Allosteric Research Articles

Addressing Clinical Limitations of Glutaminase Inhibitors: Novel Strategies for Osimertinib-Resistant Lung Cancer by Exploiting Glutamine Metabolic Dependency.

Overcoming acquired resistance to Osimertinib remains a critical challenge in treating NSCLC. This research indicates that Osimertinib-resistant cells exhibit a strong dependence on glutamine metabolism. However, targeting GLS1 shows limited anticancer effects, probably because it cannot fully block the glutamine metabolic pathway. The investigation reveals that a more effective strategy involves simultaneously inhibiting both ASCT2 and GLS1. After confirming the efficacy of this dual-targeting approach against Osimertinib-resistant cells in preclinical models, the potential of utilizing a broad-spectrum glutamine metabolism antagonist is further explored to achieve superior antitumor efficacy. DON, broad-spectrum glutamine antagonist, presents toxicity issues. Herein, the high NQO1 expression in Osimertinib-resistant NSCLC cells is leveraged to design an NQO1-responsive DON prodrug, 10e (LBJ-10e). This prodrug demonstrates superior safety compared to natural DON and greater antitumor activity against resistant tumors compared to the clinical phase II drug DRP104. These findings may address the clinical limitations of GLS1 allosteric inhibitors and underscore prodrug strategies in effectively treating Osimertinib-resistant lung cancer, providing a foundation for future clinical trials.

Distinct mechanisms drive divergent phenotypes in hypertrophic and dilated cardiomyopathy associated TPM1 variants.

Hypertrophic and dilated cardiomyopathies (HCM and DCM, respectively) are inherited disorders that may be caused by mutations to the same sarcomeric protein but have completely different clinical phenotypes. The precise mechanisms by which point mutations within the same gene bring about phenotypic diversity remain unclear. Our objective has been to develop a mechanistic explanation of diverging phenotypes in two TPM1 mutations, E62Q (HCM) and E54K (DCM). Drawing on data from the literature and experiments with stem cell-derived cardiomyocytes expressing the TPM1 mutations of interest, we constructed computational simulations that provide plausible explanations of the distinct muscle contractility caused by each variant. In E62Q, increased calcium sensitivity and hypercontractility was explained most accurately by a reduction in effective molecular stiffness of tropomyosin and alterations in its interactions with the actin thin filament that favor the 'closed' regulatory state. By contrast, the E54K mutation appeared to act via long-range allosteric interactions to increase the association rate of the C-terminal troponin I mobile domain to tropomyosin/actin. These mutation-linked molecular events produced diverging alterations in gene expression that can be observed in human engineered heart tissues. Modulators of myosin activity confirmed our proposed mechanisms by rescuing normal contractile behavior in accordance with predictions.

Antibodies disrupt bacterial adhesion by ligand mimicry and allosteric interference.

A critical step in infections is the attachment of many microorganisms to host cells using lectins that bind surface glycans, making lectins promising antimicrobial targets. Upon binding mannosylated glycans, FimH, the most studied lectin adhesin of type 1 fimbriae in E. coli , undergoes an allosteric transition from an inactive to an active conformation that can act as a catch-bond. Monoclonal antibodies that alter FimH glycan binding in various ways are available, but the mechanisms of these antibodies remain unclear. Here, we use cryoEM, mass spectrometry, binding assays, and molecular dynamics simulations to determine the structure-function relationships underlying antibody-FimH binding. Our study reveals four distinct antibody mechanisms of action: ligand mimicry by an N-linked, high-mannose glycan; stabilization of the ligand pocket in the inactive state; conformational trapping of the active and inactive states; and locking of the ligand pocket through long-range allosteric effects. These structures reveal multiple mechanisms of antibody responses to an allosteric protein and provide blueprints for new antimicrobial that target adhesins.

An SH3-binding allosteric modulator stabilizes the global conformation of the AML-associated Src-family kinase, Hck.

