Towards a Precision Medicine Approach and In Situ Vaccination against Prostate Cancer by PSMA-Retargeted oHSV.

Prostate specific membrane antigen (PSMA) is a specific high frequency cell surface marker of prostate cancers. Theranostic approaches targeting PSMA show no major adverse effects and rule out off-tumor toxicity. A PSMA-retargeted oHSV (R-405) was generated which both infected and was cytotoxic exclusively for PSMA-positive cells, including human prostate cancer LNCaP and 22Rv1 cells, and spared PSMA-negative cells. R-405 in vivo efficacy against LLC1-PSMA and Renca-PSMA tumors consisted of inhibiting primary tumor growth, establishing long-term T immune response, immune heating of the microenvironment, de-repression of the anti-tumor immune phenotype, and sensitization to checkpoint blockade. The in situ vaccination protected from distant challenge tumors, both PSMA-positive and PSMA-negative, implying that it was addressed also to LLC1 tumor antigens. PSMA-retargeted oHSVs are a precision medicine tool worth being additionally investigated in the immunotherapeutic and in situ vaccination landscape against prostate cancers.

Telomerase-based GX301 cancer vaccine in patients with metastatic castration-resistant prostate cancer: a randomized phase II trial.

Debate is around the optimal immunization regimen for cancer vaccines since too intense vaccination schedules may exhaust reactive lymphocytes. GX301 is a telomerase-based cancer vaccine whose safety and immunological effects were tested in a phase I trial applying an eight administrations schedule. Main objective of this study was to comparatively analyse safety and immunological response to three GX301 regimens in metastatic castration-resistant prostate cancer patients with response/disease stability after docetaxel chemotherapy. This was a multicentre, randomized, parallel-group, open-label trial registered with EudraCT (2014-000095-26) and ClinicalTrials.gov (NCT02293707, 2014). Ninety-eight patients were randomized to receive either eight (regimen 1), four (regimen 2) or two (regimen 3) vaccine administrations. Sixty-three patients were assessable for the primary immunological end-point. Vaccine-specific immune responses were evaluated by intracellular staining for IFN, elispot and cytotoxic assay at 90 and 180 days from baseline. No major side effects were recorded. A 54% overall immune responder rate was observed with 95% of patients showing at least one vaccine-specific immune response. Rate of immunological responders and number of immunizations were proportionally related, suggesting superiority of regimens 1 and 2 over regimen 3. Overall survival did not differ among regimens in both immunological responders and non-responders and was inversely associated (P = 0.002) with increase in the number of circulating CD8 + T regulatory cells at 180 days. These data indicate that GX301 cancer vaccine is safe and immunogenic in metastatic castration-resistant prostate cancer patients. Schedules with high number of administrations should be preferred in future studies due to their better immunological outcome.

Analysis of allergen profile in patients sensitized to canine allergen and potential Can f 5 cross-reactivity with human PSA.

Can f 5 allergy and possible cross-reactivity with human semen in which there are significant amounts of prostate-specific antigen (PSA) are particularly interesting aspects of allergy to dog. The objective of the study was to confirm cross-reactivity between human PSA and Can f 5 in a study of canine sensitised women. A total of 100 women (aged 18-73, 41 on average) with a positive history of animal fur allergy or positive skin prick tests to canine allergens were selected. Levels of Immunoglobulin E (IgE) specific to Can f 1, Can f 2, Can f 3, Can f 5 were determined. Patients with increased concentration of sIgE Can f 5 were selected for further inhibition testing using polystyrene microplate ELISA test coated with human PSA. In the studied population, allergy to Can f 5 dominated (52.3% of patients with increased concentration of canine-specific IgE were allergic to this allergenic component). In all analyzed cases, the concentration of IgE Can f 5 decreased after incubation on the ELISA plate coated with human PSA. The minimum decrease in concentration was 10.44%, the maximum was 37.73%, the average decrease was 21.6%. No statistically significant influence of the presence or absence of allergenic sIgE Can f 5 in blood serum on the occurrence of symptoms after intercourse was found. The study confirmed the moderate ability of Can f 5 to cross-react with human PSA sIgE, which may be clinically significant in some women. At the same time, symptoms of an allergy to male semen do not constitute a typical clinical presentation of allergy to Can f 5.

Phase I study of a multitargeted recombinant Ad5 PSA/MUC-1/brachyury-based immunotherapy vaccine in patients with metastatic castration-resistant prostate cancer (mCRPC).

