Abstract
Introduction: More effective, less toxic treatments for recurrent ovarian cancer are needed. Up to 80% of ovarian cancers show activation of the Src kinase and up to 40% show MEK/MAPK pathway activation. Therapies targeting signaling pathways that drive proliferation have shown promise in ovarian cancers but the development of drug resistance presents a major problem. Saracatinib, is a potent inhibitor of Abl and Src family kinases and selumetinib, is a potent, selective, ATP-uncompetitive inhibitor of MEK 1/2 kinases. Our earlier work has shown that resistance to saracatinib monotherapy emerges within weeks in cell culture and within months in xenograft models. Since oncogenic activation of both Src and MEK pathways has been observed in ovarian cancers and because resistance to Src inhibition rapidly emerges with monotherapy, we hypothesized that blocking both kinases with dual therapy might more effectively decrease ovarian cancer cell proliferation than either drug alone.
Experimental Design: Dual Src and MEK blockade effects on cell cycle and survival were assayed in ovarian cancer lines, and a primary human ovarian cancer tumor cultured immediately in vitro. Cells enriched for the stem cell marker, aldeflour dehydrogenase 1 (ALDH1+) were sorted by flow cytometry. Drug effects on tumor sphere formation and on cell cycle profiles of ALDH+ cells and those lacking this activity were analyzed. In vivo experiments are ongoing.
Results: Src activity is greater in ovarian cancer lines than normal human ovarian epithelial cells (HOSE). In saracatinib naïve ovarian cancer cells, Src inhibition led to partial growth arrest after 48 hrs of treatment. However, resistance of ovarian cancer cells to saracatinib monotherapy emerged rapidly, over a period of weeks of continuous drug exposure in vitro and in vivo. Activation of the MEK and Akt pathways developed with prolonged saracatinib treatment. While each drug alone caused a partial growth arrest in saracatinib naïve lines, treatment with both saracatinib and the MEK inhibitor drug, selumetinib, caused synergistic cell cycle arrest which was mediated by p27 binding to cylin E/cdk2. Combination treatment decreased Src, Akt and MAPK activities. Combination treatment was required for cell cycle arrest in saracatinib resistant cells. Combination treatment also induced apoptosis by an increase in cleaved PARP and induced autophagy as shown by an increase in LC3-II. Src and MEK activities were increased in ALDH1+ sorted populations compared to the ALDH- population and combination treatment decreased the % of ALDH1+cells and the % of sphere-generating cells.
Conclusions: MEK inhibition augments the effects of saracatinib by blocking bypass activation of MEK and Akt pathways, increasing cell cycle arrest, impairing survival and targeting self-renewing subpopulations. These data support further pre-clinical and clinical evaluation of combined saracatinib and MEK inhibition in ovarian cancer.
Citation Format: Fiona Simpkins, Diana Azzam, Joyce M. Slingerland. Preclinical efficacy of combined MEK and Src inhibition: MEK inhibitor reverses resistance to Src inhibitor therapy in ovarian cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: From Concept to Clinic; Sep 18-21, 2013; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2013;19(19 Suppl):Abstract nr B50.
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