Prostate-specific membrane antigen (PSMA) targeted singlet oxygen delivery via endoperoxide tethered ligands.

Singlet oxygen is the primary agent responsible for the therapeutic effects of photodynamic therapy (PDT). In this work, we demonstrate that singlet oxygen release due to thermal endoperoxide cycloreversion can be targeted towards specific features of selected cancer cells, and this targeted singlet oxygen delivery can serve as an effective therapeutic tool. Thus, cytotoxic singlet oxygen can be delivered regioselectively into prostate specific membrane antigen (PSMA) overexpressing lymph node carcinoma (LNCaP) cells. However, unlike typical photodynamic processes, there is no need for light or oxygen. The potential of the approach is exciting, considering the limitations on the availability of light and oxygen in deep-seated tumors.

Biodistribution and internal radiation dosimetry of a companion diagnostic radiopharmaceutical, [68Ga]PSMA-11, in subcutaneous prostate cancer xenograft model mice.

[68Ga]PSMA-11 is a prostate-specific membrane antigen (PSMA)-targeting radiopharmaceutical for diagnostic PET imaging. Its application can be extended to targeted radionuclide therapy (TRT). In this study, we characterize the biodistribution and pharmacokinetics of [68Ga]PSMA-11 in PSMA-positive and negative (22Rv1 and PC3, respectively) tumor-bearing mice and subsequently estimated its internal radiation dosimetry via voxel-level dosimetry using a dedicated Monte Carlo simulation to evaluate the absorbed dose in the tumor directly. Consequently, this approach overcomes the drawbacks of the conventional organ-level (or phantom-based) method. The kidneys and urinary bladder both showed substantial accumulation of [68Ga]PSMA-11 without exhibiting a washout phase during the study. For the tumor, a peak concentration of 4.5 ± 0.7 %ID/g occurred 90 min after [68Ga]PSMA-11 injection. The voxel- and organ-level methods both determined that the highest absorbed dose occurred in the kidneys (0.209 ± 0.005 Gy/MBq and 0.492 ± 0.059 Gy/MBq, respectively). Using voxel-level dosimetry, the absorbed dose in the tumor was estimated as 0.024 ± 0.003 Gy/MBq. The biodistribution and pharmacokinetics of [68Ga]PSMA-11 in various organs of subcutaneous prostate cancer xenograft model mice were consistent with reported data for prostate cancer patients. Therefore, our data supports the use of voxel-level dosimetry in TRT to deliver personalized dosimetry considering patient-specific heterogeneous tissue compositions and activity distributions.

Darolutamide Potentiates the Antitumor Efficacy of a PSMA-targeted Thorium-227 Conjugate by a Dual Mode of Action in Prostate Cancer Models.

PURPOSE: Androgen receptor (AR) inhibitors are well established in the treatment of castration-resistant prostate cancer and have recently shown efficacy also in castration-sensitive prostate cancer. Although most patients respond well to initial therapy, resistance eventually develops, and thus, more effective therapeutic approaches are needed. Prostate-specific membrane antigen (PSMA) is highly expressed in prostate cancer and presents an attractive target for radionuclide therapy. Here, we evaluated the efficacy and explored the mode of action of the PSMA-targeted thorium-227 conjugate (PSMA-TTC) BAY 2315497, an antibody-based targeted alpha-therapy, in combination with the AR inhibitor darolutamide. EXPERIMENTAL DESIGN: The in vitro and in vivo antitumor efficacy and mode of action of the combination treatment were investigated in preclinical cell line-derived and patient-derived prostate cancer xenograft models with different levels of PSMA expression. RESULTS: Darolutamide induced the expression of PSMA in androgen-sensitive VCaP and LNCaP cells in vitro, and the efficacy of darolutamide in combination with PSMA-TTC was synergistic in these cells. In vivo, the combination treatment showed synergistic antitumor efficacy in the low PSMA-expressing VCaP and in the high PSMA-expressing ST1273 prostate cancer models, and enhanced efficacy in the enzalutamide-resistant KUCaP-1 model. The treatments were well tolerated. Mode-of-action studies revealed that darolutamide induced PSMA expression, resulting in higher tumor uptake of PSMA-TTC, and consequently, higher antitumor efficacy, and impaired PSMA-TTC-mediated induction of DNA damage repair genes, potentially contributing to increased DNA damage. CONCLUSIONS: These results provide a strong rationale to investigate PSMA-TTC in combination with AR inhibitors in patients with prostate cancer.