BACKGROUND: Antitumor vaccines targeting tumor-associated antigens (TAAs) can generate antitumor immune response. A novel vaccine platform using adenovirus 5 (Ad5) vectors [E1-, E2b-] targeting three TAAs-prostate-specific antigen (PSA), brachyury, and MUC-1-has been developed. Both brachyury and the C-terminus of MUC-1 are overexpressed in metastatic castration-resistant prostate cancer (mCRPC) and have been shown to play an important role in resistance to chemotherapy, epithelial-mesenchymal transition, and metastasis. The transgenes for PSA, brachyury, and MUC-1 all contain epitope modifications for the expression of CD8+ T-cell enhancer agonist epitopes. We report here the first-in-human trial of this vaccine platform. METHODS: Patients with mCRPC were given concurrently three vaccines targeting PSA, brachyury, and MUC-1 at 5×1011 viral particles (VP) each, subcutaneously every 3 weeks for a maximum of three doses (dose de-escalation cohort), followed by a booster vaccine every 8 weeks for 1 year (dose-expansion cohort only). The primary objective was to determine the safety and the recommended phase II dose. Immune assays and clinical responses were evaluated. RESULTS: Eighteen patients with mCRPC were enrolled between July 2018 and September 2019 and received at least one vaccination. Median PSA was 25.58 ng/mL (range, 0.65-1006 ng/mL). The vaccine was tolerable and safe, and no grade >3 treatment-related adverse events or dose-limiting toxicities (DLTs) were observed. One patient had a partial response, while five patients had confirmed PSA decline and five had stable disease for >6 months. Median progression-free survival was 22 weeks (95% CI: 19.1 to 34). Seventeen (100%) of 17 patients mounted T-cell responses to at least one TAA, whereras 8 (47%) of 17 patients mounted immune responses to all three TAAs. Multifunctional T-cell responses to PSA, MUC-1, and brachyury were also detected after vaccination in the majority of the patients. CONCLUSIONS: Ad5 PSA/MUC-1/brachyury vaccine is well tolerated. The primary end points were met and there were no DLTs. The recommended phase II dose is 5×1011 VP. The vaccine demonstrated clinical activity, including one partial response and confirmed PSA responses in five patients. Three patients with prolonged PSA responses received palliative radiation therapy. Further research is needed to evaluate the clinical benefit and immunogenicity of this vaccine in combination with other immuno-oncology agents and/or palliative radiation therapy. TRIAL REGISTRATION NUMBER: NCT03481816.

PSA-Targeted Alpha-, Beta-, and Positron-Emitting Immunotheranostics in Murine Prostate Cancer Models and Nonhuman Primates.

PURPOSE: Most patients with prostate cancer treated with androgen receptor (AR) signaling inhibitors develop therapeutic resistance due to restoration of AR functionality. Thus, there is a critical need for novel treatment approaches. Here we investigate the theranostic potential of hu5A10, a humanized mAb specifically targeting free PSA (KLK3). EXPERIMENTAL DESIGN: LNCaP-AR (LNCaP with overexpression of wildtype AR) xenografts (NSG mice) and KLK3_Hi-Myc transgenic mice were imaged with 89Zr- or treated with 90Y- or 225Ac-labeled hu5A10; biodistribution and subcellular localization were analyzed by gamma counting, PET, autoradiography, and microscopy. Therapeutic efficacy of [225Ac]hu5A10 and [90Y]hu5A10 in LNCaP-AR tumors was assessed by tumor volume measurements, time to nadir (TTN), time to progression (TTP), and survival. Pharmacokinetics of [89Zr]hu5A10 in nonhuman primates (NHP) were determined using PET. RESULTS: Biodistribution of radiolabeled hu5A10 constructs was comparable in different mouse models. Specific tumor uptake increased over time and correlated with PSA expression. Treatment with [90Y]/[225Ac]hu5A10 effectively reduced tumor burden and prolonged survival (P ≤ 0.0054). Effects of [90Y]hu5A10 were more immediate than [225Ac]hu5A10 (TTN, P < 0.0001) but less sustained (TTP, P < 0.0001). Complete responses were observed in 7 of 18 [225Ac]hu5A10 and 1 of 9 mice [90Y]hu5A10. Pharmacokinetics of [89Zr]hu5A10 were consistent between NHPs and comparable with those in mice. [89Zr]hu5A10-PET visualized the NHP-prostate over the 2-week observation period. CONCLUSIONS: We present a complete preclinical evaluation of radiolabeled hu5A10 in mouse prostate cancer models and NHPs, and establish hu5A10 as a new theranostic agent that allows highly specific and effective downstream targeting of AR in PSA-expressing tissue. Our data support the clinical translation of radiolabeled hu5A10 for treating prostate cancer.

