Abstract
Cervical cancer is caused by a persistent infection with high-risk types of HPV and an accumulation of (epi)genetic alterations in host cell. Acquisition of anchorage-independent growth represents a critical hallmark during HPV-induced carcinogenesis, thereby yielding the most valuable biomarkers for early diagnosis and therapeutic targets. In a previous study, we found that miR-193a-3p and miR-193b-3p were involved in anchorage-independent growth. This study aimed to delineate the role of miR-193a/b-3p in HPV-induced carcinogenesis and to identify their target genes related to anchorage-independent growth. Cell viability and colony formation were assessed in SiHa cancer cells and HPV-16 and -18 immortalized keratinocytes upon miR-193a/b-3p overexpression. Both miRNAs reduced cell growth of all three cell lines in low-attachment conditions and showed a minor effect in adherent conditions. Online target predicting programs and publicly available expression data were used to find ca ndidate mRNAs targets of miR-193a/b-3p. Seven targets showed reduced mRNA expression upon miR-193a/b-3p overexpression. For 3 targets Western blot analysis was also performed, all showing a reduced protein expression. A direct interaction was confirmed using luciferase assays for 6 genes: LAMC1, PTK2, STMN1, KRAS, SOS2, and PPP2R5C, which are PIK3/AKT regulators. All 6 targets were overexpressed in cervical cancers and/or precursor lesions. Together with an oberserved downregulation of phosphorylated-AKT upon miR-193a/b-3p overexpression, this underlines the biological relevance of miR-193a/b-3p downregulation during HPV-induced cervical carcinogenesis.
In conclusion, downregulation of miR-193a-3p and miR-193b-3p is functionally involved in the acquisition of HPV-induced anchorage independence by targeting regulators of the PIK3/AKT pathway.
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