Replication of hepatitis C virus in non-hepatic epithelial and mouse hepatic cells
View Patent ↗Cells and cell lines which replicate HCV of non-hepatic human and non human origin are disclosed. Also provided are methods of using such cells and cell lines to identify anti-HCV agents for the treatment of HCV infection.
1. A hepatitis C virus (HCV) replicating cell line, wherein said cell line is a mouse cell line comprising an HCV genome.
2. The cell line of claim 1 , wherein the mouse cell line comprises mouse cells of hepatic origin.
3. The cell line of claim 2 , wherein the mouse cells are Hepa1-6 cells.
4. The cell line of claim 2 , wherein the mouse cells are AML12 cells.
5. The cell line of claim 1 , wherein said HCV genome is a HCV subgenome.
6. The cell line of claim 1 , wherein said HCV genome is obtained from a second cell line which replicates HCV.
7. A method of producing the cell line of claim 1 , comprising:
a) obtaining total RNA from a human non-hepatic cell culture that replicates HCV, said total RNA comprising a selection marker which renders cells expressing said RNA resistant to a selection agent;
b) introducing the total RNA into mouse cells;
c) selecting those cells which grow in the presence of said selection agent and replicate HCV; and
d) generating a cell line from the cells of step c).
8. A method for screening test compounds which inhibit HCV replication, comprising:
a) culturing the cell line of claim 1 in the presence and absence of a test compound; and
b) assaying HCV replication levels in the presence and absence of said test compound, wherein a reduced HCV replication level in the presence of said test compound is indicative that said test compound inhibits HCV replication.
9. A method for screening test compounds which modulate the antiviral response induced by interferon alpha (IFN-α) comprising
a) culturing the cell line of claim 1 in the presence and absence of a test compound;
b) contacting the cells of step a) with IFN-α; and
c) measuring the HCV replication level in the presence and absence of said compound thereby identifying agents which modulate the antiviral response mediated by IFN-α as a function of altered HCV levels.
10. The method of claim 9 , wherein the antiviral response is enhanced.
11. The method of claim 9 , wherein the antiviral response is inhibited.
12. A hepatitis C virus (HCV) replicating cell line, wherein said cell line is a
human non-hepatic cell line,
wherein said cell line comprises genomic HCV RNA, and
wherein said genomic HCV RNA consists of HCV RNA obtainable from a second HCV replicating human cell line.
13. The cell line of claim 12 , wherein the human non-hepatic cell line comprises epithelial cells.
14. The cell line of claim 13 , wherein the human epithelial cells are HeLa cells.
15. The cell line of claim 12 , wherein said HCV genome is a HCV subgenome.
16. The cell line of claim 15 , wherein said second cell line is a Huh7 derived cell line.
17. The cell line of claim 12 , wherein said second cell line is a Huh7 derived cell line.
18. The cell line of claim 12 , wherein said RNA from the second cell line is the total RNA.
19. A method for producing the cell line of claim 12 , comprising:
a) obtaining total RNA from a human hepatic cell culture that replicates HCV, said total RNA comprising a selection marker which renders cells expressing said RNA resistant to a selection agent;
b) introducing the total RNA into human non-hepatic cells;
c) selecting those cells which grow in the presence of said selection agent and replicate HCV; and
d) generating a cell line from the cells of step c).
20. A method for screening test compounds which inhibit HCV replication, comprising:
a) culturing the cell line of claim 12 in the presence and absence of a test compound; and
b) assaying HCV replication levels in the presence and absence of said test compound, wherein a reduced HCV replication level in the presence of said test compound is indicative that said test compound inhibits HCV replication.
21. A method for screening test compounds which modulate the antiviral response induced by interferon alpha (TEN-α) comprising
a) culturing the cell line of claim 12 in the presence and absence of a test compound;
b) contacting the cells of step a) with TEN-α; and
c) measuring the HCV replication level in the presence and absence of said compound thereby identifying agents which modulate the antiviral response mediated by IFN-α as a function of altered HCV levels.
22. The method of claim 21 , wherein the antiviral response is enhanced.
23. The method of claim 21 , wherein the antiviral response is inhibited.