Targeting both BDNF/TrkB pathway and delta-secretase for treating Alzheimer's disease.

Alzheimer's disease (AD) is the most common dementia, and no disease-modifying therapeutic agents are currently available. BDNF/TrkB signaling is impaired in AD and is associated with prominent delta-secretase (δ-secretase, also known as asparaginyl endopeptidase or legumain) activation, which simultaneously cleaves both APP and Tau and promotes Aß production and neurofibrillary tangles (NFT) pathologies. Here we show that the optimized δ-secretase inhibitor (#11a) or TrkB receptor agonist (CF3CN) robustly blocks δ-secretase activity separately, and their combination synergistically blunts δ-secretase, exhibiting promising therapeutic efficacy in 3xTg AD mouse model. The optimal δ-secretase inhibitor reveals demonstrable brain exposure and oral bioavailability, suppressing APP N585 and Tau N368 cleavage by δ-secretase. Strikingly, CF3CN treatment evidently escalates BDNF levels. Both #11a and CF3CN display strong in vivo PK/PD properties and ability to suppress δ-secretase activity in the brain. Orally administrated CF3CN strongly activates TrkB that triggers active Akt to phosphorylate δ-secretase T322, preventing its proteolytic activation and mitigating AD pathologies. #11a or CF3CN significantly diminishes AD pathogenesis and improves cognitive functions with the combination exhibiting the maximal effect. Thus, our data support that these derivatives are strong pharmaceutical candidates for the treatment of AD.

Cell Surface GRP94 as a Novel Emerging Therapeutic Target for Monoclonal Antibody Cancer Therapy.

Glucose-regulated protein 94 (GRP94) is an endoplasmic reticulum (ER)-resident member of the heat shock protein 90 (HSP90) family. In physiological conditions, it plays a vital role in regulating biological functions, including chaperoning cellular proteins in the ER lumen, maintaining calcium homeostasis, and modulating immune system function. Recently, several reports have shown the functional role and clinical relevance of GRP94 overexpression in the progression and metastasis of several cancers. Therefore, the current review highlights GRP94's physiological and pathophysiological roles in normal and cancer cells. Additionally, the unmet medical needs of small chemical inhibitors and the current development status of monoclonal antibodies specifically targeting GRP94 will be discussed to emphasize the importance of cell surface GRP94 as an emerging therapeutic target in monoclonal antibody therapy for cancer.

Novel ligands and modulators of triggering receptor expressed on myeloid cells receptor family: 2015-2020 updates.

Introduction: Triggering receptors expressed on myeloid cells (TREMs) are inflammatory amplifiers with defined pathophysiological role in various infectious diseases, acute and chronic aseptic inflammations, and a variety of cancers, depicting TREMs as prominent therapeutic targets.Areas covered: Herein, updates from 2015 to 2020 are discussed to divulge the TREM ligands, as well as their peptide blockers, claimed to modulate their expression. The article also presents different strategies employed during the last five years to block interactions between TREMs and their ligands to treat various disease conditions by modulating their expression and activity.Expert opinion: There has been significant progress in the discovery of novel ligands and modulators of TREMs in the last five years that mainly revolved around the function of TREM molecules. A few peptides showed encouraging results to modulate the expression and activity of TREMs in preclinical studies, and these peptides are currently under clinical investigation. Based on the findings so far in several careful studies, we expect novel therapeutics in the near future which could have the ability to treat various disease conditions associated with TREM expression.

Targeted Phototherapy for Malignant Pleural Mesothelioma: Near-Infrared Photoimmunotherapy Targeting Podoplanin.

Malignant pleural mesothelioma (MPM) has extremely limited treatment despite a poor prognosis. Moreover, molecular targeted therapy for MPM has not yet been implemented; thus, a new targeted therapy is highly desirable. Near-infrared photoimmunotherapy (NIR-PIT) is a recently developed cancer therapy that combines the specificity of antibodies for targeting tumors with toxicity induced by the photoabsorber after exposure to NIR-light. In this study, we developed a new phototherapy targeting podoplanin (PDPN) for MPM with the use of both NIR-PIT and an anti-PDPN antibody, NZ-1. An antibody-photosensitizer conjugate consisting of NZ-1 and phthalocyanine dye was synthesized. In vitro NIR-PIT-induced cytotoxicity was measured with both dead cell staining and luciferase activity on various MPM cell lines. In vivo NIR-PIT was examined in both the flank tumor and orthotopic mouse model with in vivo real-time imaging. In vitro NIR-PIT-induced cytotoxicity was NIR-light dose dependent. In vivo NIR-PIT led to significant reduction in both tumor volume and luciferase activity in a flank model (p < 0.05, NIR-PIT group versus NZ-1-IR700 group). The PDPN-targeted NIR-PIT resulted in a significant antitumor effect in an MPM orthotopic mouse model (p < 0.05, NIR-PIT group versus NZ-1-IR700 group). This study suggests that PDPN-targeted NIR-PIT could be a new promising treatment for MPM.

