Prostate-specific Antigen Immunoassays Research Articles

ENHANCEMENT OF SURFACE PLASMON RESONANCE USING COLLOIDAL GOLD NANOPARTICLES EMBEDDED IN A SILICA LAYER

This paper presents a strategy for the signal enhancement of surface plasmon resonance biosensors using colloidal gold nanoparticles and a silica layer. We describe the method for the deposition of a silica-stabilized gold nanoparticle layer on a gold film, namely an enhanced surface plasmon resonance chip. This chip shows significant changes in its surface plasmon resonance signals when biomolecules are attached to its surface as compared to a normal gold surface. These characteristics are closely related to the surface plasmon resonance effect as determined using prostate-specific antigen. The detection limit of the enhanced surface plasmon resonance chip is determined to be 0.01 ng/mL for a prostate-specific antigen immunoassay. The use of an enhanced surface plasmon resonance chip makes it possible to enhance signals 1000-fold compared to the signals obtained by conventional surface plasmon resonance sensing. The enhancement of the surface plasmon resonance spectral shift results from the coupling of the surface and particle plasmons through the application of a silica-stabilized gold nanoparticle layer on the gold surface.

Le Prostate Health Index (PHI) a-t-il une place dans l’arsenal du dépistage biologique du cancer de la prostate ? Étude préliminaire

Summary Objectives The Prostate Health Index (PHI) is calculated from concentrations of 3 different proteins: the total Prostate Specific Antigen (PSA), the free PSA and [–2]proPSA (p2PSA).We compared the PHI with 3 different markers: The total PSA, the free PSA and the free PSA/total PSA rate. Patients and methods When the total PSA was measured between 2 and 10 ng/mL, both the p2PSA and the free PSA immunoassays were performed. Statistical analysis searched a correlation between new (the p2PSA and the PHI) and usual (the total PSA, the free PSA and the free PSA/total PSA rate) markers. Results The p2PSA is correlated with the total PSA, the free PSA and the free PSA/total PSA rate but we observed a great dispersion. The PHI is not correlated with the total PSA neither with the free PSA nor with the free PSA/total PSA rate. Conclusion These results show that the PHI is an independent biomarker. A clinical study will show if the PHI is an interesting clinical biomarker.

Power-Free Microchip Immunoassay of PSA in Human Serum for Point-of-Care Testing

For point-of-care testing (POCT), a sandwich-typed immunoassay on poly(dimethylsiloxane) (PDMS) microfluidic chips was developed by employing two original technologies: a power-free microchip and laminar flow-assisted dendritic amplification (LFDA). Sequential injection of reagents of immunoassay was carried out by the power-free microchip. In addition, for the amplification of a fluorescent signal, LFDA exploited the interaction between FITC-labeled streptavidin and biotinylated anti-streptavidin on the interface of the two streams of laminar flow during the sequential injection of the reagents of the immunoassay. The microfluidic immunoassay was finished within 21 min with a sample volume of 500 nL. The calibration curve of a quantitative analysis satisfied the concentration range of the cut-off values of prostate-specific antigen (PSA) in serum. The limit of detection (LOD) reached 520 pg/mL (16 pM) of PSA in human female serum. We believe that this microfluidic immunoassay and device are promising for POCT.

Simultaneous Detection of Single Molecules and Singulated Ensembles of Molecules Enables Immunoassays with Broad Dynamic Range

We report a method for combining the detection of single molecules (digital) and an ensemble of molecules (analog) that is capable of detecting enzyme label from 10(-19) M to 10(-13) M, for use in high sensitivity enzyme-linked immunosorbent assays (ELISA). The approach works by capturing proteins on microscopic beads, labeling the proteins with enzymes using a conventional multistep immunosandwich approach, isolating the beads in an array of 50-femtoliter wells (Single Molecule Array, SiMoA), and detecting bead-associated enzymatic activity using fluorescence imaging. At low concentrations of proteins, when the ratio of enzyme labels to beads is less than ∼1.2, beads carry either zero or low numbers of enzymes, and protein concentration is quantified by counting the presence of "on" or "off" beads (digital regime). (1) At higher protein concentrations, each bead typically carries multiple enzyme labels, and the average number of enzyme labels present on each bead is quantified from a measure of the average fluorescence intensity (analog regime). Both the digital and analog concentration ranges are quantified by a common unit, namely, average number of enzyme labels per bead (AEB). By combining digital and analog detection of singulated beads, a linear dynamic range of over 6 orders of magnitude to enzyme label was achieved. Using this approach, an immunoassay for prostate specific antigen (PSA) was developed. The combined digital and analog PSA assay provided linear response over approximately four logs of concentration ([PSA] from 8 fg/mL to 100 pg/mL or 250 aM to 3.3 pM). This approach extends the dynamic range of ELISA from picomolar levels down to subfemtomolar levels in a single measurement.

