ring atoms. The term "aryl" may be used interchangeably with the terms "aryl ring", "aromatic ring", "aryl group", and "aromatic group". An aryl group typically has six to fourteen ring atoms. An "aryl group" also includes an aromatic ring fused to a non- aromatic carbocylic ring. Examples of aryl groups include phenyl, naphthyl, anthracenyl, 1,2-dihydronaphthyl, 1,2,3,4-tetrahydronaphthyl, fluorenyl, indanyl, indenyl and the like. A "substituted aryl group" is substituted at any one or more substitutable ring atom, which is a ring carbon atom bonded to a hydrogen. "Arylene" is a bivalent aryl group, i.e. , having two point of attachment to the remainder of the molecule.

The terms "cycloalkyl" and "cycloaliphatic" refer to a 3 - 12 membered saturated or unsaturated cyclic hydrocarbon radical. It can be monocyclic, bicyclic (e.g. , a bridged bicyclic ring), polycyclic (e.g. , tricyclic), or fused. For example, monocyclic C₃-C₆- cycloalkyl means a radical having from 3 to 6 carbon atoms arranged in a monocyclic ring. A C₃-C₆-cycloalkyl includes, but is not limited to, cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. "Cycloalkylene" is a bivalent cycloalkyl group, i.e. , having two point of attachment to the remainder of the molecule.

"Heterocycloalkyl" means a saturated or unsaturated non-aromatic 3 to 12 membered ring radical optionally containing one or more double bonds. It can be monocyclic, bicyclic (e.g. , a bridged bicyclic ring), tricyclic, or fused. The heterocycloalkyl contains 1 to 4 heteroatoms, which may be the same or different, selected from N, O or S. The

heterocycloalkyl ring optionally contains one or more double bonds and/or is optionally fused with one or more non-aromatic carbocyclic rings, aromatic rings (e.g. , phenyl ring) or heteroaryl rings. "5- or 6-membered monocyclic heterocycloalkyl" means a radical having from 5 or 6 atoms (including 1 to 3 heteroatoms) arranged in a monocyclic ring. Examples of heterocycloalkyl include, but are not limited to, morpholinyl, thiomorpholinyl,

pyrrolidinonyl, pyrrolidinyl, piperidinyl, piperazinyl, hydantoinyl, valerolactamyl, dihydroimidazole, dihydrofuranyl, dihydropyranyl, dihydropyridinyl, dihydropyrimidinyl, dihydrothienyl, dihydrothiophenyl, dihydrothiopyranyl, tetrahydroimidazole,

tetrahydrofuranyl, tetrahydropyranyl, tetrahydrothienyl, tetrahydropyridinyl,

tetrahydropyrimidinyl, tetrahydrothiophenyl, and tetrahydrothiopyranyl.

"Heterocycloalkylene" is a bivalent heterocycloalkyl group, i.e., having two point of attachment to the remainder of the molecule.

The term "heteroaryl", "heteroaromatic", "heteroaryl ring", "heteroaryl group",

"hetero aromatic ring", and "heteroaromatic group", are used interchangeably herein.

"Heteroaryl" when used alone or as part of a larger moiety as in "heteroaralkyl" or

"hetero arylalkoxy", refers to aromatic ring groups having five to fourteen ring atoms selected from carbon and at least one (typically 1 to 4, more typically 1 or 2) heteroatoms (e.g., oxygen, nitrogen or sulfur). "Heteroaryl" includes monocyclic rings and polycyclic rings in which a monocyclic heteroaromatic ring is fused to one or more other aromatic or

heteroaromatic rings. "Heteroarylene" is a bivalent heteroaryl group, i.e. , having two point of attachment to the remainder of the molecule.

"Monocyclic 5- or 6-membered heteroaryl" means a monocyclic aromatic ring system having five or six ring atoms selected from carbon and at least one (typically 1 to 3, more typically 1 or 2) heteroatoms (e.g., oxygen, nitrogen or sulfur). Examples of monocyclic 5-6 membered heteroaryl groups include furanyl (e.g. , 2-furanyl, 3-furanyl), imidazolyl (e.g., N- imidazolyl, 2-imidazolyl, 4-imidazolyl, 5-imidazolyl), isoxazolyl (e.g., 3-isoxazolyl, 4- isoxazolyl, 5-isoxazolyl), oxadiazolyl (e.g., 2-oxadiazolyl, 5-oxadiazolyl), oxazolyl (e.g., 2- oxazolyl, 4-oxazolyl, 5-oxazolyl), pyrazolyl (e.g. , 3-pyrazolyl, 4-pyrazolyl), pyrrolyl (e.g. , 1- pyrrolyl, 2-pyrrolyl, 3-pyrrolyl), pyridyl (e.g., 2-pyridyl, 3-pyridyl, 4-pyridyl), pyrimidinyl (e.g., 2-pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl), pyridazinyl (e.g., 3-pyridazinyl), thiazolyl (e.g., 2-thiazolyl, 4-thiazolyl, 5-thiazolyl), isothiazolyl, triazolyl (e.g., 2-triazolyl, 5-triazolyl), tetrazolyl (e.g., tetrazolyl), and thienyl (e.g. , 2-thienyl, 3-thienyl). Examples of polycyclic aromatic heteroaryl groups include carbazolyl, benzimidazolyl, benzothienyl, benzofuranyl, isobenzofuranyl, indolyl, benzotriazolyl, benzothiazolyl, benzoxazolyl, quinolinyl, isoquinolinyl, indazolyl, isoindolyl, acridinyl, or benzisoxazolyl. A "substituted heteroaryl group" is substituted at any one or more substitutable ring atom, which is a ring carbon or ring nitrogen atom bonded to a hydrogen.

The term "fused" as used herein refers to any combination of two or more cycloalkyl, heterocycloalkyl, aryl, and/or heteroaryl rings that share two adjacent ring atoms.

The term "bridged" as used herein refers to two carbocyclic refers to any combination of two cycloalkyl or heterocycloalkyl rings that share three or more adjacent ring atoms.

If a group is described as being "substituted", a non-hydrogen substituent is in the place of a hydrogen substituent on a carbon, sulfur or nitrogen of the substituent. Thus, for example, a substituted alkyl is an alkyl wherein at least one non-hydrogen substituent is in the place of a hydrogen substituent on the alkyl substituent. To illustrate, monofluoroalkyl is alkyl substituted with a fluoro substituent, and difluoroalkyl is alkyl substituted with two fluoro substituents. It should be recognized that if there is more than one substitution on a substituent, each non-hydrogen substituent can be identical or different (unless otherwise stated).

If a group is described as being "optionally substituted", the substituent can be either (1) not substituted, or (2) substituted.

If a list of groups are collectively described as being optionally substituted by one or more of a list of substituents, the list can include: (1) unsubstitutable groups, (2) substitutable groups that are not substituted by the optional substituents, and/or (3) substitutable groups that are substituted by one or more of the optional substituents.

If a group is described as being optionally substituted with up to a particular number of non-hydrogen substituents, that group can be either (1) not substituted; or (2) substituted by up to that particular number of non-hydrogen substituents or by up to the maximum number of substitutable positions on the substituent, whichever is less. Thus, for example, if a group is described as a heteroaryl optionally substituted with up to 3 non-hydrogen substituents, then any heteroaryl with less than 3 substitutable positions would be optionally substituted by up to only as many non-hydrogen substituents as the heteroaryl has

substitutable positions.

Unless otherwise indicated, suitable substituents for substituted alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups include the groups represented by halogen, -CN, -OR^c, -NR^aR^b, -C(=0)OR^c, -OC(=0)OR^c, -C(=0)NR^eR^f, -NR^dC(=0)R^c, -NR^d(C=0)OR^c, -0(C=0)NR^eR^f, -NR^d(C=0)NR^eR^f, -C(=0)R^c, (C C₆)alkyl, cycloalkyl, cycloalkyl(C

C₄)alkyl, heterocycloalkyl, heterocycloalkyl(Ci-C₄)alkyl, aryl, aryl(Ci-C₄)alkyl, heteroaryl, and heteroaryl(Ci-C₄)alkyl, wherein R^a and R^b are each independently selected from -H and (Ci-C₆)alkyl, optionally substituted with 1 to 3 substituents independently selected from halogen, hydroxy, -NR^gR^h and (Ci-C3)alkoxy; R^c is -H or (Ci-C₆)alkyl, optionally substituted with 1 to 3 substituents independently selected from halogen, -NR^gR^h, hydroxy and (Cp C₃)alkoxy; R^dis -H or (Ci-C₆)alkyl, optionally substituted with 1 to 3 substituents independently selected from halogen, -NR^gR^h, hydroxy and (Ci-C₃)alkoxy; and R^e and R^f are each independently selected from -H and (Ci-C₆)alkyl optionally substituted with 1 to 3 substituents independently selected from halogen, -NR^gR^h, hydroxy and (Ci-C₃)alkoxy; or R^e and R , together with the nitrogen to which they are attached, form a 3-8 membered ring optionally substituted with 1 to 3 substituents independently selected from halogen, -NR^gR^h, -CN, (C C₆)alkyl, halo(Ci-C₆)alkyl, (C C₃)alkoxy, halo(C C₃)alkoxy, and (C

C₃)alkoxy(Ci-C₆)alkyl. Each of the (C C₆)alkyl, cycloalkyl, cycloalkyl(C C₃)alkyl, heterocycloalkyl, heterocycloalkyl(Ci-C₃)alkyl, aryl, aryl(Ci-C₃)alkyl, heteroaryl and heteroaryl(Ci-C₃)alkyl substituents is optionally substituted with halogen, -N0₂, -CN, -NR^dC(=0)R^c, -NR^gR^h, (C C₄)alkyl, (C C₄)haloalkyl, (C C₄)alkoxy(Ci-C₄)alkyl, (C C₄)alkoxy, and (Ci-C₄)haloalkoxy, wherein R^g and R^h are each independently selected from - H, (Ci-C₆)alkyl, halo(Ci-C₆)alkyl, hydroxy(Ci-C₆)alkyl and (Ci-C₃)alkoxy(Ci-C₆)alkyl. Suitable substituents for a substituted alkyl, cycloalkyl, heterocycloalkyl can also include =0. Alternatively, suitable substituents for substituted alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups include alkyl, haloalkyl, alkoxy, haloalkoxy, cyano, and halogen.

