140 Inhibition of AKT Kinase Activity Decreases Replication of Human Respiratory Syncytial Virus

Abstract

BackgroundHuman Respiratory Syncytial Virus (RSV) is a leading cause of pediatric pulmonary disease and severe RSV infection predisposes to wheezing later in life. RSV infection has also been shown to be an environmental trigger for asthma. We are investigating whether targeting host factors important for RSV infection is a viable antiviral strategy. Lowering viral burden through these therapies will result in decreased severity of infection and may also prevent the occurrence of pathologic sequelae.MethodsInhibition of AKT by chemical inhibitors, siRNA, or dominant-negative mutants, was tested for activity against RSV replication in cultured cells. We examined the effect of viral protein expression on Akt activation and downstream signal transduction by Western blot and promoter assay. In addition, we examined the effect of Akt on specific viral processes (entry, macromolecular synthesis, and assembly) and proteins both in vitro and in RSV-infected cells, using kinase assays, Western blotting, and qRT-PCR.ResultsWe found that AKT inhibition decreases RSV protein expression and viral titers. Expression of RSV NS2 protein activates AKT, leading to NFkB-dependent transcription, and inhibition of AKT blocks this effect. Activated AKT also phosphorylates RSV P protein at a specific site. Interestingly, AKT inhibitors that target the pleckstrin homology (PH) domain of AKT showed decreased efficacy against RSV compared to those that target AKT kinase activity.ConclusionsInhibition of AKT can effectively decrease RSV replication in culture, likely by decreasing P phosphorylation and thus viral protein transcription and expression. Activation of AKT during RSV infection likely involves the NS2 protein and does not depend on the PH domain of AKT. AKT inhibitors have been found to be safe and efficacious in clinical trials for a number of different cancers; thus, AKT inhibition may be a potential therapeutic treatment for severe RSV infection.