Elucidating the role of the polypeptide N-acetylgalactosaminyltransferase 3 (GALNT3) and its closest homolog-GALNT6 in mediating aberrant O-glycosylation associated with ovarian cancer dissemination. Elucidating the role of the polypeptide N-acetylgalactosaminyltransferase 3 (GALNT3) and its closest homolog-GALNT6 in mediating aberrant O-glycosylation associated with ovarian cancer dissemination

Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy, thus understanding molecular changes associated with EOC progression could lead to the identification of essential therapeutic targets. Glycosylation is a post-translational modification (PTM) that participates in major pathophysiology events, including tumorigenesis. Aberrant mucin-type or O-glycosylation in cancer is frequently attributed to altered expression of different N-acetylgalactosaminyltransferases (GalNAc-Ts). We have previously identified the N-acetylgalactosaminyltransferase 3 (GALNT3) gene as a potential EOC oncogene, possibly implicated in aberrant O-glycosylation in EOC cells. Literature reports are suggestive for genetic and functional redundancy between different GalNAc-Ts, and our previous data were indicative for an induction of the GALNT6 expression upon GALNT3 suppression in EOC cells. Here we show that GALNT3 gene knockout (KO) in the A2780s EOC cells leads to strong GALNT6 upregulation. Moreover, A2780s cell migration, invasion, proliferation and cell cycle analysis were significantly reduced in double GALNT3/T6 gene KO cells compared to single GALNT3 gene KO. Furthermore, MUC1 expression was downregulated in both GALNT3 KO and GALNT3/6 KO, with a more prominent decrease observed in the double KO clone. In vivo studies supported the observed functional redundancy imposed by GALNT6, as animals injected with double GALNT3/T6 KO EOC cell clones displayed a significant increase in survival rates compared to those injected with the control and the GALNT3 KO clones. Collectively our data strongly suggest for possible functional redundancy of the two GalNAc-Ts (GALNT3 and T6) in EOC, with the perspective to use both of these genes as EOC markers and/or therapeutic targets. Overall design: To better understand the molecular mechanisms of GALNT3 and GALNT6 genes action in epithelial ovarian cancer (EOC), we employed the Agilent Whole Human Genome microarrays, containing ~ 44,000 genes to identify global gene expression changes upon GALNT3 suppression and double GALNT3/6 suppression in A2780s EOC cells. We compared the gene expression of the selected CRISPR-Cas9-mediated GALNT3 -knockout clones (GAL3 KO-1 and GAL3 KO-2) and the double gene Knockout GALNT3/6 clones (GAL3/6 KO-1 and GAL3/6 KO-2) against the corresponding control (C) clones .