Serum-based ALYGNSA immunoassay for the prostate cancer biomarker, total prostate-specific antigen (tPSA)

Prostate-specific antigen (PSA) is a serum glycoprotein overproduced by the prostate in prostate cancer (> or = 4 ng/mL in the bloodstream). An immunoassay for total PSA (tPSA) was developed using the ALYGNSA method to enhance capture antibody orientation and a limit of detection of 0.63 ng/mL was reported, a limit 15-fold lower than a commercial tPSA ELISA assay. This ALYGNSA assay, however, was performed using only buffer-based proteins and blocking agents (Mackness et al., Anal Bioanal Chem 396:681-686, 2010). To improve the clinical application of this system, a serum-based tPSA ALYGNSA was developed employing human serum. This assay also resulted in a limit of detection of 0.63 ng/mL of tPSA protein. The findings reported here provide support for the clinical application of this assay for diagnosis, progression, treatment, and possible recurrence of prostate cancer.

Nanoparticle-based bio-barcode assay redefines “undetectable” PSA and biochemical recurrence after radical prostatectomy

We report the development of a previously undescribed gold nanoparticle bio-barcode assay probe for the detection of prostate specific antigen (PSA) at 330 fg/mL, automation of the assay, and the results of a clinical pilot study designed to assess the ability of the assay to detect PSA in the serum of 18 men who have undergone radical prostatectomy for prostate cancer. Due to a lack of sensitivity, available PSA immunoassays are often not capable of detecting PSA in the serum of men after radical prostatectomy. This new bio-barcode PSA assay is approximately 300 times more sensitive than commercial immunoassays. Significantly, with the barcode assay, every patient in this cohort had a measurable serum PSA level after radical prostatectomy. Patients were separated into categories based on PSA levels as a function of time. One group of patients showed low levels of PSA with no significant increase with time and did not recur. Others showed, at some point postprostatectomy, rising PSA levels. The majority recurred. Therefore, this new ultrasensitive assay points to significant possible outcomes: (i) The ability to tell patients, who have undetectable PSA levels with conventional assays, but detectable and nonrising levels with the barcode assay, that their cancer will not recur. (ii) The ability to assign recurrence earlier because of the ability to measure increasing levels of PSA before conventional tools can make such assignments. (iii) The ability to use PSA levels that are not detectable with conventional assays to follow the response of patients to adjuvant or salvage therapies.

Point-of-care fluorescence immunoassay for prostate specific antigen

Background Prostate specific antigen (PSA) is widely used as a clinical marker for diagnosis, screening, and prognosis of prostate cancer. A fluorescence (FL) dye-incorporated immunochromatographic assay (ICA) was developed for detection of PSA concentration in whole blood or serum. Methods Whole blood or serum mixed with FL-conjugated detector was loaded onto a cartridge and incubated for 12 min. The FL intensity of the cartridge was then measured in a laser FL scanner. The analytical performance of the FL-ICA system was evaluated by precision and recovery tests. The comparability of the new method was examined with an automated analyzer. Results A reliable correlation between area ratio ( A T/ A C), reflecting FL intensity of test/control line, and PSA concentration was observed ( r = 0.998). The CVs of intra- and inter-assay precision in a range of 2.5–8 μg/l were < 6.0% and 4.5%, respectively, and analytical recovery of the FL-ICA system fell within 6.5% at the tested samples. When the FL-ICA method was compared with Abbott AxSYM and Bayer Centaur analyzers, there were strong correlations ( r = 0.993 and r = 0.992, p < 0.0001). Conclusion The FL-ICA system with point-of-care-testing (POCT) appeared to be an easy, fast and suitable method for measurement of PSA concentration in whole blood, and needs no accessory equipment to separate serum.

Ultrasensitive immunoassay for prostate specific antigen using scanning tunneling microscopy-based electrical detection

We characterized a vertically configured electrical detection system that used scanning tunneling microscopy (STM) to detect antigen-antibody binding. This technique could be used to easily construct a multiple measurement system in a protein chip. We utilized immunocomplexes comprised of our model protein, prostate specific antigen (PSA), corresponding antibody fragments, and gold nanoparticle-antibody conjugates. The electrical tunneling current between the STM tip and these complexes exhibited a peaklike pulse, the frequency of which depended on the surface density of the bound complexes. We could therefore quantitatively measure PSA concentrations as low as 10fg∕mL using periodogram analysis of this peak frequency.

