Method for replicating the hepatitis c virus

Abstract

A process for screening anti-HCV agents, including the steps of introducing a compound to be tested for its anti-HCV properties with transformed Vero/6418 cells, having a nucleic acid selected from the group consisting of a nucleic acid coding for the structural and non-structural proteins of the HCV, a nucleic acid coding for the non-structural proteins of the HCV, and a replicon containing a resistance gene to an antibiotic and a nucleotide sequence coding for the non-structural proteins of the HCV; extracting total RNA of the cells; and analyzing any reduction of the rate, of synthesis of the RNA of the HCV of the cells relative to a control value corresponding to the rates of synthesis of the RNAs of the HCV of the cells in the absence of the tested compound.

1 . A process for screening anti-HCV agents, comprising the steps of:
 introducing a compound to be tested for its anti-HCV properties with transformed Vero/6418 cells, comprising a nucleic acid selected from the group consisting of:   a nucleic acid coding for the structural and non-structural proteins of the HCV, a nucleic acid coding for the non-structural proteins of the HCV, and   a replicon containing a resistance gene to an antibiotic and a nucleotide sequence coding for the non-structural proteins of the HCV;   extracting total RNA of said cells; and   analyzing any reduction of the rate, of synthesis of the RNA of the HCV of said cells relative to a control value corresponding to the rates of synthesis of the RNAs of the HCV of the cells in the absence of said tested compound.   
     
     
         2 . The process according to  claim 1 , wherein said nucleic acid is one of SEQ ID NO: 1. and SEQ ID NO: 3. 
     
     
         3 . The process according to  claim 1 , wherein said nucleic acid is a replicon containing a resistance gene to hygromycin B. 
     
     
         4 . The process according to  claim 1 , wherein said cells are those deposited on the 13 Apr. 2001 at the CNCM under the number 1-2658. 
     
     
         5 . A process for preparing Vero/G418 cells comprising a nucleic acid selected from the group consisting of:
 a nucleic acid coding for structural and non-structural proteins of HCV,   a nucleic acid coding for non-structural proteins of HCV, and   a replicon containing a hygromycin B resistance gene and a nucleotide sequence coding for the non-structural proteins of the HCV, said process comprising the steps of:   inserting into said Vero/G418 cells a nucleotide sequence comprising one of SEQ ID NO: 1. and SEQ ID NO: 3 or a recombinant vector comprising one of SEQ ID NO: 1 and SEQ ID NO: 3; and   subjecting the cells thus obtained to increasing concentrations of hygromycin B.   
     
     
         6 . The process according to  claim 5 , wherein said nucleic acid is one of SEQ ID NO: 1 and SEQ ID NO: 3. 
     
     
         7 . The process according to  claim 5 , wherein said recombinant vector is inserted into said Vero/G418 cells, and said recombinant vector is selected from the group consisting of a plasmid, a cosmid, a phage and virus DNA. 
     
     
         8 . The process according to  claim 5 , wherein the concentration of hygromycin B increases from 800 to 1000 μg/ml.