Methylation profiling of urothelial carcinoma in bladder biopsy and urine

Objective: To test DNA methylation profiling in detection of urothethial carcinoma in urine. Study Design: Thirty-three bladder specimens were analyzed for the DNA methylation status of p14^ARF, p16^INK4a, RASSF1, APC, GSTP, E-Cad and CyclinD2 genes to determine if there is a difference in gene methylation between benign and malignant cases. Urine samples were analyzed in a feasibility study. Finally, methylation profiles of urine samples were obtained and compared with follow-up biopsy diagnoses. Results: We found methylated genes in 18% benign, 37% urothelial carcinoma in situ and 93% infiltrating urothelial carcinoma cases (p = 0.001). Methylation profiles from the 18 urine samples revealed a significantly higher prevalence of methylated genes in carcinoma cases than benign cases (100% vs. 50%, p = 0.025). We analyzed methylation profiles in 37 cytologically atypical urine samples with malignant or benign diagnosis on surgical follow-up and found that only APC (55% in malignant vs. 0% in benign, p = 0.025) and CyclinD2 were differentially methylated (35% in malignant vs. 0% in benign, p = 0.2) while p14^ARF, p16^INK4a, RASSF1, GSTP and E-Cad had similar methylation profiles. Conclusion: These results suggest that methylation of p14^ARF, p16^INK4a, RASSFl, GSTP and E-Cad genes may not accurately identify carcinoma, but methylated APC and CyclinD2 might be useful biomarkers for urothelial carcinoma in urine.