While ATP-site inhibitors for protein-tyrosine kinases are often effective drugs, their clinical utility can be limited by off-target activity and acquired resistance mutations due to the conserved nature of the ATP-binding site. However, combining ATP-site and allosteric kinase inhibitors can overcome these shortcomings in a double-drugging framework. Here we explored the allosteric effects of two pyrimidine diamines, PDA1 and PDA2, on the conformational dynamics and activity of the Src-family tyrosine kinase Hck, a promising drug target for acute myeloid leukemia. Using 1H-15N HSQC NMR, we mapped the binding site for both analogs to the SH3 domain. Despite the shared binding site, PDA1 and PDA2 had opposing effects on near-full-length Hck dynamics by hydrogen-deuterium exchange mass spectrometry, with PDA1 stabilizing and PDA2 disrupting the overall kinase conformation. Kinase activity assays were consistent with these observations, with PDA2 enhancing kinase activity while PDA1 was without effect. Molecular dynamics simulations predicted selective bridging of the kinase domain N-lobe and SH3 domain by PDA1, a mechanism of allosteric stabilization supported by site-directed mutagenesis of N-lobe contact sites. Cellular thermal shift assays confirmed SH3 domain-dependent interaction of PDA1 with wild-type Hck in myeloid leukemia cells and with a kinase domain gatekeeper mutant (T338M). These results identify PDA1 as a starting point for Src-family kinase allosteric inhibitor development that may work in concert with ATP-site inhibitors to suppress the evolution of resistance.

Targeting Breast Cancer: Novel Dihydropyrimidinones As Potent Eg5 Inhibitors

Introduction: Breast cancer remains a formidable health concern for women, necessitating the development of potent anticancer agents with improved safety profiles. Dihydropyrimidinones (DHPM), pyrazole, and benzofuran scaffolds have emerged as promising targets due to their diverse pharmacological profiles. In this study, we employed a scaffold hopping approach to design a novel DHPM-Pyrazole-Benzofuran core. A series of compounds (3a–3j) were synthesized using the Biginelli protocol, and their characterization was performed using various techniques such as FTIR, 1H NMR, and Mass spectroscopy. Methods: Molecular docking studies against kinesin spindle protein Eg5 (1Q0B) performed to find superior binding interactions compared to the prototype Eg5 inhibitor Monastrol. Anti breast cancer potential of these compounds was screened against the breast adrenocarcinoma MCF-7 cell line using an SRB assay. Results: Compound 3j showed good growth inhibitory activity (GI50=24.08μM) compared to Monastrol (GI50=32μM) employed as a positive control. Moreover, Compound 3j exhibited strong interactions with amino acids GLU-116 and ARG-119 with Eg5 protein 1Q0B. Conclusion: Compound 3j fits well at the allosteric site of Eg5 protein 1QOB. Compound 3j emerged as the most cytotoxic, displaying significant and impressive growth inhibitory activity (GI50=24.08μM).

Sex-mediated effects of transglutaminase 2 inhibition on endothelial function in human resistance arteries from diabetic and non-diabetic patients.

Transglutaminase 2 (TG2) is an enzyme with multiple conformations. In its open conformation, TG2 exhibits transamidase activity linked to fibrosis, arterial remodeling, and endothelial dysfunction, a process enhanced by high glucose in endothelial cells. However, the closed conformation of TG2 contributes to transmembrane signaling and nitric oxide (NO)-dependent vasorelaxation. LDN 27219, a reversible allosteric inhibitor, stabilizes TG2 in its closed conformation. &#160;We examined whether pharmacological modulation of TG2 into its closed conformation induces vasorelaxation and enhances endothelium-dependent and independent relaxation in resistance arteries from age-matched diabetic (n=14) and non-diabetic patients (n=14) (age 71 (SEM: ±2).</p> &#160;Subcutaneous arteries (diameter 133-1013 µm) were isolated from abdominal fat biopsies. TG2 mRNA expression and transamidase activity were assessed via RT-qPCR and 5-biotin(amido)pentylamine (5-BP) incorporation, while vascular reactivity was measured using wire myography. TG2 mRNA was highly expressed without significant differences between the groups and LDN 27219 induced concentration-dependent vasorelaxation in arteries from both groups. Sex-specific analysis revealed that potentiation of acetylcholine-induced vasorelaxation by LDN 27219 was driven by increased TG2 expression in non-diabetic females, while no effect was observed in arteries from non-diabetic males. Among diabetic patients, LDN 27219 increased maximal acetylcholine-induced vasorelaxation in males only. LDN 27219 did not affect endothelium-independent relaxation to sodium nitroprusside in either group.</p> In conclusion, TG2 is expressed in human resistance arteries, and LDN 27219 induced vasorelaxation, selectively enhancing ACh relaxation in non-diabetic females, likely due to increased TG2 expression. This finding underscores the importance of sex differences in TG2 modulation of vasorelaxation.

