gw-5074 and viroxime

RNA viruses can rapidly mutate and acquire resistance to drugs that directly target viral enzymes, which poses serious problems in a clinical context. Therefore, there is a growing interest in the development of antiviral drugs that target host factors critical for viral replication, since they are unlikely to mutate in response to therapy. We recently demonstrated that phosphatidylinositol-4-kinase IIIβ (PI4KIIIβ) and its product phosphatidylinositol-4-phosphate (PI4P) are essential for replication of enteroviruses, a group of medically important RNA viruses including poliovirus (PV), coxsackievirus, rhinovirus, and enterovirus 71. Here, we show that enviroxime and GW5074 decreased PI4P levels at the Golgi complex by directly inhibiting PI4KIIIβ. Coxsackievirus mutants resistant to these inhibitors harbor single point mutations in the non-structural protein 3A. These 3A mutations did not confer compound-resistance by restoring the activity of PI4KIIIβ in the presence of the compounds. Instead, replication of the mutant viruses no longer depended on PI4KIIIβ, since their replication was insensitive to siRNA-mediated depletion of PI4KIIIβ. The mutant viruses also did not rely on other isoforms of PI4K. Consistently, no high level of PI4P could be detected at the replication sites induced by the mutant viruses in the presence of the compounds. Collectively, these findings indicate that through specific single point mutations in 3A, CVB3 can bypass an essential host factor and lipid for its propagation, which is a new example of RNA viruses acquiring resistance against antiviral compounds, even when they directly target host factors.

Topics: 1-Phosphatidylinositol 4-Kinase; Animals; Antiviral Agents; Benzimidazoles; Cell Line, Tumor; Chlorocebus aethiops; Drug Resistance, Viral; Enterovirus B, Human; HeLa Cells; Humans; Indoles; Oximes; Phenols; Phosphatidylinositol Phosphates; Phosphotransferases (Alcohol Group Acceptor); Point Mutation; RNA Interference; RNA, Small Interfering; Sulfonamides; Viral Nonstructural Proteins; Virus Replication

Enviroxime is an antienterovirus compound that targets viral protein 3A and/or 3AB and suppresses a step in enterovirus replication by unknown mechanism. To date, four antienterovirus compounds, i.e., GW5074, Flt3 inhibitor II, TTP-8307, and AN-12-H5, are known to have similar mutations in the 3A protein-encoding region causing resistance to enviroxime (a G5318A [3A-Ala70Thr] mutation in poliovirus [PV]) and are considered enviroxime-like compounds. Recently, antienterovirus activity of a phosphatidylinositol 4-kinase III beta (PI4KB) inhibitor, PIK93, was reported, suggesting that PI4KB is an important host factor targetable by antienterovirus compounds (N. Y. Hsu et al., Cell 141:799-811, 2010). In this study, we analyzed the inhibitory effects of previously identified enviroxime-like compounds (GW5074 and AN-12-H5) and a newly identified antienterovirus compound, T-00127-HEV1, on phosphoinositide (PI) kinases. We found that T-00127-HEV1 inhibited PI4KB activity with a higher specificity for than other PI kinases, in contrast to GW5074, which had a broad specificity for PI kinases. In contrast, AN-12-H5 showed no inhibitory effect on PI4KB activity and only moderate inhibitory effects on PI 3-kinase activity. Small interfering RNA (siRNA) screening targeting PI kinases identified PI4KB is a target of GW5074 and T-00127-HEV1, but not of AN-12-H5, for anti-PV activity. Interestingly, T-00127-HEV1 and GW5074 did not inhibit hepatitis C virus (HCV) replication, in contrast to a strong inhibitory effect of AN-12-H5. These results suggested that PI4KB is an enterovirus-specific host factor required for the replication process and targeted by some enviroxime-like compounds (T-00127-HEV1 and GW5074) and that enviroxime-like compounds may have targets other than PI kinases for their antiviral effect.

Topics: 1-Phosphatidylinositol 4-Kinase; Antiviral Agents; Benzimidazoles; Enterovirus; HEK293 Cells; Humans; Indoles; Oximes; Phenols; Poliomyelitis; Poliovirus; Species Specificity; Sulfonamides; Viral Proteins; Virus Replication