Establishment and use of a cell line expressing HSV-1 thymidine kinase to characterize viral thymidine kinase-dependent drug-resistance.
To understand the mechanisms of antiviral drug resistance and to have a system to examine the cytotoxicity of herpes simplex virus type 1 (HSV-1) inhibitors that are thymidine kinase (TK)-dependent, we have constructed a plasmid pFTK1 by inserting a DNA fragment containing the TK gene of HSV-1 strain F into the eukaryotic expression vector pcDNA3.1/His A. TK-deficient 143B cells were transfected with this vector and neomycin-resistant cells were selected. Cell survival in HAT medium and TK activity of the cell lysates were examined to ascertain HSV-1 TK expression. A cell line expressing the viral TK gene, FTK143B (FTK), was established and used for characterization of two laboratory-derived TK-deficient drug-resistant HSV-1 mutants of strain F. The antiviral activities of several drugs, mostly nucleoside analogues, were compared in the Vero, 143B and FTK cell culture systems. We showed that both mutant viruses lost their resistance to acyclovir and to other HSV-1 TK-dependent compounds in FTK cells but not in Vero and 143B cells. Significantly increased cytotoxicity of ganciclovir and (E)-5-(2-bromovinyl)-2'-deoxyuridine was also observed in the FTK cells. This HSV-1 TK gene-transfected cell model is a useful tool to rapidly determine HSV-1 drug resistance at the viral TK level.
Kim
Jee Hyun
JH
Pharmaceutical Screening Center, Korea Research Institute of Chemical Technology, Taejon 305-600, South Korea.
Park
Jong Beak
JB
Bae
Pan Kee
PK
Kim
Hae Soo
HS
Kim
Do Wan
DW
Ahn
Jeong Keun
JK
Lee
Chong-Kyo
CK
D000818:Animals; D000998:Antiviral Agents / Q000493:pharmacokinetics / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line; D002522:Chlorocebus aethiops; D003588:Cytopathogenic Effect, Viral; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism; D024882:Drug Resistance, Viral; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006801:Humans; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D018139:Simplexvirus / Q000201:enzymology / Q000235:genetics*; D013937:Thymidine Kinase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D014709:Vero Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
D000998:Antiviral Agents; D004279:DNA, Viral; D014764:Viral Proteins; D013937:Thymidine Kinase