Compounds having one or more chiral centers can exist in various stereoisomeric forms. Stereoisomers are compounds that differ only in their spatial arrangement. When a disclosed compound is named or depicted by structure without indicating stereochemistry, it is understood that the name or the structure encompasses all possible stereoisomers, geometric isomers, or a combination thereof.

When a geometric isomer is depicted by name or structure, it is to be understood that the geometric isomeric purity of the named or depicted geometric isomer is at least 60%, 70%, 80%, 90%, 99%, or 99.9% pure by weight. Geometric isomeric purity is determined by dividing the weight of the named or depicted geometric isomer in the mixture by the total weight of all of the geomeric isomers in the mixture.

Racemic mixture means 50% of one enantiomer and 50% of is corresponding enantiomer. When a compound with one chiral center is named or depicted without indicating the stereochemistry of the chiral center, it is understood that the name or structure encompasses both possible enantiomeric forms (e.g., both enantiomerically-pure, enantiomerically-enriched or racemic ) of the compound. When a compound with two or more chiral centers is named or depicted without indicating the stereochemistry of the chiral centers, it is understood that the name or structure encompasses all possible diasteriomeric forms (e.g., diastereomerically pure, diastereomerically enriched andequimolar mixtures if one or more diastereomers e.g., racemic mixtures) of the compound.

Enantiomeric and diastereomeric mixtures can be resolved into their component enantiomers or stereoisomers by well-known methods, such as chiral -phase gas

chromatography, chiral-phase high performance liquid chromatography, crystallizing the compound as a chiral salt complex, or crystallizing the compound in a chiral solvent.

Enantiomers and diastereomers also can be obtained from diastereomerically- or

enantiomerically-pure intermediates, reagents, and catalysts by well-known asymmetric synthetic methods.

When a compound is designated by a name or structure that indicates a single enantiomer, unless indicated otherwise, the compound is at least 60%, 70%, 80%, 90%, 99% or 99.9% optically pure (also referred to as "enantiomerically pure"). Optical purity is the weight in the mixture of the named or depicted enantiomer divided by the total weight in the mixture of both enantiomers.

When the stereochemistry of a disclosed compound is named or depicted by structure, and the named or depicted structure encompasses more than one stereoisomer (e.g. , as in a diastereomeric pair), it is to be understood that one of the encompassed stereoisomers or any mixture of the encompassed stereoisomers are included. It is to be further understood that the stereoisomeric purity of the named or depicted stereoisomers at least 60%, 70%, 80%, 90%, 99% or 99.9% by weight. The stereoisomeric purity in this case is determined by dividing the total weight in the mixture of the stereoisomers encompassed by the name or structure by the total weight in the mixture of all of the stereoisomers.

Included in the present teachings are pharmaceutically acceptable salts of the compounds disclosed herein. The disclosed compounds have basic amine groups and therefore can form pharmaceutically acceptable salts with pharmaceutically acceptable acid(s). Suitable pharmaceutically acceptable acid addition salts of the compounds described herein include salts of inorganic acids (such as hydrochloric acid, hydrobromic, phosphoric, nitric, and sulfuric acids) and of organic acids (such as, e.g. , acetic acid, benzenesulfonic, benzoic, methanesulfonic, and /?-toluenesulfonic acids). Compounds of the present teachings with acidic groups such as carboxylic acids can form pharmaceutically acceptable salts with pharmaceutically acceptable base(s). Suitable pharmaceutically acceptable basic salts include ammonium salts, alkali metal salts (such as sodium and potassium salts) and alkaline earth metal salts (such as magnesium and calcium salts).

As used herein, the term "pharmaceutically-acceptable salt" refers to pharmaceutical salts that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, and allergic response, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically-acceptable salts are well known in the art. For example, S. M. Berge, et al. describes pharmacologically acceptable salts in J. Pharm. Sci., 1977, 66: 1-19.

The neutral forms of the compounds of the invention are regenerated from their corresponding salts by contacting the salt with a base or acid and isolating the parent compound in the conventional manner. The parent form of the compound may differ from the various salt forms in certain physical properties, such as solubility in polar solvents. The neutral forms of compounds disclosed herein also are included in the invention.

The terms "administer", "administering", "administration", and the like, as used herein, refer to methods that may be used to enable delivery of compositions to the desired site of biological action. These methods include, but are not limited to, intraarticular (in the joints), intravenous, intramuscular, intratumoral, intradermal, intraperitoneal, subcutaneous, orally, topically, intrathecally, inhalationally, transdermally, rectally, and the like.

Administration techniques that can be employed with the agents and methods described herein are found in e.g., Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon; and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa.

As used herein, the terms "co-administration", "administered in combination with", and their grammatical equivalents, are meant to encompass administration of two or more therapeutic agents to a single subject, and are intended to include treatment regimens in which the agents are administered by the same or different route of administration or at the same or different times. In some embodiments the one or more compounds described herein will be co- administered with other agents. These terms encompass administration of two or more agents to the subject so that both agents and/or their metabolites are present in the subject at the same time. They include simultaneous administration in separate compositions,

administration at different times in separate compositions, and/or administration in a composition in which both agents are present. Thus, in some embodiments, the compounds described herein and the other agent(s) are administered in a single composition. In some embodiments, the compounds described herein and the other agent(s) are admixed in the composition.

Generally, an effective amount of a compound taught herein varies depending upon various factors, such as the given drug or compound, the pharmaceutical formulation, the route of administration, the type of disease or disorder, the identity of the subject or host being treated, and the like, but can nevertheless be routinely determined by one skilled in the art. An effective amount of a compound of the present teachings may be readily determined by one of ordinary skill by routine methods known in the art.

The term "effective amount" or "therapeutically effective amount" means an amount when administered to the subject which results in beneficial or desired results, including clinical results, e.g., inhibits, suppresses or reduces the symptoms of the condition being treated in the subject as compared to a control. For example, a therapeutically effective amount can be given in unit dosage form (e.g. , 1 mg to about 50 g per day, altnernatively from 10 mg to about 5 grams per day; and in another alternatively from 10 mg to 1 gram per day).

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

"Pharmaceutically acceptable excipient" and "pharmaceutically acceptable carrier" refer to a substance that aids the formulation and/or administration of an active agent to and/or absorption by a subject and can be included in the compositions of the present disclosure without causing a significant adverse toxicological effect on the subject. Non- limiting examples of pharmaceutically acceptable excipients include water, NaCl, normal saline solutions, lactated Ringer's, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer's solution), alcohols, oils, gelatins, carbohydrates such as lactose, amylose or starch, fatty acid esters, hydroxymethycellulose, polyvinyl pyrrolidine, and colors, and the like. Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and/or aromatic substances and the like that do not deleteriously react with or interfere with the activity of the compounds provided herein. One of ordinary skill in the art will recognize that other pharmaceutical excipients are suitable for use with disclosed compounds.

PPARS Agonists

Disclosed herein are methods of treating certain PPAR5 related conditions, diseases, and disorders with compounds having PPAR5 agonist activity (also referred to herein as "PPAR5 agonists").

Compounds with PPAR5 agonist activity (also referred to herein as "PPAR5 agonists") can be used in any of the methods disclosed herein, provided that the PPAR5 agonist is not a compound having the formula:

or a pharmaceutically acceptable salt thereof,

wherein:

ring A is selected from a cycloalkylene, heterocycloalkylene, arylene or

heteroarylene;

ring B is selected from an aryl, heteroaryl, cycloalkyl, heterocycloalkyl,

cycloalkylene,

heterocycloalkylene, arylene or heteroarylene;

each R independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic, heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, S0₃H, or acyl;

each R 22 independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic,

heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

n is from 0 to 5;

m is from 0 to 4;

X is O, NR³⁰, sulfonyl, or S;

R 30 is selected from H or aliphatic, aryl, or cycloaliphatic;

L⁵ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -L 3 N(L 4 R 3 )L 3 -; L is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene,

23 24

cycloalkylene, heterocycloalkylene or -CR R -;

23 24

R"^J and are each independently selected from H, deuterium, halogen, aliphatic, alkyl, -C(0)OR²⁵ or -C(0)NR²⁵R²⁶;

R²⁵ and R²⁶ are each independently hydrogen, aliphatic or alkyl;

Z is selected from R¹L¹C(0)- or a carboxyl bioisostere;

L¹ is a bond or -NR³⁰-;

R¹ is hydrogen, aliphatic, -OR^1A, -NR^1AR^1B, -C(0)R^1A, -S(0)₂R^1A, -C(0)OR^1A, - S(0)₂NR^1AR^1B or -C(0)NR^1AR^1B;

R^1A, R^1B are each independently hydrogen, aliphatic or alkyl;

L is selected from a bond, aliphatic, -C(O)-, alkylC(O)-, -C(0)alkyl-, or sulfonyl; L⁴ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene,

23 24

cycloalkylene, heterocycloalkylene or -CR R -;

R³ is selected from -OH, -OR^3A, -NR^3AR^3B, -C(0)R^3A, -S(0)2R^3A, -C(0)OR^3A, - S(0)₂NR^3AR^3B, -C(0)NR^3AR^3B, aliphatic, heteroaliphatic, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or R can be joined with an atom of ring B to form a fused ring system or may be joined with an atom of L to form a heterocyclic ring system; and

3 A 3B

R , R , are each independently hydrogen, aliphatic or alkyl.

In a 1^st embodiment of the disclosed methods, the PPAR5 agonist is a compound of Formula (I):

(i);

or a pharmaceutically acceptable salt thereof,

wherein:

1 7

Ar is phenyl, which is optionally substituted with from one to five R substituents independently selected from the group consisting of halogen, (Ci-Cg)alkyl, halo(Ci-Cg)alkyl, and -OR²;

2 8

Ar is phenyl, which is optionally substituted with from one to four R substituents independently selected from the group consisting of halogen, (Ci-Cg)alkyl, halo(Ci-Cg)alkyl, and -OR²;

L is a member selected from the group consisting of -CH₂S- and -CH₂0-; K is a member selected from the group consisting of a covalent bond and -OCH₂- Z is C0₂R⁶;

R¹ is selected from the group consisting of H and (Ci-C8)alkyl;

each R 2 and R 3 is a member independently selected from the group consisting of H, (C C₈)alkyl, halo(C C₈)alkyl,— X³OR⁹, aryl, aryl(C C₄)alkyl, and heteroaryl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered ring system;

R⁶ is a member selected from the group consisting of H, (Ci-C₈)alkyl, halo(Ci- C₈)alkyl,— X⁴OR²,— X⁴NR²R³, (C₂-C₈)alkenyl, (C₃-C₇)cycloalkyl, heterocyclyl, aryl (C C₄)alkyl; and aryl(C₂-C₈)alkenyl;

R⁹ is a member selected from the group consisting of H, (Ci-C₈)alkyl, halo(Ci- C₈)alkyl, aryl, aryl(Ci-C₄)alkyl, and heteroaryl;

each X³ and X⁴ is a member independently selected from the group consisting of (Cp C₄) alkylene, (C₂-C₄)alkenylene, and (C₂-C4)alkynylene.