Osteoarthritis-associated basic calcium phosphate crystals alter immune cell metabolism and promote M1 macrophage polarization.

OBJECTIVE: A number of studies have demonstrated that molecules called 'alarmins' or danger-associated molecular patterns (DAMPs), contribute to inflammatory processes in the OA joint. Metabolic reprogramming of immune cells, including macrophages, is emerging as a prominent player in determining immune cell phenotype and function. The aim of this study was to investigate if basic calcium phosphate (BCP) crystals which are OA-associated DAMPs, impact on macrophage phenotype and metabolism. METHODS: Human monocyte derived macrophages were treated with BCP crystals and expression of M1 (CXCL9, CXCL10) and M2 (MRC1, CCL13)-associated markers was assessed by real-time PCR while surface maturation marker (CD40, CD80 & CD86) expression was assessed by flow cytometry. BCP induced metabolic changes were assessed by Seahorse analysis and glycolytic marker expression (hexokinase 2(HK2), Glut1 and HIF1α) was examined using real-time PCR and immunoblotting. RESULTS: Treatment with BCP crystals upregulated mRNA levels of CXCL9 and CXCL10 while concomitantly downregulating expression of CCL13 and MRC1. Furthermore, BCP-treated macrophages enhanced surface expression of the maturation makers, CD40, CD80 and CD86. BCP-treated cells also exhibited a shift towards glycolysis as evidenced by an increased ECAR/OCR ratio and enhanced expression of the glycolytic markers, HK2, Glut1 and HIF1α. Finally, BCP-induced macrophage activation and alarmin expression was reduced in the presence of the glycolytic inhibitor, 2-DG. CONCLUSIONS: This study not only provides further insight into how OA-associated DAMPs impact on immune cell function, but also highlights metabolic reprogramming as a potential therapeutic target for calcium crystal-related arthropathies.

Grp78 Loss in Epithelial Progenitors Reveals an Age-linked Role for Endoplasmic Reticulum Stress in Pulmonary Fibrosis.

Rationale: Alveolar epithelial cell (AEC) injury and dysregulated repair are implicated in the pathogenesis of pulmonary fibrosis. Endoplasmic reticulum (ER) stress in AEC has been observed in idiopathic pulmonary fibrosis (IPF), a disease of aging.Objectives: To investigate a causal role for ER stress in the pathogenesis of pulmonary fibrosis (PF) and therapeutic potential of ER stress inhibition in PF.Methods: The role of ER stress in AEC dysfunction and fibrosis was studied in mice with tamoxifen (Tmx)-inducible deletion of ER chaperone Grp78, a key regulator of ER homeostasis, in alveolar type II (AT2) cells, progenitors of distal lung epithelium, and in IPF lung slice cultures.Measurements and Main Results:Grp78 deletion caused weight loss, mortality, lung inflammation, and spatially heterogeneous fibrosis characterized by fibroblastic foci, hyperplastic AT2 cells, and increased susceptibility of old and male mice, all features of IPF. Fibrosis was more persistent in more severely injured Grp78 knockout (KO) mice. Grp78 KO AT2 cells showed evidence of ER stress, apoptosis, senescence, impaired progenitor capacity, and activation of TGF-ß (transforming growth factor-ß)/SMAD signaling. Glucose-regulated protein 78 is reduced in AT2 cells from old mice and patients with IPF, and ER stress inhibitor tauroursodeoxycholic acid ameliorates ER stress and fibrosis in Grp78 KO mouse and IPF lung slice cultures.Conclusions: These results support a causal role for ER stress and resulting epithelial dysfunction in PF and suggest ER stress as a potential mechanism linking aging to IPF. Modulation of ER stress and chaperone function may offer a promising therapeutic approach for pulmonary fibrosis.

