Methylation profiling of urothelial carcinoma in bladder biopsy and urine.
OBJECTIVE
To test DNA methylation profiling in detection of urothelial carcinoma in urine.


METHODS
Thirty-three bladder specimens were analyzed for the DNA p16INK4a, 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 andfound 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 p14ARF, p16INK4a, RASSF1, GSTP and E-Cad had similar methylation profiles.


CONCLUSIONS
These results suggest that methylation of p14ARF, p16INK4a, RASSF1, GSTP and E-Cad genes may not accurately identify carcinoma, but methylated APC and CyclinD2 might be useful biomarkers for urothelial carcinoma in urine.
Pu
Robert T
RT
Department of Pathology, Division of Cytopathology, Johns Hopkins Medical Institutions, Baltimore, Maryland, USA. robertpu@umich.edu
Laitala
Lauren E
LE
Clark
Douglas P
DP
D000368:Aged; D064173:Anaphase-Promoting Complex-Cyclosome; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002277:Carcinoma / Q000175:diagnosis* / Q000235:genetics* / Q000652:urine; D056742:Cyclin D2; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics / Q000378:metabolism; D016213:Cyclins / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000235:genetics / Q000652:urine; D016172:DNA Fingerprinting / Q000379:methods* / Q000639:trends; D019175:DNA Methylation; D003951:Diagnostic Errors / Q000517:prevention & control; D005188:False Negative Reactions; D005240:Feasibility Studies; D005819:Genetic Markers / Q000235:genetics; D006801:Humans; D008875:Middle Aged; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D043743:Ubiquitin-Protein Ligase Complexes / Q000235:genetics / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000175:diagnosis* / Q000235:genetics* / Q000652:urine; D019459:Urothelium / Q000378:metabolism / Q000473:pathology*
C080336:CCND2 protein, human; D015820:Cadherins; D056742:Cyclin D2; D019941:Cyclin-Dependent Kinase Inhibitor p16; D016213:Cyclins; D005819:Genetic Markers; C411762:RASSF1 protein, human; D025803:Tumor Suppressor Protein p14ARF; D025521:Tumor Suppressor Proteins; D004247:DNA; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome