Prostate-specific Antigen Isoforms Research Articles

The natural progression of patients with an IsoPSA value and its predictive ability of clinically significant prostate cancer on biopsy.

325 Background: IsoPSA is a structure-based serum assay exploring the spectrum of possible prostate-specific antigen (PSA) isoforms. It was shown to outperform total and percent-free PSA in detecting clinically significant prostate cancer (csPCa) (grade group (GG) ≥2 on biopsy). IsoPSA reduced unnecessary biopsies and magnetic resonance imaging (MRI). We sought to compare the outcomes of eventual biopsy and imaging of surveilled patients with an initially normal or high IsoPSA, thus assessing IsoPSA’s prospective predictive ability of csPCa. Methods: We performed a single-center retrospective review of patients who underwent IsoPSA testing from 2017-present. Data was dichotomized into patients with normal (≤6) and high IsoPSA (>6). We collected the outcomes of any consequent IsoPSA and PSA test, prostate biopsy and MRI. We calculated the statistical IsoPSA’s characteristics for the prediction of csPCa on biopsy. Results: The median follow-up time of 811 patients who underwent IsoPSA testing was 18 months (IQR, 16.5-20). Among 443 patients with initial low IsoPSA, 5 (1.1%) had a csPCa on a subsequent biopsy, 19 (4.3%) subsequent high IsoPSA, 122 (27.5%) rising PSA, and 22 (5%) csPCa on MRI. Among 368 patients with initial high IsoPSA, 105 (28.5%) had a csPCa on a subsequent biopsy and 106 (28.8%) on an MRI. The sensitivity of IsoPSA to predict csPCa was 95.5%, and the NPV was 94.8%. Among 124 patients with high IsoPSA and initial negative biopsy, 110 had a subsequent negative and 14 a positive biopsy (10GG1 (8.1%), 4 ≥GG2 (3.2%)), with a respective median IsoPSA of 7.2 and 9.6 (p=0.007). An IsoPSA>10 generated an OR of csPCa of 7.2 (95%CI 2.1-25.2, p=0.0005). Conclusions: In 18-month follow-up, 1.1% of patients with normal IsoPSA developed csPCa compared to 28.5% of patients with high IsoPSA. In the cohort of patients with high IsoPSA and initially negative biopsy, 3.2% eventually developed csPCa, however, having a significantly higher IsoPSA than those who remained negative. The odds of having csPCa were 7 times higher with IsoPSA>10. [Table: see text]

Immuno-analytical characteristics of PSA and derived biomarkers (total PSA, free PSA, p2PSA)

Prostate-specific antigen (PSA) is the recommended tumor marker for individual screening and follow-up of prostate cancer. This paper reviews main structural and physiological data about prostate specific antigen isoforms: total PSA, free PSA, [-2]proPSA (also named p2PSA). It describes the pre-, per- and post-analytical conditions for these different parameters. It presents the interpretation of results and derived calculated indices (free/total PSA ratio, Prostate Health Index or PHI) for the management of prostate cancer (initial diagnosis and follow-up).

Utility of Prostate-Specific Antigen Isoforms and Prostate Health Index in the Diagnosis of Metastatic Prostate Cancer.