Maternal exposure to cadmium impairs cognitive development of male offspring by targeting the Coronin-1a signaling pathway.

Direct exposure to cadmium (Cd) may induce persistent impairment in learning and memory. However, the outcomes of maternal exposure on the neurological development of offspring are much less clear, and the underlying mechanism leading to toxicity remains undisclosed. Following chronic exposure of female rats during gestation and lactation, low level of Cd was detectable in the cerebral cortex but not in the hippocampus of F1 male offspring. The synapses and neurites in hippocampus were destroyed by high Cd exposure level as evidenced by abnormal morphology and cognitive behavior deficit lasting from childhood to adulthood. The membrane glycoprotein M6a (GPM6A) regulates the filopodium formation, neurite outgrowth and synaptogenesis, and is a possible target which Cd acts upon. The signaling pathway Coronin-1a (CORO1A), Ras-related C3 botulinum toxin substrate 1 (RAC1) and p21-activated kinase 1 (PAK1) promotes GPM6A-induced filopodium formation. Our results showed that maternal exposure dramatically down-regulated the level of CORO1A as well as the expression of downstream effectors RAC1, PAK1 and GPM6A. CORO1A-knockdown by siRNA caused decreases in the expression of RAC1, PAK1 and GPM6A; and siRNA targeting combined with Cd insult further decreased the expression of these proteins. Following CORO1A overexpression, the neurites were lengthened with increased expression of all the effector proteins in SH-SY5Y cells exposed to Cd, confirming the significance of CORO1A in mediating the Cd neurotoxicity. These findings may help to disclose how Cd impairs the learning and cognitive development in children, and facilitate finding of potential therapeutic targets for the treatment of Cd poisoning.

TLR7 (Toll-Like Receptor 7) Facilitates Heme Scavenging Through the BTK (Bruton Tyrosine Kinase)-CRT (Calreticulin)-LRP1 (Low-Density Lipoprotein Receptor-Related Protein-1)-Hx (Hemopexin) Pathway in Murine Intracerebral Hemorrhage.

Background and Purpose- Heme and iron are considered to be key factors responsible for secondary insults after intracerebral hemorrhage (ICH). Our previous study showed that LRP1 (low-density lipoprotein receptor-related protein-1)-Hx (hemopexin) facilitates removal of heme. The TLR7 (Toll-like receptor 7)-BTK (Bruton tyrosine kinase)-CRT (calreticulin) pathway regulates the expression of LRP1-Hx. This study is designed to clarify whether TLR7 activation facilitates heme scavenging and to establish the potential role of the BTK-CRT-LRP1-Hx signaling pathway in the pathophysiology of ICH. Methods- ICH was induced by stereotactic, intrastriatal injection of type VII collagenase. Mice received TLR7 agonist (imiquimod) via intraperitoneal injection after ICH induction. TLR7 inhibitor (ODN2088), BTK inhibitor (LFM-A13), and CRT agonist (thapsigargin) were given in different groups to further evaluate the underlying pathway. Mice were randomly divided into sham, ICH+vehicle (normal saline), ICH+Imiquimod (2.5, 5, and 10 µg/g), ICH+ODN2088, ICH+LFM-A13, ICH+thapsigargin, and ICH+ODN2088+thapsigargin. Imiquimod was administered twice daily starting at 6 hours after ICH; ODN2088 was administered by intracerebroventricular injection at 30 minutes, and LFM-A13 or thapsigargin was administered by intraperitoneal injection at 3 hours after ICH induction. Neurological scores, cognitive abilities, as well as brain edema, blood-brain barrier permeability, hemoglobin level, brain expression of TLR7/BTK/CRT/LRP1/Hx were analyzed. Results- Low dosage imiquimod significantly attenuated hematoma volume, brain edema, BBB permeability, and neurological deficits after ICH. Imiquimod also increased protein expressions of TLR7, BTK, CRT, LRP1, and Hx; ODN2088 reduced TLR7, BTK, CRT, LRP1, and Hx expressions. Conclusions- TLR7 plays an important role in heme scavenging after ICH by modulating the BTK-CRT-LRP1-Hx pathway. TLR7 may offer protective effects by promoting heme resolution and reduction of brain edema after ICH.

