Stabilized hnf4a sarna compositions and methods of use

Abstract

The invention relates to saRNA targeting an HNF4a transcript and therapeutic compositions comprising said saRNA. Methods of using the therapeutic compositions are also provided.

1 . A synthetic isolated small activating RNA (saRNA) comprises at least one chemical modification, wherein the saRNA up-regulates the expression of a target gene, wherein the target gene is HNF4a, wherein the saRNA is double-stranded and has a formula of:
   Passenger (Sense or SS): 5′ overhang1−NT1−(XXX-NT2)n−overhang2 3′,
     Guide (Antisense or AS): 3′ overhang3−NT1′−(YYY-NT2′)n−overhang4 5′,   (I)
   
       wherein:
 each strand is 14-30 nucleotides in length, 
 each of overhang1, overhang2, overhang3 and overhang4 independently represents an oligonucleotide sequence comprising 0-5 nucleotides, 
 NT1 and NT1′ represent an oligonucleotide sequence comprising 0-20 nucleotides, and wherein NT1 is complementary to NT1′, 
 each of XXX-NT2 and YYY-NT2′ independently represents a motif of consecutive nucleotides, wherein the first 3 consecutive nucleotides have the same chemical modification, followed by an oligonucleotide sequence comprising 0-20 nucleotides, and wherein XXX is complementary to YYY, and NT2 is complementary to NT2′, 
 each of NT1, NT2, NT1′, and NT2′ comprises at least one chemical modification, and n is a number between 1 and 5. 
 
     
     
         2 . The saRNA of  claim 1 , wherein the guide strand of the saRNA having formula (I) comprises a sequence that is at least 80% complementary to a targeted sequence located in the TSS core on the template strand of the target gene. 
     
     
         3 . The saRNA of  claim 1 , wherein at least one nucleotide on the passenger strand and its complementary nucleotide on the guide strand have different sugar modifications. 
     
     
         4 . The saRNA of  claim 3 , wherein NT1, NT2, NT1′, and NT2′ have alternating sugar modifications. 
     
     
         5 . The saRNA of  claim 4 , wherein NT1, NT2, NT1′, and NT2′ have alternating 2′-OMe and 2′-F modifications. 
     
     
         6 . The saRNA of  claim 5 , wherein the 3 consecutive nucleotides of XXX have 2′-OMe modifications and the 3 consecutive nucleotides of YYY have 2′-F modifications, or wherein the 3 consecutive nucleotides of XXX have 2′-F modifications and the 3 consecutive nucleotides of YYY have 2′-OMe modifications. 
     
     
         7 . The saRNA of  claim 1 , wherein the modification of XXX or YYY is different from the modifications of the immediately adjacent nucleotides on both sides of XXX or YYY. 
     
     
         8 . The saRNA of  claim 1 , wherein overhang1, overhang2, overhang3 and/or overhang4 comprises uu, an inverted dT, or an inverted abasic nucleoside. 
     
     
         9 . A conjugate comprising a synthetic isolated small activating RNA (saRNA) of  claim 1  and a carbohydrate moiety. 
     
     
         10 . The conjugate of  claim 9 , wherein the carbohydrate moiety is a N-Acetyl-Galactosamine (GalNAc) moiety. 
     
     
         11 . The conjugate of  claim 10 , wherein the GalNAc moiety is a triantennary GalNAc-cluster. 
     
     
         12 . The conjugate of  claim 10 , wherein the GalNAc moiety is attached to the saRNA via a linker. 
     
     
         13 . The conjugate of  claim 12 , wherein the linker is NH2C6. 
     
     
         14 . The conjugate of  claim 12 , wherein the linker is attached to the passenger strand of the saRNA. 
     
     
         15 . The conjugate of  claim 14 , wherein the linker is attached to the 3′ end or 5′ end of the passenger strand. 
     
     
         16 . The conjugate of  claim 15 , wherein a phosphorothioate linkage is located between the linker and the 3′ end or 5′ end of the passenger strand. 
     
     
         17 . The conjugate of  claim 14 , wherein the linker is attached to an internal nucleotide of the passenger strand. 
     
     
         18 . A pharmaceutical composition comprising the saRNA of  claim 1  and at least one pharmaceutically acceptable excipient. 
     
     
         19 . A pharmaceutical composition comprising the conjugate of  claim 9  and at least one pharmaceutically acceptable excipient. 
     
     
         20 . A method of delivering an saRNA to cells comprising administering the conjugate of  claim 9  to cells without any transfection agents. 
     
     
         21 . A method of up-regulating the expression of a target gene, comprising administering the saRNA of  claim 1 , wherein the target gene is HNF4a. 
     
     
         22 . A method of up-regulating the expression of a target gene, comprising administering the conjugate of  claim 9 , wherein the target gene is HNF4a.