In a specific embodiment, the compound of Formula (I) has the structure of Formula

(la):

or a pharmaceutically acceptable salt thereof.

In a 2^nd embodiment of the disclosed methods, the PPAR5 agonist is a compound of Formula (II):

(II);

or a pharmaceutically acceptable salt thereof,

wherein:

X represents a COOH (or a hydrolysable ester thereof),

X^I is O or S, and the depicted bond with a dashed line is a single bond;

X represents O, S; R 1 and R 2 independently represent H, CH₃, OCH₃ or halogen;

n is 1 or 2;

one of Y and Z is N and the other is S or O;

y represents 1 or 2; and

each R independently represents CF₃ or halogen.

In a specific embodiment, the compound of Formula (II) has the structure of Formula

(Ila)

or a pharmaceutically acceptable salt thereof.

In a 3^rd embodiment of the disclosed methods, the PPAR5 agonist is a compound of Formula (III):

(HI);

or a pharmaceutically acceptable salt thereof,

wherein:

A is a saturated or unsaturated hydrocarbon chain having from 3 to 5 atoms, forming five- to seven-membered ring;

T is selected from the group consisting of -C(0)OH, -C(0)NH₂, and tetrazole;

G¹ is selected from the group consisting of -(CR'R²)^, -ZiCR¹^²)^, -(CR^_nZ-, -(CR^^CR^ -;

Z is O, S or NR;

n is 0, 1, or 2;

r and s are independently 0 or 1 ;

R 1 and R 2" are independently selected from the group consisting of hydrogen, halo, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted lower alkoxy, and lower perhaloalkyl or together may form an optionally substituted cycloalkyl; 1 2 3

X X\ and X^J are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted cycloalkyl, halogen, perhaloalkyl, hydroxy, optionally substituted lower alkoxy, nitro, cyano, and NH₂;

G₂ is selected from the group consisting of a saturated or unsaturated heterocycloalkyl linker, optionally substituted with X₄ and X5;

X⁴ and X⁵ are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, halogen, lower perhaloalkyl, hydroxy, optionally substituted lower alkoxy, nitro, cyano, NH₂, and C0₂R, or X₄ and X5 together may form a carbocycle;

R is selected from the group consisting of optionally substituted lower alkyl and hydrogen;

G3 is selected from the group consisting of a bond, a double bond,— (CR³R⁴)_m— , carbonyl, and— (CR³R⁴)_mCR³=CR⁴— ;

m is 0, 1, or 2;

R³ and R⁴ are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted lower alkoxy, optionally substituted aryl, lower perhaloalkyl, cyano, and nitro;

G⁴ is selected from the group consisting of hydrogen, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted cycloheteroalkyl, optionally substituted cycloheteroaryl, optionally substituted cycloalkenyl, and— N=(CR⁵R⁶); and

R⁵ and R⁶ are independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted cycloalkenyl, and optionally substituted cycloheteroalkyl.

In a specific embodiment, the compound of Formula (III) has the structure of Formula

(Ilia):

(Ilia);

or a pharmaceutically acceptable salt thereof. In another embodiment, the compound of Formula (III) has the structure of Formula

(nib):

(nib);

or a pharmaceutically acceptable salt thereof.

Methods of Treatment

Provided herein are methods of activating PPAR5.

Such methods can include contacting a PPAR5 protein with an effective amount of a compound or composition provided herein, thereby activating PPAR5. In some

embodiments, the contacting is performed in vitro. In other embodiments, the contacting is performed within a subject, such as a human subject, for example by administering a PPAR agonist disclosed herein to the subject. In some embodiments, the compound or composition is administered to a healthy subject. In some embodiments, the subject is a sedentary or immobilized subject. In other embodiments, the subject is an exercising subject, such as one who exercises for at least 20 minutes, at least 30 mintues, at least 45 mintures, or at least 60 minutes, at least 2, at least 3, or at least 4 days per week. In some embodiments, a healthy subject is also an exercising subject.

In some examples, contacting a PPAR5 protein in vitro or in vivo with an effective amount of one or more compounds or compositions provided herein, increases PPAR5 activity by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 100%, at least 200%, at least 300%, at least 400%, or even at least 500%, for example as compared to an amount of PPAR5 activity in the absence of the compound/composition. Methods of measuring PPAR5 activity are known, and specific examples are provided herein (e.g., measuring expression of PPAR5 at the protein or nucleic acid level, measuring Beta oxidation levels, creatine kinase levels, pentose phosphate shunt in liver, blood glucose levels and methods provided in Wang et al., PLos Biol. 2(10):e294, 2004 and Lee et al., PNAS 103:3444-9, 2006).

In some embodiments, the subject recovers from acute injury following

administration of the PPAR agonist. In some embodiments, activating PPAR5 within the subject by administration of a PPAR5 agonist (or composition containing a PPAR5 agonist) increases or maintains muscle mass or muscle tone (such as a skeletal or cardiac muscle) in the subject (such as in a healthy subject or a sedentary subject). For example, activating PPAR5 within the subject can increase muscle mass, muscle tone, or both, in the subject. In some examples, administering an effective amount of one or more PPAR5 agonist compounds or compositions provided herein increases muscle mass, muscle tone, or both, by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 100%, at least 200%, at least 300%, at least 400%, or even at least 500%, for example as compared to an amount of PPAR5 activity in the absence of the

compound/composition. Methods of measuring muscle mass and muscle tone are known, and specific examples are provided herein (e.g. , see methods provided in WO 2009/086526).

In other embodiments, activating PPAR5 within the subject (such as a healthy subject or a sedentary subject) maintains muscle mass, muscle tone, or both, in the subject. In some examples, administering an effective amount of one or more PPAR5 agonist compounds or compositions maintains muscle mass, muscle tone, or both, such that the amount of muscle mass, muscle tone or both, does not change by more than 1%, for example no more than 2%, no more than 3%, no more than 4%, no more than 5%, no more than 6%, no more than 7%, no more than 8%, no more than 9%, no more than 10%, or no more than 15%, for example as compared to an amount of muscle mass, muscle tone, or both in the absence of the compound/composition. Methods of measuring muscle mass and muscle tone are known, and specific examples are provided herein (e.g. , see methods provided in WO 2009/086526).

Thus, PPAR5 agonists and compositions containing such can be used to increase or maintain muscle mass or muscle tone (or both) in a subject. For example, the disclosed PPAR agonists and compositions containing such can be used to increase or maintain muscle mass or muscle tone (or both) in a subject following an injury, following a period of immobilization (for example confinement to a bed or wheelchair) or immobilization of a body part (for example immobilization of an appendage or joint due to a broken bone, joint replacement, tendon tear, surgery, and the like), which events can result in a loss of muscle mass and/or muscle tone. The method includes administering to the subject a therapeutically effective amount of one or more compounds provided herein. In some embodiments, the subject is a sedentary or immobilized subject. In other embodiments, the subject is an exercising subject. Methods of treating a PPAR5-related disease or condition in a subject in need thereof also are provided. The methods can include administering to the subject a therapeutically effective amount of one or more compounds or compositions provided herein.

In preferred embodiments, the ΡΡΑΡνδ-related disease is a mitochondrial disease. Examples of mitochondrial diseases include, but are not limited to, Alpers's Disease, CPEO- Chronic progressive external ophthalmoplegia , Kearns-Sayra Syndrome (KSS), Leber Hereditary Optic Neuropathy (LHON), MELAS-Mitochondrial myopathy,

encephalomyopathy, lactic acidosis, and stroke-like episodes, MERRF-Myoclonic epilepsy and ragged-red fiber disease, NARP-neurogenic muscle weakness, ataxia, retinitis pigmentosa, and Pearson Syndrome.

In other embodiments, the PPAR5-related disease is a vascular disease (such as a cardiovascular disease or any disease that would benefit from increasing vascularization in tissues exhibiting impaired or inadequate blood flow). In other embodiments, the PPAR5- related disease is a muscular disease, such as a muscular dystrophy. Examples of muscular dystrophy include but are not limited to Duchenne muscular dystrophy, Becker muscular dystrophy, limb-girdle muscular dystrophy, congenital muscular dystrophy,

facioscapulohumeral muscular dystrophy, myotonic muscular dystrophy, oculopharyngeal muscular dystrophy, distal muscular dystrophy, and Emery-Dreifuss muscular dystrophy.

In some embodiments, the PPAR5-related disease or condition is a demyelinating disease, such as multiple sclerosis, Charcot-Marie-Tooth disease, Pelizaeus-Merzbacher disease, encephalomyelitis, neuromyelitis optica, adrenoleukodystrophy, or Guillian-Barre syndrome.

In yet other embodiments, the PPAR5-related disease is a muscle structure disorder. Examples of a muscle structure disorders include, but are not limited to, Bethlem myopathy, central core disease, congenital fiber type disproportion, distal muscular dystrophy (MD), Duchenne & Becker MD, Emery- Dreifuss MD, facioscapulohumeral MD, hyaline body myopathy, limb-girdle MD, a muscle sodium channel disorders, myotonic chondrodystrophy, myotonic dystrophy, myotubular myopathy, nemaline body disease, oculopharyngeal MD, and stress urinary incontinence.

In still other embodiments, the PPAR5-related disease is a neuronal activation disorder, Examples of neuronal activation disorders include, but are not limited to, amyotrophic lateral sclerosis, Charcot-Marie-Tooth disease, Guillain-Barre syndrome, Lambert-Eaton syndrome, multiple sclerosis, myasthenia gravis, nerve lesion, peripheral neuropathy, spinal muscular atrophy, tardy ulnar nerve palsy, and toxic myoneural disorder. In other embodiments, the PPAR5-related disease is a muscle fatigue disorder.