Objective The current study was undertaken to investigate the utility of total prostate-specific antigen (tPSA), its isoform [-2] proPSA (p2PSA), and prostate health index (PHI) in the diagnosis of metastatic prostate cancer (PCa). Materials and Methods This study was conducted from March 2016 to May 2019. Eighty-five subjects who were diagnosed with PCa for the first time, following transrectal ultrasound-guided prostate biopsy, were included in the study. The prebiopsy blood samples were analyzed in Beckman Coulter Access-2 Immunoanalyzer for tPSA, p2PSA, and free PSA (fPSA), and the calculated parameters included %p2PSA, %fPSA, and PHI. Mann-Whitney's U test was used as test of significance, and p -value less than 0.05 was considered statistically significant. Results Of the 85 participants, 81.2% ( n = 69) had evidence of metastasis, both clinically and pathologically. The median tPSA (ng/mL), p2PSA (pg/mL), %p2PSA, and PHI were significantly higher in the group with evidence of metastasis (46.5 vs. 13.76; 198.0 vs. 35.72; 3.25 vs. 1.51; 237.58 vs. 59.74, respectively). The sensitivity (%), specificity (%), negative predictive value (%), and positive predictive value (%) to diagnose metastatic PCa of tPSA at a cutoff of 20 ng/mL, PHI at a cutoff of 55, and %p2PSA at a cutoff of 1.66 were 92.7, 98.5, and 94.2; 37.5, 43.7, and 62.5; 54.5, 87.5, and 71.4; and 86.4, 88.3, and 91.5, respectively. Conclusion Using tests such as %p2PSA and PHI in the standard armamentarium for the diagnosis of metastatic PCa in addition to PSA will help in selecting the appropriate treatment strategy, including active surveillance.

Clinical validation of IsoPSA, a single parameter, structure-focused assay for improved detection of prostate cancer: A prospective, multicenter study.

IsoPSA is a blood-based test that assesses prostate cancer (CaP) risk by partitioning and detecting cancer-specific structural isoforms of prostate specific antigen (PSA) with an aqueous 2- phase system. To validate the diagnostic performance of IsoPSA for High-Grade CaP and Any CaP risk on biopsy in men age ≥ 50 with total PSA ≥ 4 ng/ml. Prospective, multicenter study of 888 men scheduled for prostate biopsy at 8 academic and community sites between August 2015 and August 2020. IsoPSA test. Receiver operating characteristic and likelihood ratio analysis used to validate diagnostic performance for previously established IsoPSA Index cutoffs for High-Grade CaP (Gleason Score ≥ 7) and Any CaP (Gleason Score ≥ 6), compare IsoPSA to total PSA and % free PSA, and evaluate subgroups (total PSA 4-10 ng/ml, total PSA > 10 ng/ml, biopsy naïve, prior negative biopsy). The disease prevalence was 35.6% (High-Grade CaP) and 58.9% (Any CaP). The area under the receiver operating characteristic curve was 0.783 (High-Grade CaP) and 0.770 (Any CaP). IsoPSA outperformed total PSA and % free PSA on area under the receiver operating characteristic curve, specificity, positive and negative predictive value at similar sensitivity. Using selected IsoPSA Index cutoffs, an estimated 46% (High-Grade CaP) and 42% (Any CaP) of biopsies could be avoided in low-risk patients. IsoPSA displayed statistically informative likelihood ratio-based predictive characteristics. IsoPSA maintained accuracy in clinically relevant subgroups. IsoPSA diagnostic performance and predictive value is validated for High-Grade CaP and Any CaP in men age ≥ 50 with total PSA ≥ 4 ng/ml undergoing diagnostic biopsy. IsoPSA outperforms total and % free PSA in discriminating the risk of prostate cancer on biopsy. IsoPSA has the potential to reduce unnecessary biopsies and improve the risk-benefit ratio for CaP early detection.

Early prediction of prostate cancer biochemical recurrence and identification of disease persistence using PSA isoforms and human kallikrein-2.

The role of isoforms of prostate specific antigen (PSA) and other kallikrein-related markers in early detection of biochemical recurrence (BCR) after radical prostatectomy (RP) is not well known and serum PSA is currently used in preoperative risk nomograms. The aim of this research was to study pre- and early postoperative levels of important PSA isoforms and human kallikrein-2 to determine their ability to predict BCR and identify disease persistence (DP). This study included 128 consecutive patients who underwent RP for clinically localized prostate cancer. PSA, fPSA, %fPSA, [-2]proPSA, PHI and hK2 were measured preoperatively, at 1 and 3 months after RP. We determined the ability of these markers to predict BCR and identify DP. The DP and BCR rate were 11.7%and 20.3%respectively and the median follow up was 64 months (range 3-76 months). Preoperatively, the independent predictors of BCR were PSA (p-value 0.029), [-2]proPSA (p-value 0.002) and PHI (p-value 0.0003). Post-RP, PSA was the single marker correlating with BCR, both at one (p-value 0.0047) and 3 months (p-value 0.0004). PSA, fPSA, [-2]proPSA and PHI significantly correlated to DP at 1 and 3 months post-RP (p-value < 0.05), although PSA had the most significant existing correlation (p-value < 0.0001). [-2]proPSA and PHI are preoperative predictors of BCR and DP that outperform the currently used serum PSA. At the early postoperative period there is no additional benefit of the other markers tested.