Utility of a Fifth-Generation Ultrasensitive Prostate-Specific Antigen Assay for Monitoring Prostate Cancer Patients after Radical Prostatectomy with 3 Years of Follow-Up.

BACKGROUND: We investigated an ultrasensitive prostate-specific antigen (uPSA) immunoassay (MesoScale; lower limit of detection (LLD) of 0.0035 pg/mL) to monitor patients with prostate cancer (PCa) following radical prostatectomy (RP) and to examine whether changes in PSA in the conventionally undetectable range (<1 pg/mL) can predict biochemical relapse (BCR). METHODS: We measured uPSA in serial serum samples (N = 100) collected from 20 RP cases with a third-generation ELISA (LLD of 1 pg/mL) and the fifth-generation MesoScale assay. We analyzed the PSA nadir changes to classify patients into BCR or non-BCR groups, observed the trends in PSA kinetics, and associated BCR status with clinicohistopathological features. RESULTS: The ELISA could quantify PSA in only 38% of the RP samples, detecting BCR in 7 of 20 patients with PCa. The MesoScale assay quantified PSA in all samples, showing 8 of 20 patients with BCR. However, there was no significant difference between the median time to BCR detection based on ELISA (1016 days) compared with MesoScale data (949 days). Gleason scores were higher in the BCR groups compared with non-BCR. There was no significant difference for other clinicohistopathological parameters. CONCLUSIONS: The uPSA MesoScale technology could track miniscule changes in serum PSA in the range of 0.003-1 pg/mL in all RP cases. However, PSA kinetics and nadir at concentrations <2 pg/mL fluctuated, and increases below this range could not reliably suggest signs of BCR. Instead, ultrasensitive fifth-generation PSA assays may hold clinical potential for measuring the low concentrations of PSA in women for various medical contexts.

Systemic Inflammatory Markers Are Predictive of the Response to Brachytherapy in the Prostate.

We analyzed the influence of the neutrophil/lymphocyte ratio (NLR) and platelet/lymphocyte ratio (PLR) on the biochemical recurrence (BCR) in low-intermediate risk prostate cancer (PCa). A total of 604 patients treated with exclusive brachytherapy for low- and intermediate-risk cancers were included in this study. No patient received either androgen deprivation or brachytherapy as a boost. BCR was defined according to the Phoenix definition (nadir prostatic specific antigen (PSA) +2). The median follow-up was 60 months (IQR 44-48 months). An NLR > 3 was more frequent in statin users (p = 0.025), but not in diabetics (p = 0.079). In univariate analysis (UVA) and multivariate analysis (MVA), a NLR > 3 (MVA p = 0.03), as well as Cancer of the Prostate Risk Assessment (CAPRA) low- vs. intermediate-risk (MVA p = 0.04), were predictive of BCR. When combining the NLR score with the CAPRA risk group, CAPRA intermediate risk patients with an NLR ≤ 3 (n = 157) had the worst (p = 0.0276) BCR rates, with a 5-year recurrence-free survival (p = 0.004, Bonferroni correction for six comparisons p = 0.024). We were able to identify a subgroup of PCa patients with CAPRA intermediate-risk and an NLR ≤ 3 who had worse BCR. This is in contrast to most other cancers, which have a worse prognosis when the NLR is high.

Upconversion luminescence-infrared absorption nanoprobes for the detection of prostate-specific antigen.

Aiming to the ongoing challenge of accurate and sensitive detection for cancer biomarkers, antibody-functionalized NaYF4:Yb3+, Er3+@SiO2 nanorods were developed as upconversion luminescence (UCL)-infrared absorption (IRA) nanoprobes. Benefiting from the shielding effect of the SiO2 shell, an enhanced UCL was achieved. Additionally, an IRA detection signal was introduced by the Si-O-Si bonds of SiO2. Its mutual verification with UCL signal was favorable for ensuring the accuracy of the assay. A UCL-IRA sandwich detection method was established for the detection of the prostate-specific antigen. The UCL intensity at 542 nm and IRA at 1095 cm-1 were chosen for quantitative assay. The method has high sensitivity (0.05 pg mL-1) and selectivity. The range of detection (200 fg mL-1-200 ng mL-1) was singnificantly broadened compared with that of single-readout UCL or IRA detection. The assay performance of human serum samples demonstrated the practicability of the method in clinical cancer diagnosis. Graphical abstract.