Examples of muscle fatigue disorders include, but are not limited to chronic fatigue syndrome, diabetes (type I or II), glycogen storage disease, fibromyalgia, Friedreich's ataxia, intermittent claudication, lipid storage myopathy, MELAS, mucopolysaccharidosis, Pompe disease, and thyrotoxic myopathy.

In some embodiments, the ΡΡΑΡνδ-related disease is a muscle mass disorder.

Examples of muscle mass disorders include, but are not limited to, cachexia, cartilage degeneration, cerebral palsy, compartment syndrome, critical illness myopathy, inclusion body myositis, muscular atrophy (disuse), sarcopenia, steroid myopathy, and systemic lupus erythematosus.

In other embodiments, the ΡΡΑΡνδ-related disease is a beta oxidation disease.

Examples of beta oxidation diseases include, but are not limited to, systemic carnitine transporter, carnitine palmitoyltransferase (CPT) II deficiency, very long- chain acyl- CoA dehydrogenase (LCHAD or VLCAD) deficiency, trifunctional enzyme deficiency, medium - chain acyl - CoA dehydrogenase (MCAD) deficiency, short - chain acyl- CoA dehydrogenase (SCAD) deficiency, and riboflavin - responsive disorders of β-oxidation (RR -MADD).

In some embodiments, the PPAR5-related disease is a vascular disease. Examples of vascular diseases include, but are not limited to, peripheral vascular insufficiency, peripheral vascular disease, intermittent claudication, peripheral vascular disease (PVD), peripheral artery disease (PAD), peripheral artery occlusive disease (PAOD), and peripheral obliterative arteriopathy.

In other embodiments, the PPAR5-related disease is an ocular vascular disease.

Examples of ocular vascular diseases include, but are not limited to, age-related macular degeneration (AMD), stargardt disease, hypertensive retinopathy, diabetic retinopathy, retinopathy , macular degeneration, retinal haemorrhage, and glaucoma.

In yet other embodiments, the PPAR5-related disease is a muscular eye disease. Examples of muscular eye diseases include, but are not limited to, strabismus (crossed eye/wandering eye/walleye ophthalmoparesis), progressive external ophthalmoplegia, esotropia, exotropia, a disorder of refraction and accommodation, hypermetropia, myopia, astigmatism, anisometropia, presbyopia, a disorders of accommodation, or

internal ophthalmoplegia.

In still other embodiments, the PPAR5-related disease is a metabolic disease selected from hyperlipidemia, dyslipidemia, hypercholesterolemia, hypertriglyceridemia, HDL hypocholesterolemia, LDL hypercholesterolemia and/or HLD non-cholesterolemia, VLDL hyperproteinemia, dyslipoproteinemia, apolipoprotein A-I hypoproteinemia, atherosclerosis, disease of arterial sclerosis, disease of cardiovascular systems, cerebrovascular disease, peripheral circulatory disease, metabolic syndrome, syndrome X, obesity, diabetes (type I or II), hyperglycemia, insulin resistance, impaired glucose tolerance, hyperinsulinism, diabetic complication, cardiac insufficiency, cardiac infarction, cardiomyopathy, hypertension, Nonalcoholic fatty liver disease (NAFLD), Nonalcoholic steatohepatitis (NASH), thrombus, Alzheimer disease, neurodegenerative disease, demyelinating disease, multiple sclerosis, adrenal leukodystrophy, dermatitis, psoriasis, acne, skin aging, trichosis, inflammation, arthritis, asthma, hypersensitive intestine syndrome, ulcerative colitis, Crohn's disease, or pancreatitis.

In one embodiment, the PPAR5-related disease is a cancer selected from a cancer of the colon, large intestine, skin, breast, prostate, ovary, or lung.

Pharmaceutical Compositions and Administration Thereof

A. Additional Therapeutic Agents

Pharmaceutical compositions are disclosed that include one or more PPAR5 agonists, and typically at least one additional substance, such as an excipient, a known therapeutic other than those of the present disclosure, and combinations thereof. In some embodiments, a PPAR5 agonist can be used in combination with other agents known to have beneficial, additive or synergistic activity with the PPAR5 agonists. For example, disclosed compounds can be administered alone or in combination with: one or more other PPAR5 agonists, such as a thiazolidinedione, including rosiglitazone, pioglitazone, troglitazone, and combinations thereof, or a sulfonylurea agent or a pharmaceutically acceptable salt thereof, such as tolbutamide, tolazamide, glipizide, carbutamide, glisoxepide, glisentide, glibornuride, glibenclamide, gliquidone glimepiride, gliclazide and the pharmaceutically acceptable salts of these compounds, or muraglitazar, farglitazar, naveglitazar, netoglitazone, rivoglitazone, K- 111, GW-677954, (-)-Halofenate, acid, arachidonic acid, clofbrate, gemfibrozil, fenofibrate, ciprofibrate, bezafibrate, lovastatin, pravastatin, simvastatin, mevastatin, fluvastatin, indomethacin, fenoprofen, ibuprofen, and the pharmaceutically acceptable salts of these compounds.

In one embodiment, disclosed compounds may be administered in combination with dexamphetamine, amphetamine, mazindole, or phentermine; and administered in

combination with medicaments having an anti-inflammatory effect. B. Excipients and Dosage Forms

The particular mode of administration and the dosage regimen will be selected by the attending clinician, taking into account the particulars of the case (e.g. the subject, the disease, the disease state involved, the particular treatment, and whether the treatment is prophylactic). Treatment can involve daily or multi-daily or less than daily (such as weekly or monthly etc.) doses over a period of a few days to months, or even years. However, a person of ordinary skill in the art would immediately recognize appropriate and/or equivalent doses looking at dosages of approved compositions for treating a PPAR5 related disease using the disclosed PPAR5 agonists for guidance.

Working Examples

Skeletal muscle relies on the resident progenitor cells, the satellite cells, for postnatal growth and regeneration. Therefore, maintaining an adequate number and proper function of satellite cells is critical for muscle to appropriately response to damage. While endurance exercise promotes adaptive responses in the muscle, including an increase in the satellite cell number, it is not known whether transcriptionally directed "endurance exercise training" has similar effects. Here it is shown that mice harboring constitutively active PPAR5 in skeletal muscle displayed an accelerated regenerative process in muscle after an acute injury. Gene expression analyses showed earlier resolution of the inflammatory response and induction of myogenic markers, indicating that PPAR5 activation induces a temporal shift in the regenerative process. Notably, a significant increase in the number of satellite cells was found in mice with constitutively active PPAR5 expressed in skeletal muscle, consistent with the observed increase in proliferating cell number after the injury. PPAR5 activation induced the expression of FGF1, which is known to be involved in muscle development and regeneration. In particular, PPAR5 up-regulates FGFla isoform, which may be responsible for supporting cell proliferation and reestablishment of vasculature to augment the

regenerative process. Furthermore, the restoration of fiber integrity was improved in wild- type mice after acute treatment with the PPAR5 synthetic ligand, GW501516. Collectively, these findings allude to the therapeutic potential of PPAR5, to accelerate the recovery from acute muscle injury.

Activation of peroxisome proliferator activated receptor δ (PPAR5) induces a fiber type switch toward a more oxidative phenotype, altering both metabolic and functional output of the muscle (Wang et al., PLoS Biol 2(10):e294. Erratum in: PLoS Biol. 2005 Jan;3(l):e61 (2004); Luquet et al, FASEB J 17(15):2299-2301 (2003)). Specifically, PPAR5-mediated muscle remodeling translates into supernatural physical endurance, and protection against diet-induced obesity and symptoms of metabolic disorders that ensue (Wang et al, PLoS Biol 2(10):e294. Erratum in: PLoS Biol. 2005 Jan;3(l):e61 (2004); Wang et al, Cell 113: 159-170 (2003)). Furthermore, pharmacological activation of PPAR5 and exercise training synergistically enhance oxidative fibers and running endurance (Narkar VA et al. , Cell 134(3):405-415 (2008)). Exercise confers a myriad of healthful benefits to the body, including improvement of atrophic and disease conditions (Nicastro et al, Braz J Med Biol Res 44(11): 1070-9 (2011); Markert et al, Muscle Nerve 43(4):464-78 (2011)). Recently, endurance exercise alone has been shown to improve ageing induced decrease in satellite cell number and their myogenic capacity (Shefer et al., PLoS One 5(10):el3307 (2010)).

It is demonstrated herein that both genetic and pharmacological activation of PPAR5 promote muscle regeneration in an acute thermal injury mouse model. PPAR5 activation during regeneration expedites resolution of inflammatory response and restoration of contractile proteins. Interestingly, acute pharmacological activation of PPAR5 by oral administration of a synthetic ligand, GW501516, is sufficient to confer similar benefits during muscle regeneration after an acute injury. Based on these observations, a novel role of PPAR5 during adult muscle regeneration and its use as a therapeutic target to enhance regenerative efficiency of skeletal muscle is provided.

EXEMPLIFICATION Example 1

Experimental Procedures

A. Animals

VP16-PPAR5 mice (Wang et al, Cell 113: 159-170 (2003)) were bred to CB6F1 strain (Jackson Laboratories) and used as heterozygotes in experiments. The non-transgenic littermates served as controls. All experiments were performed when animals were 8 weeks of age. Nestin-GFP mice (Mignone et al, J Comp Neurol 469(3):311-324 (2004)) were kindly provided by Dr. Fred Gage at the Salk Institute for Biological Studies. B. Freeze burn injury

TA muscles were injured according to previously published methods with a few modifications (Brack et ah, Science 317(5839):807-810 (2007)). A stainless steel lg weight (Mettler- Toledo) equilibrated to the temperature of dry ice was placed directly on the exposed TA for 10 seconds. Following the thermal injury, incision was closed using

VetBond (3M). All injury procedures were performed on the left leg, and the right leg was used as control.

C. Histology

Animals were perfused with 15 mL of ice-cold PBS followed immediately by 20 mL of 10% saline buffered formalin. TA muscles were excised and immersed in 4%

paraformaldehyde for at least 48 hours at 4 °C. Tissues were dehydrated in series of solutions with increasing percentage of ethanol. Dehydrated tissues were cleared in xylene and allowed for paraffin to permeate over night at 60 °C. Tissues were then embedded in plastic molds.

