20–25% Lower Concentrations of Total and Free Prostate-Specific Antigen (PSA) after Calibration of PSA Assays to the WHO Reference Materials – Analysis of 1098 Patients in Four Centers

Aim: To examine the potential clinical implications of the recalibration of total prostate-specific antigen (PSA) and free PSA (fPSA) assays to the World Health Organization (WHO) standard materials. Material and methods: Data from 1098 patients with or without clinically detected prostate cancer (PCa) from four independent cohort studies were compared using commercial assays calibrated to the traditional Hybritech® PSA (PSA-Hyb) and fPSA (fPSA-Hyb) standards and to the WHO 96/670 (PSA-WHO) and 96/668 (fPSA-WHO) standards. The Access® Immunoassay System (Beckman Coulter, Inc.) was used in all studies. Results: All studies showed 20% to 25% lower PSA and fPSA test results with the WHO-standardized assays. No significant change in %fPSA (fPSA/PSA × 100) was observed. Continuing to use the traditional clinical PSA cutoffs obtained with the Hybritech standard after changing to the PSA-WHO standard could result in up to one-third of prostate cancer cases being missed. Conclusions: Manufacturers should fully inform laboratories about a calibration change and its clinical impact. Laboratory reports for PSA measurements should indicate the assay’s manufacturer and which calibration standard was used to avoid misleading information concerning PCa risk (Int J Biol Markers 2009; 24: 65-9) New WHO PSA standards 66 stable reference material – a principle called metrological traceability (5). Traditionally, new reference standards are also linked to modified interpretive criteria to reflect the change in analyte concentration of the new standard (4, 6-9). Besides the metrological traceability, antibody specificity is important for immunoassay comparability (10). DISCORDANT PSA ReSULTS To date, several manufactures have changed their PSA calibration to the new WHO reference materials. However, numerous studies have documented discordant results for PSA, fPSA, and %fPSA among commercial assays aligned or not aligned with the WHO standards (11-19). A large prospective comparison study using the original Hybritech-calibrated PSA assay demonstrated a 16% to 20% lower PSA concentration when a WHO 96/670-calibrated PSA assay (Centaur, Bayer/Siemens) was used (14). Another comparison with 5 different PSA and fPSA assays (one with cPSA instead of fPSA) showed significant differences for tPSA values but even larger variance when analyzing the %fPSA values regardless of WHO calibration (18). A large comparison tPSA study on artificial samples by Roddam et al (20) showed >10% variations between tPSA assays. While the WHO-calibrated Elecsys assay (Roche Diagnostics) was reported to yield about 20% lower results on artificial samples (20), the results of 596 clinical samples showed 7% higher concentrations compared with the Hybritech-calibrated PSA assay (18). This shows that not only calibration differences lead to these discordances but also sample types (21). All such variations could result in misinterpretation of individual PSA and %fPSA results, with erroneous clinical decisions regarding the need for a prostate biopsy (22). Thus, the adoption of the WHO standard by many commercial PSA assays appears to have compromised the timely detection of PCa by maintaining traditional interpretive criteria while significantly lowering test results in many patients. Nevertheless, the primary cause of the discordance in PSA values observed between current PSA tests appears to be the use of different calibration materials. In response to this, Beckman Coulter now provides optional calibration of their Access Hybritech PSA to the WHO 96/670 (PSA-WHO) standard and of their fPSA assay to the WHO 96/668 (fPSA-WHO) standard in addition to their conventional Access Hybritech calibrations (PSAHyb and fPSA-Hyb). However, the conventional PSA-Hyb calibration has been used for more than 20 years and was used to establish the widely accepted PSA cutoffs used in clinical practice today. These cutoffs were established in clinical studies using the manual predecessor Hybritech Tandem PSA assays (1). SePARATe STUDy ReSULTS Recent data from 4 independent studies demonstrate the differences in PSA results and potential clinical consequences when using PSA-WHO and fPSA-WHO assays compared with PSA-Hyb and fPSA-Hyb assays without making the appropriate adjustments in the clinical decision limits (23-26). The aim of this summary and commentary is to inform medical practitioners of the important clinical implications of recalibration of PSA and fPSA assays to the WHO standard materials. In the first study, Vignati and colleagues prospectively measured PSA and fPSA in 201 consecutive patients with the PSA-Hyb and PSA-WHO calibrated Access assays (i.e., the assays were performed using different calibrators but the same assay method on the Beckman Coulter Access) (26). Within the PSA range of 2.5 to 20 ng/mL, the slopes of the regression lines (slope) for PSA-WHO and fPSA-WHO were 0.767 and 0.737, respectively, with a 0.966 slope for %fPSA (Tab. I). Therefore, this study showed 23% lower PSA-WHO results and 26% lower fPSA-WHO results, while %fPSA was not significantly affected when the WHO standards were used in both the PSA and fPSA assays. The authors calculated the clinical impact when continuing to use the traditional 4 ng/mL PSA cutoff for recommending a prostate biopsy with the PSA-WHO assay. Of the 201 patients, 22.4% (n=45) would not have been classified as TABLe I PATIENT NUMBERS AND REGRESSION EqUATIONS FOR THE 4 STUDIES AND THE SUMMARy Study group Vignati et al (26) Stephan et al (25) Fillée et al (23) Jansen et al (24) Summary of all studies Patients (n) 201 641 150 106 1098 (n=992)§ PSA equation WHO=0.763 WHO=0.747 WHO=0.819 WHO=0.796 WHO=0.750 * PSA-Hyb + 0.039 * PSA-Hyb + 0.049 * PSA-Hyb 0.009 * PSA-Hyb + 0.007 * PSA-Hyb + 0.0575 fPSA equation WHO=0.737 WHO=0.776 WHO=0.782 Not measured WHO=0.769 * fPSA-Hyb + 0.012 * fPSA-Hyb + 0.001 * fPSA-Hyb 0.0015 * fPSA-Hyb + 0.003 %fPSA equation WHO=0.966 WHO=1.018 WHO=0.91 Not calculated WHO=0.988 * %fPSA-Hyb + 0.246 * %fPSA-Hyb + 0.064 * %fPSA-Hyb 0.46 * %fPSA-Hyb + 0.320§ The different Passing-Bablok regression equations for PSA, fPSA and %fPSA are given for the 4 investigated studies while study No. (24) only measured PSA. §The summary data for fPSA and %fPSA are calculated for 992 patients only.