Cidofovir Diphosphate Inhibits Adenovirus 5 DNA Polymerase via Both Non-Obligate Chain Termination and Direct Inhibition, and Polymerase Mutations Confer Cidofovir Resistance on Intact Virus [Antiviral Agents]

Human adenovirus (AdV) can cause fatal disease in immune suppressed individuals, but treatment options are limited - in part because the antiviral cytidine analog, cidofovir (CDV), is nephrotoxic. The investigational agent brincidofovir (BCV) is orally bioavailable, non-nephrotoxic, and generates the same active metabolite, cidofovir diphosphate (CDVpp). However, its mechanism of action against AdV is poorly understood. We have therefore examined the effect of CDVpp on DNA synthesis by a purified AdV5 DNA polymerase (pol). CDVpp was incorporated into nascent DNA strands, and promoted a non-obligate form of chain termination (i.e., AdV5 pol can extend, albeit inefficiently, a DNA chain even after the incorporation of a first CDVpp molecule). Moreover, unlike a conventional mismatched base pair, misincorporated CDVpp was not readily excised by the AdV5 pol. At elevated concentrations, CDVpp inhibited AdV5 pol in a manner consistent with both chain termination and direct inhibition of pol activity. Finally, a recombinant AdV5 virus was constructed, containing pol mutations (V303I, T87I) that were selected following extended passage of wild-type AdV5 in the presence of BCV. This virus had a 2.1-fold elevated EC50 for BCV, and 1.9 fold increased EC50 for CDV - thus confirming that viral resistance to BCV and CDV can be attributed to mutations in the viral pol. These findings show that the anti-AdV5 activity of CDV and BCV is mediated through the viral DNA pol, and that their antiviral activity may occur via both (non-obligate) chain termination and (at high concentration) direct inhibition of AdV5 pol activity.

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