Abstract 5399: Growth inhibition, cytotoxicity and signaling studies of PI3K & MEK inhibitor combinations in colorectal carcinoma cell lines

Abstract

Abstract MAPK and PI3K signalling pathways are frequently activated in human cancers, and represent promising therapeutic targets. Previous studies suggest that combined targeting of these pathways may be necessary for optimal therapeutic activity, hence the aim of this study was to evaluate the MEK inhibitors, ARRY-142886 and PD 0325901, alone and in combination with the dual mTOR/PI3K inhibitor, NVP-BEZ235, or the pan class I PI3K inhibitor, GDC-0941, in colorectal cancer cell lines. Growth inhibition, survival and signal transduction were measured using the Sulforhodamine B assay, clonogenicity and western blotting, respectively. Median effect analysis revealed that all MEK/PI3K inhibitor combinations exhibited marked synergistic growth inhibition in both HCT116 and HT29 cell lines. GDC-0941 displayed the greatest synergy in combination with either MEK inhibitor in the HCT116 cell line, and preliminary results suggest that this is also the case in the HT29 cell line. At concentrations up to 10 µM only NVP-BEZ235 was cytotoxic after 72 hours exposure in either colorectal cancer cell line with an LC50 of 0.51 µM in the HCT116 cell line and an LC50 of 0.44 µM in the HT29 cell line. However, no increase in cytotoxicity was observed with MEK/PI3K inhibitor combinations in the HCT116 cell line, and studies in the HT29 cell line are ongoing. Western blotting using HCT116 cell lysates revealed that NVP-BEZ235 exhibits stronger inhibition of AKT and 4EBP1 phosphorylation, and similar inhibition of S6 phosphorylation, compared to GDC-0941. Both PD 0325901 and ARRY-142886 inhibited ERK phosphorylation. However, no clear or consistent additional effect on S6, ERK or 4EBP1 phosphorylation was observed when the MEK and PI3K inhibitors were combined. These studies confirm that MEK and PI3K inhibitors are predominantly cytostatic, as opposed to cytotoxic, when used alone or in combination. The dual mTOR/PI3K inhibitory action of NVP-BEZ235 may increase its ability to inhibit 4EBP1 phosphorylation, and thereby protein translation. Furthermore, the lower level of synergy exhibited by NVP-BEZ235 in combination with MEK inhibitors, compared to GDC-0941, may be due to the inhibition of mTOR and thereby 4EBP1 phosphorylation. Hence studies have commenced using the mTOR inhibitor, rapamycin, in combination with GDC-0941. In the HCT116 cell line, preliminary results suggest that the combination of GDC-0941 and rapamycin is considerably more growth inhibitory than either compound alone. These studies confirm that dual targeting of PI3K and MEK can induce synergistic growth inhibition; however, the detailed effects of specific inhibitors should be investigated to identify optimal combinations. The research was funded by grants from the Medical Research Council UK and UCB Celltech Ltd. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5399.