Abstract 3906: Combined inhibition of PI3K isoforms and mTOR kinase is critical for cancer stem cell inhibition by VS-5584

Abstract

Abstract Cancer stem cells (CSCs) represent a subpopulation of cancer cells that have tumor-initiating capability, are particularly resistant to chemotherapy, and can mediate tumor recurrence both locally and at metastatic sites. As such, these cells represent a critical challenge for effective treatment of cancer. High-throughput screening for small molecules that preferentially target CSCs identified dual inhibitors of the PI3K/mTOR pathway, suggesting the importance of this signaling pathway for CSC biology. Here we demonstrate that ablation of mTOR or individual PI3K isoforms by isoform-specific siRNA is not sufficient to reduce the proportion of CSCs. In contrast, combined knock down of PI3K isoforms and mTOR effectively reduced the proportion of CSCs in tumor cell lines. VS-5584 is a potent and selective pan-PI3K/TORC1/TORC2 inhibitor with equipotency against all four human Class I PI3K isoforms and the mTOR kinase. We demonstrate that VS-5584 preferentially targets CSCs in multiple orthogonal assays both in vitro and in human tumor xenograft models. Cancer stem cells express high levels of aldehyde dehydrogenase, and an Aldefluor assay that measures activity of this enzyme was used to identify CSCs. VS-5584 decreased the percentage of Aldefluor+ cells across multiple breast cancer cell lines. We demonstrated that VS-5584 preferentially induced apoptosis in Aldefluor+ SUM159 cells relative to Aldefluor- cells as measured by Annexin V and Caspase 3/7 assays. In contrast, paclitaxel induced more apoptosis in Aldefluor- than Aldefluor+ cells and enriched the percentage of CSCs. Another characteristic of CSCs is their enhanced ability to efflux cytotoxic agents. This drug-resistant CSC population, called side population (SP), can be monitored by exclusion of Hoechst dye. VS-5584 effectively eliminated the SP CSCs across multiple cancer types, while cisplatin and etoposide increased this subpopulation. Furthermore, ex vivo treatment of primary breast and ovarian tumor specimens with VS-5584 decreased the proportion of CSCs as measured by the Aldefluor assay and cell surface markers. Significantly, VS-5584 also targets CSCs in vivo in MDA-MB-231 triple negative and MCF7 ER+ breast cancer xenograft models as evidenced by decreases in the percentage of Aldefluor+ cells, tumorsphere-forming efficiency, and tumor-initiating capability in an in vivo limiting dilution re-implantation assay. Consistent with the notion that combined inhibition of PI3K isoforms and mTOR is critical for exerting a strong anti-CSC effect, the mTORC1-selective inhibitor everolimus did not reduce CSCs in the MCF7 xenograft model. The potent anti-CSC activities in primary patient cancer tissue and in xenograft models provide strong rationale for the clinical development of VS-5584 in combination with agents targeting the bulk tumor to potentially achieve durable clinical responses for cancer patients. Citation Format: Vihren N. Kolev, Quentin G. Wright, David T. Weaver, Jennifer E. Ring, Christian M. Vidal, Mahesh V. Pavdal, Jonathan A. Pachter, Qunli Xu. Combined inhibition of PI3K isoforms and mTOR kinase is critical for cancer stem cell inhibition by VS-5584. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3906. doi:10.1158/1538-7445.AM2014-3906