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Antimicrobial Agents and Chemotherapy, February 2008, p. 586-597, Vol. 52, No. 2
0066-4804/08/$08.00+0     doi:10.1128/AAC.01172-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Cidofovir and (S)-9-[3-Hydroxy-(2-Phosphonomethoxy)Propyl]Adenine Are Highly Effective Inhibitors of Vaccinia Virus DNA Polymerase When Incorporated into the Template Strand

Department of Medical Microbiology and Immunology, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada,¹ San Diego Veterans Medical Research Foundation and the Division of Infectious Disease, University of California, San Diego, La Jolla, California 92093²

Received 5 September 2007/ Returned for modification 12 October 2007/ Accepted 28 November 2007

The acyclic nucleoside phosphonate drug (S)-9-[3-hydroxy-(2-phosphonomethoxy)propyl]adenine [(S)-HPMPA], is a broad-spectrum antiviral and antiparasitic agent. Previous work has shown that the active intracellular metabolite of this compound, (S)-HPMPA diphosphate [(S)-HPMPApp], is an analog of dATP and targets DNA polymerases. However, the mechanism by which (S)-HPMPA inhibits DNA polymerases remains elusive. Using vaccinia virus as a model system, we have previously shown that cidofovir diphosphate (CDVpp), an analog of dCTP and a related antiviral agent, is a poor substrate for the vaccinia virus DNA polymerase and acts to inhibit primer extension and block 3'-to-5' proofreading exonuclease activity. Based on structural similarities and the greater antiviral efficacy of (S)-HPMPA, we predicted that (S)-HPMPApp would have a similar, but more pronounced effect on vaccinia polymerase than CDVpp. Interestingly, we found that (S)-HPMPApp is a good substrate for the viral enzyme, exhibiting K_m and V_max parameters comparable to those of dATP, and certainly not behaving like CDVpp as a functional chain terminator. Metabolic experiments indicated that (S)-HPMPA is converted to (S)-HPMPApp to a much greater extent than CDV is converted to CDVpp, although both drugs cause identical effects on virus DNA replication at their 50% effective concentration. Subsequent studies showed that both compounds can be faithfully incorporated into DNA, but when CDV and (S)-HPMPA are incorporated into the template strand, both strongly inhibit trans-lesion DNA synthesis. It thus appears that nucleoside phosphonate drugs exhibit at least two different effects on DNA polymerases depending upon in what form the enzyme encounters the drug.

Published ahead of print on 3 December 2007.