Reclassification prediction of first-year protocol biopsy on active surveillance of prostate cancer by p2PSA-related parameters: from PRIAS-JAPAN.

There is no useful predictive marker for reclassification on active surveillance. Thus, we aimed to investigate thresholds of [-2] proPSA (p2PSA)-related parameters to predict reclassification of the first-year protocol biopsy (1-year PBx) and evaluate the influence of clinical decision-making. This was an observational, prospective cohort study conducted at 19 Japanese institutes. The inclusion criteria included clinical stage T1c/T2, prostate-specific antigen (PSA) levels ≤10 ng/mL, PSA density <0.2 ng/ml/cc, one or two positive biopsy cores, and Gleason score (GS) ≤6 (GS ≦7 for patients aged ≥70 years) at diagnostic biopsy. All participants were required to receive a blood-sampling test on a protocol visit at inclusion and at the 1-year PBx. PSA and PSA isoforms (free PSA, p2PSA) were measured, and parameters (%free PSA, %p2PSA, phi) were calculated. Multivariable logistic regression models were used to predict the reclassification risk. To assess the predictive power and thresholds for reclassification, we plotted Receiver Operating Characteristic (ROC) curves. Decision curve analysis (DCA) was used to evaluate the variables that yielded a net clinical benefit. A total of 135 patients were included, and 36 patients were reclassified on the 1-year PBx. Multivariate analyses showed that %p2PSA and phi at inclusion and p2PSA, %p2PSA, and phi before the 1-year PBx were significant predictors of reclassification at the 1-year PBx. The ROC analysis showed an optimal cutoff point, sensitivity, and specificity of %p2PSA and phi before the 1-year PBx of 1.64, 86%, 49% and 35.92, 89%, 47%, respectively. The DCA showed that phi before the 1-year PBx had the highest net benefit. The study limitation was its single-arm observational design. %p2PSA and phi before the 1-year PBx had a good prediction power. phi is the most useful indicator for clinical decision-making on active surveillance. This study is registered atthe Japan Trial Register with ID UMIN000009876 ( https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000011573 ).

Predictive Factors of Missed Clinically Significant Prostate Cancers in Men with Negative Magnetic Resonance Imaging: A Systematic Review and Meta-Analysis.

We systematically reviewed the literature on predictive factors for clinically significant prostate cancer diagnosis after prebiopsy negative magnetic resonance imaging in prostate cancer naïve patients. The MEDLINE® and Scopus® databases were searched up to March 2019. The review protocol was published in the PROSPERO database (CRD42019125549). The clinical factors and markers studied were age, prostate specific antigen, prostate specific antigen isoforms, prostate specific antigen density, PCA3, prostate volume, family history, ethnicity and risk calculators. The primary objective was to determine their predictive ability for clinically significant prostate cancer diagnosis. Secondary objectives included meta-analysis of the negative predictive value of prebiopsy negative magnetic resonance imaging when combined with these predictive factors. A total of 16 studies were eligible for inclusion. Few studies reported negative predictive value of magnetic resonance imaging combined with a marker. Prostate specific antigen density was the best studied and the strongest predictor of clinically significant prostate cancer in men with prebiopsy negative magnetic resonance imaging. There were 8 studies (1,015 patients) eligible for meta-analysis of the added value of prostate specific antigen density less than 0.15 ng/ml/ml to magnetic resonance imaging in reducing the risk of missing clinically significant prostate cancer. When combined with prostate specific antigen density, overall magnetic resonance imaging negative predictive value increased from 84.4% to 90.4% in cancer naïve patients. The increase was from 82.7% to 88.7% in biopsy naïve and from 88.2% to 94.1% in previous negative biopsy subgroups. The use of prostate specific antigen density less than 0.15 ng/ml/ml in the presence of prebiopsy negative magnetic resonance imaging was the most useful factor to identify men without clinically significant prostate cancer who could avoid biopsy.

