Selective sensitization of Ras-mutant (Ras-m) cancer cells to DNA-damaging chemotherapy by Wee1 inhibition with AZD1775.

e13517 Background: Human cancers harboring Ras-mutated (m) oncogenes are resistant to current EGFR-targeted therapies. Given the many patients with Ras-m cancers, limited treatment options available, and their poor outcomes, there is a great need to develop treatment modalities that effectively target these tumor types. We previously reported that Ras-m tumors activate the ATR/Chk1 DNA damage checkpoint through signaling from wild-type (wt) Ras proteins, thus conferring resistance to DNA damaging chemotherapeutic agents in vitro and in vivo.1 Consequently, we hypothesized that abrogating the ATR/Chk1 DNA damage checkpoint by downstream Wee1 kinase inhibition would selectively sensitize Ras-m and BRaf-m tumors to DNA damaging chemotherapy. Methods: Toassess whether a Wee1 kinase inhibitor (KI) enhanced the cytotoxicity of DNA damaging agents in Ras-m and Raf-m cells, we utilized a selected panel of colon cancer cell lines with the following mutational backgrounds (2-3 cell lines/group): 1) Ras-m/p53-m; 2) Ras wt/p53-m; 3) Ras-m/p53 wt and 4) BRaf-m/p53-m. 4 log dose response cell viability assays were used to determine the single agent IC50 for the topoisomerase 1 inhibitor, SN38, and the Wee 1 KI AZD1775. Cells were then treated with the two agents at doses ranging in a 5-fold difference below each individual IC50 (0.04X, 0.2X). For all viability assays, cells were plated in 96-well plates in triplicate; viability was determined by MTT assay. Results: AZD1775 treated Ras-m/p53-m cells were ~10-100 fold more sensitive to SN38. Notably, Wee1 inhibition did not sensitize Ras wt/p53-m or Ras-m/p53 wt cells to SN38. Wee1 inhibition in BRaf-m/p53-m cells led to a modest ~3-5 fold sensitization to SN38, with the exception of one cell line which showed ~ 270 fold sensitization. Conclusions: The Wee1 KI AZD1775 strongly enhances the antitumor effect of SN38 in this panel of Ras-m/p53-m cancer cells but not in cancer cells where either p53 alone or Ras alone were mutated. Although this is a limited panel of cell lines, our findings support further testing of this combination in human tumors harboring concomitant Ras and p53 mutations. Supported by NYU GI Cancer Fund. 1. Cancer Cell. 2014 Feb 10;25(2):243-56