Evolutionary Adaptations in Biliverdin Reductase B: Insights into Coenzyme Dynamics and Catalytic Efficiency.

Biliverdin reductase B (BLVRB) is a redox regulator that catalyzes nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductions of multiple substrates, including flavins and biliverdin-β. BLVRB has emerging roles in redox regulation and post-translational modifications, highlighting its importance in various physiological contexts. In this study, we explore the structural and functional differences between human BLVRB and its hyrax homologue, focusing on evolutionary adaptations at the active site and allosteric regions. Using NMR spectroscopy, we compared coenzyme binding, catalytic turnover, and dynamic behavior between the two homologues. Despite lacking the arginine "clamp" present in human BLVRB, hyrax BLVRB still undergoes conformational changes in response to the oxidative state of the coenzyme. Mutations at the allosteric site (position 164) show that threonine at this position enhances coenzyme discrimination and allosteric coupling in human BLVRB, while hyrax BLVRB does not display the same allosteric effects. Relaxation experiments revealed distinct dynamic behaviors in hyrax BLVRB, with increased flexibility in its holo form due to the absence of the clamp. Our findings suggest that the evolutionary loss of the active site clamp and modifications at position 164 in hyrax BLVRB alter the enzyme's conformational dynamics and coenzyme interactions. Identified similarities and differences underscore how key regions modulate catalytic efficiency and suggest that coenzyme isomerization may represent the rate-limiting step in both homologues.

Abstract PR004: Brain penetrant allosteric EGFR inhibitors for NSCLC

Abstract Approximately 30-50% of Non-Small Cell Lung Cancer (NSCLC) patients already have CNS disease at time of diagnosis or else develop it during treatment, and there remains a need for better outcomes in these patients. The 3rd-generation covalent EGFR inhibitor osimertinib is an effective therapy for NSCLC patients, improving upon 1st-generation inhibitors gefitinib and erlotinib, and achieving enhanced brain penetration relative to those agents. However, the survival benefit relative to the 1st-generation inhibitors is observed primarily in patients with exon19 deletions rather than the L858R mutation. We developed a series of mutant-selective allosteric EGFR inhibitors that specifically target the L858R mutation (and subsequent resistance mutations), with high selectivity over wt EGFR and across the kinome. Early compounds including the isoindolinone JBJ-09-063 demonstrated good in vivo efficacy in mouse tumor models, despite limited bioavailability. Diversification and optimization of the chemistry, involving a scaffold hopping approach and the replacement of the thiazole amide with benzimidazole, delivered a set of compounds with promising potency and rodent PK. A subset of these achieved good brain exposure in mice and efficacy in an intracranial H1975 brain metastasis model. The mutant-selectivity and ability to co-bind with osimertinib makes an allosteric EGFR inhibitor an ideal partner for combination therapy, with the potential to deliver enhanced outcomes in L858R-driven NSCLC, especially for patients with CNS disease. Citation Format: David Scott, Courtney Cullis, Tyler Beyett, Frederic Feru, Thomas Gero, Krista Gipson, Praful Gokhalle, Nathanael Gray, David Heppner, Sampson Huang, Pasi Jänne, Sandeepraj Pusalkar, Michael Eck. Brain penetrant allosteric EGFR inhibitors for NSCLC [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Optimizing Therapeutic Efficacy and Tolerability through Cancer Chemistry; 2024 Dec 9-11; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(12_Suppl):Abstract nr PR004

