Modulators of DUX4 for Regulation of Muscle Function

Abstract

Disclosed herein are methods and compositions for the treatment of facioscapulohumeral muscular dystrophy and other muscle diseases or disorders. In some cases, the methods and compositions involve the use of methyltransferase inhibitors to inhibit or repress DUX4 expression in muscle cells. Further disclosed herein are methods and cell-based assays for screening compounds for the treatment of facioscapulohumeral muscular dystrophy and other muscle diseases.

1 - 42 . (canceled) 
     
     
         43 . A kit comprising:
 a) a pharmaceutical composition comprising a compound of Formula (I)   
       
         
           
           
               
               
           
         
       
       or a pharmaceutically acceptable salt thereof, wherein:
 A is selected from C 5-12  carbocycle and 5- to 12-membered heterocycle; 
 B is selected from C 5-12  carbocycle and 5- to 12-membered heterocycle; 
 C is selected from bond, C 5-12  carbocycle, and 5- to 12-membered heterocycle; 
 D is selected from bond, C 5-12  carbocycle, and 5- to 12-membered heterocycle; 
 each of L 1  and L 2  is independently selected from bond, 
 —O—, —S—, —N(R 51 )—, N(R 51 )CH 2 —, 
 —C(O)—, —C(O)O—, —OC(O)—, —OC(O)0 2 -, 
 —C(O)N(R 51 )—, —C(O)N(R 51 )C(O)—, —C(O)N(R 51 )C(O)N(R 51 )—, —N(R 51 )C(O)—, —N(R 51 )C(O)N(R 51 )—, —N(R 51 )C(O)O—, —OC(O)N(R 51 )—, 
 —C(NR 51 )—, —N(R 51 )C(NR 51 )—, —C(NR 51 )N(R 51 )—, —N(R 51 )C(NR 51 ) N(R 51 )—, —S(O) 2 , 
 —OS(O)—, —S(O)O—, —S(O)—, —OS(O) 2 —, 
 —OS(O) 2 O—, —N(R 51 )S(O) 2 —, —S(O) 2 N(R 51 )—, 
 —N(R 51 )S(O)—, —S(O)N(R 51 )—, —N(R 51 )S(O) 2 N(R 51 )—, and —N(R 51 )S(O)N(R 51 )— or from alkylene, alkenylene, alkynylene, heteroalkylene, heteroalk-enylene, and heteroalkynylene, each of which is optionally substituted with one or more R 50 , 
 R 50  is, at each occurrence, independently selected from: 
 halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ), 
 —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , 
 —S(═O) 2 R 52   2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , 
 —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)NR 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , 
 —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , 
 —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , 
 —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ); 
 C 1-10  alkyl, C 1-10  alkenyl, and C 2-10  alkynyl, each of which is independently optionally substituted at each occurrence with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O)N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , 
 —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , 
 —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , 
 —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , 
 —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , 
 —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , 
 —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), 
 —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ) C 3-12  carbocycle, and 3- to 12-membered heterocycle; and 
 C 3-12  carbocycle and 3- to 12-membered heterocycle, wherein each C 3-12  carbocycle and 3- to 12-membered heterocycle in R 50  is independently optionally substituted with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 1-6  alkyl, C 1-6  haloalkyl, C 2-6  alkenyl, and C 2-6  alkynyl; 
 R 51  is independently selected at each occurrence from: 
 hydrogen, —C(O) R 52 , —C(O)OR 52 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 ; C 1-6  alkyl, C 2-6  alkenyl, and C 2-6  alkenyl, each of which is independently optionally substituted at each occurrence with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 3-12  carbocycle and 3- to 12-membered heterocycle; and 
 C 3-12  carbocycle and 3- to 12-membered heterocycle, wherein each C 3-12  carbocycle and 3- to 12-membered heterocycle in R 51  is independently optionally substituted with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 1-6  alkyl, C 1-6  haloalkyl, C 1-6  alkenyl, and C 2-6  alkynyl; 
 R 52  is independently selected at each occurrence from hydrogen; and C 1-20  alkyl, C 2-20  alkenyl, C 2-20  alkynyl, 1- to 6-membered heteroalkyl, C 3-12  carbocycle, and 3- to 12-membered heterocycle, each of which is optionally substituted by halogen, —CN, —NO 2 , —NH 2 , —NHCH 3 , —NHCH 2 CH 3 , ═O, —OH, —OCH 3 , —OCH 2 CH 3 , C 3-12  carbocycle, or 3- to 6-membered heterocycle; 
 R 53  and R 54  are taken together with the nitrogen atom to which they are attached to form a heterocycle, optionally substituted with one or more R 50 ; 
 each of R A , R B  and R C  is independently selected from R 50 ; 
 R D  is, at each occurrence, independently selected from hydrogen or R 50 ; and 
 each of m, n, p, and q is independently an integer from 0-12; and 
 b) packaging materials. 
 