Early Diagnosis of Prostate Cancer from the Perspective of Chinese Physicians.

Prostate cancer (PCa) is the seventh most diagnosed cancer and the tenth leading cause of cancer mortality in China. Unlike the USA, both incidence and mortality continue to increase. In China, PCa is often diagnosed at a locally advanced or metastatic stage, resulting in a high mortality-to-incidence ratio. Implementing regular screening using a well-validated biomarker may result in the earlier diagnosis of localized disease. Furthermore, it is important to be able to distinguish between low-grade and high-grade disease, to avoid subjecting patients to unnecessary biopsies, undertreatment of significant disease, or overtreatment of indolent disease. While prostate-specific antigen (PSA) is commonly used in PCa screening around the world, its relationship to PCa is still unclear and results vary widely across different studies. New biomarkers, imaging techniques and risk predictive models have been developed in recent years to improve upon the accurate detection of high-grade PCa. Blood- and urine-based biomarkers, such as PSA isoforms, prostate cancer antigen 3, or mRNA transcripts, have been used to improve the detection of high-grade PCa. These markers have also been used to create risk predictive models, which can further improve PCa detection. Furthermore, multiparametric magnetic resonance imaging is becoming increasingly accessible for the detection of PCa. Because of ethnic variations, biomarkers and risk predictive models validated in Western populations cannot be directly applied to Chinese men. Validation of new biomarkers and risk predictive models in the Chinese population may improve PCa screening and reduce mortality of this disease in China.

PSA screening—a matter of debate?

Prostate cancer (PCa) is the most common cancer type in men. Overall, the goal of prostate-specific antigen (PSA)-based screening is to identify early-stage disease that can be treated successfully. However, evidence supporting an overall survival advantage in men undergoing PSA screening is controversial, leading to a lack of consensus on guidelines. The present review article provides an overview of the most important studies on PSA screening including the latest updates of large trials. In addition, modern concepts in early detection of PCa that can be added to PSA measurement alone such as PSA isoforms or mpMRI are discussed. Lastly, recommendations for PCa screening and early detection from the European and German Societies of Urology issued in 2019 are presented.

Clinical Validation of IsoPSA™, a Single Parameter, Structure Based Assay for Improved Detection of High Grade Prostate Cancer

Current prostate specific antigen markers to detect prostate cancer are limited by low specificity for high grade disease. IsoPSA™ is a blood based, structure focused assay which predicts risk by partitioning the isoforms of prostate specific antigen that are linked to cancer in an aqueous 2-phase reagent system. We validated the clinical performance of this assay for identifying high grade disease in a new contemporary biopsy cohort. We performed a multicenter prospective validation in 271 men scheduled for prostate biopsy at a total of 7 academic and community centers who were enrolled between May 2017 and March 2018. Blood samples were obtained for assay prior to biopsy. The discrimination power of the assay to detect high grade prostate cancer (Gleason 7 or greater) was evaluated by ROC analysis and compared to prior results. Clinical performance was further improved by comparison with multiparametric magnetic resonance imaging-ultrasound vs transrectal ultrasound guided biopsies. The assay AUC was 0.784 for high grade vs low grade cancer/benign histology, which was superior to the AUCs of total prostate specific antigen and percent free prostate specific antigen. If 1,000 patients were biopsied, the assay would have reduced the number of unnecessary biopsies from 705 to 402 (43%) with only 22 missed high grade cancers, of which 7 would have been Gleason sum 4 + 3 or higher. Subset analysis of multiparametric magnetic resonance imaging guided biopsy produced a substantial improvement of the AUC to 0.831. Validation of the structure based IsoPSA assay demonstrated statistical concordance with previously reported results and verified its superior performance vs concentration based prostate specific antigen and the free-to-total prostate specific antigen ratio. The assay improvement in detecting high grade prostate cancer using multiparametric magnetic resonance imaging-ultrasound guided biopsy may help define a new diagnostic paradigm.

