Abstract 4078: Investigating chemoresistance mechanisms in colorectal cancer stem cells

Abstract

Abstract Treatment of advanced colorectal cancer (CRC) is impacted by a lack of selectivity, the development of drug resistance and overall tumour heterogeneity. Cancer stem cells (CSCs) are a subpopulation of tumour cells believed to be responsible for tumour recurrence due to their ability to self-renew, differentiate and evade current treatments. This project explores the responses of CRC CSCs to chemotherapy and aims to identify therapeutic targets of the CSC population. We used serial colonosphere culture to enrich for CSCs from two human CRC cell lines, HCT116 and SW480. Limiting dilution analysis showed that CSC enriched cell lines have significantly higher colonosphere formation ability compared to their respective parental cells. Further characterization of CSC cell lines was performed via qPCR and flow cytometry to determine the expression of colorectal CSC markers. A significant upregulation in CD166 and EpCAM gene expression was seen in HCT-CSC and SW-CSCs compared to parental cell lines. In vitro assays were used to determine cellular responses to three commonly employed chemotherapeutics, 5-fluorouracil, cisplatin and epirubicin. A limiting dilution analysis demonstrated that cells treated with chemotherapy for 48 hours had reduced colonosphere forming potential. Cell viability in response to chemotherapy was analyzed via PrestoBlue assays in both monolayer and 3D culture. CSC and parental cell lines responded similarly in a dose-dependent manner to treatment in monolayer, but CSC colonospheres were more resistant to high doses of chemotherapy than their respective parental cells. ABC transporter expression plays a major role in the development of multi-drug resistance so the expression of ABCB1, ABCC1 and ABCG2 was examined via qPCR. There was a significant upregulation in ABCB1 and ABCG2 in HCT-CSCs compared to HCT-P cells. SW-CSCs also had significant upregulation in ABCB1 compared to SW-P cells. Other resistance mechanisms may also be key targets for therapeutic intervention including the functional CSC marker aldehyde dehydrogenase (ALDH). The detoxification effects of ALDH could have a protective effect against the surges of reactive oxygen species resulting from chemotherapy treatment. The expression of ALDH and ABC transporter genes was also explored in response to chemotherapy. The goal of this project is to elucidate drug resistance mechanisms in colorectal CSCs, in order to target and eradicate the tumour cells responsible for metastases and recurrent disease. Our analysis demonstrates that colorectal CSCs possess heightened protective mechanisms in comparison to other cancer cells originating from the same tumour. Future experiments will further investigate these resistance mechanisms to determine their therapeutic potential. Note: This abstract was not presented at the meeting. Citation Format: Stacey J. Butler, Lisa Richardson, Nathan Farias, Brenda L. Coomber. Investigating chemoresistance mechanisms in colorectal cancer stem cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4078. doi:10.1158/1538-7445.AM2015-4078