Active Receptor Research Articles

Mechanosensitive release of ATP in the urinary bladder mucosa.

The urinary bladder mucosa (urothelium and suburothelium/lamina propria) functions as a barrier between the content of the urine and the underlying bladder tissue. The bladder mucosa is also a mechanosensitive tissue that releases signaling molecules that affect functions of cells in the bladder wall interconnecting the mucosa with the detrusor muscle and the CNS. Adenosine 5'-triphosphate (ATP) is a primary mechanotransduction signal that is released from cells in the bladder mucosa in response to bladder wall distention and activates cell membrane-localized P2X and P2Y purine receptors on urothelial cells, sensory and efferent neurons, interstitial cells, and detrusor smooth muscle cells. The amounts of ATP at active receptor sites depend significantly on the amounts of extracellularly released ATP. Spontaneous and distention-induced release of ATP appear to be under differential control. This review is focused on mechanisms underlying urothelial release of ATP in response to mechanical stimulation. First, we present a brief overview of studies that report mechanosensitive ATP release in bladder cells or tissues. Then, we discuss experimental evidence for mechanosensitive release of urothelial ATP by vesicular and non-vesicular mechanisms and roles of the stretch-activated channels PIEZO channels, transient receptor potential vanilloid type 4, and pannexin 1. This is followed by brief discussion of possible involvement of calcium homeostasis modulator 1, acid-sensing channels, and connexins in the release of urothelial ATP. We conclude with brief discussion of limitations of current research and of needs for further studies to increase our understanding of mechanotransduction in the bladder wall and of purinergic regulation of bladder function.

Abstract A005: Microbind affinity blood filter platform technology with affinity for heparan sulfate rapidly removes circulating epithelial tumor and cancer stem cells while downregulating VEGF A in patients with advanced and refractory solid tumors

Abstract Background: Vascular endothelial-derived growth factor A(VEGFA) has been proven to be integral in the molecular pathogenesis of metastasis and tumor growth. It is known that VEGF is responsible for encoding a heparin-binding protein, which exists as a disulfide-linked homodimer which serves to promote the propagation and movement of vascular endothelial cells. This is essential for both pathological and physiological angiogenesis. CETC and cancer stem cells are potent secretors of VEGFA which is an essential cytokine that facilitates new vascular vessel formation, thus further enabling metastatic tumor growth. Methods: We developed a new extracorporeal microbind affinity blood filter platform technology with active heparane sulfate receptors (onco seraph). Heparin is cleaved into short chains which are permanently covalently bound to ultrahigh molecular weight polyethylene (UHMWPE) surfaces to create a new composition of matter. Liquid biopsy established high capacity for the rapid removal of CETC and PDAC cancer stem cells. Onco seraph was investigated in a dose escalation phase I/II clinical trial in adult patients with advanced and relapsed/refractory solid (what) and/or PDAC after standard of care and other lines of therapy failed in preventing disease progression. Results: To date 12 patients have been treated with onco seraph. No patient experienced treatment related adverse effects. Best responses include complete responses and stable disease at a low oncoseraph dose. Greatest efficacy was observed for patients whose liquid biopsy revealed significant and rapid reduction of CETC and cancer stem cells, followed by greater than 50% reduction in circulating VEGFA. Conclusions: This data supports the conclusion that the onco seraph platform technology is safe and has a high therapeutic value as an extracorporeal treatment for metastatic solid tumors refractory to standard of care. Citation Format: Sanja ILIC, Vedran Premuzic, Robert Ward. Microbind affinity blood filter platform technology with affinity for heparan sulfate rapidly removes circulating epithelial tumor and cancer stem cells while downregulating VEGF A in patients with advanced and refractory solid tumors [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A005.

Unveiling Wide Spectrum Therapeutic Implications and Signaling Mechanisms of Valsartan in Diverse Disorders: A Comprehensive Review.

