Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.731457
Title: Targeting splice factor kinases in chronic myeloid leukaemia
Author: Alabouh, Hanan A.
ISNI:       0000 0004 6497 0049
Awarding Body: University of the West of England
Current Institution: University of the West of England, Bristol
Date of Award: 2017
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Abstract:
Alternative splicing of pre-mRNA is a ubiquitous and versatile mechanism controlling the expression of human genes. In humans, approximately 93% of multi-exonic genes are alternatively spliced, producing functionally distinct proteins. Dysregulation of alternative splicing results in the overexpression of the oncogene splice factor SRSF1, which has been implicated in various types of tumours. Initial research has shown that the alternatively spliced isoforms of vascular endothelial growth factor (VEGF) (pro- angiogenic) promotes tumorigenesis. VEGF is regulated through serine-arginine (SR)- specific protein kinase SRPK1 and SRSF1. Additionally, the activity and intracellular localisation of the SRSF1 is regulated by the kinases SRPK1 and Clk1. Therefore, it is worthly to target SRPK1 and SRSF1 in chronic myelogenous leukaemia cell line (K562 cells). Chronic myeloid leukaemia (CML) is a myeloproliferative disorder of stem cell origin. It arises from BCR-ABL1 fusion forming the Philadelphia chromosome. This fusion leads to a constitutively activated tyrosine kinase. Treatment of CML was revolutionised by tyrosine kinase inhibitors (TKIs). Despite the success of TKI in eliminating CML, drug resistance develops in patients over time. Allogenic bone marrow transplantation (BMT) is the only curative therapy, however the mortality rates increase due to infection. The precise role of SRPK1 and Clk1 in haematological malignancies is unclear. This study has sought to target SRPK1, Clk1, TK and SRSF1 through inhibition or knockdown to identify the effects of alternative splicing on the expression of CASPASE-9, RON and RUNX1 in K562 cells. Additionally, this study would xxi investigate the drug efficacy of imatinib (TK inhibitor) used in conjunction with either SPHINX (SRPK1 inhibitor) or TG003 (Clk1 inhibitor) in K562 cells. In this study, it may identify novel therapeutic targets that could be beneficial for patients with leukaemia. This study demonstrated that targeting SRPK1 and SRSF1 through inhibition or knockdown had little or no significant positive effect on the alternative splicing of CASPASE-9, RON and RUNX1 in K562 cells. Comparatively, targeting Clk1 with the selective inhibitor TG003, resulted in significantly positive effects on the alternative splicing for both CASPASE-9 and RON and RUNX1. This was confirmed by increased apoptosis of K562 cells. Knockdown studies of Clk1, showed a significant effect on RUNX1 only, eliciting the production of the Runx1 isoform. Finally, co-treatment of imatinib (TK inhibitor) and SPHINX elicited an effect on alternative splicing resulting in an overall shift to the pro-apoptotic isoforms using minimal doses of the drugs. Additionally, combinations of kinase inhibitors (imatinib and TG003) exhibited a more effective shift in RON splicing towards Ron isoforms. This suggests that co- therapy may offer a potential therapy for CML patients. The introduction of tyrosine kinase inhibitors has successfully improved the prognosis of CML. However, intrinsic and acquired resistance to imatinib is becoming an ever- increasing clinical problem. This study, for the first time, has demonstrated novel therapeutic target to Clk1 that affects the alternative splicing of CASPASE-9, RON and RUNX1 in K562 cells. Targeting of the genes is essential in the treatment of CML as cell differentiation, migration, invasion and apoptosis are affected. This study not only shows that the Clk1 selective inhibitor TG003 is beneficial on alternative splicing of xxii CASPASE-9 but conjucated inhibitors of imatinib and SPHINX induce a greater effect on the splicing of CASPASE-9 in K562 cells. Compared to the use of imatinib alone, co-treatment using both drugs maybe more effective in the treatment of CML and therefore may circumvent acquired drug resistance seen in patients with leukaemia.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.731457  DOI: Not available
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