Dottorato in MORFOGENESI E INGEGNERIA TISSUTALE

Titolo della tesi: FULL-LENGTH DACH1 ISOFORM PLAYS AN ONCOGENIC ROLE AND PROMOTES RADIORESISTANCE IN PROSTATE CANCER

Human Dachshund homologue 1 (DACH1) is a key component of the Retinal Determination Gene Network (RDGN), originally identified in Drosophila melanogaster eye specification. Evidence has shown that tumors frequently display aberrant expression of RDGN genes, but currently the role of DACH1 in human cancers remains still controversial. Moreover, the specific functional roles of DACH1 splicing variants identified in human tissues have not been explored so far. Therefore, the aim of this study was to investigate DACH1 functional expression in prostate cancer (PCa) and its role in radiotherapy, focusing on the 100-kDa full-length isoform. In the present work we studied, by immunohistochemical staining, the expression of DACH1 in normal and tumor prostate from 30 patients and found both nuclear and cytoplasmic expression of DACH1 only in tumor glands, whereas in normal glands cytoplasmic DACH1 has not been observed. Moreover, by immunoblotting, we demonstrated that normal and tumor glands differed in DACH1 splicing variants expression. Flow cytometry analysis showed that normal human prostate cell line RWPE-1 over-expressed DACH1 compared to prostate cancer PC3 cells, though Western Blot analysis revealed that RWPE-1 cells strongly expressed the ~52/57 kDa protein, while PC3 cells expressed only the 100 kDa full-length protein. The stable over-expression of 100-kDa DACH1 isoform in PC3 cell line induced increased aggressiveness features. Interestingly, full-length DACH1 promoted radioresistance in PC3 cells, suggesting a pro-oncogenic role of this specific DACH1 isoform. Confirming this role, when we transfected full-length DACH1 isoform in normal RWPE-1 cells, we observed an increased cell proliferation, AKT-pathway activation and formation of spheroids in anchorage-independent culture condition. Taken together, these findings offer a comprehensive molecular model and highlight the oncogenic role of full-length DACH1 isoform in PCa, suggesting that it could represent a novel diagnostic marker in PCa patients.