CRISPR/Cas9, a new approach to successful knockdown of ABCB1/P-glycoprotein and reversal of chemosensitivity in human epithelial ovarian cancer cell line | ||
| Iranian Journal of Basic Medical Sciences | ||
| مقاله 10، دوره 21، شماره 2، اردیبهشت 2018، صفحه 181-187 اصل مقاله (1.14 M) | ||
| نوع مقاله: Original Article | ||
| شناسه دیجیتال (DOI): 10.22038/ijbms.2017.25145.6230 | ||
| نویسندگان | ||
| Leyla Norouzi-Barough1؛ Mohammad reza Sarookhani* 2؛ Rasoul Salehi3؛ Mohammadreza Sharifi3؛ Sahar Moghbelinejad4 | ||
| 1Department of Molecular Medicine, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran | ||
| 2Cellular and Molecular Research Center, Qazvin University of Medical Sciences, Qazvin, Iran | ||
| 3Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran | ||
| 4Department of Biochemistry and Genetic, School of Medicine, Qazvin University of Medical Sciences, Qazvin, Iran | ||
| چکیده | ||
| Objective(s): Multidrug resistance (MDR) is a major obstacle in the successful chemotherapy of ovarian cancer. Inhibition of P-glycoprotein (P-gp), a member of ATP-binding cassette (ABC) transporters, is a well-known strategy to overcome MDR in cancer. The aim of this study was to investigate the efficiency and ability of CRISPR/Cas9 genome editing technology to knockdown ABCB1 gene expression in adriamycin resistant (A2780/ADR) ovarian cancer cell line and evaluate the sensitivity changes to doxorubicin. Materials and Methods: Three single-guide RNAs (sgRNAs) targeting the fourth and fifth exons of human ABCB1 gene were designed in this study. Expression level of ABCB1 was detected using quantitative real time PCR (qRT-PCR) after co-transfection of all three sgRNAs into A2780/ADR cell line and subsequent antibiotic selection. Drug sensitivity to doxorubicin was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results: The results showed that CRISPR/Cas9 system could significantly reduce the expression of P-gp. The dramatic decline in ABCB1 gene expression was associated with increased sensitivity of cells transfected with sgRNAs to doxorubicin. Conclusion: Based on the results of this study, it is concluded that the CRISPR-based systems, used in the present study, effectively down-regulated the target gene and acted as an ideal and cost-effective tool for gene editing of A2780/ADR cell line resulting in restoration of nonmalignant phenotype. | ||
| کلیدواژهها | ||
| CRISPR؛ Doxorubicin؛ Drug resistance؛ Ovarian cancer؛ P-glycoprotein | ||
| مراجع | ||
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