Abstract 202: p53 pathway restoring small molecule Prodigiosin targets chemotherapy-resistant colorectal cancer stem cells in vitro and in vivo via p73 activation

Abstract

Abstract Tumor suppressor p53 is frequently mutated or inactivated in colorectal cancer while p53 family member p73 is rarely mutated in cancer cells. Small molecules that activate p73 can elicit a p53-like tumor suppressive function and represent a novel approach for p53 pathway restoration. Colorectal tumors contain a small population of cancer stem cells (CSCs) capable of self-renewal that contributes to tumor maintenance and resistance to therapy. Targeting CSCs could improve treatment response and prolong patient survival. We tested the hypothesis that small molecule-mediated p53 pathway restoration via p73 activation can target CSCs. We have previously shown that small molecule Prodigiosin restores the p53 pathway in vitro and in vivo via activation of p73. We tested the effects of Prodigiosin on colonosphere formation, a functional in vitro model of CSC self-renewal. Prodigiosin significantly inhibits colonosphere formation and growth of multiple human colorectal cancer cell lines independent of p53 status. Aldefluor positive (Aldefluor(+)) cells represent the CSC population as they formed greater number of colonospheres and xenograft tumors upon passage, and were resistant to chemotherapeutic drugs 5-Fluorouracil and Irinotecan compared to Aldefluor negative cells. Prodigiosin prevented colonosphere formation and reduced the viability of sorted Aldefluor(+) HCT116, DLD1 and SW480 cells. Next, we tested the effects of Prodigiosin on CSC-mediated tumor growth and self-renewal in a tumor xenograft model. Prodigiosin significantly reduces the growth of tumors initiated with sorted Aldefluor(+) SW480 and DLD1 cells and prevents the passage of these tumors. No measurable toxic effects were observed in mice during the anti-CSC effects of Prodigiosin as determined by serum chemistry analysis and body weight measurement. Thus, Prodigiosin inhibits CSC self-renewal in vitro and in vivo. Mechanistic studies were performed to evaluate Prodigiosin-mediated p53-pathway restoration in CSCs. In a p53-reponsive luciferase reporter assay, Prodigiosin induced p53-pathway transcription in colonospheres, sorted Aldefluor(+) cells and tumor xenografts initiated with sorted Aldefluor(+) cells. Western blot analysis revealed that Prodigiosin elevated p73 protein levels in sorted CSCs. Prodigiosin elevated protein levels of p53/p73-regulated as well as p73-regulated p53-independent target genes in sorted CSCs. Sub-G1 analysis indicated that Prodigiosin induced cell death in colonospheres. Prodigiosin-mediated cell death in CSCs involves induction of Caspase-8 and PARP cleavage. In conclusion, Prodigiosin targets both non-CSCs and CSCs via p73-mediated p53 pathway restoration making it an attractive agent for colorectal cancer therapy. Our study has previously unrecognized implications for the role of p73 in cancer stem cell biology. Citation Format: Varun Vijay Prabhu, Bo Hong, Joshua E. Allen, Shengliang Zhang, David T. Dicker, Wafik S. El-Deiry. p53 pathway restoring small molecule Prodigiosin targets chemotherapy-resistant colorectal cancer stem cells in vitro and in vivo via p73 activation. [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 202. doi:10.1158/1538-7445.AM2014-202