5607
Shaimaa Ali Abdelhalim Mahmoud Gad
Role Of Exosomes In The Development Of Drug Resistance In Cancer
melanoma, vemurafenib, exosomes, microRNA, CDC7 and resistance
Melanoma expressing BRAFV600E mutation causes 60–80% of skin cancer-related deaths. Vemurafenib is an effective gene-targeted therapy for BRAFV600E-associated melanomas. Despite the progression-free survival correlated with BRAF inhibition by vemurafenib, resistance often emerges immediately after its antitumor effect. Tumor microenvironment has been widely accepted to participate in drug resistance in cancer. Among the extracellular vesicles, exosomes have been identified as nanovesicles (30-120 nm) that secreted to facilitate the intercellular crosstalk. Exosomes shuttle various bioactive molecules, such as proteins, DNA and microRNAs (miRNAs), from a donor cell to induce genetic modifications in a recipient cell. MicroRNAs are non-coding RNA molecules that function as gene regulators at post-transcriptional level. Exosomal miRNAs have been reported to control the expression of various genes that are involved in oncogenic signal transduction in cancer, with a subsequent alterations in cell growth, cell cycle control and apoptosis programs. Accordingly, this study aimed to investigate the role of exosomal miRNAs in promoting vemurafenib resistance in BRAFV600E-associated melanoma. In this study, vemurafenib-sensitive (parental) (A375 and WM983B) and their respective resistant (A375-NRASQ61K and WM983B-BR) melanoma cells were used; all of which harbor mutant BRAFV600E. Microarray analysis followed by qPCR were performed to identify the differentially expressed exosomal miRNAs in resistant versus parental melanoma cells. Then, transfection with miRNA mimics and inhibitors was used to investigate their functional significance; and western blot analysis was performed to identify the expression of miRNAs target genes, which was then verified in human melanoma samples. The current results showed that resistant melanoma cells displayed persistent cell growth and p-ERK expression after vemurafenib treatment, when compared to their parental cells. In addition, resistant melanoma cells differentially express a total of 31 exosomal miRNAs versus their respective parental cells. Overexpression of miR-3613-3p in A375-NRASQ61K suppressed p-ERK and CDC7 expression, and decreased resistant colonies count. However, knockdown of miR-630 expression in WM983B-BR did not affect gene expression, but reduced resistant colonies count. Inhibition of CDC7 in A375-NRASQ61K by selective CDC7 inhibitor, TAK-931, provoked cell growth arrest, reduced colonies count and suppressed the expression of p-ERK, CDC7 and its downstream effector p-MCM2. CDC7 showed higher expression in melanoma human tissues than normal skin. Conclusively, these findings suggest that exosomal miRNAs could be used as therapeutic targets to circumvent the challenge of resistance in melanoma; and selective CDC7 inhibition could be used as an alternative therapy to BRAFV600E inhibition in vemurafenib-resistant melanoma patients
2019
Ph.d
Cairo
Pharmacy