关键词: Cosmc, T-合酶, 结肠癌, O-糖基化, 外泌体, miRNAs

Abstract: Objective  To explore the effects of aberrant O-glycosylation modifications induced by the knockout of Cosmc or T-synthase genes on the expression profiles of miRNAs in exosomes derived from colorectal cancer cells and to reveal  the molecular mechanisms of O-glycosylation in the development of colorectal cancer and  identify potential biomarkers  for early diagnosis and treatment. Methods This research specifically targets the Cosmc or T-synthase genes in the human colorectal cancer cell line HCT116 to create stable cell lines exhibiting abnormal O-glycosylation with CRISPR/Cas-9 gene editing technology. Exosomes originating from these colorectal cancer cells were isolated and authenticated. A microarray chip equipped with primer sequences for 16 miRNAs closely associated with colorectal cancer was employed to assess the differential expression of miRNAs within these exosomes with fluorescent quantitative polymerase chain reaction (PCR). And then, a cohort of miRNAs that exhibited significant and consistent changes in expression levels across the exosomes from both cell lines was selected. These miRNAs were further validated independently with traditional fluorescent quantitative PCR. Subsequently, data from  The Cancer Genome Atlas Program（TCGA） database containing patient information on colorectal cancer was harnessed. Employing R programming language, Gene Set Enrichment Analysis (GSEA) was conducted on the upregulated miRNA to investigate the downstream pathways significantly impacted and the malignant biological behaviors they may influence. Results  The absence of either Cosmc or T-synthase genes results in the dysregulation of O-glycosylation in colorectal cancer cells, leading to the exposure of Tn antigens. This, in turn, affects the expression levels of specific miRNAs in exosomes derived from these cells. Specifically, the expression of hsa-miR-125b-1-3p was downregulated, while that of hsa-miR-218-5p was upregulated. Notably, hsa-miR-218-5p were found to be closely associated with the epithelial-mesenchymal transition (EMT) process in tumor cells, which is a key mechanism in cancer progression. Conclusion  It elucidates that the aberrant O-glycosylation mediated by the knockout of Cosmc or T-synthase genes significantly influences the expression of certain miRNAs in exosomes from colorectal cancer cells, potentially affect the EMT process in colorectal cancer and thereby promoting distant metastasis. Given the inherent stability and detectability advantages of colorectal cancer-derived exosomes, the altered expression levels of miRNAs within these exosomes may serve as indicators of the stated of abnormal O-glycosylation  in colorectal cancer. These findings suggest that exosomal miRNAs have potential as biomarkers for monitoring disease progression and therapeutic efficacy. Consequently, this could pave the way for more personalized diagnostic and treatment strategies tailored to individual colorectal cancer patients, enhancing the precision and effectiveness of clinical management.

Key words: Cosmc, T-synthase, colon cancer, O-glycosylation, exosome, miRNAs