Valsartan is an orally active non-peptide angiotensin receptor antagonist, an effective and well-tolerated anti-hypertensive drug. Besides its antihypertensive action, it has clinical implications in many other disorders, like heart failure (HF), arrhythmia, chronic kidney disease (CKD), diabetic complications (DM), atherosclerosis, etc. Besides angiotensin receptor blocking activity, valsartan reduces circulating levels of biochemical markers, such as hs-CRP, which is responsible for its anti-inflammatory and anti-oxidant activity. Moreover, valsartan also acts by inhibiting or inducing various signalling pathways, such as inducing autophagy via the AKT/mTOR/S6K pathway or inhibiting the TLR/NF-kB pathway. The current review exhaustively discusses the therapeutic implications of valsartan with specific emphasis on the mechanism of action in various disorders. The article provides a detailed spectrum of the therapeutic profile of valsartan and will likely be very useful to researchers working in the relevant research areas.

Exploration and identification of diabetes targets in nursing: CDH1 and DVL1.

Diabetes is a chronic disease caused by absolute or relative insufficiency of insulin secretion and impaired insulin utilization. CDH1 and DVL1 role in diabetes and its nursing care is unclear. The diabetes dataset GSE21321 and GSE19790 profiles were downloaded from the gene expression omnibus (GEO) database. Perform differentially expressed genes (DEGs) screening, weighted gene co-expression network analysis (WGCNA), protein-protein interaction (PPI) network construction and analysis, functional enrichment analysis, gene set enrichment analysis (GSEA), immune infiltration analysis, and Comparative Toxicogenomics Database (CTD) analysis. Gene expression heat map was drawn. TargetScan was used to screen the miRNA that regulates central DEGs. 1983 DEGs were obtained. According to Gene Ontology (GO) analysis, they were mainly enriched in signal regulation, catenin complexes, and signal receptor binding. In Kyoto Encyclopedia of Gene and Genome (KEGG) analysis, they were mainly concentrated in the Rap1 signaling pathway, cAMP signaling pathway, and Hippo signaling pathway. The DEGs are mainly enriched in cell signaling, Wnt signaling vesicles, growth factor activity, and the interaction between neural active ligands and receptors. In the enrichment project of Metascape, BMP signaling pathways and cell population proliferation can be seen in the GO enrichment project. The soft threshold power in WGCNA is set to 5. A total of 15 modules were generated. Core gene expression heatmap showed that core genes (CTNNB1, CDH1, DVL1) were highly expressed in diabetes samples. CTD analysis showed thatCTNNB1, CDH1, DVL1were associated with weight gain, inflammation, and necrosis. CDH1 and DVL1 are highly expressed in diabetes and may become molecular targets for diabetes and its care.

A head-to-head comparison of MM/PBSA and MM/GBSA in predicting binding affinities for the CB1 cannabinoid ligands.

