The mammalian target of rapamycin (mTOR) exerts a critical role in the regulation of cell growth and cell survival, primarily through forming two complexes with raptor (mTOR complex 1; mTORC1) or rictor (mTOR complex 2; mTORC2). The mTOR axis is dysregulated in many types of human cancers, thus becoming an attractive cancer therapeutic target. Accordingly, mTOR inhibitors including the conventional allosteric inhibitors (rapamycin and its analogs; rapalogs) and novel mTOR kinase inhibitors (e.g., INK128) have been developed. However, single-agent activity of these inhibitors in the majority of cancer types is limited or modest at best. Thus, it is crucial to identify determinants that impact cell response to mTOR-targeted cancer therapy. In this study, we found that inhibition of GSK3 with both a small molecule inhibitor (e.g., SB216765) and gene knockdown substantially antagonized the suppressive effects of mTOR inhibitors (e.g., rapamycin and INK128) on the growth of cancer cells, including G1 arrest. In agreement, activation of GSK3 by expressing a constitutively activated form of GSK3[[Unsupported Character - Symbol Font &#61538;]] sensitized cells to mTOR inhibitors. Moreover, higher basal levels of GSK3 activity from 14 lung cancer cell lines were significantly associated with better cell responses to both rapamycin and INK128 (P < 0.05). These data together indicate that GSK3 plays a critical role in determining cell response to mTOR inhibitors. Interestingly, GSK3 inhibition did not alter the effects of mTOR inhibitors on suppression of mTOR signaling and cap-dependent translation, but rescued downregulation of cyclin D1 and c-Myc caused by mTOR inhibitors, suggesting that mTOR inhibitors decrease cyclin D1 and c-Myc levels independent of inhibition of mTOR-mediated protein translation. Instead, we found that inhibition of mTOR (e.g., by INK128) decreased the stabilities of both cyclin D1 and c-Myc. Moreover, inhibition of proteasome (e.g., with MG132) prevented cyclin D1 and c-Myc reduction by INK128. Hence it is apparent that mTOR inhibition facilitates cyclin D1 and c-Myc degradation, leading to reduction of cyclin D1 and c-Myc. Together, these findings suggest a novel mechanism by which mTOR signaling positively regulates the stability of cyclin D1 and c-Myc. Our current findings thus warrant further investigation of the potential impact of GSK3 inhibition on the efficacy of mTOR inhibitors in the clinic. (This study was supported by the Georgia Cancer Coalition Distinguished Cancer Scholar award and NIH/NCI R01 CA118450. SS Ramalingam, FR Khuri and SY Sun are Georgia Cancer Coalition Distinguished Cancer Scholars)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2803. doi:1538-7445.AM2012-2803