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Targeting the splice factor kinase CLK1 in prostate cancer cells

Uzor, Simon Ogbonnaya

Targeting the splice factor kinase CLK1 in prostate cancer cells Thumbnail


Authors

Simon Ogbonnaya Uzor simon.uzor@uwe.ac.uk



Abstract

Prostate cancer is the most rampant diagnosed cancer and the second leading cause of cancer related death in men between middle age and old age. There is a need to identify the molecular genetic processes that underpin prostate cancer and to search for new treatments. Alternative splicing affects over 94% of human genes and aberrant splicing is implicated in prostate cancer. Splicing is regulated by splice factors and protein kinases (the latter including SRPK1 and CLK1). The CLK1 protein kinase specifically regulates alternative splicing by phosphorylation of SR proteins within the nucleus, particularly in nuclear speckles (splice factor storage sites). CLK1 is also found to be overexpressed during malignant prostate cell transformation-; therefore targeting the splice factor kinase CLK1 by its specific chemical inhibitor (TG003) could be therapeutically useful. We confirm that CLK1 expression is itself regulated through alternative splicing: skipping of exon 4 or retention of intron 4 results in truncated, inactive CLK1. The aims of this project are to study CLK1 alternative splicing and to explore the effects of targeting the splice factor kinase CLK1 in prostate cancer.
Prostate cancer cell lines (androgen independent PC3 and DU145 cells) were treated independently for 24, 48 and 72hrs with varying concentrations (10nM -100μM) of the
benzothiazole compound (TG003) a specific inhibitor of CLK1. Results suggest that chemical inhibition of CLK1 with TG003 treatment suppressed growth and induced apoptosis in prostate cancer cell lines, as well as causing decreased cell migration and invasion. Similar effects were observed when CLK1 expression was reduced with an siRNA. There was also increased E-cadherin expression with TG003 treatment; in
contrast, vimentin expression was reduced suggesting reversal of endothelialmesenchymal transition following TG003 treatment. RT-PCR analysis revealed that TG003 treatment altered CLK1’s own splicing by altering exon 4 skipping rates in a iv dose dependent manner, suggesting that a feedback loop mechanism contributes to the regulation of CLK1 expression. In vivo mouse work with PC3 cell line xenografts
showed a highly significant reduction in tumour growth and volume following intraperitoneal TG003 administration (10 and 50μM). In conclusion, the findings presented in this thesis suggest that targeting CLK1 may bring considerable anticancer benefits.

Thesis Type Thesis
APA6 Citation Uzor, S. O. Targeting the splice factor kinase CLK1 in prostate cancer cells. (Thesis). University of the West of England. Retrieved from https://uwe-repository....ribe.com/output/1490733
Keywords Prostate cancer, alternative splicing, CLK inhibition, TG003
Additional Information Additional Information : PhD thesis submitted on Friday 10th May, 2019. Viva due on Wednesday 26th June, 2019
Corporate Creators : Tertiary Education Trust Fund (TETFund), Nigeria.

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