Relevance of the prostate-specific antigen (PSA) nanotest compared to the classical PSA test in the organized mass screening of prostate cancer

To assess the reliability of a new measurement of prostate-specific antigen (PSA) using a blotting-paper assay (nanotest) compared to the standard PSA immunoassay. The PSA level was measured in 205 men volunteers (median age 70 years, range 41-75) using a nanotest and a standard PSA immunoassay, collected at the same time; 30 microL of capillary blood placed on to a blotting paper were collected for the nanotest and sent by mail to the same laboratory for the two assays. The results were compared statistically using the Spearman test, the intraclass correlation coefficient and the Bland-Altman test. The nanotest threshold for an abnormal PSA level was 78 pg/mL, which corresponded to a standard PSA value of 3 ng/mL, with a sensitivity of 100%. There was a significant correlation (r = 0.98, Spearman test; P < 0.001) between the nanotest and the standard PSA assay. The intraclass correlation coefficient was 0.87. The Bland-Altman test showed a good agreement between the nanotest and the standard PSA assay, but there was an increasing proportional difference with increasing PSA value. There was a very high correlation between the nanotest and the standard PSA assay, especially for standard PSA levels of <5 ng/mL. Economic and clinical studies are indicated to confirm the utility of the nanotest in organized mass screening of prostate cancer.

Synthesis and characterization of europium(III) nanoparticles for time-resolvedfluoroimmunoassay of prostate-specific antigen

Recent advances in the fabrication and bioconjugation of nanometre-sized lanthanide(III)chelate particles have led to robust high specific activity labels. This paper describes thesynthesis and characterization of lanthanide(III) nanoparticle labels and the use ofa nanoparticle in a bioaffinity assay system. Two europium(III) nanoparticleswere prepared using an extremely simple, inexpensive and fast agglomerationstrategy. A silica-stabilized nanoparticle was synthesized from hydrophobictris(dibenzoylmethane)–mono(phenanthroline) and tris(dibenzoylmethane)–mono(5-aminophenanthroline)europium(III) chelates in aqueous solution. In addition, a naphthoyltrifluoroacetone:tri-n-octylphosphineoxide:sodium dodecyl sulfate europium(III) complex was agglomerated inwater. The particle sizes ranged from 62 to 140 nm in diameter. The silica-stabilized particlewas further coated with a monoclonal antibody. The analytical performance of thebioconjugated nanoparticle label was evaluated in a model sandwich immunoassay ofprostate-specific antigen. The detection limit of human prostate-specific antigen was28 ng l−1, 850 fM, in a microtiter plate format using time-resolved fluorometry. The coefficient ofvariation ranged from 1 to 9%. The novel nanoparticle label improves the specific activityof existing lanthanide(III) nanoparticle labels and simplifies the preparation route. Inaddition, prepared high-density nanoparticle labels using lanthanide(III) chelates orother specific fluorochromes have potential applications in a number of otherfields.

A Systematic Method for Selection of Promising Serum Protein Biomarkers to Improve Prostate Cancer (PCa1) Detection

Immunoassays for prostate-specific antigen (PSA) are used routinely as an aid to detect prostate cancer (PCa). PSA permits early detection of PCa but lacks specificity (1)(2). In a typical screening population of African American men ≥40 years of age and other men ≥50 years of age, ∼4% of the men with a PSA concentration >4 μg/L, which is the upper reference limit for PSA, will have prostate cancer (1). Most of these cancers can be detected using PSA. At issue, however, is the comorbidity of men with benign prostatic hyperplasia whose age and PSA concentrations are within these ranges. For example, 50% of men >60 years of age have benign prostatic hyperplasia (3), and many will have PSA >4 μg/L. The low specificity of PSA measurement as a test for PCa necessitates many prostate biopsies, of which ∼75% are negative and theoretically unnecessary. Additional biomarkers may improve sensitivity and specificity of PSA. We theorized that bioinformatics and analysis of the existing scientific literature could identify promising protein biomarker candidates that, if combined with PSA, might improve clinical sensitivity and especially specificity. The work flow we used to select candidate biomarkers is diagramed in the flow chart in Fig. 1⇓ . We searched Medline and other relevant databases from Nerac Corporation (Tolland, CT) for publications that appeared in 1999–2004 with the terms prostate cancer and markers or biomarkers or early detection or aggressiveness or staging or grading. We also used prostate-specific genes, prostate-specific proteins, and several well-known institutions and investigators in the field of prostate cancer research as search terms. Once compiled, the titles and abstracts were read to find proteins or genes documented to be differentially regulated in tissue or serum or plasma PCa samples compared with normal or benign samples. A list of >2700 citations was …