Abstract A008: Evaluation of natural and synthetic compounds on epithelial to mesenchymal transition in triple negative breast cancer

Abstract Epithelial to mesenchymal transition (EMT) is a biological process through which epithelial cells undergo cytoskeletal changes and transform into a mesenchymal phenotype. This phenotypical transformation leads to enhanced migratory capabilities and increased resistance to apoptosis and drug therapies. The EMT characterized in part by an increase in vimentin, a mesenchymal marker, and a decrease in E-cadherin, an epithelial marker, plays a significant role in cancer metastasis and progression. The MEK5/ERK5 pathway is essential in regulating cell survival, proliferation, and migration. Inhibition of the MEK5/ERK5 pathway by small molecule inhibitors decreases cellular proliferation and impedes migration of cancer cells, including triple negative breast cancer (TNBCs) cells. A type III allosteric inhibitor of MEK5 designed by our collaborators reverses the EMT and induces a mesenchymal to epithelial transition (MET). Using a TNBC cell line, MDA-MB-231, genetically engineered with RFP tagged vimentin, we established a cellular assay to evaluate analogues of our lead type III inhibitor to further investigate the role of the MEK5-ERK pathway in the EMT, design optimal compounds that induce an MET, and explore structure-activity relationships for kinase inhibition. This research will help identify targets for EMT and optimize compound modification to improve potency and efficiency. Citation Format: Asef Faruk, Saloni Patel, Ramez Hallak, Patrick Flaherty, Jane Cavanaugh. Evaluation of natural and synthetic compounds on epithelial to mesenchymal transition in triple negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Optimizing Therapeutic Efficacy and Tolerability through Cancer Chemistry; 2024 Dec 9-11; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(12_Suppl):Abstract nr A008.

Bioavailability and molecular recognition between secondary metabolites of Euphorbia hirta L. and 5-lipoxygenase using fragment molecular orbital method and pair interaction energy decomposition analysis

Is presented the bioavailability analysis for seven secondary metabolites from Euphorbia hirta L. (Euphorbiaceae). The bioactivity score, the molecular similarity analysis for each secondary metabolite, and the molecular docking 5-LOX/metabolite complex are also presented. For the similarity analysis, it is referenced the structure of allosteric LOX-5 inhibitor 3-acetyl-11-keto-beta-boswellic acid (AKBA). The fragment molecular orbital method (FMO) and pair interaction energy decomposition analysis (PIEDA) were applied to evaluate the interaction energy between secondary metabolites and the 5-LOX active site. Log P values of all the secondary metabolites studied were found to be higher than 5, contrary to what was expected with Lipinski’s rules of five. The reference LOX-5 ligand has a high electrostatic potential similarity with most secondary metabolites studied, with values from 0.667 to 0.730. Both AKBA and metabolites ligands were wedged between the two domains of LOX-5, and the main residues contact were LEU-66, ARG-68, VAL-110, HIS-130, ILE-126, GLN, 129, and LYS-133. The PIEDA analysis showed that stabilization in the 5-LOX active site is mainly dominated by hydrophobic interactions, with values up to -35 Kcal/mol. Due to the low presence of terminal-OH groups, electrostatic origin, and charge transfer interaction energies are also important but have values of less than 20 Kcal/mol.

Combined treatment with ruxolitinib and MK-2206 inhibits ERα activity by inhibiting MAPK signaling in BT474 breast cancer cells.