The substantial increase in the number of active and inactive-state CB1 receptor experimental structures has provided opportunities for CB1 drug discovery using various structure-based drug design methods, including the popular end-point methods for predicting binding free energies-Molecular Mechanics/Poisson-Boltzmann Surface Area (MM/PBSA) and Molecular Mechanics/Generalized Born Surface Area (MM/GBSA). In this study, we have therefore evaluated the performance of MM/PBSA and MM/GBSA in calculating binding free energies for CB1 receptor. Additionally, with both MM/PBSA and MM/GBSA being known for their highly individualized performance, we have evaluated the effects of various simulation parameters including the use of energy minimized structures, choice of solute dielectric constant, inclusion of entropy, and the effects of the five GB models. Generally, MM/GBSA provided higher correlations than MM/PBSA (rMM/GBSA = 0.433 - 0.652 vs. rMM/PBSA = 0.100 - 0.486) regardless of the simulation parameters, while also offering faster calculations. Improved correlations were observed with the use of molecular dynamics ensembles compared with energyminimized structures and larger solute dielectric constants. Incorporation of entropic terms led to unfavorable results for both MM/PBSA and MM/GBSA for a majority of the dataset, while the evaluation of the various GB models exerted a varying effect on both the datasets. The findings obtained in this study demonstrate the utility of MM/PBSA and MM/GBSA in predicting binding free energies for the CB1 receptor, hence providing a useful benchmark for their applicability in the endocannabinoid system as well as other G protein-coupled receptors. The study utilized the docked dataset (Induced Fit Docking with Glide XP scoring function) from Loo et al., consisting of 46 ligands-23 agonists and 23 antagonists. The equilibrated structures from Loo et al. were subjected to 30ns production simulations using GROMACS 2018 at 300K and 1atm with the velocity rescaling thermostat and the Parinello-Rahman barostat. AMBER ff99SB*-ILDN was used for the proteins, General Amber Force Field (GAFF) was used for the ligands, and Slipids parameters were used for lipids. MM/PBSA and MM/GBSA binding free energies were then calculated using gmx_MMPBSA. The solute dielectric constant was varied between 1, 2, and 4 to study the effect of different solute dielectric constants on the performance of MM/PB(GB)SA. The effect of entropy on MM/PB(GB)SA binding free energies was evaluated using the interaction entropy module implemented in gmx_MMPBSA. Five GB models, GBHCT, GBOBC1, GBOBC2, GBNeck, and GBNeck2, were evaluated to study the effect of the choice of GB models in the performance of MM/GBSA. Pearson correlation coefficients were used to measure the correlation between experimental and predicted binding free energies.

Molecular Docking Studies of Synthesized Pyridazinone Scaffolds as Non-Nucleoside Reverse Transcriptase Inhibitors (NNRTIs)

Pyridazinone derivatives, 6-aryl-pyridazinone (2a–f) and 2-(N-substituted)-6-aryl-pyridazinone (3a–h) derivatives were synthesized and assessed for cytotoxicity action using brine shrimp assessment method and in silico molecular studies. In silico molecular study of pyridazine derivatives used as NNRTIs and Doravirine is used as reference drug. The compounds were investigated for docking modes onto probable binding sites with HIV reverse transcriptase. The majority of these demonstrated favorable binding interactive connections with the active receptor domain. The majority of the compounds exhibited a remarkable docked binding affinity score when compared with the reference drugs. Among the synthesized pyridazine derivatives, compounds 3a and 3c–h exhibited a good docking score. For the designed compounds, in silico ADME assessment indicated the favorable physicochemical properties to be a suitable drug candidate. The synthesized compounds could serve as a novel alternative in designing safe and effective anti-HIV options with reverse transcriptase inhibitor potential. In the cytotoxicity study, all the compounds were evaluated at dose levels 10, and 20 (μg/mL), and potassium dichromate was used as a reference. Compounds 2c and 2e have shown potent lethality through LC50 2.23 and 3.20 μg respectively. Various compounds, including 2a, 2b, 2d, and 2f have demonstrated significant cytotoxicity. Their LC50 values are 7.58, 6.76, 8.91, and 4.46 μg, respectively. Additionally, compounds 3b and 3d exhibited potent lethality with LC50 values of 4.023 and 4.20 μg. Other compounds, namely 3g, 3f, 3c, 3h, 3a, and 3e, displayed noteworthy cytotoxicity ability13.91, 12.58, 11.91, 11.76, 10.58, 9.76, and 7.46 μg, respectively having LC50 values. This study supports the use of in silico molecular design and the brine shrimp bioassay as a suitable, reliable, and simple method for assessing the bioactivity of compounds. It provides further evidence for their use in medicine.