Capillary Electrophoresis Analysis of Prostate-Specific Antigen (PSA).

The Capillary Electrophoresis (CE) profile of isoforms (peaks) of a glycoprotein can be useful to show alterations in its posttranslational modifications (PTMs) linked to diseases. These changes can modify the electrophoretic mobility of these isoforms in a minor extent and, therefore, very reproducible CE methods are needed to detect them. In this chapter, a method for the analysis of prostate-specific antigen (PSA) by Capillary Zone Electrophoresis (CZE) with UV detection is detailed. High reproducibility in the separation of a large number of PSA isoforms is achieved by performing capillary conditioning in acid media and by using a background electrolyte (BGE) at pH8.0 formulated with decamethonium bromide and urea.

Abstract IA08: Early detection of high-risk localized prostate cancer

Abstract Despite prostate cancer being the second leading cause of cancer death amongst U.S. men and prevalent use of serum prostate specific antigen (PSA) as an FDA-approved biomarker, PCa screening and early detection is highly controversial. Widespread PSA screening and prostate biopsy led to a profound stage migration, with the majority of newly diagnosed cases being clinically localized and low grade. Thus PSA-screening programs, typically using a threshold of 4.0ng/mL for biopsy referral, result in substantial overdiagnosis, estimated at 15-84%. Additionally, although active surveillance (AS) is an emerging management option for men with presumed indolent, low-grade (Gleason score 6, grade group I) PCa, most U.S. men with such disease still undergo curative treatment via radical prostatectomy (RP) or radiation therapy (RT), which have well-documented side effects that substantially impact quality of life. Hence, serum PSA-based screening has also led to PCa overtreatment, where screening-detected tumors that never would have caused clinical symptoms are treated definitively. Lastly, as a result of PSA screening, an estimated ~1,000,000 prostate biopsies are performed annually, which are associated with infection requiring antibiotics in ~6% of patients and hospitalization in ~1% of biopsied patients. Despite appeals to lower the commonly used cutoff for biopsy in the U.S. or the development of multivariate models that incorporate PSA and other clinical factors to provide individualized PCa risk estimates, PSA is still commonly used as a dichotomous test, with referral for biopsy if PSA is &amp;gt; 4.0ng/mL. Several modifications of serum PSA, including free PSA, PSA velocity, various PSA isoforms, and related proteins including KLK2, have been proposed as biomarkers and show clinical utility to further define an individual’s risk for PCa. More recently, urine-based biomarkers, such as the long noncoding RNA (lncRNA) transcript PCA3, as well as germline single-nucleotide polymorphisms (SNPs) that are associated with increased PCa risk, have also been proposed as biomarkers. Likewise, advances in multiparametric MRI are also enabling the noninvasive detection and targeted biopsy of prostate cancer. Lastly, numerous tissue-based prognostic assays are available to identify undersampled or unsampled aggressive prostate cancer from diagnostic material. Here, I will review the underlying challenges inherent in the early detection of aggressive, high-risk localized prostate cancer, focusing on both the high prevalence of disease as well as multifocality. In addition, I will review the clinical utility of available approaches, as well as implementation issues. Importantly, enormous advances have been made in our understanding of the prostate cancer genome and transcriptome, which have been incompletely leveraged in early detection strategies. These findings will be discussed in the context of improved detection of high-risk localized prostate cancer, which has the potential to dramatically impact the lives of millions of U.S. men. Citation Format: Scott A. Tomlins. Early detection of high-risk localized prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr IA08.