Triple-positive breast cancer (TPBC) is a type of breast cancer that overexpresses estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor-2 (HER2). Dysregulation of ER signaling has been implicated in the pathogenesis of breast cancer. ERα activation triggers the production of second messengers, including cAMP, leading to the activation of signals such as PI3K/AKT or Ras/MAPK. Ruxolitinib is a specific inhibitor of JAK1/JAK2. MK-2206 is an allosteric inhibitor of the Akt. The limitations of the use of ruxolitinib and MK-2206 as single agents necessitate the development of combination therapies with other drugs. This study is the first to investigate the effects of combining ruxolitinib with MK-2206 on MAPK and PI3K/AKT signaling in BT474 breast cancer cells. In addition, this work aimed to increase the anticancer effects of cotreatment with MK-2206 and ruxolitinib. Ruxolitinib, MK-2206, and their combination reduced cell viability in a dose- and time-dependent manner, as determined by MTT assays after 48 h of treatment. Colony formation and wound healing assays demonstrated that MK-2206 exhibited a synergistic anti-proliferative effect. The effects of ruxolitinib, MK-2206, and their combination on PI3K/AKT and MAPK signaling were assessed via western blotting. Ruxolitinib and MK-2206 combined treatment inhibit cell death in BT474 cells by downregulating ERα, Src-1, ERK1/2, SAPK/JNK, and c-Jun. Our results revealed the relationships among the ERα, PI3K/AKT, and MAPK signaling pathways in ER+ breast cancer cells. Understanding the interactions among ERα, PI3K-AKT-mTOR, and MAPK could lead to novel combination therapies.

Multiple mechanisms of allosteric regulation of the luteninizing hormone receptor

The regulatory effects of luteinizing hormone (LH) and chorionic gonadotropin (CG) are realized through the activation of the G-protein coupled LH/CG receptor (LH/CG-R). The result of this is the activation of various types of G proteins, which leads to stimulation (Gs) or inhibition (Gi) of the cAMP-dependent pathway and stimulation of calcium signaling (Gq/11, Gi), and the recruitment of β-arrestins, which prevent G protein signaling through receptor internalization and downregulation, but can also activate the mitogen-activated protein kinase cascade. Despite a certain similarity in the effects of LH and CG, there are differences between them both in efficiency and in the pattern of regulation of LH/CG-R. This is a consequence of differences in the affinity of LH and CG to the orthosteric site of the receptor, as well as differences at the level of allosteric regulation of the receptor, which is due to the presence of a C-terminal extension in the β-subunit of CG, including sites for O-glycosylation, and the variability of N-glycosylation of α- and β-subunits of gonadotropins. Moreover, the number of N-glycans, the degree of their branching and charge differ, which leads to different efficiency of activation of intracellular cascades, affecting the physiological response of the reproductive system to gonadotropins. Of great importance is the formation of homodi(oligo)meric complexes of LH/CG-R and its heterocomplexes with the follicle-stimulating hormone receptor, where protomers allosterically influence the efficiency of LH/CG-R activation and the bias of signal transduction. Taking into account the large number of allosteric sites in LH/CG-R, the development of low-molecular allosteric regulators is underway, including agonists based on thieno[2,3-d]-pyrimidine and peptides derived from the cytoplasmic loops of LH/CG-R. These regulators can become prototypes of drugs for correcting the functions of the reproductive system. This review is devoted to the analysis of data on the similarities and differences in the signaling and physiological effects of gonadotropins with LH activity, the role of allosteric mechanisms in this, and the prospects for creating allosteric regulators of LH/CG-R.

Intricate Structure-Function Relationships: The Case of the HtrA Family Proteins from Gram-Negative Bacteria.

Proteolytic enzymes play key roles in living organisms. Because of their potentially destructive action of degrading other proteins, their activity must be very tightly controlled. The evolutionarily conserved proteins of the HtrA family are an excellent example illustrating strategies for regulating enzymatic activity, enabling protease activation in response to an appropriate signal, and protecting against uncontrolled proteolysis. Because HtrA homologs play key roles in the virulence of many Gram-negative bacterial pathogens, they are subject to intense investigation as potential therapeutic targets. Model HtrA proteins from bacterium Escherichia coli are allosteric proteins with reasonably well-studied properties. Binding of appropriate ligands induces very large structural changes in these enzymes, including changes in the organization of the oligomer, which leads to the acquisition of the active conformation. Properly coordinated events occurring during the process of HtrA activation ensure proper functioning of HtrA and, consequently, ensure fitness of bacteria. The aim of this review is to present the current state of knowledge on the structure and function of the exemplary HtrA family proteins from Gram-negative bacteria, including human pathogens. Special emphasis is paid to strategies for regulating the activity of these enzymes.

Disruption of LEDGF/p75-directed integration derepresses antisense transcription of the HIV-1 genome.