Self-assembled biotin-phenylalanine nanoparticles for the signal amplification of surface plasmon resonance biosensors.

A strategy for amplifying the signal of surface plasmon resonance (SPR) biosensors is reported. Biotinylated phenylalanine (Biotin-Phe) monomers were rapidly self-assembled into nanoparticles in a mild environment. The self-assembled nanoparticles were then used as the carriers of streptavidin-antibody complexes by the streptavidin-biotin interaction. The signal was amplified because of the high molecular weight of the nanoparticle-streptavidin-antibody conjugate. With prostate-specific antigen as a model analyte, the target concentration as low as 1 pg mL-1 was readily measured. The results of the nanoparticle-enhanced SPR biosensor for analysis of serum samples are well consistent with those achieved by the enzyme-linked immunosorbent assays. This work is valuable for designing of various optical and electronic biosensors through the streptavidin-biotin interaction. Graphical abstract.

Electrochemical assays for determination of H2O2 and prostate-specific antigen based on a nanocomposite consisting of CeO2 nanoparticle-decorated MnO2 nanospheres.

A nanocomposite consisting of CeO2 nanoparticle-decorated MnO2 nanospheres (CeO2@MnO2) was synthesized for the first time via a hydrothermal method. CeO2@MnO2 was exploited to construct an electrochemical assays for detecting H2O2 and prostate-specific antigen (PSA) with square wave voltammetry (SWV). The electrochemical results proved that CeO2@MnO2 owned a better electrocatalytic effect towards H2O2 reduction than pure MnO2 NS and CeO2 NP due to the synergistic effect between MnO2 NS and CeO2 NP. Under optimized conditions, CeO2@MnO2-based assay can be applied to detect H2O2 in the range 1 to 3.0 × 103 µmol L-1. The label-free electrochemical immunoassay based on CeO2@MnO2 displayed linearly with concentrations of PSA from 0.005 to 50.0 ng mL-1. The electrochemical assays also possessed acceptable sensitivity, selectivity, and stability. The study showed that CeO2@MnO2 hold great potential as a biosensing platform and the clinical determination of tumor markers in human serum. Graphical abstract A nanocomposite consisting of CeO2 nanoparticles decorated MnO2 nanospheres (CeO2 @MnO2) was firstly synthesized via a hydrothermal method. CeO2@MnO2 was firstly exploited to construct electrochemical assays for detecting H2O2 and prostate-specific antigen (PSA) with square wave voltammetry (SWV), respectively. The electrochemical results proved that CeO2@MnO2 owned better electrocatalysis towards H2O2 reduction than pure MnO2 NS and CeO2 NP due to the synergistic effect between MnO2 NS and CeO2 NP. Under optimized conditions, CeO2@MnO2 based assay relative to the H2O2 system can be applied to detect H2O2 with range from 1 to 3.0 × 103 µmol L-1. The label-free electrochemical immunoassay based on CeO2@MnO2 relative to the H2O2 system displayed linearly with concentrations of PSA from 0.005 to 50.0 ng mL-1. The electrochemical assays also possessed acceptable sensitivity, selectivity and stability. The study showed that CeO2@MnO2 hold great potential for biosensing platform and the clinic determination of tumor markers in human serum.

In Pursuit of Stability Enhancement of a Prostate Cancer Targeting Antibody Derived from a Transgenic Animal Platform.

Accelerated timelines necessitate the discovery of fully human antibodies as biotherapeutics using transgenic animals with a notion that such mAbs bypass humanization. A transgenic animal derived mAb (PCa75) targeted against a prostate cancer antigen had several 'unusual residues' (rare somatic hypermutations, rSHM, with positional frequency of <1%) that resulted in compromised biophysical properties (Tm = 61 °C and intrinsic stability ΔGu = 24.3 kJ/mol) and a sub-optimal immunogenicity profile. In our quest for quality medicine, we pursued antibody engineering strategies to enhance the stability of PCa75. PCa62, an engineered variant of PCa75, retained function while significantly improving the drug-like attributes of the molecule (Tm = 75 °C and intrinsic stability ΔGu = 63.5 kJ/mol). rSHM is rather prevalent, 18 out the 21 approved transgenic animal-derived antibodies have at least one 'unusual residue'. Thus, engineering of rSHM remains critical to enhance the stability and minimize immunogenicity risk of biotherapeutics.

