Amino acid changes within conserved region III of the herpes simplex virus and human cytomegalovirus DNA polymerases confer resistance to 4-oxo-dihydroquinolines, a novel class of herpesvirus antiviral agents.
The 4-oxo-dihydroquinolines (PNU-182171 and PNU-183792) are nonnucleoside inhibitors of herpesvirus polymerases (R. J. Brideau et al., Antiviral Res. 54:19-28, 2002; N. L. Oien et al., Antimicrob. Agents Chemother. 46:724-730, 2002). In cell culture these compounds inhibit herpes simplex virus type 1 (HSV-1), HSV-2, human cytomegalovirus (HCMV), varicella-zoster virus (VZV), and human herpesvirus 8 (HHV-8) replication. HSV-1 and HSV-2 mutants resistant to these drugs were isolated and the resistance mutation was mapped to the DNA polymerase gene. Drug resistance correlated with a point mutation in conserved domain III that resulted in a V823A change in the HSV-1 or the equivalent amino acid in the HSV-2 DNA polymerase. Resistance of HCMV was also found to correlate with amino acid changes in conserved domain III (V823A+V824L). V823 is conserved in the DNA polymerases of six (HSV-1, HSV-2, HCMV, VZV, Epstein-Barr virus, and HHV-8) of the eight human herpesviruses; the HHV-6 and HHV-7 polymerases contain an alanine at this amino acid. In vitro polymerase assays demonstrated that HSV-1, HSV-2, HCMV, VZV, and HHV-8 polymerases were inhibited by PNU-183792, whereas the HHV-6 polymerase was not. Changing this amino acid from valine to alanine in the HSV-1, HCMV, and HHV-8 polymerases alters the polymerase activity so that it is less sensitive to drug inhibition. In contrast, changing the equivalent amino acid in the HHV-6 polymerase from alanine to valine alters polymerase activity so that PNU-183792 inhibits this enzyme. The HSV-1, HSV-2, and HCMV drug-resistant mutants were not altered in their susceptibilities to nucleoside analogs; in fact, some of the mutants were hypersensitive to several of the drugs. These results support a mechanism where PNU-183792 inhibits herpesviruses by interacting with a binding determinant on the viral DNA polymerase that is less important for the binding of nucleoside analogs and deoxynucleoside triphosphates.
Thomsen
Darrell R
DR
Infectious Disease Biology, Pharmacia Corporation, Kalamazoo, Michigan 49001, USA.
Oien
Nancee L
NL
Hopkins
Todd A
TA
Knechtel
Mary L
ML
Brideau
Roger J
RJ
Wathen
Michael W
MW
Homa
Fred L
FL
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002522:Chlorocebus aethiops; D003587:Cytomegalovirus / Q000187:drug effects*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics*; D024882:Drug Resistance, Viral; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D018259:Herpesvirus 1, Human / Q000187:drug effects*; D018258:Herpesvirus 2, Human / Q000187:drug effects*; D006801:Humans; D009154:Mutation; D019384:Nucleic Acid Synthesis Inhibitors; D011804:Quinolines / Q000494:pharmacology*; D014709:Vero Cells
D000998:Antiviral Agents; D004791:Enzyme Inhibitors; D019384:Nucleic Acid Synthesis Inhibitors; C470975:PNU 182171; C455641:PNU183792; D011804:Quinolines; D004259:DNA-Directed DNA Polymerase