Disruption of HIV-1 Integrase (IN) interactions with the host-factor Lens Epithelium-Derived Growth Factor (LEDGF)/p75 leads to decreased, random integration, increased latent infection, and described here, accumulation of HIV-1 antisense RNA (asRNA). asRNA increase was observed following interruptions of IN-LEDGF/p75 interactions either through pharmacologic perturbations of IN-LEDGF/p75 by treatment with allosteric HIV-1 integrase inhibitors (ALLINIs) or in cell lines with LEDGF genetic knockout. Additionally, by impairing Tat-dependent HIV transcription, asRNA abundance markedly increases. Illumina sequencing characterization of asRNA transcripts in primary T cells infected in the presence of ALLINIs showed that most initiate from within the HIV-1. Overall, loss of IN-LEDGF/p75 interactions increase asRNA abundance. Understanding the relationship between ALLINIs, integration sites, asRNA, and latency could aid in future therapeutic strategies.

Role of Computer-Aided Design and Development in Modern Pharmaceuticals

The integration of molecular biology and computational technologies has transformed modern drug discovery and development processes. Advanced computational methodologies, particularly artificial intelligence (AI) and machine learning (ML), have reshaped traditional drug development approaches through unprecedented access to ligand property data, target binding interactions, three-dimensional protein structures, and virtual libraries containing billions of drug-like molecules. AI and deep learning (DL) implementations have enhanced multiple stages of drug discovery, from target identification to lead optimization. These computational advances, backed by improved hardware capabilities and sophisticated algorithms, now enable targeting of previously "undruggable" proteins. This review presents modern computational approaches in pharmaceutical development, including strategies for challenging protein targets through covalent regulation, allosteric inhibition, protein-protein interaction modulation, and targeted protein degradation. AI-driven methods have accelerated drug discovery pipelines, reduced development costs, and improved clinical trial success rates. The transition from traditional broad-spectrum approaches to precision medicine, supported by computational tools, has enabled personalized therapeutic strategies. Current limitations in computer-aided drug design persist, yet the combination of computational predictions with experimental validation continues to advance therapeutic development. Recent developments in quantum computing and advanced neural networks promise to further enhance drug discovery efficiency and success rates in the coming decades.

P114 Deucravacitinib treatment did not affect immune response to SARS-Cov-2 vaccines and infection in patients with plaque psoriasis: results from the Phase 3 POETYK long-term extension trial

Abstract Introduction Deucravacitinib, an oral, selective, allosteric TYK2 inhibitor, is approved in the US, EU, and other countries for the treatment of adults with moderate-to-severe plaque psoriasis who are candidates for systemic therapy. Deucravacitinib was efficacious and well tolerated in the global, 52-week, Phase 3 POETYK PSO-1 (NCT03624127) and POETYK PSO-2 (NCT03611751) trials. Patients completing the parent trials could enrol in the ongoing, open-label POETYK long-term extension (LTE) (NCT04036435), which overlapped with the peak of the global COVID-19 (SARS-CoV-2) pandemic. We investigated serological responses and predictors of seroconversion to SARS-CoV-2 vaccination and/or infection during deucravacitinib treatment in the POETYK LTE. Methods POETYK LTE started in August 2019 and overlapped with the COVID-19 pandemic, which emerged in December 2019. The first infection in the POETYK program was reported 24 March 2020, the first vaccination occurred on 17 December 2020, and samples were collected through 2 August 2023. This analysis included patients who (i) were fully vaccinated with an mRNA vaccine or a non-mRNA vaccine or had a reported SARS-CoV-2 infection during the trial and (ii) had their first serum sample available ≥15 days after the second mRNA dose or ≥30 days and up to 229 days after a non-mRNA vaccine or infection. Spike RBD antibody level ≥0.8 U/mL and nucleocapsid antibody level ≥1.0 cut-off index were used as measures of seroconversion; nucleocapsid antibody levels ≥1.0 U/mL were used as an indicator of prior SARS-CoV-2 infection. Results 596 (87.8%) patients were vaccinated (mRNA vaccine, n = 498; non-mRNA vaccine, n = 98). Baseline characteristics were similar between patients who were vaccinated (n = 406), infected (n = 83), and both vaccinated and infected (n = 190). Seroconversion occurred in 99.2% of mRNA vaccine recipients and 98.9% of non-mRNA vaccine recipients [mean RBD antibody levels, 9085.1 and 4277.9 U/mL, respectively; range, 0.4–75 000 U/mL (seroconversion, ≥0.8 U/mL titre)]. Seroconversion occurred in 100% of infected unvaccinated patients (mean RBD antibody level, 3663.8 U/mL), 99.1% of noninfected vaccinated patients (6384.2 U/mL), and 100% of infected vaccinated patients (23 636.2 U/mL). The mean duration between reported infection and sample collection was 177 days. RBD antibody levels remained high for &amp;gt;24 weeks in mRNA vaccine recipients, regardless of the time after vaccination or infection. Age, body mass index, and sex were not predictors of antibody levels or seroconversion. Conclusions In the POETYK LTE trial, &amp;gt;98% of patients mounted a serologic response to SARS-CoV-2 vaccination and/or infection, with more robust responses in vaccinated than unvaccinated patients. Deucravacitinib did not impact immune responses to vaccines that protect against COVID-19.