5119 Structural basis for glucocorticoid receptor multimerization

Abstract Disclosure: E. Estébanez-Perpiñá: None. Abstract ENDO-2024 - BostonStructural basis for glucocorticoid receptor multimerization Andrea Alegre-Martí[1]*, Alba Jimenez-Panizo2*, Agustina Lafuente3, Montserrat Abella[1], Thomas A Johnson2, Paloma Pérez4, Juan Fernández-Recio5, Diego M. Presman3, Gordon L. Hager2, Pablo Fuentes-Prior[1], Eva Estébanez-Perpiñá[1] [1]Department of Biochemistry and Molecular Biomedicine, Institute of Biomedicine of the University of Barcelona (IBUB), Faculty of Biology, University of Barcelona (UB), 08028 Barcelona, Spain.2National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-5055, USA.3IFIBYNE, UBA-CONICET, Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Buenos Aires, C1428EGA, Argentina.4Instituto de Biomedicina de Valencia (IBV)-CSIC, 46010, Valencia, Spain.5Instituto de Ciencias de la Vid y del Vino (ICVV), CSIC - Universidad de La Rioja - Gobierno de La Rioja, 26007 Logroño, Spain. Abstract: The glucocorticoid receptor (GR) is a ubiquitously e2xpressed ligand-regulated transcription factor essential for life and one of the most targeted proteins in drug discovery due to its powerful anti-inflammatory actions. The functional oligomeric state of the full-length receptor, which is essential for its transcriptional activity in cells, remains disputed. Here we present a new crystal structure of agonist-bound ancient GR-LBD in a large cell, along with a thorough analysis of previous structural work. The building block of the current structure is a homodimer we previously identified in GR-LBD crystals and its biological relevance has been verified by studying a battery of GR point mutants including crosslinking assays in solution and quantitative fluorescence microscopy in live cells. Several mutually exclusive multimeric assemblies of this dimer in the crystal highlight the versatility of GR-LBD for self-association and reveal implications for the conformation of the active full-length receptor. Our results underscore the relevance of non-canonical dimerization modes for GR-LBD, especially of contacts made by key residues such as Tyr545, Pro637 and Asp641. Of note, a non-conservative mutation of the latter, p.Asp641Val, causes Chrousos syndrome in humans. Understanding relevant quaternary assemblies of the GR is pivotal not only to understand and predict the therapeutic outcome of major blockbuster drugs but also to lessen their deleterious side effects and open new avenues for drug design. Presentation: 6/3/2024

Transgenic Drosophila Expressing Active Human LH Receptor in the Gonads Exhibit a Decreased Fecundity: Towards a Platform to Identify New Orally Active Modulators of Gonadotropin Receptor Activity.

The gonadotropins luteinizing hormone (LH) and follicle-stimulating hormone (FSH) and their receptors are major regulators of reproduction in mammals and are absent in insects. We previously established transgenic Drosophila lines expressing a constitutively active human LH receptor variant (LHRD578Y) and the wild-type receptor (LHRwt; inactive in the absence of an agonist). That study showed that ubiquitously expression of LHRD578Y-but not of LHRwt-resulted in pupal lethality, and targeted expression in midline cells resulted in thorax/bristles defects. To further study the Drosophila model for an in vivo drug screening platform, we investigated here whether expressing LHRD578Y in the fly gonads alters reproduction, as shown in a transgenic mice model. The receptor was expressed in somatic cells of the gonads using the tissue-specific traffic jam-Gal4 driver. Western blot analysis confirmed receptor expression in the ovaries. A fecundity assay indicated that the ectopic expression of LHRD578Y resulted in a decrease in egg laying compared to control flies carrying, but not expressing the transgene (~40% decrease in two independent fly lines, p < 0.001). No significant reduction in the number of laid eggs was seen in flies expressing the LHRWT (<10% decrease compared to non-driven flies, p > 0.05). The decreased egg laying demonstrates a phenotype of the active receptor in the fly gonads, the prime target organs of the gonadotropins in mammals. We suggest that this versatile Drosophila model can be used for the pharmacological search for gonadotropin modulators. This is expected to provide: (a) new mimetic drug candidates (receptor-agonists/signaling-activators) for assisted reproduction treatment, (b) blockers for potential fertility regulation, and (c) leads relevant for the purpose of managing extra gonadotropic reported activities.

