Abstract 1358: Promoter hypermethylation of KISS1R, KiSS1, SEPT9 and CSAD as a prognostic biomarker panel to assess the metastatic potential of muscle invasive bladder tumors
Authors: Beatrice Stubendorff, Kerstin Wilhelm, Kathleen Posselt, James Catto, Arndt Harmann, Susanne Fuessel, Vladimir Novotny, Mieczyslaw Gajda, Heiko Wunderlich, Marc-Oliver Grimm, Kerstin Junker
Journal: Cancer Research (2014)

Abstract Introduction: Adjuvant chemotherapy improves the outcome of bladder cancer patients at risk for the development of metastases. However, there are no parameters available that allow an individual risk assessment. The aim of this project is to identify a specific DNA methylation pattern that provides a reliable tool for prognosis assessment of muscle invasive bladder tumors and to understand metastasis associated processes regulated by DNA methylation. Materials &amp; methods: Based on our previous microarray analysis we validated differentially methylated genes in a larger cohort of primary bladder tumors (n=100) in correlation to metastasis. Candidate promoter methylation and gene expression were assessed by quantitative analysis of DNA methylation using real-time PCR and qRT-PCR. Protein expression was determined by immunohistochemical analysis for KiSS1 and KISS1R on tissue microarrays from 2 independent patient cohorts comprising 290 muscle-invasive bladder carcinomas. For analyzing the gene function in tumor cell behavior, gene expression was knocked down by siRNA transfection of bladder cancer cells. Changes in proliferation and migration were measured using real-time cell monitoring (xCELLigence, Roche). Invasion assay was performed using matrigel coated cell culture inserts. Results: Methylation differences were confirmed for KISS1R, KiSS1, SEPT9 (p&amp;lt;0.05) and CSAD (p&amp;lt;0.01). mRNA expression differences between patient groups were detected for KISS1R and SEPT9v3. Protein expression of KISS1R was significantly reduced in correlation to positive lymph nodes (p&amp;lt;0.001) and distant metastases (p&amp;lt;0.05). Knock down of SEPT9v3 resulted in increased cell proliferation (77%), migration (15%) and invasion (39%) compared to negative control. Conclusion: Muscle-invasive bladder tumors show significant differences in the DNA methylation pattern in correlation to the metastatic potential. KISS1R is characterized by strongest association with the metastatic risk of primary tumors on methylation as well as RNA and protein expression level. Reduced expression of SEPT9v3 seems to induce cell proliferation, migration and invasion. The combination of candidate genes KISS1R, KiSS1, CSAD and SEPT9 could serve as prognostic marker panel for determination of the metastatic risk of primary bladder tumors. Citation Format: Beatrice Stubendorff, Kerstin Wilhelm, Kathleen Posselt, James Catto, Arndt Harmann, Susanne Fuessel, Vladimir Novotny, Mieczyslaw Gajda, Heiko Wunderlich, Marc-Oliver Grimm, Kerstin Junker. Promoter hypermethylation of KISS1R, KiSS1, SEPT9 and CSAD as a prognostic biomarker panel to assess the metastatic potential of muscle invasive bladder tumors. [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 1358. doi:10.1158/1538-7445.AM2014-1358