Paraffin embedded tissue blocks were sectioned at 7 μιη thick on Leica Jung 2500 Microtome. Sections were stained with hematoxylin and counter stained with 1% eosin. Slides were dried and mounted with Entellan mounting media (EMS). Three random non- overlapping fields were photographed for analysis. Regenerating fiber number was measured by counting the number of discernible muscle fibers with centralized myonuclei (Ge et ah, Am J Physiol Cell Physiol 297 (6):C1434- 1444 (2009)). Regenerating fiber cross sectional area (CSA) was measured using Image J software.

D. Evans Blue dye staining

Injured animals were injected with Evans Blue dye according published protocol

(Hamer et al, J Anat 200(Pt l):69-79 (2002)). Sterile 1% w/v Evans Blue dye in PBS was intraperitoneally injected at 1% volume relative to the body mass of an animal. Seven hours after the injection, injured TA muscles were harvested and snap-frozen by isopentane quenching in liquid nitrogen. Frozen sections were cut in ΙΟμιη thickness, fixed in ice-cold acetone, dipped in xylene and mounted with DPX. Proportion of the stained area over the total area was measured using ImageJ software. E. BrdU Labeling

50 mg/kg body weight of BrdU (Sigma) was injected intraperitoneally as solution of 10 mg/mL BrdU in saline. TA muscles were harvested at 7 days after injury and processed for paraffin sections as described above. BrdU incorporation was visualized using the BrdU Labeling and Detection Kit I (Roche) and BrdU+ nuclei were counted and represented as a proportion of total nuclei in a field.

F. RT-QPCR

Whole or partial tissues were homogenized by Polytron probe homogenizer in Trizol reagent (Invitrogen). Total RNA was extracted from the homogenates according to the manufacturer's protocol. One microgram of DNase-treated total RNA was reverse transcribed using Superscript II Reverse Transcriptase (Invitrogen) according to the manufacturer's instructions. cDNAs were diluted 1/40 with ddH₂0 and used as templates in RT-QPCR reactions with SYBRGreenER qPCR SuperMix detection system (Invitrogen). Samples were prepared in technical triplicates and relative mRNA levels were calculated by using the standard curve methodology and normalized against GAPDH mRNA levels in the same samples.

G. Myofiber Isolation

Either whole or partial gastrocnemius muscle was digested in 2% collagenase I (Sigma) in DMEM with 10% FBS for 60 minutes at 37 °C. Muscle tissue was further mechanically digested by triturating with fire polished wide bore Pasteur pipet. Liberated fibers were washed in two changes of PBS with 10%FBS and finally mounted on glass slides with Vectashield mounting media (Vector Labs).

H. Isolation of satellite cells

Satellite cells were harvested from TA of 8 weeks old animals according to published protocols with some modifications (Day et al. (2007) Nestin-GFP reporter expression defines the quiescent state of skeletal muscle satellite cells. Dev Biol 304(l):246-259). Muscles were removed and washed briefly in DMEM on ice. They were then minced to fine slurry with razor blade on 60mm culture dish over ice. Minced muscles were transferred to one well of a 6-well plate containing 5 ml of 450KPU/ml pronase in DMEM. The tissues were digested at 37°C/5% C0₂ for 60 minutes. After digestion, tissues were vigorously triturated 20 times through 10ml serological pipet. Digested tissues were filtered through 40 micron cell strainer and washed with equal volume of DMEM with 20% horse serum. Cells were spun down at lOOOg for 10 minutes and resuspended in sorting buffer (DMEM with 10% FBS). Cells were separated from larger debris by 20%/60% Percoll gradient (Yablonka-Reuveni Z et al. (1987) Isolation and clonal analysis of satellite cells from chicken pectoralis muscle. Dev Bio 119: 252-259). GFP positive cells were sorted on BD FACSAria II sorter.

Example 2

Muscle specific activation of PPAR5 confers regenerative advantage While it has been shown that the majority of the metabolic genes are down regulated in this model, PPAR5 expression was induced over 2 fold at 2 days after the injury (Warren et al. (2007) Mechanisms of skeletal muscle injury and repair revealed by gene expression studies in mouse models. J Physiol. 582.2: 825-841, Figure 1A). This injury dependent up- regulation of PPAR5 strongly suggested a possible role for PPAR5 during the early part of the regenerative proces s .

Freeze burn injury was used to elicit the regenerative program, which has been shown to model the standard course of regenerative response, including satellite cell activation (Karpati and Molnar. "Muscle fibre regeneration in human skeletal muscle diseases." In: Schiaffino S, Partridge T (eds). Skeletal muscle repair and regeneration. Springer, Dordrecht, 2008). Additionally, since the injury is directly applied to the surface of the muscle, it is highly localized and reproducible.

Using Evans Blue dye uptake as a marker of myofiber damage, fiber integrity was histologically assessed. The freeze burn injury does not incapacitate the animals and the damaged fibers restore original cross sectional area by 21 days after the injury (Figure IB). By comparing the proportion of stained fibers within the cross sectional area (CSA) of the injured muscle 5 days after the injury, the degree of existing damage was quantified. At 5 days after the injury, VP16-PPAR5 (TG) animals show significantly less dye uptake, thus increased fiber intactness, over the wildtype (WT) animals (Figure 1C). While 14% of the total CSA shows dye uptake, only 5% of the total CSA of TG muscle show dye uptake (n=8 WT; n=5 TG; p=0.001) (Figure ID). At 12 and 36 hours after the injury, however, both WT and TG animals showed similar proportions of stained area (50.6% and 47.4% (p=0.67), and 38.5% and 43.3% (p=0.23), respectively) (Figures IE and IF). Similar level of dye uptake shortly after the injury shows that both WT and TG animals initially sustain similar degree of damage from the injury and suggests that PPAR5 activation does not confer protection from damage. Instead, the reduction in Evans Blue dye uptake observed 5 days after the injury suggests that the muscle specific PPAR5 activation promotes restoration of fiber integrity after the injury.

The morphological hallmarks of regenerating fibers was determined for a detailed analysis of the process. H&E stained transverse sections through the injured area were examined at 3, 5 and 7 days post injury. At 3 days after the injury, both WT and TG animals showed similar degrees of degeneration defined as necrosing fibers surrounded by infiltrating monocytes (Figure 1G). No regenerating fibers, characterized by small, round shape and centralized nuclei, were discernible at this time point in WT animals, but a notable few were seen in TG animals (arrows, Figure 1G). By day 5 after the injury, obvious differences begin to emerge. In WT animals, small regenerating fibers were visible but necrosing fibers and monocytes were still prevalent at the site of the injury (arrowheads, Figure 1G). While in the TG animals, the injury site harbors orderly arrangement of small regenerating fibers. Quantification of regenerating fiber number and CSA reveals that by 5 days post injury, TG animals show significant regenerative advantage over their WT counterparts. Both CSA of the regenerating fibers and the number of regenerating fibers were significantly greater for TG animals at 43.5% (n=5 or 6; p<0.03) and 33.0% (n=l l or 12; p<0.001), respectively (Figures IB and 1C). By day 7 post injury, the damage site appears architecturally similar between WT and TG animals, where both show a field of immature regenerating fibers without the infiltrating immune cells . However, quantification of the regenerating fibers revealed a regenerative advantage of the TG animals in the number of nascent regenerating fibers (Figure 1H). At 21 days after the injury, both WT and TG animals have restored their fiber size and number to that of the uninjured level (Figure 1J). These data demonstrate that the muscle specific activation of PPAR5 sufficiently bestows regenerative advantage, most prominently observed in the early stages of the regenerative process. Example 3

PPAR5 activation leads to temporal shift,

thus increased efficiency, of the regenerative process

Skeletal muscle regeneration is an intricately orchestrated process involving a variety of cell types. For example, immune cells, both neutrophils and macrophages, are necessary for the proper progression of regenerative process (Zacks et al., Muscle Nerve 5:152-161 (1982); Grounds et al., Cell Tissue Res 250:563-569 (1987); Teixeira et al. , Muscle Nerve 28(4):449-459 (2003); Summan et al., Am J Physiol Regul Integr Comp Physiol 290:R1488- R1495 (2006); Contreras-Shannon et al., Am J Physiol Cell Physiol 292:C953-967 (2007); Segawa et al., Exp Cell Res 314(17):3232-3244 (2008)). Additionally, various cytokines are necessary to promote chemotaxis of monocytes and also to directly regulate the activities of myogenic cells (Warren et al., Am J Physiol Cell Physiol 286(5):C1031-1036 (2004);

Yahiaoui et al, J Physiol 586:3991-4004 (2008); Chazaud et al, JCB 163(5): 1133- 1143 (2003)). Therefore, the temporal expression profiles of genes associated with various aspects of the regenerative process was determined.

Global, injury specific gene expression changes, were identified in VP16-PPAR5 animals by microarray. Comparing the gene expression profiles of injured TG to WT 3 days post-injury, 3257 genes that changed expression pattern, of those, 1375 of them were down regulated and 1882 were up regulated. Interestingly, genes involved in myogenesis and remodeling were robustly up-regulated by PPAR5 activation while those involved in inflammatory response were down regulated in injured TG muscles (Figure 2A).

Additionally, genes involved in developmental processes, angiogenesis and anti-apoptotic processes emerged from the analysis (Figure 2A). Relative expressions of regeneration markers reveal down-regulation of early makers (inflammatory genes) and up-regulation of regenerative/remodeling genes (myogenic, vascularization, ECM genes) in TG animals 3 days post injury (Figure 2B). Collectively, PPAR5 activation appears to control a network of genes involved directly in myogenesis and also in remodeling and repair processes after the injury.

Underlying phasic progression of the regenerative program is a temporally coordinated gene expression of a variety of contributing processes. In order to validate and temporally expand the microarray data, expression of CD68 (inflammation) and MyoD (myogenesis) were measured by Q-PCR at several time points over 7 days after injury

(Figures 2C and 2D). A temporal shift in the expression patterns of regenerative markers for TG animals compared to their WT littermates was observed. TG animals showed rapid induction of CD68 whose expressions peaked sooner and were subsequently down regulated earlier than in the WT animals. Interestingly, inflammatory markers studied here peaked at similar levels between the two genotypes, which indicates that TG animals do not completely suppress their inflammatory responses. Instead, it appears that the TG animals respond and resolve their inflammatory responses more efficiently, which is consistent with the accelerated restoration of muscle morphology observed. TG animals also show higher expression of perinatal myosin heavy chain gene, Myh8, at 7 days post injury, indicating more efficient reassembly of the contractile properties (Figure 2E). PPAR5 activation leads to a temporal shift in the expression patterns of regenerative markers, which together with the histology data, shows a role of PPAR5 in increasing regenerative efficiency.