Membrane trafficking of synaptic adhesion molecules.

Synapse formation and stabilization are aided by several families of adhesion molecules, which are generally seen as specialized surface receptors. The function of most surface receptors, including adhesion molecules, is modulated in non-neuronal cells by the processes of endocytosis and recycling, which control the number of active receptors found on the cell surface. These processes have not been investigated extensively at the synapse. This review focuses on the current status of this topic, summarizing general findings on the membrane trafficking of the most prominent synaptic adhesion molecules. Remarkably, evidence for endocytosis processes has been obtained for many synaptic adhesion proteins, including dystroglycans, latrophilins, calsyntenins, netrins, teneurins, neurexins, neuroligins and neuronal pentraxins. Less evidence has been obtained on their recycling, possibly because of the lack of specific assays. We conclude that the trafficking of the synaptic adhesion molecules is an important topic, which should receive more attention in the future.

Discovery of a Potent, Orally Active, and Long-Lasting P2X7 Receptor Antagonist as a Preclinical Candidate for Delaying the Progression of Chronic Kidney Disease.

Chronic kidney disease (CKD) is a condition characterized by functional deterioration with sustained inflammation and progressive fibrosis of the kidneys affecting over 800 million people worldwide. The P2X7 receptor (P2X7R) plays a key role in CKD progression. Our previous P2X7R antagonists demonstrated good efficacy for treating kidney injury but were limited by low oral exposure and short half-life, restricting their application. This study reports the optimization of P2X7R antagonists for better oral pharmacokinetics. The candidate compound 13a with the respective IC50 of 34.86 and 25.28 nM against human and murine P2X7R, administered orally at 10 mg/kg in mice, exhibits a remarkably long half-life of 161.64 h, with a high exposure of 1,163,980.55 μg·h/L. Oral administration of 13a (0.3 or 1.0 mg/kg, twice weekly) significantly reduced renal injury and fibrosis in unilateral ureteral obstruction and adenine diet-induced mice models, highlighting its potential for delaying the progression of CKD.

Analysis of EGFR binding hotspots for design of new EGFR inhibitory biologics.

The epidermal growth factor (EGF) receptor (EGFR) is activated by the binding of one of seven EGF-like ligands to its ectodomain. Ligand binding results in EGFR dimerization and stabilization of the active receptor conformation subsequently leading to activation of downstream signaling. Aberrant activation of EGFR contributes to cancer progression through EGFR overexpression/amplification, modulation of its positive and negative regulators, and/or activating mutations within EGFR. EGFR targeted therapeutic antibodies prevent dimerization and interaction with endogenous ligands by binding the ectodomain of EGFR. However, these antibodies have had limited success in the clinic, partially due to EGFR ectodomain resistance mutations, and are only applicable to a subset of patients with EGFR-driven cancers. These limitations suggest that alternative EGFR targeted biologics need to be explored for EGFR-driven cancer therapy. To this end, we analyze the EGFR interfaces of known inhibitory biologics with determined structures in the context of endogenous ligands, using the Rosetta macromolecular modeling software to highlight the most important interactions on a per-residue basis. We use this analysis to identify the structural determinants of EGFR targeted biologics. We suggest that commonly observed binding motifs serve as the basis for rational design of new EGFR targeted biologics, such as peptides, antibodies, and nanobodies.

Immunophenotyping of Peripheral Blood Cells in Patients with Chronic Lymphocytic Leukemia Treated with Ibrutinib.