Gliclazide alters macrophages polarization state in diabetic atherosclerosis in vitro via blocking AGE-RAGE/TLR4-reactive oxygen species-activated NF-kß nexus.

Hyperglycemic milieu in diabetes mellitus stimulates macrophages for exaggerated pro-inflammatory cytokine response, particularly IL-1ß, IL-6, and TNF-α. Although hyperglycemia causes macrophages to produce pro-inflammatory cytokines, AGEs (advanced glycation end products) active inflammation, produced as a result of chronic hyperglycemia, inducers cause polarization of macrophages into pro-inflammatory M1 phenotype. AGEs in diabetes accelerate atherosclerotic plaque initiation and progression via promoting macrophages polarization towards pro-inflammatory state. Gliclazide (Glz) is a well known antidiabetic drug with excellent safety profile. Its repurposing in the management of diabetes-associated late complications has tremendous merit. The present study demonstrated that Glz retards diabetic atherosclerotic progression, and cytokines storm in a concentration dependent manner over a concentration range of 1-100 µM than those of AGEs (200 µg/ml)-treated cells through a mechanism that alters macrophage M1 polarization state. Glz exerted these beneficial effects, independent of its antidiabetic effect. Glz pretreatment significantly (P < 0.05) inhibited the AGEs-induced pro-inflammatory mediators (NO•, reactive oxygen species, i-NOS), and production of pro-inflammatory cytokines, including IL-1ß, IL-6, and TNF-α. It also significantly (P < 0.05) promoted the production of anti-inflammatory cytokines (IL-10 and TGF-ß) in RAW 264.7 mouse macrophages. Glz pretreatment also effectively abated the AGEs-induced RAGE (~2-fold decrease), and CD86 surface marker expressions (P < 0.001 at 100 µM) on macrophages by inhibiting the NF-kß activation in a concentration dependent manner (1-100 µM) (P < 0.001). In conclusion, our data demonstrates that Glz alleviates the diabetic atherosclerosis progression by ameliorating the AGEs-mediated M1 pro-inflammatory phenotype via blocking AGE-RAGE/TLR4-reactive oxygen species -activated NF-kß nexus in macrophages.

Immunomodulatory Effect of a Salvia plebeia R. Aqueous Extract in Forced Swimming Exercise-induced Mice.

This study investigated the immunomodulatory effect of Salvia plebeia R. aqueous extract (FIE-SP, SPW) in forced swimming exercise-induced mice and the immunostimulatory effects on Raw264.7 cells. Mice were randomly assigned to four groups: the control group (CON), the forced swimming test group (FST), and two FIE-SP groups (low and high dose of FIE-SP). Compared with the control group, the FIE-SP groups showed significantly increased ratios of T lymphocyte surface markers CD4+/CD8+ and major histocompatibility complex (MHC)I/MHCII, as well as increased concentrations of immunoglobulin (Ig)A and IgG. FIE-SP groups significantly increased Th1 cytokines and decreased Th2 cytokines compared with negative control exercise-induced mice. Conversely, the immunostimulatory effects of FIE-SP significantly increased phagocytic activities, nitric oxide (NO) production, and pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin (IL)-1ß in Raw264.7 cells. Furthermore, FIE-SP increased natural killer (NK) cell activities and cytokines (IL-12) in splenocytes compared with the CON group. These results indicated that FIE-SP supplementation could prevent imbalanced immune states and produce immunostimulatory effects to support innate immunity.