Prostate-Specific Antigen: a Review of Assay Techniques, Variability and Their Clinical Implications

Prostate-specific antigen (PSA) is used worldwide in the detection and management of prostate cancer. Its discovery revolutionised the care of patients with prostate cancer, but there remains a question of why inter-assay variability exists and how significantly this affects the decision-making process in the care of patients with the disease. We conducted a literature review to evaluate the two areas: inter-assay variability and its relation to clinical practice. Our results show there are internationally recognised standard assays, and the variability of assays are reliant on local/national calibration to a set standard preparation and rely on the measurement of different isoforms of PSA on an equimolar basis. Inter-assay variability exists and can be as high as 8%, which has differing impacts on decision making for diagnostic procedures and treatment modalities depending on the clinical circumstances and PSA level.

The role of prostate cancer biomarkers in undiagnosed men.

This article intends to review biomarkers derived from blood, urine, and tissue that can aid in the diagnosis of prostate cancer (PCa). PCa screening requires tools that complement prostate-specific antigen (PSA) with a higher specificity for clinically significant disease. Novel blood biomarkers, such as the Prostate Health Index (phi) and 4Kscore, utilize isoforms of PSA to more accurately predict high-grade PCa than traditional tools such as PSA and the percentage free-to-total PSA. Several gene products associated with PCa can be detected in the urine through commercially available assays. PCa antigen 3 (PCA3), though approved for repeat biopsy decisions, appears inferior to other biomarkers such as phi for identifying aggressive disease. However, combinations of PCA3 with other urine assays have shown promising results. One tissue-based hypermethylation test, named ConfirmMDx, can also be used to determine the need for repeat biopsy in men with a prior negative biopsy. Several biomarkers have been developed to aid in the screening and diagnosis of PCa. Such tests are often indicated in men with moderately elevated PSA or history of a prior negative biopsy. Their use facilitates reduction of unnecessary biopsies without sacrificing the early diagnosis of clinically significant PCa.

The Single-parameter, Structure-based IsoPSA Assay Demonstrates Improved Diagnostic Accuracy for Detection of Any Prostate Cancer and High-grade Prostate Cancer Compared to a Concentration-based Assay of Total Prostate-specific Antigen: A Preliminary Report

BackgroundIsoPSA is a serum-based assay that predicts prostate cancer (PCa) risk by partitioning isoforms of prostate-specific antigen (PSA) with an aqueous two-phase reagent. ObjectivesTo determine the diagnostic accuracy of IsoPSA in identifying the presence or absence of PCa and the presence of high-grade disease in a contemporary biopsy cohort. Design, setting, and participantsMulticenter prospective study of 261 men scheduled for prostate biopsy at five academic and community centers in the USA enrolled between August 2015 and December 2016. InterventionPerformance of the IsoPSA assay. Outcome measurements and statistical analysisDiscrimination power was evaluated using receiver operating characteristic (ROC) analysis. The outcome of the IsoPSA assay was transformed into risk probability using logistic regression. Decision curve analysis (DCA) was used to compare the net benefit of IsoPSA against other clinical protocols. Results and limitationsThe overall prevalence was 53% for any PCa and 34% for high-grade PCa. The area under the ROC curve was 0.79 for any cancer versus none and 0.81 for high-grade PCa versus low-grade PCa/benign histology. In this preliminary study, DCA revealed a superior net benefit of IsoPSA against no biopsy, all biopsy, and the modified Prostate Cancer Prevention Trial Risk Calculator 2.0. At a cutoff selected to recommend biopsy, IsoPSA demonstrated a 48% reduction in false-positive biopsies; at a cutoff selected to identity men at low risk of high-grade disease, there was a 45% reduction in the false-positive rate. ConclusionThe structure-based IsoPSA assay outperformed concentration-based PSA measurement, and provided a net benefit against other protocols. Once validated, clinical use of IsoPSA could significantly reduce unnecessary biopsies while identifying patients needing treatment. Patient summaryThe IsoPSA assay outperformed prostate-specific antigen in predicting the overall risk of prostate cancer and the risk of clinically significant cancer in a preliminary study. The IsoPSA assay could assist in determining the need for prostate biopsy for patients.