Chronic lymphocytic leukemia (CLL) is a B-cell-derived hematologic malignancy whose progression depends on active B-cell receptor (BCR) signaling. Despite the spectacular efficacy of Ibrutinib, an irreversible inhibitor of Bruton tyrosine kinase (BTK), resistance can develop in CLL patients, and alternative therapeutic strategies are therefore required. Cancer immunotherapy has revolutionized cancer care and may be an attractive approach in this context. We speculated that characterizing the immune responses of CLL patients may highlight putative immunotherapeutic targets. Here, we used high-dimensional spectral flow cytometry to compare the distribution and phenotype of non-B-cell immune populations in the circulating blood of CLL patients treated with Ibrutinib displaying a complete response or secondary progression. Although no drastic changes were observed in the composition of their immune subsets, the Ibrutinib-resistant group showed increased cycling of CD8+ T cells, leading to their overabundance at the expense of dendritic cells. In addition, the expression of 11 different surface checkpoints was similar regardless of response status. Together, this suggests that CLL relapse upon Ibrutinib treatment may not lead to major alterations in the peripheral immune response.

Biological variability of human intraepithelial lymphocytes throughout the human gastrointestinal tract in health and coeliac disease.

Intraepithelial lymphocytes are the first line of defence of the human intestinal immune system. Besides, their composition is altered on patients with coeliac disease (CD), so they are considered as biomarkers with utility on their diagnose and/or monitoring. Our aim is to address their variability through the human gastrointestinal tract in health and characterized them in further depth in the coeliac duodenum. Intraepithelial lymphocytes were isolated from human gastric, duodenal, ileal and colonic biopsies, then stained with specific antibodies and acquired by flow cytometry. Our results confirmed that the profile of Intraepithelial lymphocytes change through the length of the human gastrointestinal tract. Besides and given the central role that Interleukin-15 (IL-15) elicits on CD pathogenesis; we also assessed the expression of its receptor revealing that there was virtually no functional IL-15 receptor on duodenal Intraepithelial lymphocytes. Nevertheless and contrary to our expectations, the active IL-15 receptor was not increased either on Intraepithelial lymphocytes from CD patients. IL-15 might require additional stimulus to activate intraepithelial lymphocytes. These findings may provide novel tools to aid on a CD diagnosis and/or monitoring, at the time that provide the bases to perform functional studies in order of getting a deeper insight in the specific function that Intraepithelial lymphocytes elicit on CD pathogenesis.

Recommended Tool Compounds for the Melanocortin Receptor (MCR) G Protein-Coupled Receptors (GPCRs).

The melanocortin receptors are a centrally and peripherally expressed family of Class A GPCRs with physiological roles, including pigmentation, steroidogenesis, energy homeostasis, and others yet to be fully characterized. There are five melanocortin receptor subtypes that, apart from the melanocortin-2 receptor (MC2R), are stimulated by a shared set of endogenous agonists. Until 2020, X-ray crystallographic and cryo-electron microscopic (cryo-EM) structures of these receptors were unavailable, and the investigation of their mechanisms of action and putative ligand-receptor interactions was driven by site-directed mutagenesis studies of the receptors and targeted structure-activity relationship (SAR) studies of the endogenous and derivative synthetic ligands. Synthetic derivatives of the endogenous agonist ligand α-MSH have evolved into a suite of powerful ligands such as NDP-MSH (melanotan I), melanotan II (MTII), and SHU9119. This suite of tool compounds now enables the study of the melanocortin receptors and serves as scaffolds for FDA-approved drugs, means of validating stably expressing melanocortin receptor cell lines, core ligands in assessing cryo-EM structures of active and inactive receptor complexes, and essential references for high-throughput discovery and mechanism of action studies. Herein, we review the history and significance of a finite set of these essential tool compounds and discuss how they are being utilized to further the field's understanding of melanocortin receptor physiology and greater druggability.

A 'double-edged' role for type-5 metabotropic glutamate receptors in pain disclosed by light-sensitive drugs.

