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Background In Canada, patients with esophageal cancer have the 2nd worst 5-year survival rate (14%). Esophageal squamous cell carcinoma (ESCC) is the most common esophageal cancer type worldwide. Its high mortality rate is partly due to the treatment resistance acquired by patients, which has mostly been attributed to the presence of cancer stem cells (CSCs). Therefore, targeting CSCs is a promising strategy to improve treatment and survival of ESCC patients. CSCs are a subpopulation of tumor cells presenting high self-renewal and multipotent capacities, and therefore largely contribute to tumor heterogeneity in addition to treatment resistance. Aims In this study, we aim to establish human ESCC cell lines resistant to radiotherapy (TE15RR), chemotherapy (TE15CR) and both (TE15RCR), and study the relationship between resistance acquisition and CSC properties; Methods To do so, we exposed human ESCC cell line TE15 to chronic doses of radiotherapy (2 Gy/week for a total dose of 60 Gy) and/or increasing doses of chemotherapy (5-FU (1, 2, 5, 10 and 15mM)). We harvested samples at different time points to fully investigate resistance acquisition process. Then, we further validated resistance and properties of the resistant TE15 cell lines. Results We confirmed that TE15RR cells are less susceptible to radiation than control cells and showed that they are more proliferative at baseline using MTT assays. Moreover, we demonstrated that TE15RR cells comprise a higher proportion of CD24high/CD44high CSC population using flow cytometry. We also found that they express higher levels of ALDH1, another well-known CSC marker, by qPCR and Western Blot. Our preliminary observations also suggest a higher invasive phenotype in TE15RR cells in limited dilution spheroid assay than in control cells supporting that TE15RR cells display CSC-like properties. TE15CR and TE15RCR cell lines were validated using similar approaches. Finally, mass spectrometry was performed to compare the proteomic profile of the resistant cell lines to the control cell line in order to identify key players in treatment resistance acquisition. Conclusions Brief, we developed radioresistant, chemoresistant and radiochemoresistant human ESCC cell lines and demonstrated that resistance acquisition correlates with CSC enrichment. Mass spectrophotometry revealed significant proteomic differences between resistant and control cell lines, which should lead to the identification of novel targets to overcome resistance in ESCC patients. Funding Agencies Canada research chair TIER 2 |