P120 Deucravacitinib, an oral, selective tyrosine kinase 2 inhibitor, in patients with moderate-to-severe scalp psoriasis: laboratory parameter results of a Phase 3b/4, multicentre, randomized, double-blind, placebo-controlled trial (PSORIATYK SCALP)

Abstract Introduction Deucravacitinib, an oral, selective, allosteric TYK2 inhibitor, is approved in the US, EU, and other countries for treatment of adults with moderate-to-severe plaque psoriasis who are candidates for systemic therapy. Scalp psoriasis occurs in ∼80% of patients with plaque psoriasis; is associated with itching, pain, and bleeding; disproportionately reduces quality of life; and is challenging to treat with topical agents. The 52 week, Phase 3b/4, double-blind PSORIATYK SCALP trial evaluated deucravacitinib efficacy and safety in patients with moderate-to-severe scalp psoriasis, including those with more limited overall psoriasis; these patients are candidates for systemic therapy according to IPC guidelines. Here, the effect of deucravacitinib on changes in laboratory parameters associated with Janus kinase (JAK) 1,2,3 inhibitors was evaluated. Methods Adults (aged ≥18 years) with moderate-to-severe scalp psoriasis defined by more focused and objective inclusion criteria (ss-PGA ≥3, scalp surface area involvement ≥20%, PSSI ≥12) and body surface area involvement ≥3% were randomized 1:2 to oral placebo or deucravacitinib 6 mg once daily through Week 16. Changes from baseline in lipid (total cholesterol, triglycerides), chemistry [creatinine, alanine aminotransferase (ALT), aspartate aminotransferase (AST)], and haematology (lymphocytes, neutrophils, platelets, haemoglobin) parameters in blood were evaluated through Week 16 (primary endpoint). Laboratory parameter adverse events, CTCAE Grade ≥3 laboratory abnormalities, shifts from baseline in severity grade of laboratory abnormalities, and discontinuations due to laboratory abnormalities were evaluated. Results One hundred and fifty-four patients were randomized (placebo, n = 51; deucravacitinib, n = 103). No clinically meaningful mean changes from baseline or placebo were observed in any laboratory parameter, including triglycerides, ALT, and AST, through Week 16. Hypercholesterolaemia was reported in one patient in each group; an elevated liver function test was reported in one patient receiving placebo. Grade ≥3 laboratory abnormalities were rare in both groups (deucravacitinib, Grade 3 triglyceride elevation; placebo, Grade 4 triglyceride elevation). Most deucravacitinib-treated patients maintained their baseline laboratory abnormality grade or shifted to a lower grade. No treatment discontinuations or interruptions were reported due to laboratory abnormalities. No muscle-related events linked to creatine phosphokinase elevations were reported. Conclusions This laboratory parameter profile is consistent with results of the Phase 3 POETYK PSO-1, PSO-2, and long-term extension trials, in which deucravacitinib was safe and well-tolerated in patients with plaque psoriasis, including those with moderate-to-severe scalp psoriasis. Signature laboratory changes commonly seen with JAK1,2,3 inhibitors were not reported with deucravacitinib. These results support the safety profile of once-daily oral deucravacitinib in patients with moderate-to-severe scalp psoriasis including those with more limited overall plaque psoriasis.