Long-Circulating Prostate-Specific Membrane Antigen-Targeted NIR Phototheranostic Agent.

Targeted photodynamic therapy (PDT) combined with image-guided surgical resection is a promising strategy for precision cancer treatment. Prostate-specific membrane antigen (PSMA) is an attractive target due to its pronounced overexpression in a variety of tumors, most notably in prostate cancer. Recently, we reported a pyropheophorbide-based PSMA-targeted agent, which exhibited long plasma circulation time and effective tumor accumulation. To further advance PSMA-targeted photodynamic therapy by harvesting tissue-penetrating properties of the NIR light, we developed a bacteriochlorophyll-based PSMA-targeted photosensitizer (BPP), consisting of three building blocks: (1) a PSMA-affinity ligand, (2) a peptide linker to prolong plasma circulation time and (3) a bacteriochlorophyll photosensitizer for NIR fluorescence imaging and photodynamic therapy (Qy absorption maximum at 750 nm). BPP exhibited excellent PSMA-targeting selectivity in both subcutaneous and orthotopic mouse models. The nine D-peptide linker in BPP structure prolonged its plasma circulation time (12.65 h). Favorable pharmacokinetic properties combined with excellent targeting selectivity enabled effective BPP tumor accumulation, which led to effective PDT in a subcutaneous prostate adenocarcinoma mouse model. Overall, bright NIR fluorescence of BPP enables effective image guidance for surgical resection, while the combination of its targeting capabilities and PDT activity allows for potent and precise image-guided photodynamic treatment of PSMA-expressing tumors.

Association of Acute, High-dose Cadmium Exposure with Alterations in Vascular Endothelial Barrier Antigen Expression and Astrocyte Morphology in the Developing Rat Central Nervous System.

Clinical and experimental studies have demonstrated the neurotoxic and behavioural effects of cadmium. However, the exact pathophysiological mechanism(s) of cadmium neurotoxicity on the human central nervous system (CNS) is not completely understood. A rat blood-brain barrier (BBB) endothelial marker, the endothelial barrier antigen (EBA), has been identified and we have shown previously that an anti-EBA IgG1 antibody exclusively recognizes barrier-competent microvessels in the rat CNS and peripheral nervous system (PNS). Endothelial cells of peripheral tissues or brain regions possessing fenestrated microvascular endothelia do not display immunoreactivity for EBA. Here, we describe the application of sequential indirect immunofluorescence with anti-EBA, and an antibody directed against glial fibrillary acidic protein (GFAP), to evaluate the immunoreactivity patterns and morphological alterations in BBB microvessels and astrocytes, following a single, high dose of cadmium in normal, term-delivered young rats. We detected a moderate reduction in immunoreactivity and number of microvessels labelled by the anti-EBA in the forebrain, cerebellum and midbrain in cadmium-exposed rats compared with normal controls. We observed weakly GFAP-reactive astrocytes displaying cell bodies with ill-defined borders and blurred cytoplasm within the white and grey matter of cadmium-exposed brains. The astrocyte nuclei were markedly enlarged, intensely hyperchromatic and exhibited chromatin condensation with nuclear fragmentation. This study indicates for the first time that EBA is involved in, and could serve as a potentially useful marker for studying, cadmium neurotoxicity in the rat model system.

EZH2 Inhibition in Ewing Sarcoma Upregulates GD2 Expression for Targeting with Gene-Modified T Cells.

