Abstract 5014: Differences in DNA methylation pattern of primary bladder tumours correlate with their metastatic potential
Authors: Beatrice Stubendorff, Ewa Dudziec, James Catto, Jimsgene Sanjmyatav, Mieczyslaw Gaida, Heiko Wunderlich, Marc-Oliver Grimm, Kerstin Junker
Journal: Cancer Research (2012)

Abstract Introduction: The prognosis for patients with metastatic bladder cancer is poor with a 5-year survival rate of only 6%. Early prediction of the metastatic risk in primary tumours can lead to improvements in prognosis and therapy. Currently, there are no parameters available that enable an individual risk assessment for patients with metastatic bladder cancer. Therefore, identification of new reliable prognostic markers for early prediction of tumour spread is required. The aim of this project is to determine whether changes in DNA methylation correlate with the metastasis risk and to identify a specific DNA methylation pattern that provides a reliable tool for prognosis assessment. Materials &amp; methods: Genomic DNA was isolated from 23 invasive bladder tumour tissues with and without lymph node metastases. For enrichment of methylated fragments genomic DNA was incubated with 5-Methylcytosin antibody. Input and IP were labelled with Cy3 and Cy5. Labelled DNA was hybridizes onto CpG island microarrays. Cluster analysis enabled selection of differentially methylated candidate genes. The actual methylation state was confirmed by cleavage of genomic DNA with methylation dependent restriction enzyme (McrBc) followed by quantitative PCR. Results: In total, 201 promoter CpG islands show highly significant differences in DNA methylation between metastasised and non-metastasised primary bladder tumours (p&amp;lt;0,01). In metastasised tumours 74 CpG islands are hypermethylated and 127 CpG islands are hypomethylated. Data base research for identification of candidate genes resulted in 61 hypermethylated and 58 hypomethylated genes that can clearly assigned to the identified promoter associated CpG islands. Validation of methylation level of candidate gene RASSF1 resulted in higher methylation in metastatic tumours (mean methylation: 38%) than in non-metastatic tumours (mean methylation: 26%). RASSF1 methylation shows strong correlation to T stage (r=0,65). Methylation of candidate gene KISS1R shows significant differences between patient groups (p=0,001). Mean methylation in metastatic tumours is 48% and in non-metastatic tumours 22%. Conclusion: Primary bladder tumours show significant differences in their DNA methylation pattern in correlation to their metastatic potential. KISS1R methylation is strongly associated with the metastatic risk of primary tumours. Further analysis will demonstrate the correlation between changes in DNA methyaltion and expression of corresponding genes. The impact of DNA methylation on tumour behaviour shall be determined in functional analysis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5014. doi:1538-7445.AM2012-5014