Development of an optical microfiber immunosensor for prostate specific antigen analysis using a high-order-diffraction long period grating.

Fiber-optic biosensors are of great interest to many bio/chemical sensing applications. In this study, we demonstrate a high-order-diffraction long period grating (HOD-LPG) for the detection of prostate specific antigen (PSA). A HOD-LPG with a period number of less than ten and an elongated grating pitch could realize a temperature-insensitive and bending-independent biosensor. The bio-functionalized HOD-LPG was capable of detecting PSA in phosphate buffered saline with concentrations ranging from 5 to 500 ng/ml and exhibited excellent specificity. A limit of detection of 9.9 ng/ml was achieved, which is promising for analysis of the prostate specific antigen.

Genetic signature of prostate cancer mouse models resistant to optimized hK2 targeted α-particle therapy.

Hu11B6 is a monoclonal antibody that internalizes in cells expressing androgen receptor (AR)-regulated prostate-specific enzyme human kallikrein-related peptidase 2 (hK2; KLK2). In multiple rodent models, Actinium-225-labeled hu11B6-IgG1 ([225Ac]hu11B6-IgG1) has shown promising treatment efficacy. In the present study, we investigated options to enhance and optimize [225Ac]hu11B6 treatment. First, we evaluated the possibility of exploiting IgG3, the IgG subclass with superior activation of complement and ability to mediate FC-γ-receptor binding, for immunotherapeutically enhanced hK2 targeted α-radioimmunotherapy. Second, we compared the therapeutic efficacy of a single high activity vs. fractionated activity. Finally, we used RNA sequencing to analyze the genomic signatures of prostate cancer that progressed after targeted α-therapy. [225Ac]hu11B6-IgG3 was a functionally enhanced alternative to [225Ac]hu11B6-IgG1 but offered no improvement of therapeutic efficacy. Progression-free survival was slightly increased with a single high activity compared to fractionated activity. Tumor-free animals succumbing after treatment revealed no evidence of treatment-associated toxicity. In addition to up-regulation of canonical aggressive prostate cancer genes, such as MMP7, ETV1, NTS, and SCHLAP1, we also noted a significant decrease in both KLK3 (prostate-specific antigen ) and FOLH1 (prostate-specific membrane antigen) but not in AR and KLK2, demonstrating efficacy of sequential [225Ac]hu11B6 in a mouse model.

Multifaceted Bioanalytical Methods for the Comprehensive Pharmacokinetic and Catabolic Assessment of MEDI3726, an Anti-Prostate-Specific Membrane Antigen Pyrrolobenzodiazepine Antibody-Drug Conjugate.

Complex biotherapeutic modalities, such as antibody-drug conjugates (ADC), present significant challenges for the comprehensive bioanalytical characterization of their pharmacokinetics (PK) and catabolism in both preclinical and clinical settings. Thus, the bioanalytical strategy for ADCs must be designed to address the specific structural elements of the protein scaffold, linker, and warhead. A typical bioanalytical strategy for ADCs involves quantification of the Total ADC, Total IgG, and Free Warhead concentrations. Herein, we present bioanalytical characterization of the PK and catabolism of a novel ADC. MEDI3726 targets prostate-specific membrane antigen (PMSA) and is comprised of a humanized IgG1 antibody site-specifically conjugated to tesirine (SG3249). The MEDI3726 protein scaffold lacks interchain disulfide bonds and has an average drug to antibody ratio (DAR) of 2. Based on the structural characteristics of MEDI3726, an array of 4 bioanalytical assays detecting 6 different surrogate analyte classes representing at least 14 unique species was developed, validated, and employed in support of a first-in-human clinical trial (NCT02991911). MEDI3726 requires the combination of heavy-light chain structure and conjugated warhead to selectively deliver the warhead to the target cells. Therefore, both heavy-light chain dissociation and the deconjugation of the warhead will affect the activity of MEDI3726. The concentration-time profiles of subjects dosed with MEDI3726 revealed catabolism of the protein scaffold manifested by the more rapid clearance of the Active ADC, while exhibiting minimal deconjugation of the pyrrolobenzodiazepine (PBD) warhead (SG3199).

