Abstract 4394: Molecular interplay between Tank-binding kinase (TBK1) and Yes-associated protein (YAP1) inKRASmutant NSCLC

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality in the United States. Lung adenocarcinomas are highly correlated with smoking and are characterized by mutations in KRAS , EGFR , BRAF and other oncogenes. KRAS mutations are widespread in adenocarcinomas among smokers and known to be a key player in various downstream signaling pathways contributing to the tumorigenesis. Recently, a non-canonical IκB kinase, Tank Binding Kinase 1 (TBK1), has been found to contribute in KRAS mutant cancers. TBK1 has well documented functions in immune response, cell survival and in mitosis. While it has been suggested that TBK1-mediated regulation of Akt signaling might facilitate oncogenesis, the molecular mechanisms underlying TBK1 function downstream of KRAS is not fully elucidated. Yes associated protein 1 (YAP1) is an oncogenic component of the Hippo signaling cascade which could promote KRAS mediated oncogenesis and could substitute for the loss of Kras in mouse models of pancreatic cancer. In our present study, we demonstrate a unique and novel interplay between TBK1 and the oncogenic Hippo effector molecule, YAP1. YAP1 and its paralog, TAZ are known transcriptional co-activators that function to maintain organ size during development and is often activated in cancers. We find that TBK1 could physically interact with YAP1 and phosphorylate it at T110, T114, S128 and S131 residues in vitro . Knocking down (KD) or knock out (KO) of TBK1 resulted in a significant elevation of YAP1 expression at the protein level; surprisingly, without any effect at the mRNA level. Interestingly, the upregulation of YAP1 upon depletion of TBK1 was restricted to KRAS mutant NSCLC cell-lines and not in EGFR mutant cell lines. This elevation of YAP1 upon TBK1 KD was mainly observed in the nucleus; notably, there were only minimal changes in the levels of MST and LATS, raising the possibility that these changes occur independent of the classic hippo signaling pathway. Treatment with cycloheximide, an inhibitor of protein-translation, could not diminish the elevated level of YAP1 protein in the TBK1 depleted cells, indicating that the increased YAP1 level is due to enhanced protein stability, probably as a result of post-translational modification(s). Depletion of TBK1 also resulted in the induction of EMT-like features, promoting cell-migration in scratch assays and elevated the proportion of stem-like side-population cells, probably in a YAP1-dependent manner. An unbiased RNA-Seq analysis in A549 and H460 cells indicated that MAP kinase (MAPK) pathway is activated upon TBK1 KD, which might also be involved in the YAP1 protein regulation. Our recent experiments further support this argument. The in-depth molecular mechanism(s) by which TBK1 regulates YAP1 in KRAS mutant cells are under investigation, and we hypothesize that this regulatory event contributes to KRAS mediated oncogenesis in NSCLC. Citation Format: Biswarup Saha, Neha Jaiswal, Namrata Bora-Singhal, Srikumar Chellappan. Molecular interplay between Tank-binding kinase (TBK1) and Yes-associated protein (YAP1) in KRAS mutant NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4394.