Chimeric antigen receptor (CAR) engineering of T cells allows one to specifically target tumor cells via cell surface antigens. A candidate target in Ewing sarcoma is the ganglioside GD2, but heterogeneic expression limits its value. Here we report that pharmacological inhibition of Enhancer of Zeste Homolog 2 (EZH2) at doses reducing H3K27 trimethylation, but not cell viability, selectively and reversibly induces GD2 surface expression in Ewing sarcoma cells. EZH2 in Ewing sarcoma cells directly binds to the promoter regions of genes encoding for two key enzymes of GD2 biosynthesis, and EZH2 inhibition enhances expression of these genes. GD2 surface expression in Ewing sarcoma cells is not associated with distinct in vitro proliferation, colony formation, chemosensitivity, or in vivo tumorigenicity. Moreover, disruption of GD2 synthesis by gene editing does not affect its in vitro behavior. EZH2 inhibitor treatment sensitizes Ewing sarcoma cells to effective cytolysis by GD2-specific CAR gene-modified T cells. In conclusion, we report a clinically applicable pharmacological approach for enhancing efficacy of adoptively transferred GD2-redirected T cells against Ewing sarcoma, by enabling recognition of tumor cells with low or negative target expression.

Targeting surface nucleolin induces autophagy-dependent cell death in pancreatic cancer via AMPK activation.

Pancreatic cancer remains one of the deadliest human cancers despite current advances in conventional therapeutics including surgery and adjuvant therapies. Here, we showed that LZ1, a peptide derived from a snake venom cathelicidin, significantly inhibited growth of pancreatic cancer cells by inducing autophagy-dependent cell death both in vitro and in vivo. The LZ1-induced cell death was blocked by pharmacological or genetic inhibition of autophagy. In orthotopic model of pancreatic cancer, systemic administration of LZ1 (1-4 mg/kg) exhibited remarkable antitumor efficacy, significantly prolonged mice survival, and showed negligible adverse effects by comparison with gemcitabine (20 mg/kg). Mechanistic studies revealed that LZ1 acts through binding to nucleolin, whose expression on cell surface is frequently increased in pancreatic cancer cells. LZ1 binding triggers degradation of surface-expressed nucleolin. This leads to activation of 5'-AMP kinase which results in suppression of mTORC1 activity and induction of autophagic flux. These data suggest that LZ1, targeting nucleolin-AMPK-autophagy axis, is a promising lead for the development of therapeutic agents against pancreatic cancer.

Targeting PFKFB3 sensitizes chronic myelogenous leukemia cells to tyrosine kinase inhibitor.

Resistance to the BCR-ABL tyrosine kinase inhibitor (TKI) remains a challenge for curing the disease in chronic myeloid leukemia (CML) patients as leukemia cells may survive through BCR-ABL kinase activity-independent signal pathways. To gain insight into BCR-ABL kinase activity-independent mechanisms, we performed an initial bioinformatics screen and followed by a quantitative PCR screen of genes that were elevated in CML samples. A total of 33 candidate genes were identified to be highly expressed in TKIs resistant patients. Among those genes, 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), controlling the limiting step of glycolysis, was found to be strongly associated with TKIs resistance. PFKFB3 knockdown or pharmacological inhibition of its kinase activity markedly enhanced the sensitivity of CML cells to TKIs. Furthermore, pharmacological inhibition of PFKFB3 inhibited CML cells growth and significantly prolonged the survival of both allograft and xenograft CML mice. ChIP-seq data analysis combined with subsequent knockdown experiment showed that the Ets transcription factor PU.1 regulated the elevated expression of PFKFB3 in TKIs-resistant CML cells. Therefore, our results showed that targeting PFKFB3 sensitizes CML cells to TKIs and PFKFB3 may be a potential BCR-ABL kinase activity-independent mechanism in CML.

ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies: an infectious diseases perspective (Agents targeting lymphoid or myeloid cells surface antigens [II]: CD22, CD30, CD33, CD38, CD40, SLAMF-7 and CCR4).