Alteration of Surface Glycoproteins After Photodynamic Therapy.

BACKGROUND: Cell membranes have been identified as an important intracellular cancer treatment target, since the glycoconjugates present on the cell surface are involved in numerous cell functions. Photodynamic therapy (PDT) is a therapeutic modality employed in the treatment of tumors that uses visible light to activate a photosensitizer. OBJECTIVE: This study analyzed the expression of surface carbohydrates after PDT with two different photosensitizers, 5-aminolevulinic acid (ALA) and Photosan-3. METHODS: Mice were injected subcutaneously with 2 × 105 B16 cells. After 7-10 days, the presence of a tumor with a diameter of 3.6 mm was observed. Photosan-3® and 5-aminolevulinic acid-ALA were used in the PDT treatment. Control animals (not submitted to either laser treatment or photosensitizer injection) and treated animals were euthanized 15 days post-treatment. The tumors were irradiated with a red diode laser, λ = 655 nm, energy density of 10 J.cm-2, and power density of 45 mW.cm-2. After 2 weeks of treatment with PDT, the mice were euthanized, the tumors were collected, and the cell surfaces were labeled with lectins concanavalin A (ConA) and wheat germ agglutinin (WGA). RESULTS: Fluorescence microscopy analysis of the cell surfaces with lectins ConA and WGA showed the presence of α-mannose and α-glucose. CONCLUSIONS: The combined effects of either Photosan-3 or ALA and red laser light on melanoma suggest an inhibitory glycosylation action from PDT on the surface of B16-F10 cells.

Precision therapeutic targeting of human cancer cell motility.

Increased cancer cell motility constitutes a root cause of end organ destruction and mortality, but its complex regulation represents a barrier to precision targeting. We use the unique characteristics of small molecules to probe and selectively modulate cell motility. By coupling efficient chemical synthesis routes to multiple upfront in parallel phenotypic screens, we identify that KBU2046 inhibits cell motility and cell invasion in vitro. Across three different murine models of human prostate and breast cancer, KBU2046 inhibits metastasis, decreases bone destruction, and prolongs survival at nanomolar blood concentrations after oral administration. Comprehensive molecular, cellular and systemic-level assays all support a high level of selectivity. KBU2046 binds chaperone heterocomplexes, selectively alters binding of client proteins that regulate motility, and lacks all the hallmarks of classical chaperone inhibitors, including toxicity. We identify a unique cell motility regulatory mechanism and synthesize a targeted therapeutic, providing a platform to pursue studies in humans.

Arsenic trioxide induces cell cycle arrest and affects Trk receptor expression in human neuroblastoma SK-N-SH cells.

BACKGROUND: Arsenic trioxide (As2O3), a drug that has been used in China for approximately two thousand years, induces cell death in a variety of cancer cell types, including neuroblastoma (NB). The tyrosine kinase receptor (Trk) family comprises three members, namely TrkA, TrkB and TrkC. Various studies have confirmed that TrkA and TrkC expression is associated with a good prognosis in NB, while TrkB overexpression can lead to tumor cell growth and invasive metastasis. Previous studies have shown that As2O3 can inhibit the growth and proliferation of a human NB cell line and can also affect the N-Myc mRNA expression. It remains unclear whether As2O3 regulates Trks for the purposes of treating NB. METHODS: The aim of the present study was to investigate the effect of As2O3 on Trk expression in NB cell lines and its potential therapeutic efficacy. SK-N-SH cells were grown with increasing doses of As2O3 at different time points. We cultured SK-N-SH cells, which were treated with increasing doses of As2O3 at different time points. Trk expression in the NB samples was quantified by immunohistochemistry, and the cell cycle was analyzed by flow cytometry. TrkA, TrkB and TrkC mRNA expression was evaluated by real-time PCR analysis. RESULTS: Immunohistochemical and real-time PCR analyses indicated that TrkA and TrkC were over-expressed in NB, and specifically during stages 1, 2 and 4S of the disease progression. TrkB expression was increased in stage 3 and 4 NB. As2O3 significantly arrested SK-N-SH cells in the G2/M phase. In addition, TrkA, TrkB and TrkC expression levels were significantly upregulated by higher concentrations of As2O3 treatment, notably in the 48-h treatment period. Our findings suggested that to achieve the maximum effect and appropriate regulation of Trk expression in NB stages 1, 2 and 4S, As2O3 treatment should be at relatively higher concentrations for longer delivery times;however, for NB stages 3 and 4, an appropriate concentration and infusion time for As2O3 must be carefully determined. CONCLUSION: The present findings suggested that As2O3 induced Trk expression in SK-N-SH cells to varying degrees and may be a promising adjuvant to current treatments for NB due to its apoptotic effects.

