Nucleic acids for targeting multiple regions of the HCV genome
Compositions and methods effective for modulating Hepatitis C viral infection are provided.
1. A composition comprising a miRNA expression cassette comprising a cluster sequence selected from the group of consisting of cluster 1, cluster 2 and cluster 5.
2. The composition of claim 1 , wherein said cluster sequence is cluster 5.
3. The composition of claim 1 or claim 2 , wherein said cluster sequence is contained in a vector.
4. The composition of claim 3 , wherein said vector is selected from the group consisting of AAV vectors, lentiviral vectors, retroviral vectors and AV vectors.
5. The composition of claim 4 , wherein said vector is an recombinant AAV vector having a serotype selected from the group consisting of AAV-2, AAV-6 and AAV 8.
6. The composition of claim 3 , wherein the expression of nucleotides is under the control of a single promoter.
7. The composition of claim 6 , wherein said promoter is a tissue-specific promoter.
8. The composition of claim 7 , wherein said promoter is a liver-specific alpha-one antitrypsin promoter, said composition additionally comprising a liver-specific enhancer and a polyadenylation signal.
9. The composition of claim 8 , wherein said enhancer is the apolipoprotein E hepatic control region.
10. The composition of claim 8 , wherein said polyadenylation signal is the bovine growth hormone polyadenylation signal.
11. The composition of claim 3 , wherein said composition comprises an intron.
12. The composition of claim 11 , wherein said intron is downstream of said promoter and upstream of said cluster.
13. The composition of claim 12 , wherein said intron is derived from human growth hormone.
14. A method of reducing HCV viral load in a patient comprising administering to the liver of said patient a therapeutically effective amount of the composition of claim 3 which directs cleavage of target HCV mRNA sequences present in said patient, thereby reducing HCV viral load.
15. The method of claim 14 , wherein said composition is introduced directly into said patient.
16. The method of claim 15 , wherein said introduction is via intravenous infusion.
17. The method of claim 14 , wherein said method results in inhibition of HCV replication in said patient.
18. The method of claim 12 , wherein introducing the vector to a cell, tissue, or organ of interest comprises: ligating the miRNA cassette into a viral delivery vector to form a viral miRNA expression construct; packaging the viral miRNA expression construct into viral particles; and delivering the viral particles to the cell, tissue, or organ of interest.
19. The method of claim 18 , wherein the miRNA cassette comprising nucleic acids which target multiple regions of a HCV genome.
20. The method of claim 19 , wherein said vector is an AAV vector and the miRNA cassette is cluster 5.