BACKGROUND: The present review is part of the ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies. AIMS: To review, from an Infectious Diseases perspective, the safety profile of agents targeting CD22, CD30, CD33, CD38, CD40, SLAMF-7 and CCR4 and to suggest preventive recommendations. SOURCES: Computer-based MEDLINE searches with MeSH terms pertaining to each agent or therapeutic family. CONTENT: The risk and spectrum of infections in patients receiving CD22-targeted agents (i.e. inotuzumab ozogamicin) are similar to those observed with anti-CD20 antibodies. Anti-Pneumocystis prophylaxis and monitoring for cytomegalovirus (CMV) infection is recommended for patients receiving CD30-targeted agents (brentuximab vedotin). Due to the scarcity of data, the risk posed by CD33-targeted agents (gemtuzumab ozogamicin) cannot be assessed. Patients receiving CD38-targeted agents (i.e. daratumumab) face an increased risk of varicella-zoster virus (VZV) infection. Therapy with CD40-targeted agents (lucatumumab or dacetuzumab) is associated with opportunistic infections similar to those observed in hyper-IgM syndrome, and prevention strategies (including anti-Pneumocystis prophylaxis and pre-emptive therapy for CMV infection) are warranted. SLAMF-7 (CD319)-targeted agents (elotuzumab) induce lymphopenia and increase the risk of infection (particularly due to VZV). The impact of CCR4-targeted agents (mogamulizumab) on infection susceptibility is difficult to distinguish from the effect of underlying diseases and concomitant therapies. However, anti-Pneumocystis and anti-herpesvirus prophylaxis and screening for chronic hepatitis B virus (HBV) infection are recommended. IMPLICATIONS: Specific management strategies should be put in place to reduce the risk and/or the severity of infectious complications associated to the reviewed agents.

ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies: an infectious diseases perspective (Agents targeting lymphoid cells surface antigens [I]: CD19, CD20 and CD52).

BACKGROUND: The present review is part of the ESCMID Study Group for Infections in Compromised Hosts (ESGICH) Consensus Document on the safety of targeted and biological therapies. AIMS: To review, from an Infectious Diseases perspective, the safety profile of agents targeting CD19, CD20 and CD52 and to suggest preventive recommendations. SOURCES: Computer-based MEDLINE searches with MeSH terms pertaining to each agent or therapeutic family. CONTENT: Although CD19-targeted agents (blinatumomab or inebilizumab) are not associated with an increased risk of infection, they may cause IgG hypogammaglobulinaemia and neutropenia. The requirement for prolonged intravenous infusion of blinatumomab may increase the risk of catheter-associated bloodstream infections. Infection remains the most common non-haematological adverse effect of anti-CD20 monoclonal antibodies, including severe respiratory tract infection, hepatitis B virus (HBV) reactivation and varicella-zoster virus infection. Screening for chronic or resolved HBV infection is recommended for patients receiving anti-CD20 monoclonal antibodies. Antiviral prophylaxis should be offered for 12-18 months to hepatitis B surface antigen (HBsAg)-positive and HBsAg-negative/anti-hepatitis B core antibody (HBc)-positive patients. Anti-Pneumocystis prophylaxis should be considered in patients receiving concomitant chemotherapy, particularly steroids. Alemtuzumab (anti-CD52) increases the risk of infections, in particular among leukaemia and solid organ transplant patients. These populations benefit from anti-Pneumocystis prophylaxis, prevention strategies for cytomegalovirus infection, and screening for HBV, hepatitis C virus and tuberculosis. Antiviral prophylaxis for at least 6-12 months should be provided for HBsAg-positive patients. IMPLICATIONS: As there are limited clinical data for many of the reviewed agents, special attention must be given to promptly detect and report emerging infectious complications.

Mesenchymal stem cells show functional defect and decreased anti-cancer effect after exposure to chemotherapeutic drugs.

