MicroRNAs in normal and malignant colon stem cells and their possible role in stem cell origin of colon cancer
Date
2014
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
The role of miRNAs in colon cancer development pertaining specifically to the stem cell origin of cancer is yet to be elucidated. We hypothesized that perturbation of miR expression levels contributes to changes in target gene belonging to self-renewal pathways, which initiate colon tumorigenesis. Our miR profiling studies to identify miRs that regulate colon cancer stem cells (CSCs) were broadly divided in two parts. In one study we profiled the normal crypt bottom subsection that is enriched in stem cells and identified miR23b to be one of the miRs to have a significant differential expression. MiR23b has already been identified as having a role in renal, prostate, bladder, breast and colon cancers involving cell migration, invasion and apoptosis. Recently, a role for miR23b in ovarian CSCs and response to chemotherapy has also been elucidated. Consequently, I postulated that miR23b regulates colon CSCs. My results showed that miR23b is overexpressed in the ALDEFLUOR high sub-population of HT29 and SW480 colon cancer cells. It was shown to control the colon CSC phenotype and significantly affects proliferation, cell cycle, self-renewal, EMT, invasion and chemoresistance to the anti-cancer drug 5- FU. I validated that the colon CSC marker LGR5 is a target of miR23b, showing its role in modulating Wnt signaling. MiR23b also influences the transcription of multiple gene targets such as ATF2 and AKT2, which were identified by our extensive RNA SEQ analysis. The other aspect of the study was to identify miRs, which are differentially expressed in the CSCs as compared to the normal SCs from fresh patient samples. Normal and tumor tissue were initially screened for the expression of multiple putative CSC markers (ALDH1, LRIG1, CD166, ABCG2, BMI1, Telomerase) with an aim to decide which markers should be used alone or in combination to isolate SCs. My results indicated for the first time that SC markers ALDH1, LRIG1 and CD166 do not co-stain cells in tumors suggesting a co-existence of subpopulations of CSCs in tumor. MiR profiling identified miRs such as miR200c, miR92a, miR20a and miR93 that had a significant differential expression in ALDEFLUOR high tumor cells as compared to ALDEFLUOR high normal cells. MiR92a was also significantly upregulated in ALDEFLUOR high cells of colon cancer cell lines HT29 and regulated its proliferation. Future studies have to be done on the other newly recognized candidate miRs that show differential expression in primary tumor SCs. Understanding the role of miRs in CSCs could contribute to identification of new targets for therapeutic intervention.