Jonathan Baden

Development of a Multiplexed Urine Assay for Prostate Cancer Diagnosis

BACKGROUND Several studies have demonstrated the value of DNA methylation in urine-based assays for prostate cancer diagnosis. However, a multicenter validation with a clinical prototype has not been published. METHODS We developed a multiplexed, quantitative methylation-specific polymerase chain reaction (MSP) assay consisting of 3 methylation markers, GSTP1, RARB, and APC, and an endogenous control, ACTB, in a closed-tube, homogeneous assay format. We tested this format with urine samples collected after digital rectal examination from 234 patients with prostate-specific antigen (PSA) concentrations > or =2.5 microg/L in 2 independent patient cohorts from 9 clinical sites. RESULTS In the first cohort of 121 patients, we demonstrated 55% sensitivity and 80% specificity, with area under the curve (AUC) 0.69. In the second independent cohort of 113 patients, we found a comparable sensitivity of 53% and specificity of 76% (AUC 0.65). In the first cohort, as well as in a combined cohort, the MSP assay in conjunction with total PSA, digital rectal examination status, and age improved the AUC without MSP, although the difference was not statistically significant. Importantly, the GSTP1 cycle threshold value demonstrated a good correlation (R = 0.84) with the number of cores found to contain prostate cancer or premalignant lesions on biopsy. Moreover, samples that exhibited methylation for either GSTP1 or RARB typically contained higher tumor volumes at prostatectomy than those samples that did not exhibit methylation. CONCLUSIONS These data confirm and extend previously reported studies and demonstrate the performance of a clinical prototype assay that should aid urologists in identifying men who should undergo biopsy.

Multicenter Evaluation of an Investigational Prostate Cancer Methylation Assay

PURPOSE Prostate specific antigen tests have low specificity, which frequently results in unnecessary biopsy and typically limits screening to patients with prostate specific antigen greater than 4.0 ng/ml. We evaluated an investigational prostate cancer methylation specific polymerase chain reaction assay that detects aberrant methylation in 3 markers (GSTP1, RARbeta2 and APC) that indicate the presence of prostate cancer. MATERIALS AND METHODS The assay was evaluated in 337 post-digital rectal examination urine samples (178 cancer and 159 noncancer) collected prospectively at a total of 9 clinical sites. Samples were processed wholly or after division into equal portions. Subject prostate specific antigen was 2.0 to 10.0 ng/ml. All subjects underwent transrectal ultrasound guided needle biopsy with 6 or greater cores sampled. Detection of 1 or greater markers indicated positivity. RESULTS Methylation specific polymerase chain reaction assay performance was better in whole than in divided urine cohorts (p = 0.035). Assay AUC was 0.72 in the whole urine cohort and 0.67 in the combined population. These values were higher than those of prostate specific antigen alone using 4.0 ng/ml as the cutoff (p = 0.00 and 0.01, respectively). Moreover, the assay together with the Prostate Cancer Prevention Trial risk calculator or a standard nomogram significantly improved AUC in the whole urine cohort and the combined population vs predictive algorithms alone (p <0.05). Assay positive predictive value was 54% in whole urine cohort with prostate specific antigen 2.0 to 4.0 ng/ml and negative predictive value was 87% with prostate specific antigen 4.1 to 10.0 ng/ml. Assay positive predictive value was higher in subjects with all 3 methylation markers positive. CONCLUSIONS These data demonstrate that this investigational assay used in conjunction with current screening algorithms may potentially add value to the biopsy decision making process.

Predicting prostate biopsy result in men with prostate specific antigen 2.0 to 10.0 ng/ml using an investigational prostate cancer methylation assay.

PURPOSE The inadequacies of prostate specific antigen testing have created a need for novel markers for prostate cancer screening. The investigational ProCaM™ prostate cancer methylation assay detects aberrant methylation of DNA in cells associated with prostate cancer. We describe a large, prospective, multicenter study done to verify the performance of this assay. MATERIALS AND METHODS The assay is designed to detect epigenetic modifications in the 3 markers GSTP1, RARβ2 and APC, which are indicative of prostate cancer. A total of 232 men with cancer and 283 without cancer from 18 clinical sites were evaluated by trained operators at central testing laboratories. Study inclusion criteria were age 40 to 75 years, total prostate specific antigen between 2.0 and 10.0 ng/ml, and a digital rectal examination result. All participants signed an informed consent form and underwent transrectal ultrasound guided needle biopsy with 10 or more cores. RESULTS Assay sensitivity was 60%, specificity was 80% and the informative rate was 97%. Assay predictive accuracy was higher than that of age, digital rectal examination, family history, prostate specific antigen, prior negative biopsy and prostate volume (AUC 0.73 vs 0.52 to 0.66, p <0.038). Risk factors plus the assay improved overall predictive power (AUC 0.79, p = 0.001). A man with a positive prostate cancer methylation result was 7.7 times more likely to have high grade cancer. CONCLUSIONS The prostate cancer methylation assay correlated with positive biopsy and with Gleason score. This assay has the potential to add value to the biopsy decision making process by improving current prostate cancer screening algorithms to more accurately identify men with prostate cancer.

DETECTING PROSTATE CANCER

BACKGROUND When prostate cancer is suspected, the prostate gland is biopsied with the aid of transrectal ultrasound (TRUS). The sensitivity of prostatic biopsy is about 50%. The fusion of magnetic resonance imaging (MRI) data with TRUS enables the targeted biopsy of suspicious areas. We studied whether this improves the detection of prostate cancer. METHODS 168 men with suspected prostate cancer underwent prostate MRI after a previous negative biopsy. Suspicious lesions were assessed with the classification of the Prostate Imaging Reporting and Data System and biopsied in targeted fashion with the aid of fused MRI and TRUS. At the same sitting, a systematic biopsy with at least 12 biopsy cores was performed. RESULTS Prostate cancer was detected in 71 patients (42.3%; 95% CI, 35.05-49.82). The detection rate of fusion-assisted targeted biopsy was 19% (95% CI, 13.83-25.65), compared to 37.5% (95% CI, 30.54-45.02) with systematic biopsy. Clinically significant cancer was more commonly revealed by targeted biopsy (84.4%; 95% CI, 68.25-93.14) than by systematic biopsy (65.1%; 95% CI, 52.75-75.67). In 7 patients with normal MRI findings, cancer was detected by systematic biopsy alone. Compared to systematic biopsy, targeted biopsy had a higher overall detection rate (16.5% vs. 6.3%), a higher rate of infiltration per core (30% vs. 10%), and a higher rate of detection of poorly differentiated carcinoma (18.5% vs. 3%). Patients with negative biopsies did not undergo any further observation. CONCLUSION MRI/TRUS fusion-assisted targeted biopsy improves the detection rate of prostate cancer after a previous negative biopsy. Targeted biopsy is more likely to reveal clinically significant cancer than systematic biopsy; nevertheless, systematic biopsy should still be performed, even if the MRI findings are negative.