BACKGROUND: Mesenchymal stem cells (MSC) are used for several therapeutic applications to improve the functions of bone, cardiac, nervous tissue as well as to facilitate the repopulation of hematopoietic stem cells. MSC give rise to the non-hematopoietic stromal cells of the bone marrow and are important for the maintenance of normal hematopoiesis. Chemotherapeutic drugs used for treatment of leukemia extensively damage the stromal cells and alter their gene expression profiles. METHODS: We determined the changes in adipogenic, osteogenic differentiation, phenotypic and gene expression in MSC during treatment with chemotherapeutic drugs cytarabine, daunorubicin and vincristine. We also tested anti-cancer effects of drug treated MSC on leukemia cells. RESULTS: Treatment with the chemotherapeutic drugs resulted in functional defects in MSC, leading to reduced proliferation, osteogenic and adipogenic differentiation. The drug treated MSC also showed decreased expression of cell surface receptors, and the changes in proliferation, phenotype and differentiation defect was partially reversible after withdrawing the drugs from the cells. The drug treated MSC showed increased expression of cytokines, IL6, FGF2 and TNFA but reduced levels of differentiation markers SOX9 and ACTC1. Drug treated MSC also contributed to reduced anti-cancer effects in leukemia cells. CONCLUSIONS: Chemotherapeutic drug treatment altered the phenotype, osteogenic and adipogenic differentiation potential of MSC and modified the gene expression profile of the cells to render them more chemoprotective of the leukemic cells. Thus, additional therapeutic efforts to target the stromal cell population will help in preventing chemoresistance, disease relapse in leukemia and to maintain a healthy bone marrow stroma.

STRO-1 confers myofibroblast transdifferentiation in fibroblasts derived from oral submucous fibrosis.

BACKGROUND: STRO-1 is a mesenchymal cell marker present on all clonogenic stromal precursors. Current evidence has indicated that the pathogenesis of fibrotic changes may be mediated by stemness properties. The aim of this study was to investigate the role of STRO-1 in areca quid chewing-associated oral submucous fibrosis (OSF). METHODS: Thirty OSF specimens and ten normal buccal mucosae were examined by immunohistochemistry. The activity of STRO-1 from fibroblasts cultured from normal buccal mucosa (BMFs) and OSF (OSFFs) were measureed and the effect of arecoline, a major areca nut alkaloid, on STRO-1 in BMFs was evaluated. Compared the activities between sorted STRO-1+ cells and STRO-1- cells from OSFF were measured by collagen gel contraction, migration, invasion abilities, and the expression of α-smooth muscle actin (α-SMA) and pro-α1 (I) chain of type I collagen. RESULTS: Our results first showed that the expression of STRO-1 was more evident in areca quid chewing-associated OSF than normal buccal mucosa tissues (P < .05). Arecoline dose-dependently activated the level of STRO-1 in BMFs (P < .05). The relative expression of STRO-1 was significantly higher in OSFFs compared with BMFs (P < .05). In addition, the sorted STRO-1+ cells from OSFFs exhibited higher collagen gel contraction, migration, and invasion abilities as well as elevated expression of α-SMA and pro-α1 (I) chain of type I collagen than the negative subset (P < .05). CONCLUSION: These findings suggested that the stemness marker STRO-1 may be a crucial factor in the pathogenesis of areca quid chewing-associated OSF.

Effect of bleaching agent extracts on murine macrophages.

OBJECTIVES: The aim of this study was to evaluate the cytotoxicity and the influence of bleaching agents on immunologically cell surface antigens of murine macrophages in vitro. MATERIALS AND METHODS: RAW 264.7 cells were exposed to bleaching gel extracts (40% hydrogen peroxide or 20% carbamide peroxide) and different H2O2 concentrations after 1 and 24-h exposure periods and 1-h exposure and 23-h recovery. Tests were performed with and without N-acetyl cysteine (NAC) and buthionine sulfoximine (BSO). Cell viability was determined by MTT assay. The expression of surface markers CD14, CD40, and CD54 with and without LPS stimulation was detected by flow cytometry, while the production of TNF-α was measured by ELISA. Statistical analysis was performed using the Mann-Whitney U test (α = 0.05). RESULTS: Extracts of bleaching agents were cytotoxic for cells after a 1-h exposure; cells could not recover after 24 h. This effect can be mitigated by the antioxidant NAC and increased by BSO, an inhibitor of glutathione (GSH) synthesis. LPS stimulated expression of all surface markers and TNF-α production. Exposure to bleaching agent extracts and H2O2 leads to a reduction of TNF-α, CD14, and CD40 expression, while the expression of CD54 was upregulated at non-cytotoxic concentrations. Whereas NAC reduced this effect, it was increased in the presence of BSO. CONCLUSIONS: Extracts of bleaching agents were irreversibly cytotoxic to macrophages after a 1-h exposure. Only the expression of CD54 was upregulated. The reactions are mediated by the non-enzymatic antioxidant GSH. CLINICAL RELEVANCE: The addition of an antioxidant can downregulate unfavorable effects of dental bleaching.