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.

Arsenic trioxide induces cell cycle arrest and affects Trk receptor expression in human neuroblastoma SK-N-SH cells

BACKGROUND: Arsenic trioxide (As2O3), a drug that has been used in China for approximately two thousand years, induces cell death in a variety of cancer cell types, including neuroblastoma (NB). The tyrosine kinase receptor (Trk) family comprises three members, namely TrkA, TrkB and TrkC. Various studies have confirmed that TrkA and TrkC expression is associated with a good prognosis in NB, while TrkB overexpression can lead to tumor cell growth and invasive metastasis. Previous studies have shown that As2O3 can inhibit the growth and proliferation of a human NB cell line and can also affect the N-Myc mRNA expression. It remains unclear whether As2O3 regulates Trks for the purposes of treating NB. METHODS: The aim of the present study was to investigate the effect of As2O3 on Trk expression in NB cell lines and its potential therapeutic efficacy. SK-N-SH cells were grown with increasing doses of As2O3 at different time points. We cultured SK-N-SH cells, which were treated with increasing doses of As2O3 at different time points. Trk expression in the NB samples was quantified by immunohistochemistry, and the cell cycle was analyzed by flow cytometry. TrkA, TrkB and TrkC mRNA expression was evaluated by real-time PCR analysis. RESULTS: Immunohistochemical and real-time PCR analyses indicated that TrkA and TrkC were over-expressed in NB, and specifically during stages 1, 2 and 4S of the disease progression. TrkB expression was increased in stage 3 and 4 NB. As2O3significantly arrested SK-N-SH cells in the G2/M phase. In addition, TrkA, TrkB and TrkC expression levels were significantly upregulated by higher concentrations of As2O3 treatment, notably in the 48-h treatment period. Our findings suggested that to achieve the maximum effect and appropriate regulation of Trk expression in NB stages 1, 2 and 4S, As2O3 treatment should be at relatively higher concentrations for longer delivery times;however, for NB stages 3 and 4, an appropriate concentration and infusion time for As2O3 must be carefully determined. CONCLUSION: The present findings suggested that As2O3 induced Trk expression in SK-N-SH cells to varying degrees and may be a promising adjuvant to current treatments for NB due to its apoptotic effects.

Endoplasmic reticulum stress contributes to ferritin molecules-mediated macrophage migration via P-selectin glycoprotein ligand-1.

SCOPE: Obesity is associated with elevated serum ferritin and increased macrophage activation and infiltration; however, the causal mechanisms underlying this relationship remain undefined. METHODS AND RESULTS: Serum ferritin and soluble P-selectin glycoprotein ligand (sPSGL)-1 level were higher in obese adolescents and patients with moderate nonalcoholic fatty liver disease (NAFLD) compared with controls (all p < 0.05). Multivariate linear regression revealed that serum ferritin was independently associated with sPSGL-1 (B = 0.249, 95% confidence interval: 0.011-0.487, p = 0.041) after adjustment for covariates. The messenger (m) RNA expression of GRP78/Bip, ferritin, and PSGL-1 in leukocytes was greater in patients with nonalcoholic fatty liver disease than in controls. An animal study showed that a tunicamycin injection (an endoplasmic reticulum stress inducer) triggered serum sPSGL-1 and ferritin elevation (all p < 0.01). An in vitro study revealed that serum ferritin and apoferritin induced tumor necrosis factor-α and sPSGL-1 secretion (all p < 0.01). A wound healing assay showed that PSGL-1 blocking inhibited apoferritin-mediated macrophage migration. GRP78/Bip knockdown by the endotoxin EGF-SubA completely inhibited apoferritin-mediated macrophage migration and PSGL-1 expression at the protein and mRNA levels (all p < 0.05). CONCLUSION: ER stress associated mechanisms are required for apoferritin-/ferritin-mediated macrophage migration via the PSGL-1-dependent pathway.