Multiplexed Detection of Cancer Serum Antigens with a Quantum Dot-Based Lab-on-Bead System.

A quantum dot (QD)-based lab-on-bead system is a unique tool for multiple analysis of cancer markers in human serum samples by using a flow cytometer. In terms of specificity and sensitivity, this method is comparable with ELISA, the "gold standard" of serological in-clinic detection of single analytes. Fluorescent microspheres encoded with QDs have been used for the quantitative detection of free and total prostate-specific antigen in human serum samples. Developed multiplex assay demonstrates a clear discrimination between serum samples from control subjects and cancer patients. The proposed QD-based method is adaptable and makes it possible to develop numerous clinical tests with decreased duration and cost for early diagnosis of various diseases.

Metal-Labeled Aptamers as Novel Nanoprobes for Imaging Mass Cytometry Analysis.

Imaging mass cytometry (IMC) is an emerging imaging technology that exploits the multiplexed analysis capabilities of the CyTOF mass cytometer to make spatially resolved measurements for tissue sections. In a comprehensive view of tissue composition and marker distribution, recent developments of IMC require highly sensitive, multiplexed assays. Approaching the sensitivity of the IMC technique, we designed a novel type of biocompatible metal-labeled aptamer nanoprobe (MAP), named 167Er-A10-3.2. The small molecular probe was synthesized by conjugating 167Er-polymeric pentetic acid (167Er-DTPA) with an RNA aptamer A10-3.2. For demonstration, 167Er-A10-3.2 was applied for observing protein spatial distribution on prostatic epithelium cell of paraffin embedded Prostatic adenocarcinoma (PaC) tissue sections by IMC technology. The 167Er-A10-3.2 capitalizes on the ability of the aptamer to specifically bind target cancer cells as well as the small size of 167Er-A10-3.2 can accommodate multiple aptamer binding antigen labeled at high density. The detection signal of 167Er-A10-3.2 probe was 3-fold higher than that of PSMA antibody probe for a targeted cell under lower temperature epitope retrieval (37 °C) of PaC tissue. Furthermore, we successfully demonstrated the simultaneously staining ability of aptamer probes in IMC analysis. The successful imaging acquisition using aptamers probes in IMC technology may offer opportunity for the diagnosis of malignancies in the future.

CD8+ T Cells Impact Rising PSA in Biochemically Relapsed Cancer Patients Using Immunotherapy Targeting Tumor-Associated Antigens.

The management of men with prostate cancer (PCa) with biochemical recurrence following local definitive therapy remains controversial. Early use of androgen deprivation therapy (ADT) leads to significant side effects. Developing an alternative, clinically effective, and well-tolerated therapy remains an unmet clinical need. INO-5150 is a synthetic DNA therapy that includes plasmids encoding for prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA), and INO-9012 is a synthetic DNA plasmid encoding for interleukin-12 (IL-12). This phase 1/2, open-label, multi-center study enrolled men with PCa with rising PSA after surgery and/or radiation therapy. Patients were enrolled into one of four treatment arms: arm A, 2 mg of INO-5150; arm B, 8.5 mg of INO-5150; arm C, 2 mg of INO-5150 + 1 mg of INO-9012; and arm D, 8.5 mg of INO-5150 + 1 mg of INO-9012. Patients received study drug with electroporation on day 0 and on weeks 3, 12, and 24, and they were followed for up to 72 weeks. Sixty-two patients were enrolled. Treatment was well tolerated. 81% (50/62) of patients completed all visits. 85% (53/62) remained progression-free at 72 weeks. PSA doubling time (PSADT) was increased when assessed in patients with day 0 PSADT ≤12 months. Immunogenicity was observed in 76% (47/62) of patients by multiple assessments. Analysis indicated that CD38 and perforin co-positive CD8 T cell frequency correlated with attenuated PSA rise (p = 0.05, n = 50).

Bovine papillomavirus prostate cancer antigen virus-like particle vaccines are efficacious in advanced cancers in the TRAMP mouse spontaneous prostate cancer model.