Efficacy Against Human Prostate Cancer by Prostate-specific Membrane Antigen-specific, Transforming Growth Factor-ß Insensitive Genetically Targeted CD8+ T-cells Derived from Patients with Metastatic Castrate-resistant Disease.

Current immunotherapy has limited efficacy on metastatic castrate-resistant prostate cancer (mCRPC). We therefore sought to improve the antitumor ability of mCRPC patient-derived CD8+ T-cells by the endowment of specificity to prostate-specific membrane antigen (PSMA) and insensitivity to immunosuppressant molecule transforming growth factor-ß (TGF-ß) under the control of herpes simplex virus-1 thymidine kinase. CD8+ T-cells were collected by leukapheresis and cultured in a Food and Drug Administration-approved Cell Processing Work Station. We developed a chimeric antigen receptor retroviral construct using an anti-PSMA chimeric immunoglobulin-T-cell receptor(ζ) gene (PZ1) and dominant negative TGF-ß type II receptor (TßRIIDN), that could induce CD8+ T-cells to be PSMA reactive and insensitive to TGF-ß. Cr51 release assay was performed on PC-3 and PC-3-PSMA. The further antitumor functions of PSMA-specific, TGF-ß insensitive CD8+ T-cells was evaluated using an immunodeficient RAG-1-/- mouse model. We found PSMA-specific, TGF-ß insensitive CD8+ T-cells from mCRPC were expanded with strong expression of PZ1 and thymidine kinase genes, and their growth was not suppressed by TGF-ß. The survival of these cells decreased sharply after treatment with ganciclovir. Treatment of PSMA-specific TGF-ß, insensitive CD8+ T-cells was associated with 61.58% specific lysis on PC-3-PSMA, and significantly suppressed PC3-PSMA tumor compared with the PC3 tumor. A large amount of tumor apoptosis and CD8+ T-cell infiltration were found only in the PC3-PSMA tumor. This study verified that PSMA-specific, TGF-ß insensitive CD8+ T-cells derived from mCRPC patients could be successfully expanded and used to overcome the immunosuppressive effects of the tumor microenvironment to control PSMA-expressing PC in vitro and in vivo. This may provide a promising approach for men with mCRPC who fail androgen deprivation therapy. PATIENT SUMMARY: We investigated the role of a novel chimeric antigen receptor T-immunotherapy based on autologous metastatic castrate-resistant prostate cancer patient-derived prostate-specific membrane antigen (PSMA)-specific, transforming growth factor-ß insensitive CD8+ T-cells on PSMA-positive prostate cancer. We found that this chimeric antigen receptor T-cells could kill PSMA-positive prostate cancer specifically. The results suggest that this novel immunotherapy treatment is a potential new approach for men with metastatic castrate-resistant prostate cancer.

Translation and Clinical Development of Bispecific T-cell Engaging Antibodies for Cancer Treatment.

Bispecific T-cell Engagers (BiTE®) antibody constructs enable a polyclonal T-cell response to cell-surface tumor-associated antigens, bypassing the narrow specificities of T-cell receptors and the need for antigen presentation through the major histocompatibility complex pathways. Blinatumomab, a CD19xCD3 BiTE® antibody construct, received accelerated approval for the treatment of relapsed/refractory Philadelphia chromosome negative acute lymphoblastic leukemia. Herein we review the pharmacology, safety, and efficacy observed in studies of blinatumomab and other BiTE® antibody constructs. Quantitative systems pharmacology is envisioned as a means to optimize dosing decisions for trials in which BiTE® antibody constructs are administered as monotherapy or in combination with other immunotherapies.

