DNA hypermethylation in papillary renal cell carcinoma

Epigenetic alterations play an essential role during carcinogenesis, and earlier studies suggested that global histone modification levels are predictive for patients' outcome in various tumour entities (e.g. prostate, lung, breast and kidney cancer). So far, only smaller studies on histone H3K9 acetylation in patients with papillary urothelial neoplasm of low malignant potential (PUNLMP) and histone H2AX phosphorylation in non-muscle invasive bladder cancer exist; both studies indicate prognostic relevance for bladder cancer. We demonstrate that histone methylation levels (H3K4, H4K20) decrease from normal tissue over non-muscle invasive and muscle invasive bladder cancer to bladder cancer metastases. Histone modifications are correlated to advanced pathological stage in non-muscle and muscle invasive bladder cancer. Furthermore, H4K20me3 appeared to be predictive for bladder cancer specific survival in patients with muscle-invasive bladder cancer undergoing radical cystectomy. OBJECTIVE To investigate the pattern of DNA CpG island hypermethylation in papillary renal cell carcinoma (pRCC). MATERIAL AND METHODS DNA from pRCC (n = 32) and adjacent normal tissue (n = 15) was isolated. A quantitative methylation-specific PCR was performed to analyse the methylation pattern at APC (actin beta), CDH1 (E-cadherin), GSTP1 (glutathione S-transferase pi 1), RASSF1A (Ras association domain family member 1A) and TIMP3 (TIMP metallopeptidase inhibitor 3); a sequence of ACTB without CpG was used to normalize for DNA input and to calculate the relative amount of methylated DNA (normalized index of methylation, NIM). RESULTS RASSF1A hypermethylation was observed in most pRCC and normal samples (100 vs 94.4%), but the median NIM was significantly higher in pRCC samples (2.11 vs 0.61; P < 0.001). RASSF1A hypermethylation allowed discrimination of pRCC and normal tissue with a sensitivity of 87.5% and a specificity of 73.3% as determined via receiver operator characteristic analysis (area under curve = 0.814). Hypermethylation at APC (3.0 vs 6.7%), CDH1 (15.6 vs 0%), GSTP1 (21.9 vs 6.7%) and TIMP3 (6.3 vs 0%) was infrequent in pRCC and normal tissue. CDH1 was significantly correlated with pathological stage (P = 0.015), and patients with methylated CDH1 methylation showed a trend towards shorter recurrence-free survival (log-rank P = 0.057). The number of methylated gene sites was correlated with pathological stage (P = 0.007) and lymph node metastasis (P = 0.008). CONCLUSIONS DNA hypermethylation at RASSF1A is common in pRCC tissue irrespective of the histological subtype, but also frequently seen at lower levels in normal adjacent tissue. Aberrant hypermethylation could be a prognostic marker for pRCC.