Exampl 4

PPAR5 directs neovascularization via regulation of FGF1 This example describes adaptive responses bestowed by PPAR5 activation in the muscle which may contribute to the observed beneficial effects on regeneration.

Increased vasculature is one of the hallmarks of oxidative myofibers, which facilitates introduction of immune cells and also supports increased number of satellite cells. TG animals show increased expression of FGF1 in TA muscle (Figure 3D). Upon injury, TG animals maintain high expression of FGF1 expression (Figure 3D). Immunostaining transverse sections of uninjured TA from WT and TG animals revealed 36% increase in the number of CD31+ capillaries per field by PPAR5 activation (Figures 3A-C). Furthermore, after the injury, TG animals show increased expression of CD31, which is indicative of increased vascularity (Figures 3E-F). The induction of FGFla upon activation of PPAR delta with the GW1516 ligand was confirmed using a luciferase reporter assay (Figure 3G). FGF1 has been shown to be expressed in regenerating fibers in chronic disease models and has been implicated in myogenesis and regeneration (Oliver, Growth Factors. 1992;7(2):97- 106, 1992; Saito, 2000, Muscle Nerve. 23(4):490-7) and to increase microvasculature in adipocytes and PPAR5 directly regulates expression of FGFla isoform (Jonker et ah, Nature. 485(7398):391-4, 2012). Therefore, increased vascularity may contribute to the accelerated regenerative process observed in VP16-PPAR5 animals. Exampl

PPAR5 activation positively regulates quiescent satellite cell number

One of the first events following the injury is the proliferation of muscle resident progenitors, the satellite cells. This example describes results showing that the regenerative advantage observed in TG animals could be due to altered satellite cell homeostasis.

Nestin expression was used as a marker of satellite cells, and nestin-GFP;VP16- PPAR5 double transgenic animals were used to genetically label quiescent satellite cells(SCs) in vivo (Mignone et al, J Comp Neurol 469(3):311-324 (2004); Day et al, Dev Biol 304(1 ):246-259 (2007)). Gastrocnemius muscles were enzymatically digested to liberate individual fibers, then mounted for quantification (Figure 4A). While double transgenic animals averaged 1.01 SCs per mm of fiber length, GFP+ animals only had 0.15 SCs per mm, a 6.48 fold higher SC content on VP16- PPAR5 muscle fiber (Figure 4B).

Satellite cell activity was measured as myoblast proliferation elicited by the freeze burn injury in vivo. After the freeze burn injury, BrdU was intraperitoneally injected at 12 hrs, 24 hrs and 2 days after the injury and the muscles were harvested 7 days after the injury to calculate the ratio of BrdU+ to total nuclei. TG animals showed 40-60% increase in the number of BrdU+ proliferating cells at all three injection times (Figure 4C). Therefore, PPAR5 induced increase in the number of quiescent satellite cells yields higher number of fusion competent myoblasts, leading to the enhancement of regenerative capacity of the muscle.

Example 6

Acute pharmacological activation of PPAR5 confers regenerative advantage

Pharmacological activation of PPAR5 has been shown to induce PPAR5 target genes in fast-twitch hind limb muscles (Narkar et al, Cell 134(3):405-415 (2008)). To demonstrate that an acute pharmacological activation of PPAR5 can modulate regenerative process after injury, C57BL6J mice were treated with GW501516 (Sundai Chemicals, China) orally at 5 mg/kg for 4 days prior to and 5 days after the thermal injury to the TA.

Up-regulation of known PPAR5 target genes (PDK4, CPTlb, and catalase) was confirmed by QPCR, attesting to the successful delivery and activity of the PPAR5 ligand in the muscle (Figure 5A). While vehicle treated animals showed dye uptake in 7.6% of the cross sectional area (CSA), merely 4.9% of the muscle CSA was stained in the ligand treated animals (Figures 5B and 5C). Therefore, the drug treated animals showed 34.7% reduction in the proportion of stained area 5 days after the injury, demonstrating that pharmacological activation of PPAR5 enables accelerated restoration of myofiber integrity after the injury.

Moreover, BrdU injection at 48 hours after the injury revealed that PPAR5 activation promotes myoblast proliferation after the injury (Figure 5D). However, an increase in the number of quiescent satellite cells was not observed after 9 days or 4 weeks of ligand treatment. Since satellite cells do not undergo rapid turnover, length of ligand treatment may have been too short. Nonetheless, GW501516 treatment promoted myoblast proliferation in vivo after the injury, which may contribute to the accelerated regeneration after the injury.

The expression of inflammatory marker genes at 3 days after the injury was measured by QPCR. While the initial inflammatory responses are similarly generated with or without the PPAR5 ligand treatment at 12 hours after the injury, by 3 days after the injury, the expressions of inflammatory marker genes were significantly reduced by the PPAR5 agonist treatment (Figure 5E). This result is consistent with the known role of PPAR5 as an antiinflammatory, and also corroborates the data discussed earlier with the genetic over- expression of activated PPAR5 during muscle regeneration.

In summary, PPAR5 activation expedites skeletal muscle regeneration following an acute thermal injury. VP 16- PPAR5 transgenic animals showed increased satellite cell proliferation at the early phase of the regenerative process, which subsequently translated into increased CSA and the number of nascent regenerating fibers. Most interestingly, muscle specific over expression of PPAR5 seems to increase the resident satellite cell pool.

Increased satellite cell population on a muscle fiber seems to contribute to the accelerated resolution of the injury. These findings unveil a novel role for PPAR5 in the maintenance of skeletal muscle; as a potential therapeutic target for accelerated restoration of muscle mass after an acute injury and other atrophic conditions.

Notably, PPAR5 activation seems to promote rapid emergence of nascent fibers after the injury. There being no evidence of hyperplasia at 21 days after the injury when the regenerative process is essentially complete, it is concluded that the additional nascent fibers efficiently fuse with each other to restore mature fibers (Karpati G, Molnar MJ in Skeletal muscle repair and regeneration, eds Schiaffino S, Partridge T (Springer, Dordrecht), (2008)). While IGF-1 and myostatin seem to rely on fiber hypertrophy to augment regenerative progress, PPAR5 seems to employ a unique way to promote regeneration (Menetrey et ah, J Bone Joiny Surg Br 82(l): 131-7 (2000); Wagner et αΙ., Αηη Neurol 52(6): 832-6 (2002); Bogdanovich et al., Nature 420(6914):418-21 (2002)). Underlying this difference may be the increased number of quiescent satellite cells. Higher number of progenitor cells leads to the increase in post injury proliferating cells and consequent increase in the number of nascent fibers. While various growth factors and chemokines, including IGF-1 and myostatin, have been shown to enhance proliferation of satellite cells and promote regeneration, it is unclear whether any of them positively regulate the number of quiescent satellite cells (Husmann I et al., Cytokine Growth Factor Rev 7(3):249-258 (1996); McCroskery S et al., J Cell Biol 162(6): 1135-1147 (2003); Musaro A et al, Nat Genet 27: 195-200 (2001); Amthor H et al. , PNAS 106(18):7479-84 (2009)). The findings shown herein indicate a novel role of PPAR5 as a positive regulator of satellite cell pool. Interestingly, since rapid cell proliferation was not observed under normal conditions, PPAR5 mediated satellite cell expansion is transient and tightly regulated, most likely elicited by external stimuli, such as signals for postnatal growth and injury. In an adult muscle, satellite cell number is finite, diminishing

detrimentally in disease state and aging. It is of great therapeutic benefit if PPAR5 activation can bestow infinite abundance of satellite cell population throughout the life of an organism.

While enhancement in regenerative capacity was observed in both genetic and pharmacological models, the inherent differences in the experimental parameters is acknowledged. Orally administered GW501516 was delivered systemically, presumably activating PPAR5 in a variety of organs and cell types in the animal. However, in VP16- PPAR5 animals, activation of the PPAR5 receptors is limited to the mature muscle fibers. Additionally, genetic background of the animals may affect the efficiency of regeneration after an injury (Grounds and McGeachie, Cell Tissue Res 255(2):385-391 (1989); Roberts et al., J Anat 191:585-594 (1997)). Extramuscular effects of PPAR5 agonist administration may require further investigation when considering clinical use of GW501516 to augment muscle injury treatment. Recently, pharmacological activation of PPAR5 has been shown to improve sarcolemmal integrity in mdx mice (Miura et al., Hum mol Genet 18(23):4640-4649 (2009)).

The results herein expand previous understandings of the role of PPAR5 in muscle physiology. It is shown herein that PPAR5 not only controls running endurance and metabolic parameters in the muscle, but also its regenerative program. PPAR5 activation affects multiple facets of the regenerative program, exerting comprehensive but transient effects to expedite the progress. In view of these findings, PPAR5 may be pharmacologically targeted to enhance the regenerative capacity of the muscle after injury and possibly other degenerative conditions where satellite cell function is compromised. For example, PPAR5 activation can be used to treat other degenerative conditions such as aging induced satellite cell dysfunction and ensuing sarcopenia.

Example 7

PPAR6 activity screen

Cell Culture and Transfection: CV-1 cells were grown in DMEM+10 charcoal stripped FCS. Cells were seeded into 384-well plates the day before transfection to give a confluency of 50-80% at transfection. A total of 0.8 g DNA containing 0.64 micrograms pCMX-PPARDelta LBD, 0.1 micrograms pCMX.beta.Gal, 0.08 micrograms pGLMH2004 reporter and 0.02 micrograms pCMX empty vector was transfected per well using FuGene transfection reagent according to the manufacturer's instructions (Roche). Cells were allowed to express protein for 48 h followed by addition of compound.

Plasmids: Human PPAR5 was used to PCR amplify the PPAR5 LBD. The amplified cDNA ligand binding domain (LBD) of PPAR5 isoform was (PPAR5 amino acid 128 to C- terminus) and fused to the DNA binding domain (DBD) of the yeast transcription factor GAL4 by subcloning fragments in frame into the vector pCMX GAL (Sadowski et al. (1992), Gene 118, 137) generating the plasmids pCMX-PPARDelta LBD. Ensuing fusions were verified by sequencing. The pCMXMH2004 lucif erase reporter contains multiple copies of the GAL4 DNA response element under a minimal eukaryotic promoter (Hollenberg and Evans, 1988). pCMXpGal was generated.

