Prostate-specific mRNA detection in whole blood as an analytically validated prognostic biomarker for patients with castration-resistant prostate cancer (CRPC).

Abstract

45 Background: To estimate the association between molecular biomarkers and outcomes, robust assays are needed before qualification in prospective trials. Although tumor specific transcripts detected in blood by PCR have been associated with outcome, an analytically validated PCR platform enabling detection of CTC-associated transcripts in clinical settings remains to be identified. Here we explore detecting a panel of prostate specific transcripts in conjunction with conventional markers in CRPC. Methods: Blood was collected from 97 patients with progressive CRPC in PAXgene tubes (2.5 ml) for total RNA extraction. Five genes expressed in prostate but not in nucleated blood cells were analyzed by primer-specific RT-PCR. Fluidigm Dynamic Array platform was analytically validated for KLK3, KLK2, HOXB13, GHRL2 and FOXA1 assays. Each PCR-reaction was run in 6 replicates, detection thresholds for each gene were chosen by ROC analysis of an independent set of 56 CRPC patients versus 51 healthy volunteers, and results were reported as transcripts present or absent. CellSearch reported CTC number/7.5 ml of blood drawn into CellSave tube. Results: Prostate specific transcripts for KLK3, KLK2, HOXB13, GHRL2 and FOXA1were detected in 50 (52%), 39 (40%), 53 (55%), 28 (29%) and 47 (48%) of patients, respectively.Two or more genes were detected in 53/97 patients (55%, 95% CI 45-65%) of patients with progressive CRPC studied with a median survival time of 17 months (95% CI: 13.8, 23.5). 40/45 CRPC-patients with ≥5 CTC and 13/52 CRPC patients with <5 CTC had transcripts present in blood. CTC and the gene panel had a substantial level of agreement, kappa of 0.631 (95% CI: 0.479, 0.783). Transcript detection predicted overall survival in proportional hazards models that included LDH dichotomized at 250. The predictive accuracy (CPE) of PCR detection + LDH was 0.759 (SE 0.035), comparable to CTC-enumeration + LDH (0.771, SE 0.033). Conclusions: Detecting prostate specific mRNA in whole blood using validated RT-PCR assays, applicable to larger CRPC cohorts, provides prognostic value. The clinical utility in monitoring treatment will be prospectively explored.