Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Fan, K.
miR-375 in Merkel cell carcinoma, more than a serum biomarker.
PhD-Studium (Doctor of Philosophy); Humanmedizin; [ Dissertation ] Graz Medical University; 2020. pp. 122
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- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Becker Jürgen Christian
- Heinemann Akos
- Schrama David
- Zebisch Armin
- Altmetrics:
- Abstract:
- Dysregulation of miRNAs is found in all known cancer types and essential for carcinogenesis, metastasis and treatment responses. Certain features of miRNAs, such as circulating in the blood and stability, allow them to act as biomarker for cancer diagnosis, prognosis and monitoring. In Merkel cell carcinoma (MCC), rare and aggressive skin cancer, studies regarding miRNAs are rather limited. Thus, we performed miRNAs profiling in MCCs. miR-375 is identified and confirmed as highly abundant miRNAs in classical MCC cell lines and tumor tissues. Next, presence of miR-375 in MCC conditioned mediums and sera of two preclinical MCC models indicates it is likely present as a circulating miRNA in MCC patients. Indeed, miR-375 serum levels distinguish MCC patients with tumor burden or without, correlate with MCC tumor stage, and serve as valuable marker for MCC monitoring. Next, we explored underline mechanism of transcriptional regulation of highly expressed miR-375 in MCCs. atonal BHLH transcription factor 1 (ATOH1) is considered as deduced miR-375 regulator. High expression of ATOH1 is confirmed in cMCC cell lines and MCC tissues and exhibits similar expression pattern as miR-375. We demonstrate that ATOH1 is the inducer of miR-375 in MCC cells and fibroblasts via knockdown and overexpression experiments. Interestingly, ectopic expression of Merkel cell polyomavirus large T antigens (MCPyV LTs) and ATOH1 induce similar cell morphologic changes, and ATOH1 and miR-375 expression are both induced by MCPyV LTs. Furthermore, we examined the role of miR-375 in MCC cells via knockdown experiments using antagomirs via nucleofection. Nearly complete miR-375 knockdown in cMCC cell lines did neither change cell viability nor cell morphology. Hippo and epithelial–mesenchymal transition (EMT) related oncogenic signaling pathways, which are predicted as regulated by miR-375 in MCC cells, was only slightly altered. These observations render miR-375 unlikely as an intracellular oncogene in MCC cells. At last, we scrutinized the function of miR-375 in intercellular signaling, focusing on cancer associated fibroblasts (CAF) polarization. Exosomal MCC-derived miR-375 is transferred to fibroblasts causing their polarization towards CAF phenotype including increased expression of alpha-smooth muscle actin (α-SMA), C-X-C motif chemokine ligand 2 (CXCL2) and interleukin 1 beta (IL-1β). Coculture induced fibroblast polarization is inhibited by miR-375 antagomirs or mimicked by ectopic miR-375 expression via targeting recombination signal binding protein for immunoglobulin kappa J region (RBPJ) and p53. A series of in situ observations from MCC tumor samples are consistent with our hypothesis. Thus, we provide several lines of evidence that miR-375 expression in MCC generates a protumorigenic microenvironment by inducing fibroblast polarization. Taken together, in this thesis, we not only demonstrate that miR-375 serves as a circulating serum marker for MCC, but also explore its transcriptional mechanism and functional role in MCC cells. More importantly, we demonstrate exosomal miR-375 derived from MCC cells polarizes fibroblasts into CAF phenotype.