We used light-sensitive drugs to identify the brain region-specific role of mGlu5 metabotropic glutamate receptors in the control of pain. Optical activation of systemic JF-NP-26, a caged, normally inactive, negative allosteric modulator (NAM) of mGlu5 receptors, in cingulate, prelimbic, and infralimbic cortices and thalamus inhibited neuropathic pain hypersensitivity. Systemic treatment of alloswitch-1, an intrinsically active mGlu5 receptor NAM, caused analgesia, and the effect was reversed by light-induced drug inactivation in the prelimbic and infralimbic cortices, and thalamus. This demonstrates that mGlu5 receptor blockade in the medial prefrontal cortex and thalamus is both sufficient and necessary for the analgesic activity of mGlu5 receptor antagonists. Surprisingly, when the light was delivered in the basolateral amygdala, local activation of systemic JF-NP-26 reduced pain thresholds, whereas inactivation of alloswitch-1 enhanced analgesia. Electrophysiological analysis showed that alloswitch-1 increased excitatory synaptic responses in prelimbic pyramidal neurons evoked by stimulation of presumed BLA input, and decreased BLA-driven feedforward inhibition of amygdala output neurons. Both effects were reversed by optical silencing and reinstated by optical reactivation of alloswitch-1. These findings demonstrate for the first time that the action of mGlu5 receptors in the pain neuraxis is not homogenous, and suggest that blockade of mGlu5 receptors in the BLA may limit the overall analgesic activity of mGlu5 receptor antagonists. This could explain the suboptimal effect of mGlu5 NAMs on pain in human studies and validate photopharmacology as an important tool to determine ideal target sites for systemic drugs.

Dual-Targeting of the HER2 Cancer Receptor with an Antibody-Directed Enzyme and a Nanobody-Guided MMAE Prodrug Scaffold.

Antibody-enzyme conjugates have shown potential as tissue-specific prodrug activators by antibody-directed enzyme prodrug therapy (ADEPT), but the approach met challenges clinically due to systemic drug release. Here, we report a novel dual-targeting ADEPT system (DuADEPT) which is based on active cancer receptor targeting of both a trastuzumab-sialidase conjugate (Tz-Sia) and a highly potent sialidase-activated monomethyl auristatin E (MMAE) prodrug scaffold. The scaffold is based on a four-way junction of the artificial nucleic acid analog acyclic (L)-threoninol nucleic acid ((L)-aTNA) which at the ends of its four arms carries one nanobody targeting HER2 and three copies of the prodrug. Dual-targeting of the constructs to two proximal epitopes of HER2 was shown by flow cytometry, and a dual-targeted enzymatic drug release assay revealed cytotoxicity upon prodrug activation specifically for HER2-positive cancer cells. The specific delivery and activation of prodrugs in this way could potentially be used to decrease systemic side effects and increase drug efficacy, and utilization of Tz-Sia provides an opportunity to combine the local chemotherapeutic effect of the DuADEPT with an anticancer immune response.

Development in the Study of Natural Killer Cells for Malignant Peritoneal Mesothelioma Treatment.

Malignant peritoneal mesothelioma (MPeM) is a rare primary malignant tumor originating from peritoneal mesothelial cells. Insufficient specificity of the symptoms and their frequent reappearance following surgery make it challenging to diagnose, creating a need for more efficient treatment options. Natural killer cells (NK cells) are part of the innate immune system and are classified as lymphoid cells. Under the regulation of activating and inhibiting receptors, NK cells secrete various cytokines to exert cytotoxic effects and participate in antiforeign body, antiviral, and antitumor activities. This review provides a comprehensive summary of the specific alterations observed in NK cells following MPeM treatment, including changes in cell number, subpopulation distribution, active receptors, and cytotoxicity. In addition, we summarize the impact of various therapeutic interventions, such as chemotherapy, immunotherapy, and targeted therapy, on NK cell function post-MPeM treatment.

