The Establishment and Evaluation of Luminescent-Labelled Immunometric Assays for Prostate-Speciflc Antigen-α1-Antichymotrypsin Complexes in Serum

Abstract

Summary:Prostate-specific antigen is found in the prostate in two forms, one with a low (30 000) and one with a high (100000) relative molecular mass. The latter has recently been found to be a complex of prostatespecific antigen with aj-antichymotrypsin.Immunoluminometric assays were designed for the prostate-specific antigen-ocj-antichymotrypsin complex äs well äs for (Xi-antichymotrypsin, the former being compared with a commercially available radioimmunoassay for prostate-specific antigen (ProsChek RIA -Yang Laboratories).The precision of the immunoluminometric assays was acceptable (intra-assay Variation < 7%; inter-assay Variation < 8.5%) in the measuring ranges 0-90 g/l for the prostate-specific antigen-a r antichymotrypsin complex and 0-12 g/l for a r antichymotrypsin.The correlation between the assays for prostate-specific antigen and prostate-specific antigen-aj-antichymotrypsin complex was acceptable, showing a correlation coefficient r = 0.83 after double logarithmic transformation, or r = 0.85 using the Spearman rank correlation on 131 data pairs.Extremely high arantichymotrypsin levels (above 2 g/l) caused interference in the prostate-specific antigena r antichymötrypsin complex assay. Such levels, although rare, are encountered in pulmonary inflammatory disease. The reference ranges for the three assays were found to be äs follows: prostate-specific antigen 0.13-4.63 g/l, prostate-specific antigeii-ai-antichymotrypsin complex 0.08 -1.78 g/l, and for ai-antichymotrypsin 0.27-0.61 g/l. These values were obtained from 82 höspitalised males fpr the first two assays and from 80 males and females free from infection fpf the latter. % Purified prostate-specific antigen (M r 30000) does not react in the prostate-specific antigen-arantichymotrypsin complex as$ay; a concentration of 200 /1 generates a signal whieh is less than that from the first Standard (0.04 g/l).In three cases of metästatic cancer of the prostate, discrepancies were found in the values from the prostatespecific antigen assay (8.56, 30.0 and 107 g/l) and the prostate-specific antigen-a r antichymotrypsin complex assay (0.78, 3.72 and 1,24 jig/1). This may indieate the production of an altered prostate-specific antigen, which was unable to complex with a r antichymotrypsin.The new assay is not suitable äs a screening assay for prostatic cancer, but it may be of interest for detecting metästatic disease.

Reference Ranges for the Concentrations of Total and Complexed Plasma Prostate-Specific Antigen and Their Ratio in Patients with Benign Prostate Hyperplasia

“…was also stable for at least 5 days at 4°C, 2 months at -20°C and 2 years at -80°C. Using an assay for measuring PSA complexed to cti ACT, we comfirmed previous data [6][7][8][9][10] showing that PSA complexed to a [ACT is the major form of PSA in circulation in prostate cancer and BPH patients, account ing for 91 and 73% of the total PSA immunoreactivity, respectively [11]. We also showed that the use of a totalto-complexed PSA ratio of 0.8 as a cut-off point increased the specificity from 38 to 79% without decreasing the sen sitivity (82%) in patients with prostate cancer with a total PSA concentration <15 ng/ml [11], These results agree with others previously reported [9,12], and suggest that the use of complexed-to-total PSA ratio may help urolo gists to identify patients with organ-confined prostate cancer.…”

“…Several groups have therefore studied this problem, and their results show that calculation of the free PSA/total PSA ratio or complexed PSA/total PSA ratio instead total PSA alone provides a better discrimination between be nign prostate hyperplasia and prostate cancer [7][8][9][10][11], Measuring PSA:aiACT complex instead of free PSA has a potential advantage in test samples with low total PSA levels in which only a minimal fraction of PSA is in the free form and most of it is in the form of PSAxxi ACT complex. For example, a plasma sample with a total PSA level of 2 ng/ml where 10% of this PSA in the free form will have only 0.2 ng/ml of free PSA but 1.8 ng/ml of PSA total PSA ratio of 0.80 as a cut-off to exclude prostate can cer, we would have avoided 39 of the 40 biopsies that did not demonstrate cancer in the group of patients with PSA >4 ng/ml and normal DRE and TRUS.…”

“…assays to evaluate free PSA or PSA complexed to otiACT and have investigated their usefulness in discriminating between BPH and prostate cancer patients [7][8][9][10][11], Their results show that calculation of the free PSA/total PSA or complexed PSA/total PSA ratios provide a better discrim ination between BPH and prostate cancer than does the total PSA alone. In all studies, the complexed-to-total PSA ratio in patients with prostate cancer was significant ly higher than in men with BPH, which means that deter mining the concentrations and ratios of the different forms of PSA in serum may provide valuable information to better distinguish between hyperplasia and cancer.…”

Prostate-Specific Antigen and Its Complexes with AlpharAntichymotrypsin in the Plasma of Patients with Prostatic Disease

“…There fore, the total PSA measured by available immunoassays consists of the sum of free PSA and the PSA complexed to cii-ACT (PSA:ai-ACT), the latter representing the major portion. Several groups have developed assays to evaluate PSA complexed to arACT and have investigated its use fulness in discriminating between BPH and prostate can cer patients [9,10,12,13]. Stenman et al [9] and Christensson et al [12] found that PSA:ai-ACT is the predomi nant form of PSA in serum from patients with prostate cancer and suggested that calculation of the free/total PSA ratio provides a better discrimination between BPH and prostate cancer.…”

“…The release of active PSA in intercellular fluids or blood plasma would be expected to result in rapid inacti vation of the enzymatic activity by extracellular inhibi tors [6,7], In fact, it has been reported that in prostate cancer patients, free noncomplexed PSA is a minor frac tion in serum [8] and that PSA in blood is bound to a2-macroglobulin (a2-M) and ai-antichymotrypsin (cl-ACT) [8][9][10], but not to protein C inhibitor (PCI) [11], Both the free form and the ai-ACT-bound form are detectable by current PSA immunoassays, but the efficiency with which they are detected may differ. In contrast, the PSA bound to a2-M is probably engulfed in the oo-M structure, renOver the past several years, there has been tremendous interest in prostate-specific antigen (PSA) because it is usually present at higher than normal levels in the serum of prostate carcinoma patients and is therefore useful as a marker for diagnosing this disease and monitoring the therapy [1,2].…”