siRNA-mediated gene silencing of synuclein

Abstract

The present invention is directed to small interfering RNAs that down regulate expression of a synuclein gene and methods of using the small interfering RNAs.

1 . A small interfering RNA (siRNA) that down regulates expression of a synuclein gene.  
     
     
         2 . The siRNA of  claim 1  wherein said siRNA comprises a nucleotide sequence substantially complementary to 15-30 consecutive nucleotides of SEQ. ID. NO.: 1.  
     
     
         3 . The siRNA of  claim 1  wherein said nucleotide sequence is substantially complementary to 15-30 consecutive nucleotides of SEQ ID. NO.: 6.  
     
     
         4 . The siRNA of  claim 1  wherein said nucleotide sequence is substantially complementary to 15 to 21 consecutive nucleotides SEQ. ID. NO.:7.  
     
     
         5 . The siRNA of  claim 1  wherein said nucleotide sequence is complementary to SEQ ID NO: 7.  
     
     
         6 . The siRNA of  claim 1  wherein said synuclein gene is wild-type α-synuclein.  
     
     
         7 . The siRNA of  claim 1  wherein said synuclein gene is a mutant α-synuclein gene.  
     
     
         8 . The siRNA of  claim 7  wherein said mutant gene comprises guanine to cytosine at position 134 of SEQ. ID. NO. 1.  
     
     
         9 . The siRNA of  claim 7  wherein said mutant gene comprises guanine to adenosine at position 203 of SEQ. ID. NO. 1.  
     
     
         10 . The siRNA of  claim 7  wherein said siRNA down regulates expression of said mutant α-synuclein gene and not a wild-type α-synuclein gene.  
     
     
         11 . The siRNA of  claim 1  wherein said siRNA comprises a vector encoding said siRNA.  
     
     
         12 . The siRNA of  claim 11  wherein said vector is a viral vector.  
     
     
         13 . The siRNA of  claim 12  wherein said vector is selected from the group consisting of adenoviral, lentiviral, poliovirus, adeno-associated viral, HSV, and murine Maloney-based viral vector.  
     
     
         14 . The siRNA of  claim 11  wherein said vector comprises a constitutive promoter or a regulatable promoter operatively linked to a nucleotide sequence encoding said siRNA.  
     
     
         15 . The siRNA of  claim 11  wherein said vector comprises a Pol III H-1 promoter operatively linked to said siRNA.  
     
     
         16 . The siRNA of  claim 1  wherein said siRNA forms a hairpin structure comprising a duplex structure and a loop structure.  
     
     
         17 . The siRNA of  claim 11  wherein said vector comprises a nucleotide sequence substantially identical to SEQ ID NO: 21.  
     
     
         18 . The siRNA of  claim 1  wherein said siRNA comprises a double-stranded RNA comprising a first strand that is complementary to a mRNA encoded by SEQ ID NO: 1, or a fragment thereof; and a second strand that is complementary to said first strand.  
     
     
         19 . The siRNA of  claim 18  wherein said duplex is less than 30 nucleotides in length.  
     
     
         20 . The siRNA of  claim 17  wherein said duplex comprises SEQ ID NO: 19 and SEQ ID NO: 20.  
     
     
         21 . The siRNA of  claim 1  wherein said synuclein gene is associated with a condition amenable to treatment with the siRNA.  
     
     
         22 . The siRNA of  claim 21  wherein said condition is a neurodegenerative disease.  
     
     
         23 . The siRNA of  claim 21  wherein said condition is a synucleinopathy.  
     
     
         24 . The siRNA of  claim 22  wherein said neurodegenerative disease is Parkinson's disease.  
     
     
         25 . The siRNA of  claim 1  wherein said nucleotide sequence is complementary to any one of the SEQ ID NOS: 8-15.  
     
     
         26 . The siRNA of  claim 1  comprising a nucleotide sequence substantially identical to SEQ ID NO: 19, SEQ ID NO: 23, SEQ ID NO: 25, SEQ ID NO: 27, SEQ ID NO: 29, SEQ ID NO: 31, or SEQ ID NO: 33.  
     
     
         27 . A composition comprising a siRNA in an amount sufficient to down regulate expression of a synuclein gene, wherein said siRNA comprises a nucleotide sequence substantially complementary to 15-30 consecutive nucleotides of SEQ. ID. NO.: 1.  
     
     
         28 . A vector comprising: 
 a promoter;    a nucleotide sequence operatively linked to said promoter, said nucleotide sequence comprising: 
 15-30 consecutive nucleotides of SEQ. ID. NO.: 1.  
   wherein said nucleotide sequence encodes an siRNA that down regulates a synuclein gene.    
     
     
         29 . A method of reducing expression of a synuclein gene in a cell, said method comprising introducing into a cell a siRNA in an amount effective to down regulate expression of said synuclein gene, 
 wherein said siRNA comprises a nucleotide sequence substantially complementary to 15-30 consecutive nucleotides of SEQ. ID. NO.: 1.    
     
     
         30 . The method of  claim 29  wherein said nucleotide sequence is substantially complementary to 15 to 30 consecutive nucleotides of SEQ. ID. NO.:6.  
     
     
         31 . The method of  claim 29  wherein said nucleotide sequence is complementary to 15 to 21 consecutive nucleotides of SEQ ID NO: 7.  
     
     
         32 . The method of  claim 29  wherein said siRNA is introduced into said cell in a vector encoding said siRNA.  
     
     
         33 . The method of  claim 29  wherein in said siRNA forms a hairpin structure and a loop.  
     
     
         34 . The method of  claim 32  wherein said vector comprises a nucleotide sequence substantially identical to SEQ ID NO: 21.  
     
     
         35 . The method of  claim 29  comprising introducing said siRNA into a cell of the central nervous system or the peripheral nervous system.  
     
     
         36 . A method of reducing cell death, said method comprising introducing into a cell a siRNA in an amount effective to down regulate expression of said synuclein gene, 
 wherein said siRNA comprises a nucleotide sequence substantially complementary to 15-30 consecutive nucleotides of SEQ. ID. NO.: 1.    
     
     
         37 . A method of treating a neurodegenerative disease or a synucleinopathy in a subject comprising administering to said subject a therapeutically effective amount of a siRNA comprising a nucleotide sequence substantially complementary to 15-30 consecutive nucleotides of SEQ. ID. NO.: 1, wherein the expression of said synuclein gene is down regulated.