     
     
         44 . A method of determining D4Z4 de-repression activity of a compound in a cell, comprising:
 administering a DNA methyltransferase inhibitor to the cell, thereby inducing DUX4 expression in the cell;   administering a compound to the cell, wherein the compound is of Formula (I):   
       
         
           
           
               
               
           
         
       
       or a pharmaceutically acceptable salt thereof, wherein:
 A is selected from C 5-12  carbocycle and 5- to 12-membered heterocycle; 
 B is selected from C 5-12  carbocycle and 5- to 12-membered heterocycle; 
 C is selected from bond, C 5-12  carbocycle, and 5- to 12-membered heterocycle; 
 D is selected from bond, C 5-12  carbocycle, and 5- to 12-membered heterocycle; 
 each of L 1  and L 2  is independently selected from bond, 
 —O—, —S—, —N(R 51 )—, N(R 51 )CH 2 —, 
 —C(O)—, —C(O)O—, —OC(O)—, —OC(O)0 2 -, 
 —C(O)N(R 51 )—, —C(O)N(R 51 )C(O)—, —C(O)N(R 51 )C(O)N(R 51 )—, —N(R 51 )C(O)—, —N(R 51 )C(O)N(R 51 )—, —N(R 51 )C(O)O—, —OC(O)N(R 51 )—, 
 —C(NR 51 )—, —N(R 51 )C(NR 51 )—, —C(NR 51 )N(R 51 )—, —N(R 51 )C(NR 51 ) N(R 51 )—, —S(O) 2 , 
 —OS(O)—, —SO(O)O—, —S(O)—, —OS(O)0 2 -, 
 —OS(O) 2 O—, —N(R 51 )S(O) 2 —, —S(O) 2 N(R 51 )—, 
 —N(R 51 )S(O)—, —S(O)N(R 51 )—, —N(R 51 )S(O) 2 N(R 51 )—, and —N(R 51 )S(O)N(R 51 )— or from alkylene, alkenylene, alkynylene, heteroalkylene, heteroalk-enylene, and heteroalkynylene, each of which is optionally substituted with one or more R 50 ; 
 R 50  is, at each occurrence, independently selected from: 
 halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , 
 —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , 
 —S(═O) 2 R 52   2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , 
 —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)NR 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , 
 —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , 
 —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , 
 —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ); 
 C 1-10  alkyl, C 1-10  alkenyl, and C 2-10  alkynyl, each of which is independently optionally substituted at each occurrence with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ), —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , 
 —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , 
 —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , 
 —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , 
 —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , 
 —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , 
 —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), 
 —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 3-12  carbocycle, and 3- to 12-membered heterocycle; and 
 C 3-12  carbocycle and 3- to 12-membered heterocycle, wherein each C 3-12  carbocycle and 3- to 12-membered heterocycle in R 50  is independently optionally substituted with one or more substituents selected from halogen —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ), —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 1-6  alkyl, C 1-6  haloalkyl, C 2-6  alkenyl, and C 2-6  alkynyl; 
 R 51  is independently selected at each occurrence from: 
 hydrogen, —C(O) R 52 , —C(O)OR 52 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 ; C 1-6  alkyl, C 2-6  alkenyl, and C 2-6  alkynyl each of which is independently optionally substituted at each occurrence with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ) 2 , —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 5 —P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 3-12  carbocycle and 3- to 12-membered heterocycle; and 
 C 3-12  carbocycle and 3- to 12-membered heterocycle, wherein each C 3-12  carbocycle and 3- to 12-membered heterocycle in R 51  is independently optionally substituted with one or more substituents selected from halogen, —NO 2 , —CN, —OR 52 , —SR 52 , —N(R 52 ), —NR 53 R 54 , —S(═O)R 52 , —S(═O) 2 R 52 , —S(═O) 2 N(R 52 ) 2 , —S(═O) 2 NR 53 R 54 , —NR 52 S(═O) 2 R 52 , —NR 52 S(═O) 2 N(R 52 ) 2 , —NR 52 S(═O) 2 NR 53 R 54 , —C(O)R 52 , —C(O)OR 52 , —OC(O)R 52 , —OC(O)OR 52 , —OC(O)N(R 52 ) 2 , —OC(O)N 53 R 54 , —NR 52 C(O)R 52 , —NR 52 C(O)OR 52 , —NR 52 C(O)N(R 52 ) 2 , —NR 52 C(O)NR 53 R 54 , —C(O)N(R 52 ) 2 , —C(O)NR 53 R 54 , —P(O)(OR 52 ) 2 , —P(O)(R 52 ) 2 , —P(O)(OR 52 )(R 52 ), —P(O)(NR 52 )(R 52 ), —NR 52 P(O)(R 52 ), —P(O)(NR 52 )(OR 52 ), —P(O)(NR 52 ) 2 , ═O, ═S, ═N(R 52 ), C 1-6  alkyl, C 1-6  haloalkyl, C 1-6  alkenyl, and C 2-6  alkynyl; 
 R 52  is independently selected at each occurrence from hydrogen; and C 1-20  alkyl, C 2-20  alkenyl, C 2-20  alkynyl, 1- to 6-membered heteroalkyl, C 3-12  carbocycle, and 3- to 12-membered heterocycle, each of which is optionally substituted by halogen, —CN, —NO 2 , —NH 2 , —NHCH 3 , —NHCH 2 CH 3 , ═O, —OH, —OCH 3 , —OCH 2 CH 3 , C 3-12  carbocycle, or 3- to 6-membered heterocycle; 
 R 53  and R 54  are taken together with the nitrogen atom to which they are attached to form a heterocycle, optionally substituted with one or more R 50 ; 
 each of R A , R B  and R C  is independently selected from R 50 ; 
 R D  is, at each occurrence, independently selected from hydrogen or R 50 ; and 
 each of m, n, p, and q is independently an integer from 0-12; and 
 measuring DUX4 expression, thereby determining D4Z4 de-repression activity of the compound in the cell. 
 