Evaluation of Polymeric Nanomedicines Targeted to PSMA: Effect of Ligand on Targeting Efficiency.

Targeted nanomedicines offer a strategy for greatly enhancing accumulation of a therapeutic within a specific tissue in animals. In this study, we report on the comparative targeting efficiency toward prostate-specific membrane antigen (PSMA) of a number of different ligands that are covalently attached by the same chemistry to a polymeric nanocarrier. The targeting ligands included a small molecule (glutamate urea), a peptide ligand, and a monoclonal antibody (J591). A hyperbranched polymer (HBP) was utilized as the nanocarrier and contained a fluorophore for tracking/analysis, whereas the pendant functional chain-ends provided a handle for ligand conjugation. Targeting efficiency of each ligand was assessed in vitro using flow cytometry and confocal microscopy to compare degree of binding and internalization of the HBPs by human prostate cancer (PCa) cell lines with different PSMA expression status (PC3-PIP (PSMA+) and PC3-FLU (PSMA-). The peptide ligand was further investigated in vivo, in which BALB/c nude mice bearing subcutaneous PC3-PIP and PC3-FLU PCa tumors were injected intravenously with the HBP-peptide conjugate and assessed by fluorescence imaging. Enhanced accumulation in the tumor tissue of PC3-PIP compared to PC3-FLU highlighted the applicability of this system as a future imaging and therapeutic delivery vehicle.

Effects of four chemotherapeutic agents, bleomycin, etoposide, cisplatin, and cyclophosphamide, on DNA damage and telomeres in a mouse spermatogonial cell line.

Treatment with chemotherapeutics agents may induce persistent DNA damage in male germ cells with the possibility of long-term consequences on fertility and progeny outcome. Telomeres, specialized structures at the physical ends of chromosomes, play an important role in the maintenance of genetic stability and in the response of somatic cells to anticancer drugs. Our objective was to test the hypothesis that exposure to bleomycin, etoposide, or cisplatin (the drugs used to treat testicular cancer) or cyclophosphamide (an anticancer agent and immunosuppressant) targets telomeres in the male germ line. C18-4 spermatogonial cells were exposed to bleomycin, etoposide, cisplatin, or 4-hydroperoxycyclophosphamide (4OOH-CPA, a preactivated analog of cyclophosphamide). All four anticancer drugs induced a significant increase in DNA damage in C18-4 cells, as assessed by gamma-H2AX immunofluorescence. Interestingly, the gamma-H2AX signal was localized to telomeres after treatment with bleomycin, cisplatin, and 4OOH-CPA, but not etoposide. Mean telomere lengths, the intensity of the telomere fluorescence in situ hybridization signal, telomerase activity, and the expression of the telomerase enzyme mRNA components, Tert and Terc, were reduced by exposure to cisplatin and 4OOH-CPA, but not by bleomycin or etoposide. Thus, although all four anticancer drugs induced DNA damage in this spermatogonial cell line, telomeres were not specifically affected by etoposide and only the two alkylating agents, cisplatin and 4OOH-CPA, induced telomere dysfunction. This telomere dysfunction may contribute to infertility and developmental defects in the offspring.

Potential therapeutic use of herbal extracts in trypanosomiasis.

The aim of the present study was to evaluate the effects of crude extracts from Handroanthus impetiginosa, Ageratum conyzoides, and Ruta graveolens on Leishmania amazonensis and Trypanosoma cruzi infection in vitro. The results showed that the extracts caused significant toxicity in promastigotes and trypomastigotes. A significant decrease in the rate of cell invasion by pretreated trypomastigotes and promastigotes was also observed. The extracts caused a significant reduction of the multiplication of intracellular amastigotes of both parasites. Therefore, these herbal extracts may be potential candidates for the development of drugs for the treatment of leishmaniasis and Chagas disease.