Lipid Formulated Compositions and Methods for Inhibiting Expression of a Gene from the Ebola Virus

Abstract

The invention relates to lipid formulated double-stranded ribonucleic acid (dsRNA) for inhibiting the expression of a gene from the Ebola virus.

1 . A pharmaceutical composition for inhibiting the expression of a gene from an Ebola virus in an organism, comprising a double-stranded ribonucleic acid (dsRNA) and a lipid formulation comprising the cationic lipid of formula A: 
       
         
           
           
               
               
           
         
         where R 1  and R 2  are independently alkyl, alkenyl or alkynyl, and wherein each of R 1  and R 2  can be optionally substituted, and wherein R 3  and R 4  are independently lower alkyl or together form an optionally substituted heterocyclic ring; and 
         wherein said dsRNA comprises at least two sequences that are complementary to each other and wherein a sense strand comprises a first sequence and an antisense strand comprises a second sequence comprising a region of complementarity which is substantially complementary to at least a part of a mRNA encoding Ebola, and wherein said region of complementarity is less than 30 nucleotides in length. 
       
     
     
         2 . The pharmaceutical composition of  claim 1 , wherein said first sequence of said dsRNA consists of the sequence of SEQ ID NO:1027, and said second sequence consists of the sequence of SEQ ID NO:1028. 
     
     
         3 . The pharmaceutical composition of  claim 1 , wherein said first sequence is selected from the group consisting of the sense sequences of Table 2 and said second sequence is selected from the group consisting of the antisense sequences of Table 2. 
     
     
         4 . The pharmaceutical composition of  claim 1 , wherein said dsRNA comprises at least one modified nucleotide. 
     
     
         5 . The pharmaceutical composition of  claim 3 , wherein said dsRNA comprises at least one modified nucleotide. 
     
     
         6 . The pharmaceutical composition of  claim 4 , wherein said modified nucleotide is chosen from the group of: a 2′-O-methyl modified nucleotide, a nucleotide comprising a 5′-phosphorothioate group, and a terminal nucleotide linked to a cholesteryl derivative or dodecanoic acid bisdecylamide group. 
     
     
         7 . The pharmaceutical composition of  claim 4 , wherein said modified nucleotide is chosen from the group of: a 2′-deoxy-2′-fluoro modified nucleotide, a 2′-deoxy-modified nucleotide, a locked nucleotide, an abasic nucleotide, 2′-amino-modified nucleotide, 2′-alkyl-modified nucleotide, morpholino nucleotide, a phosphoramidate, and a non-natural base comprising nucleotide. 
     
     
         8 .- 9 . (canceled) 
     
     
         10 . A method for inhibiting the expression of a gene from an Ebola virus in a cell, the method comprising:
 (a) introducing into the cell the pharmaceutical composition of  claim 1 ; and   (b) maintaining the cell produced in step (a) for a time sufficient to obtain degradation of the mRNA transcript of a gene from the Ebola virus, thereby inhibiting expression of a gene from the Ebola virus in the cell.   
     
     
         11 . A method of treating, preventing or managing pathological processes mediated by Ebola expression comprising administering to a subject in need of such treatment, prevention or management a therapeutically or prophylactically effective amount of the pharmaceutical composition of  claim 1 . 
     
     
         12 . The pharmaceutical composition of  claim 1 , wherein the dsRNA targets the VP35 of Ebola. 
     
     
         13 . The pharmaceutical composition of  claim 1 , wherein said dsRNA, upon contact with a cell infected with Ebola virus, inhibits expression of a gene from the virus by at least 40% compared to a control. 
     
     
         14 . The pharmaceutical composition of  claim 1 , wherein said region of complementarity is 15-30 nucleotides in length. 
     
     
         15 . The pharmaceutical composition of  claim 1 , wherein said region of complementarity is 19-24 nucleotides in length. 
     
     
         16 . The pharmaceutical composition of  claim 1 , wherein R 1  and R 2  of formula A are independently selected from oleoyl, pamitoyl, steroyl, or linoleyl. 
     
     
         17 . The pharmaceutical composition of  claim 1 , wherein R 1  and R 2  of formula A are both linoleyl. 
     
     
         18 . The pharmaceutical composition of  claim 1 , wherein R 3  and R 4  of formula A are methyl. 
     
     
         19 . The pharmaceutical composition of  claim 1 , wherein the cationic lipid of formula A is 2,2-Dilinoleyl-4-dimethylaminoethyl-[1,3]-dioxolane. 
     
     
         20 . The pharmaceutical composition of  claim 1 , further comprising a neutral lipid, a sterol, a PEG, or a PEG-modified lipid. 
     
     
         21 . The pharmaceutical composition of  claim 20 , wherein said pharmaceutical composition comprises LNP09. 
     
     
         22 . The pharmaceutical composition of  claim 20 , wherein said pharmaceutical composition comprises Formulation M. 
     
     
         23 . A method of increasing life-span of, decreasing viral titre in, or sustaining platelet count in a subject infected with an Ebola virus, comprising administering to the subject the pharmaceutical composition of  claim 1  in an amount sufficient to increase the life-span of, decreasing viral titre in, or sustaining platelet count in the subject. 
     
     
         24 .- 42 . (canceled) 
     
     
         43 . A pharmaceutical composition for inhibiting the expression of a gene from an Ebola virus in an organism, comprising a double-stranded ribonucleic acid (dsRNA) and a lipid formulation comprising a compound of formula X, selected from the group consisting of: 
       
         
           
           
               
               
           
         
       
       ; and wherein said dsRNA comprises at least two sequences that are complementary to each other and wherein a sense strand comprises a first sequence and an antisense strand comprises a second sequence comprising a region of complementarity which is substantially complementary to at least a part of a mRNA encoding Ebola virus, and wherein said region of complementarity is less than 30 nucleotides in length. 
     
     
         44 . The pharmaceutical composition of  claim 43 , wherein said first sequence of said dsRNA consists of the sequence of SEQ ID NO:1027, and said second sequence consists of the sequence of SEQ ID NO:1028. 
     
     
         45 .- 72 . (canceled)