Approximately 30% of human non-small cell lung cancer (NSCLC) patients harbor a somatic KRAS mutation resulting, in aberrant activation of downstream signaling pathways that control cell proliferation, cell growth, and cell survival. Importantly, alleles of LKB1, a serine/threonine kinase that functions as a tumor suppressor, are somatically inactivated in ~30% of NSCLCs within KRAS-mutant NSCLC. The loss of LKB1 gives rise to aggressive, highly metastatic, and highly drug resistant tumors. We have previously demonstrated that the inactivation of the tumor suppressor lkb1 rendered mutant kras murine NSCLC resistant to targeted agents including BET bromodomain and kinase inhibitors. However, the mechanism by which LKB1 inactivation attenuates the efficacy of the therapies remains elusive. Consequently, investigation of the molecular-basis of drug resistance to formulate effective therapies for this KRAS-mutated and LKB1-inactivated (KRAS/LKB1) subset of patients is warranted. Using lung adenocarcinoma cells that lack LKB1 expression, we have established isogenic cells that express vector control, wild-type LKB1, or kinase-dead LKB1. Our preliminary data using these isogenic cell lines suggest the previously unrealized function of LKB1 to activate HSP90 to stabilize client signaling proteins. Furthermore, we have performed p23 immunoprecipitation (IP) followed by Western blot for HSP90 to assess the HSP90 ATPase activity. In the cells ectopically expressing wild-type LKB1, p23 co-immunoprecipitated with HSP90 showing that HSP90 is in active conformation. In contrast, few HSP90 co-immunoprecipitated with p23 in the cells that express the kinase-dead version of LKB1 or vector control, demonstrating that little HSP90 is in active conformation without LKB1 kinase activity. Using NetworKIN, a web-based bioinformatic analysis, we find putative LKB1 phosphorylation motifs on HSP90. These results suggest that LKB1 phosphorylates and activates HSP90. Consequently, the inactivation of LKB1 contributes to the destabilization of key HSP90 client signaling proteins and inadvertently promotes compensatory mechanisms to stabilize the select signaling molecules. Using our LKB1-isogenic cell lines, we demonstrated that LKB1 activates HSP90 to stabilize RAF1, resulting in MAPK pathway activation. In contrast, LKB1 inactivation led to reduced HSP90 activity, which reduced MAPK activity and elevated IGF-1R-mediated phosphatidylinositol 3-kinase (PI3-K) activity in vivo and in vitro models of mutant KRAS NSCLC. The aberrant activation of PI3-K pathway augments survival by the stabilization of the anti-apoptotic molecule, MCL1. The use of HSP90 inhibitors does not fully inhibit MAPK or PI3-K pathways due to the compensatory mechanisms chaperoning the IGF-1R axis. Importantly, we confirmed that canonical LKB1-AMPK-mTOR pathway does not fully control PI3K or MAPK signaling. Taken together, our data supports the presence of an HSP90-independent mechanism to stabilize kinases, including IGF-1R, to make KRAS/LKB1 NSCLC resistant to targeted therapies.

Citation Format: Jeffrey H. Becker, Michael P. Kahle, Margaret Soucheray, Ines Pulido, Fatima Al-Shahrour, Manuel Sanchez del Pino, Kwok-Kin Wong, Julian Carretero, Takeshi Shimamura. A new role for LKB1 to regulate Heat Shock Protein 90 activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-085.