     
     
         45 . The method of  claim 44 , wherein the DNA methyl-transferase inhibitor is a nucleoside analogue. 
     
     
         46 . The method of  claim 45 , wherein the nucleoside analogue is a ribonucleoside analogue. 
     
     
         47 . The method of  claim 45 , wherein the nucleoside analogue is a deoxyribonucleoside analogue. 
     
     
         48 . The method of  claim 45 , wherein the nucleoside analogue is an adenosine or deoxyadenosine analogue, or a guanosine or deoxyguanosine analogue, or a uridine or thymidine analogue, or a cytidine or deoxycytidine analogue. 
     
     
         49 - 52 . (canceled) 
     
     
         53 . The method of  claim 45 , wherein the nucleoside analogue is selected from Table 2. 
     
     
         54 . The method of  claim 44 , wherein the cell is a muscle lineage. 
     
     
         55 . The method of  claim 54 , wherein the muscle lineage cell is selected from the group consisting of skeletal muscle cell, muscle precursor cell, myotube, myoblast, and satellite cell. 
     
     
         56 . (canceled) 
     
     
         57 . (canceled) 
     
     
         58 . (canceled) 
     
     
         59 . (canceled) 
     
     
         60 . (canceled) 
     
     
         61 . (canceled) 
     
     
         62 . (canceled) 
     
     
         63 . A method of reducing DUX4 expression in a subject, comprising: administering a histone methyltransferase inhibitor to the subject, thereby reducing expression of DUX4, wherein the subject has symptoms of facioscapulohumeral muscular dystrophy (FSHD). 
     
     
         64 . The method of  claim 63 , wherein the reduction of expression of DUX4 occurs in skeletal muscle of the subject. 
     
     
         65 . The method of  claim 63 , wherein the subject has facioscapulohumeral muscular dystrophy (FSHD). 
     
     
         66 . The method of  claim 63 , wherein the subject is suspected of having facioscapulohumeral muscular dystrophy (FSHD). 
     
     
         67 . The method of  claim 63 , wherein the subject is human. 
     
     
         68 . The method of  claim 63 , wherein the histone methyltransferase inhibitor is administered to the subject orally, intravenously, intramuscularly, subcutaneously, or transdermally. 
     
     
         69 . The method of  claim 43 , further comprising monitoring the subject for reduction in symptoms of facioscapulohumeral muscular dystrophy (FSHD). 
     
     
         70 . The method of  claim 43 , further comprising monitoring the subject for reduction in DUX4 expression.