Sample preparation of serum to allow capillary electrophoresis analysis of prostate specific antigen isoforms

Prostate cancer is the second most frequently diagnosed cancer in men worldwide. Currently prostate specific antigen (PSA) serum concentration is the most used prostate cancer marker, but it only shows limited specificity. Because PSA glycosylation is altered by prostate cancer, detecting glycosylation changes could increase PSA specificity as a prostate cancer marker. Changes in PSA glycosylation can modify its electrophoretic- behavior and techniques such as capillary zone electrophoresis (CZE) and two-dimensional electrophoresis (2-DE) could be applied to detect changes in PSA glycosylation.Most serum PSA is complexed with alpha-1 antichymotrypsin (ACT). To have access to most of the PSA, the complexed PSA has to be released as free PSA (fPSA); in addition, this total fPSA must be purified from the serum matrix so that it can be analyzed using CZE.In this work a methodology for isolating PSA from serum for its CZE analysis was established. By using PSA standard, the effect of this methodology, which combines conditions for dissociating complexed PSA and immunoaffinity chromatographic purification, was studied. It was seen that this highly repeatable sample treatment did not noticeably alter the circular dichroism (CD) spectrum or the CZE pattern of PSA standard. Therefore, as a proof-of-concept, the developed sample treatment was applied to serum from a cancer patient with a high PSA content. The following observations can be made from these experiments: first of all, the 2-DE pattern of serum PSA remained unchanged after sample treatment; second, as hypothesized, the established sample preparation methodology made it possible to obtain the CZE pattern of PSA from serum; and third, the CZE pattern of serum PSA and of PSA standard from seminal plasma of healthy individuals, both submitted to the sample treatment method, showed some differences regarding the proportion of CZE peaks of the glycoprotein. These differences could be related to possible changes in the linkages of peptide backbone, in glycosylation or in other post-translational modifications between samples from both origins.

Evaluating serum insulin-like growth factor 1 and insulin-like growth factor binding protein 3 as markers in prostate cancer diagnosis.

Prostate-specific antigen (PSA) lacks specificity and sensitivity in discriminating prostate cancer (PCa) from benign prostatic hyperplasia (BPH) when the total PSA (tPSA) level is between 4 and 10 ng/mL. It remains to be investigated if additional tumor-associated molecules may improve the PCa diagnostic accuracy. The aim of the present study was to investigate whether serum levels of insulin-like growth factor 1 (IGF1), insulin-like growth factor binding protein 3 (IGFBP3) and their combinations with PSA may enhance the diagnosis of PCa. Serum tPSA and free PSA (fPSA) levels were measured using an automated chemiluminescence-based method. IGF1 and IGFBP3 levels were evaluated by radioimmunoassays in a prospectively and consecutively enrolled subset of 149 patients with tPSA ≤10 ng/mL made up of patients with benign prostatic hyperplasia (BPH; n = 113) and PCa (n = 36). IGF1 and IGFBP3 serum levels did not significantly differ between the PCa and BPH groups. No important correlation was found between the IGF molecules and PSA isoforms in both groups. Statistical analysis of the combination of markers indicated that only the free/total PSA ratio (f/tPSA%) was informative and independent in predicting the presence of PCa, considering that for high values of this percentage (17%) the probability of finding PCa decreased. Receiver operating characteristics areas under the curve (AUC) for IGF1 and IGFBP3 were not informative (AUC ~0.5 in both cases) contrary to the AUC for f/tPSA% (AUC = 0.689, p = 0.0002). The present study showed that neither IGF1 and IGFBP3 alone nor in combination with PSA enhance the diagnostic performance of PSA in PCa.

