Detecting DNA methylation of the BCL2, CDKN2A and NID2 genes in urine using a nested methylation specific polymerase chain reaction assay to predict bladder cancer.
OBJECTIVE
Detection of methylated DNA has been shown to be a good biomarker for bladder cancer. Bladder cancer has the highest recurrence rate of any cancer and, as such, patients are regularly monitored using invasive diagnostic techniques. As urine is easily attainable, bladder cancer is an optimal cancer to detect using DNA methylation. DNA methylation is highly specific in cancer detection. However, it is difficult to detect because of the limited amount of DNA present in the urine of patients with bladder cancer. Therefore, an improved, sensitive and noninvasive diagnostic test is needed.


METHODS
We developed a highly specific and sensitive nested methylation specific polymerase chain reaction assay to detect the presence of bladder cancer in small volumes of patient urine. The genes assayed for DNA methylation are BCL2, CDKN2A and NID2. The regions surrounding the DNA methylation sites were amplified in a methylation independent first round polymerase chain reaction and the amplification product from the first polymerase chain reaction was used in a real-time methylation specific polymerase chain reaction. Urine samples were collected from patients receiving treatment at Wolfson Medical Center in Holon, Israel.


RESULTS
In a pilot clinical study using patient urine samples we were able to differentiate bladder cancer from other urogenital malignancies and nonmalignant conditions with a sensitivity of 80.9% and a specificity of 86.4%.


CONCLUSIONS
We developed a novel methylation specific polymerase chain reaction assay for the detection and monitoring of bladder cancer using DNA extracted from patient urine. The assay may also be combined with other diagnostic tests to improve accuracy.
Oncoveda Tumor Biology Center, Medical Diagnostic Laboratories LLC, a Division of Genesis Biotechnology Group, Hamilton, New Jersey, USA. mscher@mdlab.com
Michael B
MB
Scher
Michael B
MB
Elbaum
Yakov
Y
Mogilevkin
David W
DW
Hilbert
Jack H
JH
Mydlo
A Ami
AA
Sidi
Martin E
ME
Adelson
Eli
E
Mordechai
Jason P
JP
Trama
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000652:urine*; D002135:Calcium-Binding Proteins; D015815:Cell Adhesion Molecules / Q000235:genetics* / Q000652:urine; D019175:DNA Methylation; D005260:Female; D019254:Genes, bcl-2; D019942:Genes, p16; D006801:Humans; D008297:Male; D008875:Middle Aged; D010865:Pilot Projects; D016133:Polymerase Chain Reaction / Q000379:methods; D001749:Urinary Bladder Neoplasms / Q000175:diagnosis / Q000235:genetics* / Q000652:urine
D014408:Biomarkers, Tumor; D002135:Calcium-Binding Proteins; D015815:Cell Adhesion Molecules; C114729:NID2 protein, human