(-)-Syringaresinol Exerts an Antidepressant-like Activity in Mice by Noncompetitive Inhibition of the Serotonin Transporter.

Albizia julibrissin Durazz. is one of the most popular herbs used for depression treatment, but the molecular basis for its mechanism of action has not been fully addressed. Previously, we isolated and identified two lignan glycoside derivatives that were shown to noncompetitively inhibit serotonin transporter (SERT) activity but with a relatively low inhibitory potency compared with those of conventional antidepressants. We characterized the pharmacological profile of the parental compound of these previously isolated lignan glycosides, (-)-syringaresinol (SYR), in inhibiting SERT by using biochemical, pharmacological, and behavioral approaches. SYR, as a potent inhibitor, decreases SERT Vmax but with little change in Km for its fluorescent substrate. SYR was shown to block the conformational conversion essential for substrate transport by stabilizing SERT in an outward-open and inward-closed conformation. In addition, our molecular docking and biochemical validation demonstrated that SYR binds to an allosteric site in SERT and noncompetitively inhibits SERT transport and binding activity. Furthermore, administration of SYR was indicated to exert an antidepressant-like activity and to effectively attenuate chronic unpredictable mild stress (CUMS)-induced abnormalities in behaviors and synaptic protein expression in depressive animal models. This study not only provides molecular insights into the mechanism of action of A. julibrissin in the treatment of depression, but also opens up the possibility of development of a novel class of allosteric site-targeted therapeutic agents with an underlying mechanism of action different from that of conventional antidepressants.

Structure of the SMYD2-PARP1 Complex Reveals Both Productive and Allosteric Modes of Peptide Binding.

Allosteric regulation allows proteins to dynamically respond to environmental cues by modulating activity at sites away from the catalytic center. Despite its importance, the SET-domain protein lysine methyltransferase superfamily has been understudied. Here, we present four crystal structures of SMYD2, a unique family member with a MYND domain. Our findings reveal a novel allosteric binding site with high conformational plasticity and promiscuity, capable of binding peptides, proteins, PEG, and small molecules. This site exhibits positive cooperativity with substrate binding, influencing catalytic activity. Mutations here significantly alter substrate affinity, changing the enzyme's kinetic profile. Specificity studies show interaction with PARP1 but not histones, suggesting targeted regulation. Interestingly, this site's function remains unaffected by active site changes, indicating unidirectional mechanisms. Our discovery provides novel insights into SMYD2's biochemical regulation and lays the foundation for broader research on allosteric control in lysine methyltransferases. Given SMYD2's role in various cancers, this work opens exciting avenues for designing specific allosteric inhibitors with reduced off-target effects.

Tilting the Scales toward EGFR Mutant Selectivity: Expanding the Scope of Bivalent "Type V" Kinase Inhibitors.

Binding multiple sites within proteins with bivalent compounds is a strategy for developing uniquely active agents. A new class of dual-site inhibitors has emerged targeting the epidermal growth factor receptor (EGFR) anchored to both the orthosteric (ATP) and allosteric sites. Despite proof-of-concept successes, enabling selectivity against oncogenic activating mutations has not been achieved and classifying these inhibitors among kinase inhibitors remains underexplored. This study investigates the structure-activity relationships, binding modes, and biological activity of ATP-allosteric bivalent inhibitors (AABIs). We find that AABIs selectively inhibit drug-resistant EGFR mutants (L858R/T790M and L858R/T790M/C797S) by anchoring a methyl isoindolinone moiety along the αC-helix channel of the allosteric site. In contrast, related Type I1/2 inhibitors target wild-type EGFR but are less effective against resistant mutants. This shift in selectivity demonstrates that mutant-selective AABIs classify as "Type V" bivalent inhibitors.