Impact of capillary conditioning and background electrolyte composition on capillary electrophoresis analysis of prostate specific antigen isoforms

Glycoproteins expressed in the human body can experience modifications as result of pathological situations. Detection of those changes can be useful as disease biomarkers. As a result of these modifications, size and/or electrical charge of the glycoprotein can be altered. Migration in capillary zone electrophoresis (CZE) is governed by the size to charge ratio of the analyte and therefore this separation technique can be used to monitor those modifications. At its turn, the alteration of the electrophoretical pattern of a given glycoprotein could be used as disease biomarker. To this aim, high repeatability for separation of a large number of peaks for a given glycoprotein is desirable.For prostate cancer, new markers are needed to decrease the high number of false positive results provided by the biomarkers currently used in clinics. In this sense, CZE methods for analysis of the several prostate specific antigen (PSA) peaks which this glycoprotein exhibit, called isoforms and containing one or more glycoforms, could be useful to study the PSA pattern as prostate cancer marker.In this study two complementary strategies to achieve both lot-to-lot capillary repeatability and high resolution of a large number of PSA isoforms are developed. Better performance and precision have been obtained for capillaries conditioned with HCl than for those conditioned with NaOH. Optimization of the background electrolyte (BGE) pH value to 8.0 and inclusion of 3M urea on its composition were the two factors of highest impact for enhancing resolution of the highest number of PSA peaks. Under the optimized conditions for capillary conditioning and BGE pH and composition, long-term resolution of 10 isoforms of PSA was achieved. Inter-day (n=3) %RSD was 0.55 for the ratio tm/tEOF, 1.15 for μeff, and 5.02 for % Acorr of the PSA peaks.

A portable chemiluminescence imaging immunoassay for simultaneous detection of different isoforms of prostate specific antigen in serum

A multianalyte chemiluminescence (CL) imaging immunoassay strategy for sensitive detection of different isoforms of prostate specific antigen (PSA) was developed. The microtiter plates were fabricated by simultaneously immobilizing of free-PSA (f-PSA) and total-PSA (t-PSA) capture antibody on nitrocellulose (NC) membrane. Each of the array were spotted in replicates of six spots within a spacing of 2mm. 16 or 48 detection wells were integrated on a single NC membrane and each well could be used as a microreactor and microanalysis chamber. Under a sandwiched immunoassay, the CL signals on each sensing site were collected by a charge-coupled device (CCD), presenting an array-based chemiluminescence imaging. Soybean peroxidase (SBP) was used to label f-PSA or t-PSA monoclonal antibody. With the amplification effects of two enhancers, 3-(10′-phenothiazinyl) propane-1-sulfonate (SPTZ) and 4-morpholinopyridine (MORP), the CL intensity could significantly enhanced, which improved the sensing sensitivity and detection limit. Under the optimal conditions, the linear response to the analyte concentration ranged from 0.01–36.7ng/mL and 0.02–125ng/mL for f-PSA and t-PSA, respectively. The results for the detection of forty serum samples from prostate cancer patients and cancer-free patients showed good agreement with the clinical data, suggesting that the proposed assay had acceptable accuracy. The proposed CL imaging immunoassay possess high throughput and acceptable reproducibility, stability and accuracy, which made it great potential to available to distinguish different isoforms of PSA in serum samples.

Conventional and Promising Biomarkers for Prostate Cancer their Clinical Implication and Prospective Role

Prostate cancer (PCa) is one of the most common cancer in men and most common causes of male cancerrelated deaths. Over many years, biomarkers have been extensively studied for screening, diagnosis, prediction of (PCa) behavior and outcome and for assigning the patients during treatments. Molecular biomarkers could also help scientists to attain better understanding for the molecular basis of the disease and prediction of the patient response to therapies. Early detection of PCa was made possible about 30 years ago by the introduction of prostate specific antigen (PSA) in the clinical practice. However, PCa screening remains controversial, because of the risk of over diagnosis and/or over treatment and the inability to detect a significant proportion of advanced tumors. Several novel biomarkers have shown promises in preliminary studies. This review will focus on traditional biomarkers approved by FDA as PSA, PSA isoforms, Prostate health index (phi) and prostate cancer antigen 3 (PCA3), also novel and promising PCa biomarkers as Prostate stem cell antigen (PSCA) and Urokinase plasminogen activator (uPA), emphasizing on their molecular and biochemical basis, clinical implication and prospective role.