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Influenza viruses are the cause of yearly epidemics and occasional pandemics that represent a significant challenge to public health. Current control strategies are imperfect and there is an unmet need for new antiviral therapies. Here, we report the identification of small molecule compounds able to effectively and specifically inhibit growth of influenza A and B viruses in cultured cells through targeting an assembly interface of the viral RNA-dependent RNA polymerase. Using an existing crystal structure of the primary protein–protein interface between the PB1 and PA subunits of the influenza A virus polymerase, we conducted an in silico screen to identify potential small molecule inhibitors. Selected compounds were then screened for their ability to inhibit the interaction between PB1 and PA in vitro using an ELISA-based assay and in cells, to inhibit nuclear import of a binary PB1–PA complex as well as transcription by the full viral ribonucleoprotein complex. Two compounds emerged as effective inhibitors with IC 50 values in the low micromolar range and negligible cytotoxicity. Of these, one compound also acted as a potent replication inhibitor of a variety of influenza A virus strains in Madin-Darby canine kidney (MDCK) cells, including H3N2 and H1N1 seasonal and 2009 pandemic strains. Importantly, this included an oseltamivir-resistant isolate. Furthermore, potent inhibition of influenza B viruses but not other RNA or DNA viruses was seen. Overall, these compounds provide a foundation for the development of a new generation of therapeutic agents exhibiting high specificity to influenza A and B viruses.

WC5 is a 6-aminoquinolone that potently inhibits the replication of human cytomegalovirus (HCMV) but has no activity, or significantly less activity, against other herpesviruses. Here we investigated the nature of its specific anti-HCMV activity. Structure-activity relationship studies on a small series of analogues showed that WC5 possesses the most suitable pattern of substitutions around the quinolone scaffold to give potent and selective anti-HCMV activity. Studies performed to identify the possible target of WC5 indicated that it prevents viral DNA synthesis but does not significantly affect DNA polymerase activity. In yield reduction experiments with different multiplicities of infection, the anti-HCMV activity of WC5 appeared to be highly dependent on the viral inoculum, suggesting that WC5 may act at an initial stage of virus replication. Consistently, time-of-addition and time-of-removal studies demonstrated that WC5 affects a phase of the HCMV replicative cycle that precedes viral DNA synthesis. Experiments to monitor the effects of the compound on virus attachment and entry showed that it does not inhibit either process. Evaluation of viral mRNA and protein expression revealed that WC5 targets an event of the HCMV replicative cycle that follows the transcription and translation of immediate-early genes and precedes those of early and late genes. In cell-based assays to test the effects of WC5 on the transactivating activity of the HCMV immediate-early 2 (IE2) protein, WC5 markedly interfered with IE2-mediated transactivation of viral early promoters. Finally, WC5 combined with ganciclovir in checkerboard experiments exhibited highly synergistic activity. These findings suggest that WC5 deserves further investigation as a candidate anti-HCMV drug with a novel mechanism of action.