Background: Quantitation of prostate specific antigen (PSA) and digital rectal examination remain the primary tools for detection of prostate cancer in the clinical setting. The limited specificity of PSA has dictated the need for alternative diagnostic methods. DIA was developed to identify long DNA fragments isolated from stool, indicative of DNA released from cancer cells that had escaped apoptosis. In this study we have evaluated DIA as a screening tool for prostate cancer, using free-DNA isolated from two-spin plasma preparations of patients undergoing prostatectomy.
Methods: Prior to undergoing prostatectomy, blood samples were collected from consented patients (n=127) and processed as two-spin plasma preparations. Two control groups in this study included: negative prostate biopsy patients (group 1, n=23) and post-prostatectomy patients (group 2, n=25). In all patients, free-DNA was isolated from plasma preparations over a Qiagen column, followed by incubation in a hybrid-capture technique to isolate specific DNA fragments. Primer sets for targeted sequences were multiplexed for the detection of 200bp, 1.3kb, 1.8kb and 2.4kb DNA fragments at a single locus. Four separate genetic loci were interrogated for the presence of the DNA fragment lengths using real-time PCR. All data points were considered significant and a cut-off was determined for each individual fragment length.
Results: Of the 127 patients with prostate cancer, 90 patients (70.9%) were determined to have a DNA integrity score of 7 or greater (out of 10 fragment lengths) whereas group 1 had 9 patients (39.1%) and group 2 had 2 patients (8%) with significant scores. Of the patients with a negative age-adjusted PSA and prostate cancer, 15 of 25 (60%) were positive in DIA. Sensitivity for the DIA assay was 70.9% with a specificity of 60.9% (group 1) and 92% (group 2). There was no statistically significant correlation between a positive DIA score and Gleason grade or organ confined disease.
Conclusion: DIA detected 70.9% of prostate cancers and 60% of those with a negative age-adjusted PSA while maintaining overall specificity. The variability in specificity between the two control groups may be explained by the established 19-30% detection of prostate cancer on subsequent biopsies. We present the first generation of a promising new plasma based assay which has the potential to complement PSA in the early identification of prostate cancer. Further investigation is needed to determine the utility of DIA as an alternative to prostate biopsy and as a marker to monitor patients with an elevated PSA.

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