Prostate-specific antigen immunosensing based on mixed self-assembled monolayers, camel antibodies and colloidal gold enhanced sandwich assays

Prostate-specific antigen (PSA) is a valuable biomarker for prostate cancer screening. We developed a PSA immunoassay on a commercially available surface plasmon resonance biosensor. Our PSA receptor molecule consists of a single domain antigen-binding fragment, cAbPSA-N7, derived from dromedary heavy-chain antibodies and identified after phage display. It binds PSA with a high k(on) value of 1.9x10(6) M-1 s-1, and was covalently immobilised on a gold substrate via a mixed self-assembled monolayer (SAM) of alkanethiols by using carbodiimide-coupling chemistry in 10mM acetate buffer pH 5.5 to obtain an optimal pre-concentration. The best performing and optimised mixed SAM consisted of (10%) 16-mercapto-1-hexadecanoic acid (16-MHA) for covalent cAbPSA-N7 immobilisation and (90%) 11-mercapto-1-undecanol (11-MUOH) to minimise non-specific adsorption of the analyte. In this way, two advantages are incorporated in a single coupling layer. Up to 28 fmol/mm2 of cAbPSA-N7 could be immobilised and 30% of its binding sites participate actively in PSA interaction. In addition, the optimised layer showed also optimal performance to assess physiological samples. Although PSA concentrations as low as 10 ng/ml could be detected directly, this detection limit could be enhanced to PSA levels in the sub ng/ml range by introducing a sandwich assay involving a biotinylated secondary antibody and streptavidin modified gold nanoparticles. This approach realizes the PSA detection at clinical relevant concentrations.

Soy Consumption and Phytoestrogens: Effect on Serum Prostate Specific Antigen When Blood Lipids and Oxidized Low-Density Lipoprotein are Reduced in Hyperlipidemic Men

Soy Consumption and Phytoestrogens: Effect on Serum Prostate Specific Antigen When Blood Lipids and Oxidized Low-Density Lipoprotein are Reduced in Hyperlipidemic Men

Evaluation of the clinical performance of equimolar- and skewed-response total prostate-specific antigen assays versus complexed and free PSA assays and their ratios in discriminating between benign prostatic hyperplasia and prostate cancer

Background: Prostate-specific antigen (PSA) exists in human serum in two principal forms, free PSA (fPSA) and protein-complexed PSA, predominantly PSA-ACT (α 1-antichymotrypsin). Equimolar response (EMR) total PSA (tPSA) immunoassays measure each of these forms equally while skewed-response (SKR) assays overestimate or underestimate the tPSA concentration. The advantages of EMR over SKR tPSA assays are controversial. Methods: We used five nonhuman serum-based samples each containing a different proportion of fSPA:PSA-ACT (0:100 to 100:0, %:%) and patients' serum samples from men with histologically confirmed benign prostatic hyperplasia (BPH) ( n=94) or PCA ( n=30) and a wide range of fPSA concentrations to investigate the molar response status of six tPSA assays. Receiver–operator characteristic (ROC) curve analysis was used to compare the discriminatory power of these assays in distinguishing men with BPH from those with PCA. Results: The Bayer Immuno-1 tPSA (BtPSA) assay demonstrated EMR characteristics and diagnostic accuracy similar to the Hybritech Tandem-E and Tandem-R tPSA assays. At 90% sensitivity, EMR tPSA assays had higher specificity than SKR tPSA assays. Conclusions: The BtPSA assay is an EMR tPSA assay and EMR assays provide improved diagnostic specificity over SKR tPSA assays.

