Molecular mechanisms of imatinib resistance in gastrointestinal stromal tumor with focus on microRNAs

Gastrointestinal stromal tumor (GIST) is mainly initialized by mutations in receptor tyrosine kinase genes KIT or PDGFRA. The development of imatinib, a small molecule inhibitor that targets these tyrosine kinase receptors, remarkably improved patient outcome. However, imatinib resistance remains a major therapeutic challenge in GIST therapy, and its underlying mechanisms are still not completely understood. This thesis work aimed to explore the role of microRNAs (miRNAs) and DOG1 in imatinib resistance of GIST.

In Paper I, we identified specific miRNA signatures associated with imatinib resistance, metastatic disease, KIT mutational status and survival in GIST patients treated with neoadjuvant imatinib. Importantly, we demonstrate that miR-125a-5p modulates imatinib response in the single KIT-mutated GIST882 cells through PTPN18 regulation. This study highlights the clinical impact of miRNAs in GIST patients treated with imatinib pre-operatively, and suggests the important role of miR-125a-5p and PTPN18 in imatinib resistance of GISTs.

In Paper II, we tested our hypothesis that miR-125a-5p overexpression in imatinib-resistant GISTs suppresses PTPN18 expression that subsequently leads to defective FAK dephosphorylation. Indeed, we demonstrate that silencing of PTPN18 increased FAK phosphorylation in GIST cells, and the acquired imatinib-resistant GIST882R cells exhibited higher pFAK and lower PTPN18 expressions than the imatinib-sensitive parental cells. FAK and pFAK expressions are also associated with imatinib resistance in GIST specimens. This study highlights the potential role of PTPN18 and pFAK in imatinib resistance of GIST.

In Paper III, we found that miR-320a and miR-320b are upregulated and their potential target MCL1 is downregulated in imatinib-treated GISTs. Imatinib treatment affects MCL1 and miR-320 levels in GIST882 cells, and the imatinib-resistant GIST882R cells showed higher levels of the anti-apoptotic MCL1L isoform and lower expression of miR-320a/b as compared to GIST882 cells. This study suggests that miR-320a/b and MCL1 play a role in imatinib-induced cell death and resistance in GIST.

In Paper IV, we evaluated the functional role of DOG1 in imatinib-resistant GIST48 and –sensitive GIST882 cells using specific DOG1 activator and inhibitor. We showed that DOG1 is localized in different cellular compartments in imatinib-resistant and -sensitive GIST cells. Pharmacological modulation of DOG1 activity has subtle effect on cell viability and proliferation, but may shift early apoptotic cells to late apoptotic stages in GIST48 cells. Overall, this thesis work describes the role of miRNAs in cell viability and resistance to imatinib treatment in GIST.

List of scientific papers

I. Akçakaya P, Caramuta S, Åhlén J, Ghaderi M, Berglund E, Östman A, Bränström R, Larsson C and Lui WO. microRNA expression signatures of gastrointestinal stromal tumors: associations to imatinib resistance and patient outcome. British Journal of Cancer 2014; 111(11):2091-2102
https://doi.org/10.1038/bjc.2014.548

II. Akçakaya P, Gangaev A, Lee L, Zeljic K, Hajeri P, Berglund E, Ghaderi M, Åhlén J, Bränström R, Östman A, Larsson C and Lui WO. FAK phosphorylation is regulated by PTPN18 and associated with imatinib resistance in gastrointestinal stromal tumors. [Manuscript]

III. Akçakaya P, Zeljic K, Åhlén J, Gangaev A, Ghaderi M, Caramuta S, Berglund E, Bränström R, Larsson C and Lui WO. Involvement of MCL1 and miR-320 in imatinib-induced cell death of gastrointestinal stromal tumor cells. [Manuscript]

IV. Berglund E, Akçakaya P, Berglund D, Karlsson F, Vukojević V, Lee L, Bogdanović D, Lui WO, Larsson C, Zedenius J, Fröbom R, Bränström R. Functional role of the Ca2+-activated Cl- channel DOG1/TMEM16A in gastrointestinal stromal tumor cells. Experimental Cell Research 2014; 326(2):315-325
https://doi.org/10.1016/j.yexcr.2014.05.003