Compounds: All compounds were dissolved in DMSO and diluted 1: 1000 upon addition to the cells. Compounds were tested in quadruple in concentrations ranging from 0.001 to 100 μΜ. Cells were treated with compound for 24 h followed by luciferase assay. Each compound was tested in at least two separate experiments.

Luciferase assay: Medium including test compound was aspirated and washed with PBS. 50μ1 PBS including 1 mM Mg++ and Ca++ were then added to each well. The luciferase assay was performed using the LucLite kit according to the manufacturer's instructions (Packard Instruments). Light emission was quantified by counting on a Perkin Elmer Envision reader. To measure 3-galactosidase activity 25 μΐ supernatant from each transfection lysate was transferred to a new 384 microplate. Beta-galactosidase assays were performed in the microwell plates using a kit from Promega and read in a Perkin Elmer Envision reader. The beta-galactosidase data were used to normalize (transfection efficiency, cell growth etc.) the lucif erase data.

Statistical Methods: The activity of a compound is calculated as fold induction compared to an untreated sample. For each compound the efficacy (maximal activity) is given as a relative activity compared to GW501516, a PPAR5 agonist. The EC₅₀ is the concentration giving 50% of maximal observed activity. EC₅₀ values were calculated via non-linear regression usingGraphPad PRISM (GraphPad Software, San Diego, Calif.).

Claims

We claim:

1. A method of activating PPAR5, comprising administering to the subject a

therapeutically effective amount of a PPAR5 agonist, thereby activating the PPAR5 protein, with the proviso that the PP a compound having the formula:

or pharmaceutically acceptable salts thereof,

wherein:

ring A is selected from a cycloalkylene, heterocycloalkylene, arylene or

heteroarylene;

ring B is selected from an aryl, heteroaryl, cycloalkyl, heterocycloalkyl,

cycloalkylene,

heterocycloalkylene, arylene or heteroarylene;

each R independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic, heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

each R 22 independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic,

heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

n is from 0 to 5;

m is from 0 to 4;

X is O, NR³⁰, sulfonyl, or S;

R 30 is selected from H or aliphatic, aryl, or cycloaliphatic;

L⁵ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -L 3 N(L 4 R 3 )L 3 -;

L is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -CR 23 R 24 -;

R 2"3^J and 24 are each independently selected from H, deuterium, halogen, aliphatic, alkyl, -C(0)OR²⁵ or -C(0)NR²⁵R²⁶;

R²⁵ and R²⁶ are each independently hydrogen, aliphatic or alkyl; Z is selected from R l^QO)- or a carboxyl bioisostere;

L¹ is a bond or -NR³⁰-;

R¹ is hydrogen, aliphatic, -OR^1A, -NR^1AR^1B, -C(0)R^1A, -S(0)₂R^1A, -C(0)OR^1A, -S(0)₂NR^1AR^1B or -C(0)NR^1AR^1B;

R^1A, R^1B are each independently hydrogen, aliphatic or alkyl;

L is selected from a bond, aliphatic, -C(O)-, alkylC(O)-, -C(0)alkyl-, or sulfonyl; L⁴ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene,

23 24

cycloalkylene, heterocycloalkylene or -CR R -;

R³ is selected from -OH, -OR^3A, -NR^3AR^3B, -C(0)R^3A, -S(0)2R^3A, -C(0)OR^3A, -S(0)₂NR^3AR^3B, -C(0)NR^3AR^3B, aliphatic, heteroaliphatic, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or R can be joined with an atom of ring B to form a fused ring system or may be joined with an atom of L to form a heterocyclic ring system; and

3 A 3B

R , R , are each independently hydrogen, aliphatic or alkyl. 2. The method of claim 1, wherein the PPAR5 protein is present in a subject, and contacting comprises administering the one or more PPAR5 agonists to the subject.

3. The method of claim 2, wherein activating the PPAR5 protein within the subject increases or maintains muscle mass or muscle tone in the subject.

4. A method of treating a PPAR5 related disease or condition in a subject, comprising administering to the subject in need thereof a therapeutically effective amount of one or more PPAR5 agonists,

with the proviso that the PP a compound having the formula:

or pharmaceutically acceptable salts thereof,

wherein:

ring A is selected from a cycloalkylene, heterocycloalkylene, arylene or

heteroarylene;

ring B is selected from an aryl, heteroaryl, cycloalkyl, heterocycloalkyl,

cycloalkylene, heterocycloalkylene, arylene or heteroarylene;

each R independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic, heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

each R 22 independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic,

heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

n is from 0 to 5;

m is from 0 to 4;

X is O, NR³⁰, sulfonyl, or S;

R 30 is selected from H or aliphatic, aryl, or cycloaliphatic;

L⁵ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -L 3 N(L 4 R 3 )L 3 -;

L is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -CR 23 R 24 -;

R 2"3^J and 24 are each independently selected from H, deuterium, halogen, aliphatic, alkyl, -C(0)OR²⁵ or -C(0)NR²⁵R²⁶;

R²⁵ and R²⁶ are each independently hydrogen, aliphatic or alkyl;

Z is selected from R¹L¹C(0)- or a carboxyl bioisostere;

L¹ is a bond or -NR³⁰-;

R¹ is hydrogen, aliphatic, -OR^1A, -NR^1AR^1B, -C(0)R^1A, -S(0)₂R^1A, -C(0)OR^1A, - S(0)₂NR^1AR^1B or -C(0)NR^1AR^1B;

R^1A, R^1B are each independently hydrogen, aliphatic or alkyl;

L is selected from a bond, aliphatic, -C(O)-, alkylC(O)-, -C(0)alkyl-, or sulfonyl;

L⁴ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -CR 23 R 24 -;

R³ is selected from -OH, -OR^3A, -NR^3AR^3B, -C(0)R^3A, -S(0)2R^3A, -C(0)OR^3A, - S(0)₂NR^3AR^3B, -C(0)NR^3AR^3B, aliphatic, heteroaliphatic, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or R can be joined with an atom of ring B to form a fused ring system or may be joined with an atom of L to form a heterocyclic ring system; and

R 3 A , R 3B , are each independently hydrogen, aliphatic or alkyl.

5. A method of increasing or maintaining muscle mass or muscle tone in a subject, comprising administering to the subject a therapeutically effective amount of one or more PPAR5 agonists,

with the proviso that the PP a compound having the formula:

or pharmaceutically acceptable salts thereof,

wherein:

ring A is selected from a cycloalkylene, heterocycloalkylene, arylene or

heteroarylene;

ring B is selected from an aryl, heteroaryl, cycloalkyl, heterocycloalkyl,

cycloalkylene,

heterocycloalkylene, arylene or heteroarylene;

each R independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic, heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, S0₃H, or acyl;

each R 22 independently is selected from deuterium, halogen, aryl, heteroaryl, aliphatic,

heteroaliphatic, cycloaliphatic, N0₂, OH, amino, amide, aminosulfonyl, carboxyl, carboxyl ester, alkylsulfonyl, SO₃H, or acyl;

n is from 0 to 5;

m is from 0 to 4;

X is O, NR³⁰, sulfonyl, or S;

R 30 is selected from H or aliphatic, aryl, or cycloaliphatic;

L⁵ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -L 3 N(L 4 R 3 )L 3 -;

L is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene, cycloalkylene, heterocycloalkylene or -CR 23 R 24 -;

R 2"3^J and 24 are each independently selected from H, deuterium, halogen, aliphatic, alkyl, -C(0)OR²⁵ or -C(0)NR²⁵R²⁶;

R²⁵ and R²⁶ are each independently hydrogen, aliphatic or alkyl;

Z is selected from R¹L¹C(0)- or a carboxyl bioisostere; L¹ is a bond or -NR³⁰-;

R¹ is hydrogen, aliphatic, -OR^1A, -NR^1AR^1B, -C(0)R^1A, -S(0)₂R^1A, -C(0)OR^1A, -S(0)₂NR^1AR^1B or -C(0)NR^1AR^1B;

R^1A, R^1B are each independently hydrogen, aliphatic or alkyl;

L is selected from a bond, aliphatic, -C(O)-, alkylC(O)-, -C(0)alkyl-, or sulfonyl;

L⁴ is selected from a bond, aliphatic, heteroaliphatic, arylene, heteroarylene,

23 24

cycloalkylene, heterocycloalkylene or -CR R -;

R³ is selected from -OH, -OR^3A, -NR^3AR^3B, -C(0)R^3A, -S(0)2R^3A, -C(0)OR^3A, -S(0)₂NR^3AR^3B, -C(0)NR^3AR^3B, aliphatic, heteroaliphatic, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or R can be joined with an atom of ring B to form a fused ring system or may be joined with an atom of L to form a heterocyclic ring system; and

3 A 3B

R , R , are each independently hydrogen, aliphatic or alkyl.

6. The method of claim 4, wherein the PPAR5 related disease is a vascular disease, muscular disease, demyelinating disease, or a metabolic disease.

7. The method of claim 6, wherein the muscular disease is a muscular dystrophy disease.

8. The method of claim 7, wherein the muscular dystrophy disease is Duchenne muscular dystrophy, Becker muscular dystrophy, limb-girdle muscular dystrophy, congenital muscular dystrophy, facioscapulohumeral muscular dystrophy, myotonic muscular dystrophy, oculopharyngeal muscular dystrophy, distal muscular dystrophy, or Emery-Dreifuss muscular dystrophy. 9. The method of claim 7, wherein the demyelinating disease is multiple sclerosis, Charcot-Marie-Tooth disease, Pelizaeus-Merzbacher disease, encephalomyelitis,

neuromyelitis optica, adrenoleukodystrophy, or Guillian-Barre syndrome.

10. The method of claim 7, wherein the metabolic disease is obesity,

hypertriglyceridemia, hyperlipidemia, hypoalphalipoproteinemia, hypercholesterolemia, dyslipidemia, Syndrome X, or Type II diabetes mellitus.

11. The method of claim 4, wherein the PPAR5-related disease or condition is a muscle structure disorder, a neuronal activation disorder, a muscle fatigue disorder, a muscle mass disorder, a mitochondrial disease, a beta oxidation disease, a metabolic disease, a cancer, a vascular disease, an ocular vascular disease, or a muscular eye disease.