The Proportion of Prostate-specific Antigen (PSA) Complexed to α1-Antichymotrypsin Improves the Discrimination between Prostate Cancer and Benign Prostatic Hyperplasia in Men with a Total PSA of 10 to 30 μg/L

The aim of this study was to assess the diagnostic accuracy of the proportion of prostate-specific antigen (PSA) complexed to alpha(1)-antichymotrypsin (PSA-alpha(1)ACT:PSA ratio) in the differential diagnosis of prostate cancer (CaP) and benign prostatic hyperplasia (BPH) in men with total PSA of 10-30 microg/L. We used our immunoassays (ELISAs) for total PSA and PSA-alpha(1)ACT complex to study 146 men. In 123, total PSA was between 10 and 20 microg/L; 66 of these had CaP and 57 BPH. In 23 men, total PSA was between 20 and 30 microg/L; 14 of these had CaP and 9 BPH. We calculated the area under the ROC curves (AUC) for total PSA, PSA-alpha(1)ACT complex, and PSA-alpha(1)ACT:PSA ratio, and determined the cutoff points that gave sensitivities approaching 100%. In the total PSA range between 10 and 20 microg/L, the AUC was significantly higher for the PSA-alpha(1)ACT:PSA ratio (0.850) than for total PSA (0.507) and PSA-alpha(1)ACT complex (0.710; P <0.0001). A cutoff ratio of 0.62 would have permitted diagnosis of all 66 patients with CaP (100% sensitivity) and avoided 19% of unnecessary biopsies (11 of 57 patients). In the total PSA range between 20 and 30 microg/L, the AUC for the PSA-alpha(1)ACT:PSA ratio (0.980; 95% confidence interval, 0.82-0.99) was greater than the AUC for total PSA (0.750; 95% confidence interval, 0.51-0.89; P = 0.042). In this range, a cutoff point of 0.64 would have permitted the correct diagnosis of all 14 patients with CaP and 6 of the 9 with BPH. The diagnostic accuracy of the PSA-alpha(1)ACT:PSA ratio persists at high total PSA concentrations, increasing the specificity of total PSA. Prospective studies with large numbers of patients are needed to assess whether the ratio of PSA-alpha(1)ACT to total PSA is a useful tool to avoid unnecessary prostatic biopsy in patients with a total PSA >10 microg/L.

Advanced prostatic carcinoma with low levels of serum prostate-specific antigen

The serum levels of prostate-specific antigen (PSA) represent a significant diagnostic and monitoring parameter of prostatic carcinoma (PC). The aim of the study was to establish correlation of serum PSA level in addition to grade, histological type, and clinical stage of PC in patients with normal or intermediary PSA serum level. In 37 untreated PC patients with preoperative serum PSA levels ranging between 0.1 and 9.6 ng/ml, paraffin-embedded tissue and serum samples were immunohistologically studied and immunoassay for PSA was done. The most representative was poorly differentiated PC with D stage. In serum samples from PC patients 27 (73.7%) normal (< or = 4.0 ng/ml), and 10 (27.3%) intermediate (4.1-10 ng/ml) PSA levels were found. Immunohistochemistry, in 36 PC (97.3%) had demonstrated the expression of PSA. Our study results had shown low serum PSA levels in some patients with advanced poorly differentiated PC.

Development of Region-Specific Antisera and Immunoassay for Human Prostate-Specific Antigen (PSA) with Use of Purposely Designed Synthetic Peptides

Prostate-specific antigen (PSA) immunoassay is now widely used in the clinical management of prostate cancer. However, the diversity of molecular forms of the protein caused the results of PSA immunoassay variable depending on specificity of the antibody used. The current study provided an efficient method for production of anti-PSA antisera strictly specific for the expected region of the molecule using purposely designed synthetic peptides. The synthetic peptides included PSA (42-92), (53-92), (50-70), (76-92), (174-184) and (199-210). Immunization of the peptides in rabbits afforded seventeen kinds of antisera. Binding activity to native PSA and immunohistochemistry with human prostate proved nine of them, three anti-PSA (42-92), two anti-PSA (53-92) and four anti-PSA (50-70) antisera, to be useful for further experiment. None of antisera against PSA (76-92) and (174-184) showed binding activity with native PSA and immunostaining in human prostate. Using anti-PSA (53-92) antisera with synthetic PSA (42-92) peptide as antigen and biotinyl-Gly-Gly-PSA (42-92) as labeled antigen, a novel type of PSA specific enzyme immunoassay system was successfully developed. The assay system is well defined in terms of region specificity, which is highly advantageous for its clinical use. In addition, the strict region-specificity will make the assay system useful as an analytical tool for characterization of the PSA molecule in blood. Attempts for production of PSA antisera specific to other regions of PSA molecule along this line is now in progress.

Prostate-specific antigen and human glandular kallikrein 2 are markedly elevated in urine of patients with polycystic ovary syndrome.