Crystal structure of the GluK1 ligand-binding domain with kainate and the full-spanning positive allosteric modulator BPAM538

Kainate receptors play an important role in the central nervous system by mediating postsynaptic excitatory neurotransmission and modulating the release of the inhibitory neurotransmitter GABA through a presynaptic mechanism. To date, only three structures of the ligand-binding domain (LBD) of the kainate receptor subunit GluK1 in complex with positive allosteric modulators have been determined by X-ray crystallography, all belonging to class II modulators. Here, we report a high-resolution structure of GluK1-LBD in complex with kainate and BPAM538, which belongs to the full-spanning class III. One BPAM538 molecule binds at the GluK1 dimer interface, thereby occupying two allosteric binding sites simultaneously. BPAM538 stabilizes the active receptor conformation with only minor conformational changes being introduced to the receptor. Using a calcium-sensitive fluorescence-based assay, a 5-fold potentiation of the kainate response (100 μM) was observed in presence of 100 μM BPAM538 at GluK1(Q)b, whereas no potentiation was observed at GluK2(VCQ)a. Using electrophysiology recordings of outside-out patches excised from HEK293 cells, BPAM538 increased the peak response of GluK1(Q)b co-expressed with NETO2 to rapid application of 10 mM L-glutamate with 130 ± 20 %, and decreased desensitization determined as the steady-state/peak response ratio from 23 ± 2 % to 90 ± 4 %. Based on dose–response relationship experiments on GluK1(Q)b the EC50 of BPAM538 was estimated to be 58 ± 29 μM.

Interfacial adsorption and catalysis of single-atom Pt-loaded ZrO2 surfaces for enhancing formaldehyde oxidation

The emerging transition metal single-atom catalysts (SACs) are expected to have high activity for converting detrimental formaldehyde (HCHO) under mild conditions. Currently, the exploitation of novel SACs with designed or modified support surfaces remains largely imperative. Herein, we used density functional theory to explore the catalytic performance for HCHO oxidation on various single-atom Pt-loaded ZrO2 surfaces, in which oxygen vacancy and lattice hydroxyl group were included on the surfaces. The surface modifications could enhance the co-adsorption of HCHO and O2 molecules and provide more active sites due to interfacial electron redistribution. The transition state search approach was applied to investigate the oxidation pathways on various ZrO2-supported SAC surfaces, wherein distinct dehydrogenation mechanisms and intermediates were clarified. The oxygen vacancy defects and hydroxyl groups can enhance the catalytic activity for HCHO oxidation by providing active receptor sites for the H-transfer and facilitating the CH bond breakage. Microkinetic modeling was further used to characterize the reaction rates and kinetics behavior of HCHO oxidation on the SAC surfaces. Our simulation demonstrates the excellent catalytic performance of the modified ZrO2-supported SAC in the HCHO oxidation. Meanwhile, the mechanism is also an important implication for the synthesis and design of novel SACs.

Anti-HDGF antibody targets EGFR tyrosine kinase inhibitor-tolerant cells in NSCLC patient-derived xenografts.

Constitutively active mutant epidermal growth factor receptor (EGFR) is one of the major oncogenic drivers in NSCLC. Targeted therapy using EGFR tyrosine kinase inhibitor (TKI) is a firstline option in patients that have metastatic or recurring disease. However, despite the high response rate to TKI, most patients have a partial response, and the disease eventually progresses in 10 to 19 months. It is believed that drug-tolerant cells that survive TKI exposure during the progression-free period facilitate the emergence of acquired resistance. Thus, targeting the drug-tolerant cells could improve the treatment of NSCLC with EGFR mutations. We demonstrated here that EGFR mutant patient-derived xenograft (PDX) tumors responded partially to osimertinib despite near complete inhibition of EGFR activation. Signaling in AKT/mTOR and MAPK pathways could be reactivated shortly after initial inhibition. As a result, many tumor cells escaped drug killing and regained growth following about 35 days of continuous osimertinib dosing. However, when an antibody to hepatoma-derived growth factor (HDGF) was given concurrently with osimertinib, tumors showed complete or near-complete responses.There was significant prolongation of progression-free survival (PFS) of tumor-bearing mice as well. Immunohistochemistry and western blot analysis of tumors collected in the early stages of treatment suggest that increased suppression of the AKT/mTOR and MAPK pathways could be a mechanism that results in enhanced efficacy of osimertinib when it is combined with anti-HDGF antibody.