Date of Award
5-1-2019
Thesis Type
phd
Document Type
Thesis (Restricted Access)
Divisions
science
Department
Faculty of Science
Institution
Universiti Malaya
Abstract
Anoikis is apoptosis induced when cells are detached from the extracellular matrix and neighbouring cells. Since anoikis serves as a regulatory barrier, cancer cells often acquire resistance towards anoikis during tumorigenesis to become metastatic. MicroRNAs (miRNAs) are short strand of RNA molecules regulating genes post-transcriptionally, by binding to mRNAs and reducing the expression of its target genes. This study aims to elucidate the role of a novel miRNA, miR-6744-5p, in regulating anoikis in breast cancer and identify its target gene. Anoikis resistant variant of luminal A type breast cancer MCF-7 cell line (MCF-7-AR) was generated by selecting and amplifying surviving cells after repeated exposure to growth in suspension. miRNA microarray revealed a list of dysregulated miRNAs, from which miR-6744-5p was chosen for overexpression and knockdown studies. In MCF-7, overexpression of miR-6744-5p increased anoikis as shown by viability and caspase-3/7 activity assay, inhibited cell migration as shown by wound healing assay and increased E-cadherin expression as shown by Western blotting. Knockdown of miR-6744-5p decreased anoikis, increased cell migration and decreased E-cadherin expression. In the invasive triple-negative breast cancer cell line MDA-MB-231, overexpression of miR-6744-5p promoted anoikis and inhibited cell migration but knockdown of miR-6744-5p produced no effect. Additionally, overexpression of miR-6744-5p also induced morphological changes of MDA-MB-231 cells and inhibited invasiveness of the cells in vitro in transwell invasion assay and in vivo in zebrafish larva metastasis model. Furthermore, N-acetyltransferase 1 (NAT1) has been identified and validated as the direct target of miR-6744-5p using luciferase reporter assay and western blot. Overall, this study has proven the ability of miR-6744-5p to increase anoikis in both luminal A and triple negative breast cancer cell lines, highlighting its therapeutic potential in treating breast cancer.
Note
Thesis (PhD) - Faculty of Science, Universiti Malaya, 2019.
Recommended Citation
Sharan, Malagobadan, "Elucidation of microRNA miR-6744-5p regulation of anoikis in breast cancer cell lines and potential for therapeutic applications / Sharan Malagobadan" (2019). Student Works (2010-2019). 6341.
https://knova.um.edu.my/student_works_2010s/6341