Background: PAX8 is a member of the paired box (Pax) multigene family of transcription factors, which are involved in the developmental and tissue-specific control of the expression of several genes in both vertebrates and invertebrates. Previously, several studies reported that PAX8 is expressed at high levels in specific types of tumors. In particular, PAX8 has been recently reported to be conspicuously expressed in human ovarian cancer, but the functional role of PAX8 in the carcinogenesis of this type of tumor has not been addressed. In this study, we investigated the contribution of PAX8 in ovarian cancer progression.Methods: Stable PAX8 depleted ovarian cancer cells were generated using short hairpin RNA (shRNA) constructs. PAX8 mRNA and protein were detected by RT-PCR, immunoblot and immunofluorescence. Cell proliferation, motility and invasion potential of PAX8 silenced cells were analyzed by means of growth curves, wound healing and Matrigel assays. In addition, PAX8 knockdown and control cells were injected into nude mice for xenograft tumorigenicity assays. Finally, qPCR was used to detect the expression levels of EMT markers in PAX8-overexpressing and control cells.Results: Here, we show that PAX8 plays a critical role in the migration, invasion and tumorigenic ability of ovarian cancer cells. Our results show that RNA interference-mediated knockdown of PAX8 expression in SKOV-3 ovarian cancer cells produces a significant reduction of cell proliferation, migration ability and invasion activity compared with control parental SKOV-3 cells. Moreover, PAX8 silencing strongly suppresses anchorage-independent growthin vitro. Notably, tumorigenesisin vivoin a nude mouse xenograft model is also significantly inhibited.Conclusions: Overall, our results indicate that PAX8 plays an important role in the tumorigenic phenotype of ovarian cancer cells and identifies PAX8 as a potential new target for the treatment of ovarian cancer.

High-Grade Serous Ovarian Carcinoma (HGSC) is the most incidental and lethal subtype of epithelial ovarian cancer (EOC) with a high mortality rate of nearly 65%. Recent findings aimed at understanding the pathogenesis of HGSC have attributed its principal source as the Fallopian Tube (FT). To further comprehend the exact mechanism of carcinogenesis, which is still less known, we performed a transcriptome analysis comparing FT and HGSC. Our study aims at exploring new players involved in the development of HGSC from FT, along with their signaling network, and we chose to focus on non-coding RNAs. Non-coding RNAs (ncRNAs) are increasingly observed to be the major regulators of several cellular processes and could have key functions as biological markers, as well as even a therapeutic approach. The most physiologically relevant and significantly dysregulated non-coding RNAs were identified bioinformatically. After analyzing the trend in HGSC and other cancers, MAGI2-AS3 was observed to be an important player in EOC. We assessed its tumor-suppressive role in EOC by means of various assays. Further, we mapped its signaling pathway using its role as a miRNA sponge to predict the miRNAs binding to MAGI2AS3 and showed it experimentally. We conclude that MAGI2-AS3 acts as a tumor suppressor in EOC, specifically in HGSC by sponging miR-15-5p, miR-374a-5p and miR-374b-5p, and altering downstream signaling of certain mRNAs through a ceRNA network.

Long non-coding RNAs (lncRNAs) are increasingly being identified as crucial regulators in pathologies like cancer. High-grade serous ovarian carcinoma (HGSC) is the most common subtype of ovarian cancer (OC), one of the most lethal gynecological malignancies. LncRNAs, especially in cancers such as HGSC, could play a valuable role in diagnosis and even therapy. From RNA-sequencing analysis performed between an OC cell line, SKOV3, and a Fallopian Tube (FT) cell line, FT194, an important long non-coding RNA, HAND2 Anti sense RNA 1 (HAND2-AS1), was observed to be significantly downregulated in OCs when compared to FT. Its downregulation in HGSC was validated in different datasets and in a panel of HGSC cell lines. Furthermore, this study shows that the downregulation of HAND2-AS1 is caused by promoter hypermethylation in HGSC and behaves as a tumor suppressor in HGSC cell lines. Since therapeutic relevance is of key importance in HGSC research, for the first time, HAND2-AS1 upregulation was demonstrated to be one of the mechanisms through which HDAC inhibitor Panobinostat could be used in a strategy to increase HGSC cells’ sensitivity to chemotherapeutic agents currently used in clinical trials. To unravel the mechanism by which HAND2-AS1 exerts its role, an in silico mRNA network was constructed using mRNAs whose expressions were positively and negatively correlated with this lncRNA in HGSC. Finally, a putative ceRNA network with possible miRNA targets of HAND2-AS1 and their mRNA targets was constructed, and the enriched Gene Ontology (GO) biological processes and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were identified.

Background: Ovarian cancer is the third most common cause of death among gynecologic malignancies worldwide. Understanding the biology and molecular pathogenesis of ovarian epithelial tumors is key to developing improved prognostic indicators and effective therapies. We aimed to determine the effects of PAX8 expression on the migrative, adhesive and survival capabilities of high-grade serous carcinoma cells. Methods: PAX8 depleted Fallopian tube secretory cells and ovarian cancer cells were generated using short interfering siRNA. Anoikis resistance, cell migration and adhesion properties of PAX8 silenced cells were analyzed by means of specific assays. Chromatin immunoprecipitation (ChIP) was carried out using a PAX8 polyclonal antibody to demonstrate that PAX8 is able to bind to the 5′-flanking region of the ITGB3 gene positively regulating its expression. Results: Here, we report that RNAi silencing of PAX8 sensitizes non-adherent cancer cells to anoikis and affects their tumorigenic properties. We show that PAX8 plays a critical role in migration and adhesion of both Fallopian tube secretory epithelial cells and ovarian cancer cells. Inhibition of PAX8 gene expression reduces the ability of ovarian cancer cells to migrate and adhere to the ECM and specifically to fibronectin and/or collagen substrates. Moreover, loss of PAX8 strongly reduces ITGB3 expression and consequently the correct expression of the αvβ3 heterodimer on the plasma membrane. Conclusions: Our results demonstrate that PAX8 modulates the interaction of tumor cells with the extracellular matrix (ECM). Notably, we also highlight a novel pathway downstream this transcription factor. Overall, PAX8 could be a potential therapeutic target for high-grade serous carcinoma.

The transcription factor Pax8 is expressed during thyroid development and is involved in the morphogenesis of the thyroid gland and maintenance of the differentiated phenotype. In particular, Pax8 has been shown to regulate genes that are considered markers of thyroid differentiation. Recently, the analysis of the gene expression profile of FRTL-5 differentiated thyroid cells after the silencing of Pax8 identified Wnt4 as a novel target. Like the other members of the Wnt family, Wnt4 has been implicated in several developmental processes including regulation of cell fate and patterning during embryogenesis. To date, the only evidence on Wnt4 in thyroid concerns its down-regulation necessary for the progression of thyroid epithelial tumors.