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Journal of Virology, October 2003, p. 10689-10694, Vol. 77, No. 19
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.19.10689-10694.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Dynamics of Subgenomic Hepatitis C Virus Replicon RNA Levels in Huh-7 Cells after Exposure to Nucleoside Antimetabolites

Lieven J. Stuyver,1* Tamara R. McBrayer,1 Phillip M. Tharnish,1 Abdalla E. A. Hassan,1 Chung K. Chu,2 Krzysztof W. Pankiewicz,1 Kyochi A. Watanabe,1 Raymond F. Schinazi,3 and Michael J. Otto1

Pharmasset, Inc., Tucker, Georgia 30084,1 College of Pharmacy, University of Georgia, Athens, Georgia 30602,2 Veterans Affairs Medical Center and Department of Pediatrics, Emory University School of Medicine, Decatur, Georgia 300333

Received 15 May 2003/ Accepted 10 July 2003

Treatment with antimetabolites results in chemically induced low nucleoside triphosphate pools and cell cycle arrest in exponentially growing cells. Since steady-state levels of hepatitis C virus (HCV) replicon RNA were shown to be dependent on exponential growth of Huh-7 cells, the effects of antimetabolites for several nucleoside biosynthesis pathways on cell growth and HCV RNA levels were investigated. A specific anti-HCV replicon effect was defined as (i) minimal interference with the exponential cell growth, (ii) minimal reduction in cellular host RNA levels, and (iii) reduction of the HCV RNA copy number per cell compared to that of the untreated control. While most antimetabolites caused a cytostatic effect on cell growth, only inhibitors of the de novo pyrimidine ribonucleoside biosynthesis mimicked observations seen in confluent replicon cells, i.e., cytostasis combined with a sharp decrease in replicon copy number per cell. These results suggest that high levels of CTP and UTP are critical parameters for maintaining the steady-state level replication of HCV replicon in Huh-7 cells.

* Corresponding author. Mailing address: Pharmasset, Inc., 1860 Montreal Rd., Tucker, GA 30084. Phone: (678) 395-0050. Fax: (678) 395-0030. E-mail: lstuyver{at}pharmasset.com.

Journal of Virology, October 2003, p. 10689-10694, Vol. 77, No. 19
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.19.10689-10694.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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