SGK1 AND SGK3 REGULATE THE INNATE IMMUNE SIGNALING IN RESPONSE TO DSDNA VIRAL INFECTION IN MACROPHAGES

The STING signaling pathway is essential for the innate immune response to DNA viruses and bacteria and is important in tumor immunity. STING binding to cGAMP or to synthetic agonists leads to the activation of the kinase TBK1 which phosphorylates the transcription factor IRF3 which promotes the expression of type 1 interferons such as IFNb to block viral activity. Aberrant type 1 IFN expression is associated with human diseases including autoimmunity, HIV, and cancer. Here we identify N-[4-(1H- pyrazolo[3,4-b] pyrazin-6-yl)-phenyl]-sulfonamide (Sanofi-14h), a compound with a preference for inhibition of the AGC family kinase SGK3, as an inhibitor of IFNb gene expression in response to STING stimulation of macrophages.Sanofi-14h abrogated SGK activity and also impaired activation of the critical TBK1/IRF3 pathway downstream of STING activation, notably blocking the ligand-induced interaction of STING with TBK1. Deletion of SGK1 and SGK3 in macrophages suppressed activation of IFNb transcription but did not block TBK1/IRF3 activation downstream of STING. Gene and protein expression analysis revealed that deletion of SGK1/3 in a macrophage cell line decreases basal expression of critical transcription factors required for the innate immune response, such as IRF7 and STAT1. Whereas deletion of SGK3 in mice prevents migration of classical monocytes to the blood and the spleen in response to DMXAA stimulation. Mechanistically, SGK1 and SGK3 deletion caused an expression increase in the protein and mRNA levels of NDRG1 a known SGK1/3 substrate and a negative regulator of IRF7 in fish, indicating that SGK1/3 could be regulating the levels of IRF7 through modulation of NDRG1. Additional studies reveal that other AGC kinase inhibitors block TBK1 and IRF3 activation suggesting common action on a critical regulatory node in the STING pathway. Thus, studies with Sanofi-14h have revealed both SGK-dependent and SGK- independent effects in the STING pathway and suggest a mechanism to alter type 1 IFN transcription through small molecule therapy.