Background/Objectives: Prostate cancer (PCa) is a leading cause of cancer-related deaths among men, with early detection playing a crucial role in improving outcomes. MyProstateScore 2.0 (MPS2), a novel urinary biomarker test, predicts clinically significant PCa to reduce invasive biopsy procedures. This study evaluates the analytical performance of MPS2 using both a post-digital rectal exam (DRE) and non-DRE urine samples. Methods: We assessed the reproducibility, precision, and detection limits of the eighteen MPS2 analytes. Analytical parameters including the linear range, upper and lower limits of quantification (ULOQ and LLOQ), and interference from substances commonly present in urine were evaluated. The reproducibility of the MPS2 scores was evaluated across post-DRE and non-DRE clinical urine samples. Results: MPS2 analytes demonstrated high linearity (R² ≥ 0.975) across defined quantification ranges, with PCR efficiencies of 97–105%. The limits of detection (LOD) ranged from 40 to 160 copies/reaction, while the ULOQ was determined to be 10⁶–10⁷ copies/reaction for each analyte. Precision studies showed intra-run, inter-run, and inter-instrument standard deviations ≤0.5 Crt. Among the 12 potential interfering substances, only whole blood affected the performance of MPS2. The reproducibility of the MPS2 scores was consistent across post-DRE and non-DRE urine samples, meeting the acceptance criteria. Conclusions: The analytical validation confirms that MPS2 is robust and reliable in detecting biomarkers for clinically significant PCa. These findings, coupled with previous clinical validations, support the clinical use of MPS2 as a non-invasive diagnostic tool.

Background/Objectives: Active surveillance (AS) is recommended for men with low-risk prostate cancer to minimize overtreatment while monitoring for disease progression. However, current surveillance strategies rely heavily on repeat biopsies, which are invasive and associated with morbidity. MyProstateScore 2.0—Active Surveillance (MPS2-AS) is a urine-based biomarker test developed to predict progression to Grade Group ≥ 2 (GG ≥ 2) and Grade Group ≥ 3 (GG ≥ 3) prostate cancers in men on AS. The objective of this study was to analytically validate the reproducibility and robustness of MPS2-AS analyte detection and risk score calculation across key laboratory variables. Methods: Analytical precision was evaluated using pooled urine specimens processed using the MPS2-AS laboratory workflow. Eight pooled urine samples were tested in a within-laboratory design across five days, with two runs per day, and two replicates per run. Additional reproducibility studies assessed variability across three QuantStudio™ 12K Flex Real-Time PCR Systems and three OpenArray™ chip lots. Ten RNA biomarkers were quantified by RT-PCR and used to calculate the MPS2-AS GG1-2 and GG1-3 risk scores. Variance components were estimated using hierarchical ANOVA. Results: The MPS2-AS analyte measurements demonstrated high precision across within-laboratory testing, with standard deviations ranging from 0.00 to 0.60 and coefficients of variation (%CV) from 0.00 to 4.01%. The reproducibility across qPCR instruments and OpenArray chip lots showed similar robustness, with analyte %CVs of ≤4.57% and ≤4.10%, respectively. These stable analyte measurements translated to reproducible model outputs, with %CV ≤ 10.69% for the GG1-2 risk score and ≤7.20% for the GG1-3 risk score across all tested conditions. No systematic bias was observed between runs, days, instruments, or reagent lots. Conclusions: MPS2-AS demonstrates strong analytical precision and reproducibility for quantifying urinary biomarkers and generating GG1-2 and GG1-3 risk scores. These results support the reliability of MPS2-AS for clinical laboratory implementation and its use as a non-invasive tool to inform biopsy decisions in men with Grade Group 1 prostate cancer undergoing active surveillance.