Soluble Epoxide Hydrolase Inhibitor Suppresses the Expression of Triggering Receptor Expressed on Myeloid Cells-1 by Inhibiting NF-kB Activation in Murine Macrophage.

Triggering receptors expressed on myeloid cell-1 (TREM-1) is a superimmunoglobulin receptor expressed on myeloid cells. TREM-1 amplifies the inflammatory response. Epoxyeicosatrienoic acids (EETs), the metabolites of arachidonic acid derived from the cytochrome P450 enzyme, have anti-inflammatory properties. However, the effects of EETs on TREM-1 expression under inflammatory stimulation remain unclear. Therefore, inhibition of soluble epoxide hydrolase (sEH) with a highly selective inhibitor [1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea, TPPU] was used to stabilize EETs. LPS was intratracheally injected into mice to induce pulmonary inflammation, after TPPU treatment for 3 h. Histological examination showed TPPU treatment-alleviated LPS-induced pulmonary inflammation. TPPU decreased TREM-1 expression, but not DAP12 or MyD88 expression. Murine peritoneal macrophages were challenged with LPS in vitro. We found that TPPU reduced LPS-induced TREM-1 expression in a dose-dependent manner, but not DAP12 or MyD88 expression. TPPU also decreased downstream signal from TREM-1, reducing pro-inflammatory cytokine TNF-α and IL-1ß mRNA expression. Furthermore, TPPU treatment inhibited IkB degradation in vivo and in vitro. Our results indicate that the inhibition of sEH suppresses LPS-induced TREM-1 expression and inflammation via inhibiting NF-kB activation in murine macrophage.

Gpnmb/osteoactivin: an indicator and therapeutic target in tumor and nontumorous lesions.

Non-metastatic melanoma glycoprotein B (Gpnmb), a type I transmembrane glycoprotein, was first cloned and described in low-metastatic human melanoma and xenografts in 1995. Up to now a growing number of studies have confirmed that Gpnmb is expressed not only in numerous normal tissues but also at pathological sites and malignant tissues and often connected with the invasive and metastatic phenotypes, including breast cancer. Nowadays, immunotherapeutic approaches for cancer therapy, by which monoclonal antibodies (Mabs) target tumor specific antigens, have shown great potential. Glembatumumabvedotin, also called CR011-vcMMAE, is a Mab-drug conjugate which was developed for the treatment of Gpnmb-expressing cancers. Several phase I/II studies have confirmed the safety and activity of glembatumumabvedotin in patients with advanced/metastatic breast cancer and unresectable cutaneous melanoma. Moreover, increasing numbers of studies have supported the potential roles of targeting Gpnmb with glembatumumabvedotin in patients with recurrent osteosarcoma, uveal melanoma, ALS, Gaucher disease, pancreatic ductal adenocarcinoma etc. This review will summarize the latest understanding of Gpnmb in the aspects of diagnosis, progression and prognosis of pathological disorders and neoplasms, emphasizing the clinical advances in targeting Gpnmb-expressing malignancies.

Targeting Glycoprotein NMB With Antibody-Drug Conjugate, Glembatumumab Vedotin, for the Treatment of Osteosarcoma.

BACKGROUND: Cure rates for children and young adults with osteosarcoma have remained stagnant over the past three decades. Targeting glycoprotein non-metastatic b (GPNMB) with the antibody-drug conjugate glembatumumab vedotin has improved outcomes for patients with melanoma and breast cancer. The potential utility of targeting GPNMB in osteosarcoma was explored. METHODS: GPNMB protein expression was evaluated by immunohistochemistry in human osteosarcoma tumor samples and by enzyme-linked immunosorbent assay (ELISA) in osteosarcoma cell lines. mRNA expression was measured by quantitative PCR in primary osteosarcoma samples and cell lines. Surface GPNMB expression was evaluated by flow cytometry and correlated with in vitro and in vivo cytotoxicity of glembatumumab vedotin. RESULTS: Sixty seven human osteosarcoma samples were evaluated by immunohistochemistry, including 12 samples from initial biopsy, 38 samples from definitive surgery, and 17 from the time of disease recurrence. GPNMB was expressed in 92.5% (62/67) of osteosarcoma samples. All primary osteosarcoma samples expressed high levels of GPNMB mRNA. Glembatumumab induced cytotoxic effects in 74% (14/19) of osteosarcoma cell lines, and GPNMB protein levels correlated with glembatumumab in vitro cytotoxicity (r = -0.46, P = 0.04). All osteosarcoma cell lines demonstrated surface GPNMB expression. CONCLUSIONS: GPNMB is expressed in osteosarcoma and targeting GPNMB with the antibody-drug conjugate glembatumumab vedotin demonstrates osteosarcoma cytotoxic activity. Clinical trials are indicated to assess the efficacy of targeting GPNMB in patients with osteosarcoma.