Prostate cancer is a candidate for immunotherapy because cancer cells express tissue-specific proteins that can be therapeutic targets. However, immune checkpoint inhibitors and active immunization have performed poorly in clinical trials. We developed a novel virus-like particle (VLP) vaccine composed of bovine papillomavirus L1 protein engineered to display surface docking sites. We decorated VLPs with peptides encoding T cell epitopes from two prostate cancer-associated tumor antigens, prostate stem cell antigen (PSCA), and prostatic acid phosphatase (PAP-1 and PAP-2), and a neo-antigen, stimulator of prostatic adenocarcinoma-specific T cells (SPAS-1). The VLP vaccines induced a mean frequency of antigen-specific IFN-γ secreting CD8 + T cells of 2.9% to PSCA, 9.5% to SPAS-1, 0.03% to PAP-1, and 0.03% to PAP-2 in tumor-bearing TRAMP mice. We treated TRAMP mice at 19-20 weeks of age, when mice have advanced stages of carcinogenesis, with either VLP vaccine, anti-PD1 antibody, or combination immunotherapy. The VLP vaccine alone or in combination with anti-PD1 antibody significantly reduced tumor burden, while anti-PD1 antibody had a modest non-significant therapeutic effect. All treatments significantly increased CD3 + and CD8 + T cell infiltration into tumor tissue compared to control mice, and combination therapy resulted in significantly greater CD3 + and CD8 + T cell infiltration than monotherapy. Reduction in tumor burden in vaccine-treated mice was inversely correlated with CD8 + T cell numbers in tumor tissue. No other immunotherapy has shown efficacy in this animal model of advanced prostate cancer, making bovine papillomavirus VLPs an attractive vaccine technology to test in patients with metastatic prostate cancer.

Integrated Single Microbead-Arrayed µ-Fluidic Platform for the Automated Detection of Multiplexed Biomarkers.

An automated, single microbead-arrayed µ-fluidic immunoassay (AMIA) device is innovatively devised in this study, which enables the highly sensitive and simultaneous detection of multiplex biomarkers with fully automatic operations. The AMIA platform not only achieves automated assay processing and multiplexed target detection by integrating single microbead manipulation, sample loading, multistep washing, and immunoreaction on a microfluidic chip but also confers high sensitivity due to the highly efficient signal enriching effect on a single microbead by the use of only a routine sandwich immunoreaction. As such, as low as the pg/mL level of multiplexed protein biomarkers can be simultaneously determined in a quite small volume of serum (∼20 µL is enough), which can well meet the clinical demand for disease screening and prognosis. What is more, the detection results of several clinically important biomarkers in clinical samples with the AMIA platform exhibit excellent consistency with those obtained by using a standard clinical test. Thus, in virtue of the excellent features in terms of high sensitivity, multiplexing capability, generality, and high degree of automation, the AMIA provides a practical and user-friendly platform for assaying different biomarkers in clinical diagnostics and point-of-care testing.

Ultrasensitive amperometric immunosensor for the prostate specific antigen by exploiting a Fenton reaction induced by a metal-organic framework nanocomposite of type Au/Fe-MOF with peroxidase mimicking activity.

To increase the sensitivity of electrochemical sensor, Fe-MIL-88B-NH2 (Fe-MOF) with peroxidase-like activity is designed for the construction of immunoprobe. The Fe-MOF was prepared by one-step hydrothermalf method using 2-aminoterephthalic acid and iron(III) chloride. For the immunoprobe, it was fabricated by gold nanocomposite/Fe-MOF (Au/Fe-MOF) for the immobilization of labeling antibody (the antibody was used to conjuncting with label materials). The thin layer of Methylene Blue (MB) covered by reduced graphene oxide-gold nanocomposites (Au-rGO) serves as a substrate to covalently fix coating antibodies. The MB as a redox-active species was modified on the glass carbon electrode that can give a strong amperometric signal at 0.18 V (vs. Ag/AgCl). With the participation of H2O2, Fe-MOF can induce the Fenton reaction which degrades MB covered by Au-rGO on the substrate. The rest of MB on the surface of electrode becomes oxidized thereby generating a current signal. Square wave voltammetry (SWV) was used to quantify PSA. Under optimal conditions, the immunoassay is stable, specific and reproducible. It has a lower detection limit of 0.13 pg mL-1 (S/N = 3) and a wide analytical range that extends from 0.001 to 100 ng mL-1. Graphical abstractA sandwich-type amperometric immunoassay based on Fe-MOF-induced Fenton reaction was designed for sensitive determination of prostate specific antigen.