12. The method of claim 11, wherein:

the muscle structure disorder is selected from Bethlem myopathy, central core disease, congenital fiber type disproportion, distal muscular dystrophy (MD), Duchenne & Becker MD, Emery-Dreifuss MD, facioscapulohumeral MD, hyaline body myopathy, limb- girdle MD, a muscle sodium channel disorders, myotonic chondrodystrophy, myotonic dystrophy, myotubular myopathy, nemaline body disease, oculopharyngeal MD, or stress urinary incontinence;

the neuronal activation disorder is selected from amyotrophic lateral sclerosis, Charcot-Marie-Tooth disease, Guillain-Barre syndrome, Lambert-Eaton syndrome, multiple sclerosis, myasthenia gravis, nerve lesion, peripheral neuropathy, spinal muscular atrophy, tardy ulnar nerve palsy, or toxic myoneural disorder;

the muscle fatigue disorder is selected from chronic fatigue syndrome, diabetes (type I or II), glycogen storage disease, fibromyalgia, Friedreich's ataxia, intermittent claudication, lipid storage myopathy, MELAS, mucopolysaccharidosis, Pompe disease, or thyrotoxic myopathy; the muscle mass disorder is cachexia, cartilage degeneration, cerebral palsy, compartment syndrome, critical illness myopathy, inclusion body myositis, muscular atrophy (disuse), sarcopenia, steroid myopathy, or systemic lupus erythematosus;

the mitochondrial disease is selected from Alpers's Disease, CPEO-Chronic progressive external ophthalmoplegia, Kearns-Sayra Syndrome (KSS), Leber Hereditary Optic Neuropathy (LHON), MELAS-Mitochondrial myopathy, encephalomyopathy, lactic acidosis, and stroke-like episodes, MERRF-Myoclonic epilepsy and ragged-red fiber disease, NARP-neurogenic muscle weakness, ataxia, and retinitis pigmentosa, or Pearson Syndrome; the beta oxidation disease is selected from systemic carnitine transporter, carnitine palmitoyltransferase ( CPT ) II deficiency, very long-chain acyl-CoA dehydrogenase

(LCHAD or VLCAD) deficiency, trifunctional enzyme deficiency, medium-chain acyl-CoA dehydrogenase (MCAD) deficiency, short-chain acyl-CoA dehydrogenase (SCAD) deficiency or riboflavin-responsive disorders of β -oxidation (RR -MADD);

the metabolic disease is selected from hyperlipidemia, dyslipidemia,

hypercholesterolemia, hypertriglyceridemia, HDL hypocholesterolemia, LDL hypercholesterolemia and/or HLD non-cholesterolemia, VLDL hyperproteinemia, dyslipoproteinemia, apolipoprotein A-I hypoproteinemia, atherosclerosis, disease of arterial sclerosis, disease of cardiovascular systems, cerebrovascular disease, peripheral circulatory disease, metabolic syndrome, syndrome X, obesity, diabetes (type I or II), hyperglycemia, insulin resistance, impaired glucose tolerance, hyperinsulinism, diabetic complication, cardiac insufficiency, cardiac infarction, cardiomyopathy, hypertension, Non-alcoholic fatty liver disease (NAFLD), Nonalcoholic steatohepatitis (NASH), thrombus, Alzheimer disease, neurodegenerative disease, demyelinating disease, multiple sclerosis, adrenal leukodystrophy, dermatitis, psoriasis, acne, skin aging, trichosis, inflammation, arthritis, asthma,

hypersensitive intestine syndrome, ulcerative colitis, Crohn's disease, or pancreatitis;

the cancer is a cancer of the colon, large intestine, skin, breast, prostate, ovary, or lung;

the vascular disease is selected from peripheral vascular insufficiency, peripheral vascular disease, intermittent claudication, peripheral vascular disease (PVD), peripheral artery disease (PAD), peripheral artery occlusive disease (PAOD), or peripheral obliterative arteriopathy;

the ocular vascular disease is selected from age-related macular degeneration (AMD), stargardt disease, hypertensive retinopathy, diabetic retinopathy, retinopathy , macular degeneration, retinal haemorrhage, or glaucoma; and

the muscular eye disease is selected from strabismus, progressive external

ophthalmoplegia, esotropia, exotropia, a disorder of refraction and accommodation, hypermetropia, myopia, astigmatism, anisometropia, presbyopia, a disorders of

accommodation, or internal ophthalmoplegia. 13. The method of any one of claims 1 - 12, wherein the subject is a sedentary or immobilized subject.

14. The method of any one of claims 1 - 13, wherein the subject is an exercising subject. 15. The method of claim 4, wherein the PPAR5-related disease is a mitochondrial disease.

16. The method of claim 15, wherein the mitochondrial disease is selected from the group consisting of Alpers's Disease, CPEO-Chronic progressive external ophthalmoplegia, Kearns-Sayra Syndrome (KSS), Leber Hereditary Optic Neuropathy (LHON), MELAS- Mitochondrial myopathy, encephalomyopathy, lactic acidosis, and stroke-like episodes, MERRF-Myoclonic epilepsy and ragged-red fiber disease, NARP-neurogenic muscle weakness, ataxia, and retinitis pigmentosa, and Pearson Syndrome. 17. The method of any one of claims 1 - 16, wherein administering comprises intraarticular, intravenous, intramuscular, intratumoral, intradermal, intraperitoneal, subcutaneous, oral, topical, intrathecal, inhalational, transdermal, or rectal administration.

18. The method of any of claims 1 - 17, wherein the one or more compounds

administered to the subject at a dose of about 1 mg/kg to about 10 mg/kg.

19. The method of any one of claims 1 - 18, wherein the PPAR5 agonist is a compound of Formula (I):

(I);

or a pharmaceutically acceptable salt thereof,

wherein:

Ar 1 is phenyl, which is optionally substituted with from one to five R 7 substituents independently selected from the group consisting of halogen, (Ci-C8)alkyl, halo(Ci-C8)alkyl, and -OR²;

Ar 2 is phenyl, which is optionally substituted with from one to four R 8 substituents independently selected from the group consisting of halogen, (Ci-Cg)alkyl, halo(Ci-Cg)alkyl, and -OR²;

L is a member selected from the group consisting of -CH₂S- and -CH₂0-;

K is a member selected from the group consisting of a covalent bond and -OCH₂-

Z is C0₂R⁶;

R¹ is selected from the group consisting of H and (Ci-C8)alkyl;

each R 2 and R 3 is a member independently selected from the group consisting of H, (C C₈)alkyl, halo(C C₈)alkyl,— X³OR⁹, aryl, aryl(C C₄)alkyl, and heteroaryl, or optionally, if both present on the same substituent, may be joined together to form a three- to eight-membered ring system; R⁶ is a member selected from the group consisting of H, (Ci-Cg)alkyl, halo(Cr C₈)alkyl,— X⁴OR²,— X⁴NR²R³, (C₂-C₈)alkenyl, (C₃-C₇)cycloalkyl, heterocyclyl, aryl (C C₄)alkyl; and aryl(C₂-C8)alkenyl;

R⁹ is a member selected from the group consisting of H, (Ci-C₈)alkyl, halo(Ci- Cg)alkyl, aryl, aryl(Ci-C₄)alkyl, and heteroaryl;

each X³ and X⁴ is a member independently selected from the group consisting of (Cp C₄) alkylene, (C₂-C4)alkenylene, and (C₂-C4)alkynylene.

20. The method of any one of claims 1 - 18, wherein the PPAR5 agonist is a compound of Formula (II):

(II);

or a pharmaceutically acceptable salt thereof,

wherein:

X represents a COOH (or a hydrolysable ester thereof),

X^I is O or S, and the depicted bond with a dashed line is a single bond;

X represents O, S;

R 1 and R 2 independently represent H, CH₃, OCH₃ or halogen;

n is 1 or 2;

one of Y and Z is N and the other is S or O;

y represents 1 or 2;

each R independently represents CF₃ or halogen

or a pharmaceutically acceptable salt thereof.

21. The method of any one of claims 1 - 18, wherein the PPAR5 agonist is a

of Formula (III):

(in);

or a pharmaceutically acceptable salt thereof, wherein:

A is a saturated or unsaturated hydrocarbon chain having from 3 to 5 atoms, forming a five- to seven-membered ring;

T is selected from the group consisting of -C(0)OH, -C(0)NH₂, and tetrazole;

Z is O, S or NR;

n is 0, 1, or 2;

r and s are independently 0 or 1 ;

1 2

R' and R" are independently selected from the group consisting of hydrogen, halo, optionally substituted lower alkyl, optionally substituted lower heteroalkyl, optionally substituted lower alkoxy, and lower perhaloalkyl or together may form an optionally substituted cycloalkyl;

1 2 3

X X\ and X^J are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted cycloalkyl, halogen, perhaloalkyl, hydroxy, optionally substituted lower alkoxy, nitro, cyano, and NH₂;

G₂ is selected from the group consisting of a saturated or unsaturated heterocycloalkyl linker, optionally substituted with X₄ and X5;

X⁴ and X⁵ are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, halogen, lower perhaloalkyl, hydroxy, optionally substituted lower alkoxy, nitro, cyano, NH₂, and C0₂R, or X₄ and X5 together may form a carbocycle;

R is selected from the group consisting of optionally substituted lower alkyl and hydrogen;

G3 is selected from the group consisting of a bond, a double bond,— (CR³R⁴)_m— , carbonyl, and— (CR³R⁴)_mCR³=CR⁴— ;

m is 0, 1, or 2;

R³ and R⁴ are independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted lower alkoxy, optionally substituted aryl, lower perhaloalkyl, cyano, and nitro;

G⁴ is selected from the group consisting of hydrogen, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted cycloheteroalkyl, optionally substituted cycloheteroaryl, optionally substituted cycloalkenyl, and— N=(CR⁵R⁶); and R⁵ and R⁶ are independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted cycloalkenyl, and optionally substituted cycloheteroalkyl.

The method of claim 19, wherein the compound has the structure of Formula (la):

or a pharmaceutically acceptable salt thereof.

The method of claim 20, wherein the compound has the structure of Formula (Ila):

(Ila)

or a pharmaceutically acceptable salt thereof.

24. The method of claim 21 wherein the compound has the structure of Formula (Ilia):

(Ilia);

or a pharmaceutically acceptable salt thereof. The method of claim 21 wherein the compound has the structure of Formula (Illb)

(nib);

or a pharmaceutically acceptable salt thereof.