Abstract 5608: Functional genetic screens to find modulators of sensitivity to inhibitors of the PI3K/mTOR pathway

Abstract

Abstract The PI3K/mTOR pathway is an important signaling pathway which is often hyperactivated in breast cancer by genetic events. Inhibitors of several PI3K pathway members have been developed in recent years. Clear improvements in survival have not been demonstrated in most cancer patients, including breast cancer patients. Clearly, additional targets for combination therapies and biomarkers of resistance are needed. We performed siRNA based screens, to find genes that can modulate the sensitivity of breast cancer cells to PI3K and mTOR inhibitors. Resistance screens: We screened the retroviral NKI hairpin library, consisting of 21,000 hairpins against 8,000 genes, in four breast cancer cell lines, using rapamycin (an inhibitor of mTORC1), AZ4 (a small molecule inhibitor of mTORC1 and 2) and the dual PI3K/mTOR inhibitors PI103 and NVP-BEZ235. Common hits identified in these resistance screens and validated in multiple cell lines were PTEN, GSK3A/B and ARID1A. PTEN and GSK3A/B are known members of the PI3K pathway; resistance by PTEN loss is most likely caused by hyperactivation of PI3K signaling, whereas GSK3A/B knockdown probably leads to decreased degradation of Cyclin D (published previously). Knockdown of the SWI/SNF complex component ARID1A could also induce resistance against PI3K and mTOR inhibition in various cell lines. Similar to PTEN knockdown, ARID1A downregulation caused hyperactivation of PI3K signaling. Further studies should elucidate which SWI/SNF transcriptional targets are responsible for this induction of PI3K signaling. Synthetic lethal screens: We screened the Dharmacon ‘kinome’ and ‘druggable genome’ siRNA libraries (over 8,000 genes) in HCC1954 breast cancer cells, to find genes whose knockdown enhances the toxicity of mTOR inhibitors. Few genes could reproducibly enhance the sensitivity of breast cancer cells to mTOR inhibition; most of these genes, such as RHEB and PIK3CA, are important members of the PI3K/mTOR pathway. We found knockdown of the anti-apoptotic MCL1 gene to increase sensitivity to PI3K and mTOR inhibition by causing an increase in apoptosis. Although not clearly established as a specific PI3K pathway member, MCL1 expression may be regulated by downstream proteins in the PI3K/mTOR pathway. Knockdown of the integrin α/vitronectin receptor ITGAV had a sensitizing effect on PI3K/mTOR inhibition in HCC1954 and SKBR3 breast cancer cells. The mechanism of this interaction is still under investigation but may involve upregulation of integrin function in response to PI3K/mTOR inhibition. In conclusion, most of the genes that can modulate the sensitivity of breast cancer cells to PI3K/mTOR inhibition are known members of the PI3K pathway, however ARID1A and ITGAV are new modulators of sensitivity to PI3K pathway inhibition. 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 5608. doi:1538-7445.AM2012-5608