Presence of Cleaved Synaptosomal-Associated Protein-25 and Decrease of Purinergic Receptors P2X3 in the Bladder Urothelium Influence Efficacy of Botulinum Toxin Treatment for Overactive Bladder Syndrome.

OBJECTIVES: To evaluate whether botulinum toxin A (BoNT-A) injection and Lipotoxin (liposomes with 200 U of BoNT-A) instillation target different proteins, including P2X3, synaptic vesicle glycoprotein 2A, and SNAP-25, in the bladder mucosa, leading to different treatment outcomes. MATERIALS AND METHODS: This was a retrospective study performed in a tertiary teaching hospital. We evaluated the clinical results of 27 OAB patients treated with intravesical BoNT-A injection (n = 16) or Lipotoxin instillation (n = 11). Seven controls were treated with saline. Patients were injected with 100 U of BoNT-A or Lipotoxinin a single intravesical instillation. The patients enrolled in this study all had bladder biopsies performed at baseline and one month after BoNT-A therapy. Treatment outcome was measured by the decreases in urgency and frequency episodes at 1 month. The functional protein expressions in the urothelium were measured at baseline and after 1 month. The Wilcoxon signed-rank test and ordinal logistic regression were used to compare the treatment outcomes. RESULTS: Both BoNT-A injection and Lipotoxin instillation treatments effectively decreased the frequency of urgency episodes in OAB patients. Lipotoxin instillation did not increase post-void residual volume. BoNT-A injection effectively cleaved SNAP-25 (p < 0.01). Liposome encapsulated BoNT-A decreased urothelial P2X3 expression in the five responders (p = 0.04), while SNAP-25 was not significantly cleaved. CONCLUSIONS: The results of this study provide a possible mechanism for the therapeutic effects of BoNT-A for the treatment of OAB via different treatment forms. BoNT-A and Lipotoxin treatments effectively decreased the frequency of urgency episodes in patients with OAB.

Potent functional uncoupling between STIM1 and Orai1 by dimeric 2-aminodiphenyl borinate analogs.

The coupling of ER Ca(2+)-sensing STIM proteins and PM Orai Ca(2+) entry channels generates "store-operated" Ca(2+) signals crucial in controlling responses in many cell types. The dimeric derivative of 2-aminoethoxydiphenyl borinate (2-APB), DPB162-AE, blocks functional coupling between STIM1 and Orai1 with an IC50 (200 nM) 100-fold lower than 2-APB. Unlike 2-APB, DPB162-AE does not affect L-type or TRPC channels or Ca(2+) pumps at maximal STIM1-Orai1 blocking levels. DPB162-AE blocks STIM1-induced Orai1 or Orai2, but does not block Orai3 or STIM2-mediated effects. We narrowed the DPB162-AE site of action to the STIM-Orai activating region (SOAR) of STIM1. DPB162-AE does not prevent the SOAR-Orai1 interaction but potently blocks SOAR-mediated Orai1 channel activation, yet its action is not as an Orai1 channel pore blocker. Using the SOAR-F394H mutant which prevents both physical and functional coupling to Orai1, we reveal DPB162-AE rapidly restores SOAR-Orai binding but only slowly restores Orai1 channel-mediated Ca(2+) entry. With the same SOAR mutant, 2-APB induces rapid physical and functional coupling to Orai1, but channel activation is transient. We infer that the actions of both 2-APB and DPB162-AE are directed toward the STIM1-Orai1 coupling interface. Compared to 2-APB, DPB162-AE is a much more potent and specific STIM1/Orai1 functional uncoupler. DPB162-AE provides an important pharmacological tool and a useful mechanistic probe for the function and coupling between STIM1 and Orai1 channels.