OSTEOPONTIN A POTENTIAL PROMOTER OF COLITIS ASSOCIATED CARCINOMA IN CHRONIC COLITIS – A MOLECULAR STUDY IN HUMANS
Maximilian Sehn*1, Danielle Cardoso Da Silva2, Manna Subhakankha2, Sefer Elezkurtaj3, Michael Hummel3, Britta Siegmund2, Katharina Beyer1, Michael Schumann2
1Department of General and Visceral Surgery, Berlin Institute of Health, Charité Universitätsmedizin Berlin, Freie Universität Berlin, Humboldt-Universität Zu Berlin, Berlin, Berlin, Germany; 2ChariteCentrum 13 Innere Medizin mit Gastroenterologie und Nephrologie, Berlin, Berlin, Germany; 3Institute of Pathology, Berlin Institute of Health, Charité Universitätsmedizin Berlin, Freie Universität Berlin, Humboldt-Universität Zu Berlin, Berlin, Germany, Berlin, Berlin, Germany
Introduction:
Inflammatory bowel diseases (IBD) carry a high disease burden and are frequently diagnosed in the western world. Both diseases expose patients to an increased risk for the development of colitis-associated carcinoma (CAC). Research till date provided evidence supporting a different carcinogenesis for CAC as opposed to sporadic colonic cancer. However, as most of this research relies on murine models of CAC and full transfer to the human setting remains doubtful. Osteopontin (OPN) is a member of the secreted phosphoprotein family. It previously has been described as an impactor on lymphoid cell differentiation and the development of intestinal fibrosis in IBD. A potential connection of OPN to the CAC carcinogenesis has, however, not been reported until today.
Material and Methods:
We selected 50 patients in five groups, specifically ulcerative Colitis (UC), Crohn's disease (CD), healthy control (CTRL), UC-CAC and CD-CAC. RNA was isolated from microdissections of surgical colon specimens. Amplification-free, quantitative RNA expression analysis of a defined set of 624 immunology, barrier, onco- and polarity genes was performed. Primary data analysis, differential expression and statistical analysis were done using the Benjamini-Yekutieli correction for multiple testing and the student's t-test for normally distributed data. Further analysis included grouping of expression patterns (heatmaps) using R and pathway analysis. Results were validated using cell culture and murine organoid experiments, phospho-blots and immunofluorescence. Lastly, a possible OPN mediated tumerogenic signal to the mitochondrial respiratory chain was measured by a specific assay.
Results:
OPN was identified to be the most upregulated gene in CAC tissue in both comparisons, CD to CD-CAC and UC to UC-CAC (p-adj=0.000016). Pathway analysis predicted an OPN-mediated stimulation of the NFkB and ERK pathways. The OPN-mediated activation of ERK was reproduced in cell culture experiments with HT29/B6 cells. The NFkB pathway was not activated in an OPN-dependent fashion. Mouse colonic organoid experiments as well as oxygen consumption assays showed a possible carcinogenic role of OPN via induction of epithelial-to-mesenchymal transformation (EMT).
Conclusions:
We present compelling evidence for a potential role of OPN in the colitis-associated carcinogenesis. Involved pathways include ERK and EMT. Thus, OPN and its effectors could serve as possible diagnostic targets for an early CAC detection or identification of patients at risk. OPN signaling could also be a potential therapeutic target to improve survival for this frequently treatment-refractory tumor.
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