Induction of viral mutation by incorporation of miscoding ribonucleoside analogs into viral RNA

Abstract

The present invention is directed to the identification and use of ribonucleoside analogs to induce the mutation of an RNA virus, including BVDV, HIV and HCV, or a virus which otherwise replicates through an RNA intermediate. The increase in the mutation rate of the virus results in reduced viability of progeny generations of the virus, thereby inhibiting viral replication. In addition to these methods and related compositions, the invention provides methods and combinatorial chemistry libraries for screening ribonucleoside analogs for mutagenic potential.

1 .- 18 . (canceled)  
     
     
         19 . A viral particle comprising viral genomic RNA, wherein the viral genomic RNA comprises an RNA nucleoside analog.  
     
     
         20 . The viral particle of  claim 19 , wherein the particle is an HIV particle, and an HCV particle, or a BVDV particle.  
     
     
         21 . The viral particle of  claim 19 , wherein the nucleoside analog is selected from the group consisting of N4-aminocytidine, N1-methyl-N-4-aminocytidine, 3,N4-ethenocytidine, 3-methylcytidine, 5-hydroxycytidine, N4-dimethylcytidine, 5-(2-hydroxyethyl)cytidine, 5-chlorocytidine, 5-bromocytidine, N4-methyl-N-4-aminocytidine, 5-aminocytidine, 5-nitrosocytidine, 5-(hydroxyalkyl)-cytidine, 5-(thioalkyl)-cytidine and cytidine glycol, 5-hydroxyuridine, 3-hydroxyethyluridine, 3-methyluridine, O2-methyluridine, O2-ethyluridine, 5-aminouridine, O4-methyluridine, O4-ethyluridine, O4-isobutyluridine, O4-alkyluridine, 5-nitrosouridine, 5-(hydroxyalkyl)-uridine, and 5-(thioalkyl)-uridine, 1,N6-ethenoadenosine, 3-methyladenosine, and N6-methyladenosine, 8-hydroxyguanosine, O6-methylguanosine, O6-ethylguanosine, O6-isopropylguanosine, 3,N2-ethenoguanosine, 06-alkylguanosine, 8-oxo-guanosine, 2,N3-ethenoguanosine, and 8-aminoguanosine.  
     
     
         22 . A population of cells comprising a highly variable population of replicated homologous viral nucleic acids.  
     
     
         23 . The population of cells of  claim 22 , wherein the cells are in cell culture.  
     
     
         24 . The population of cells of  claim 22 , wherein the viral nucleic acids are derived from a retrovirus or a flavivirus.  
     
     
         25 . A cell comprising a viral genomic nucleic acid, an RNA analog, a cellular mRNA analog and a viral genomic RNA analog.  
     
     
         26 . The cell of  claim 25 , wherein the viral genomic nucleic acid is integrated into the cellular genome.  
     
     
         27 . The cell of  claim 25 , wherein the viral genomic nucleic acid is a retroviral or a flaviviral nucleic acid.  
     
     
         28 . The cell of  claim 25 , wherein the viral genomic nucleic acid is an HIV nucleic acid, a pestivirus nucleic acid, or an HCV virus nucleic acid.  
     
     
         29 . The cell of  claim 25 , wherein the viral genomic nucleic acid is selected from the group consisting of a HIV-1, HIV-2, HTLV-1, HTLV-2, SIV, hepatitis A, hepatitis B, hepatitis C, BVDV, and dengue fever virus.  
     
     
         30 .- 40 . (canceled)  
     
     
         41 . A pharmaceutical composition comprising a therapeutically effective dose of an RNA nucleoside analog, wherein the analog is one that in a infected cell with a virus of interest is incorporated by a polymerase into an RNA copy of a genomic nucleic acid encoding the virus, said analog replacing a first natural occurring nucleotide having a first complementary nucleotide wherein said analog complements a second nucleotide which is other than the first nucleotide together with a pharmaceutically acceptable carrier.  
     
     
         42 . The pharmaceutical composition of  claim 41 , wherein the nucleoside analog is selected from the group consisting of N4-aminocytidine, N1-methyl-N-4-aminocytidine, 3,N4-ethenocytidine, 3-methylcytidine, 5-hydroxycytidine, N4-dimethylcytidine, 5-(2-hydroxyethyl)-cytidine, 5-chlorocytidine, 5-bromocytidine, N4-methyl-N-4-aminocytidine, 5-aminocytidine, 5-nitrosocytidine, 5-(hydroxyalkyl)-cytidine, 5-(thioalkyl)-cytidine and cytidine glycol, 5-hydroxyuridine, 3-hydroxyethyluridine, 3-methyluridine, O2-methyluridine, O2-ethyluridine, 5-aminouridine, O4-methyluridine, O4-ethyluridine, O4-isobutyluridine, O4-alkyluridine, 5-nitrosouridine, 5-(hydroxyalkyl)-uridine, and 5-(thioalkyl)-uridine, 1,N6-ethenoadenosine, 3-methyladenosine, and N6-methyladenosine, 8-hydroxyguanosine, O6-methylguanosine, O6-ethylguanosine, O6-isopropylguanosine, 3,N2-ethenoguanosine, 06-alkylguanosine, 8-oxo-guanosine, 2,N3-ethenoguanosine, and 8-aminoguanosine.  
     
     
         43 . The pharmaceutical composition of  claim 41 , wherein the pharmaceutical composition is suitable for oral administration.  
     
     
         44 . The pharmaceutical composition of  claim 41 , wherein the pharmaceutical composition is suitable for parenteral administration.  
     
     
         45 .- 66 . (canceled)  
     
     
         67 . The pharmaceutical composition of  claim 41 , wherein the RNA nucleoside analog is an enantio-specific nucleoside analog.