Prostate-specific antigen (PSA) is a well-established tumor marker of prostatic adenocarcinoma. Human glandular kallikrein 2 (hK2), another serine protease closely related to PSA, is also gaining ground as a promising diagnostic tool in prostate cancer. The expression of these 2 proteases is known to be regulated by androgens and progestins in hormonally responsive tissues, such as the male prostate and the female breast. Previously, we have shown that serum PSA levels in normal women are very low but still detectable by ultrasensitive PSA immunoassays. We have also demonstrated that some women with hyperandrogenic syndromes have elevated serum PSA levels. In this study, we have measured urinary PSA and urinary hK2 levels in 35 polycystic ovary syndrome (PCOS) patients and compared them to those of 41 age-matched controls. We found that urinary PSA levels were significantly higher (P < 0.0001) in PCOS patients (mean +/- SE = 820 +/- 344 ng/L) than in the controls (mean +/- SE = 4.3 +/- 1.8 ng/L). Similarly, the difference between urinary hK2 of patients (mean +/- SE = 8.2 +/- 3.1 ng/L) and controls (0.5 +/- 0.3 ng/L) was also significant (P < 0.001). A weak correlation was observed between urinary PSA and serum 3 alpha-androstanediol glucuronide (r(s) = 0.42, P = 0.03) as well as between urinary PSA and serum testosterone (r(s) = 0.40, P = 0.04). The results of this study indicate that urinary PSA, and possibly urinary hK2, are promising markers of hyperandrogenism in females suffering from PCOS.

Reference Reagents for Prostate-specific Antigen (PSA): Establishment of the First International Standards for Free PSA and PSA (90:10)

Prostate-specific antigen (PSA) measurements in serum by immunoassay are widely used in the screening, diagnosis, and monitoring of patients with prostate cancer although the lack of common reference reagents has led in the past to wide differences in estimates. We report here the results of a WHO international collaborative study in which two preparations of PSA representative of the main immunoreactive components in serum, free PSA and PSA 90:10, and a preparation of recombinant DNA-derived PSA were assessed as potential standards for the calibration of diagnostic immunoassays for PSA. Coded vials of the candidate materials and serum preparations containing PSA in the clinically important range were provided to the 10 laboratories in the study, and participants were asked to perform PSA assays currently in use in their laboratories. Data from 89 immunoassays by 26 different method-laboratory combinations were contributed to the study and analyzed centrally at the National Institute for Biological Standards and Control. Potency estimates of the preparations relative to the in-house calibrators were in good agreement with the target value of 1 microg of total PSA/vial, the preparation of free PSA giving 1.10 microg/vial (95% confidence interval, 0.99-1.21 microg/vial) and PSA 90:10, 1.11 microg/vial (95% confidence interval, 1.04-1.18 microg/vial). No immunoreactivity was detected in ampoules containing the recombinant material. Use of a common standard of PSA 90:10 significantly reduced the between-laboratory geometric coefficients of variation for serum samples included in the study and gave a much narrower range of potency estimates. The preparation of free PSA was established by WHO as the First International Standard for PSA (free) with an assigned content of 1 microg of total PSA per vial. In addition, the preparation of bound PSA was established as the First International Standard for PSA (90:10) with an assigned content of 1 microg of total PSA per vial.

Is ICI 182,780 an antiprogestin in addition to being an antiestrogen?

The pure antiestrogen ICI 182,780 has been shown to have antiprogestin activity in reporter gene constructs. Cell lines, naturally devoid of progesterone receptors (PR) were transfected with either the A or B forms of the human PR and a luciferase construct driven by a progesterone-response element (PRE). Because this system is an artificial one, our purpose was to determine whether these observations could be made in a human breast cancer cell line, naturally containing PR. We further evaluated the dose-response of ICI 182,780 and RU-486 (mifepristone) on PR and estrogen receptors (ER) in the presence of either progesterone, norgestrel or estradiol. These effects were measured using immunoassays for prostate-specific antigen (PSA) and human glandular kallikrein (hK2) and pS2. We found that ICI 182,780 blocked progesterone-stimulated PSA and hK2 production 100% at 10(-5) M, which decreased significantly by 10-6 M. This inhibition did not occur when norgestrel was the progestin used. RU-486 showed 100% blockade for both progestins at all concentrations used. We concluded that the antiprogestin activity of ICI 182,780 exists for progesterone only. This weak antiprogestin activity may be unlikely to have significant clinical implications.