Abstract 976: Junctional adhesion molecule-A (JAM-A) expression is downmodulated by miR-21 during colorectal cancer progression

Abstract

Abstract Introduction: MicroRNAs are small non-coding-RNAs that control cell homeostasis and are deregulated in human colorectal cancer (CRC). MiR-21 up-regulation is frequent in CRC and represents a driver for tumor initiation and progression. Junctional Adhesion Molecule A (JAM-A) is a tight junction protein involved in regulation of para-cellular permeability in epithelial and endothelial cells. JAM-A forms homodimers and activates signalling pathways regulating cell adhesion, polarity and proliferation. For this reason, monoclonal antibodies against JAM-A are currently tested in the preclinical setting as potential targeted therapies. JAM-A loss appears to be an early event in CRC initiation but very little is known about the role of JAM-A in human sporadic CRC. The aim of our study was defining the pattern of expression of JAM-A in human CRC and to the define the whether its deregulation is a consequence of aberrant microRNA expression. Methods: JAM-A expression was analyzed by immunohistochemistry in 1254 cases of human sporadic CRC. MiR-21 and JAM-A expression was analyzed in human CRC cells [RKO, HT29, HCT116, SW620, HCT15, SW480, SW837, SW48, DLD1 (both Wild-Type and Knock-Out for miR-21)] by Real-Time-PCR and Western-Blotting. MiR-21 silencing and over-expression were performed using Exiqon Locked Nucleic Acids and Ambion Pre-miR respectively. MiR-21 binding site in JAM-A 3’UTR was predicted using bio-informatics tools (RNA Hybrid) and a 236bp area encompassing the predicted seed region was cloned downstream of the Firefly Luciferase gene in PGL3 vector for Luciferase Reporter Assay. Results: JAM-A down-regulation was observed in cancer compared to normal adjacent tissues. JAM-A was progressively down-regulated in the progression from normal epithelium, dysplasia, intraepithelial and invasive cancer. Poorly differentiated cancers showed total loss of JAM-A. Shifted staining from apical to baso-lateral compartment was observed in 25% of all cancers. Silencing and re-expression of miR-21 in CRC cell lines resulted in increase and down-regulation of JAM-A protein expression respectively. Luciferase reporter assay experiments were able to define the potential seed region by which miR-21 interacts with JAM-A 3′UTR. Conclusion: Our data suggest that JAM-A expression is deregulated during human colorectal cancer progression as a consequence of miR-21 over-expression. These results highlight how miR-21 could potentially exert its’ actions on para-cellular permeability and cell polarity acting on junctional adhesion molecules. Understanding the mechanisms of JAM-A loss or re-localization may help to stratify patient's treatment and identify patients who might benefit from anti-JAM-A monoclonal antibodies. Citation Format: Andrea Lampis, Claudio Murgia, Viola Paulus-Hock, Matteo Fassan, Joanne Edwards, Paul Horgan, Owen Sansom, Luigi Terracciano, Michael Karin, Carlo Croce, Chiara Braconi, Nicola Valeri. Junctional adhesion molecule-A (JAM-A) expression is downmodulated by miR-21 during colorectal cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 976. doi:10.1158/1538-7445.AM2014-976