In this study, we describe a series of pyridone derivatives as pan-MEK/RAF nondegrading molecular glues. Through investigation of the metabolic sites of the reported MEK/RAF inhibitor 16b, rational design and systematic SAR studies led to the discovery of compound D56, which exhibits well-balanced in vitro and in vivo potency. D56 could effectively block MEK and ERK phosphorylation with IC50 values of 0.379 and 0.015 nM, respectively. Furthermore, protein-protein interaction (PPI) assays demonstrated that D56 induces MEK1-BRAF and MEK1-CRAF complexes formation at low concentrations, indicating that D56 is a potent MEK/RAF molecular glue. D56 possesses an excellent selectivity over other 332 human-related kinases at 1 μM. Most importantly, in AsPC-1, HCT116, and OCI-AML-3 mouse xenograft models, D56 achieved significant tumor growth inhibition. Taken together, these findings suggest that compound D56 is a potent pan-MEK/RAF nondegrading molecular glue for treating RAS-driven cancers.

Taste G protein-coupled receptors (GPCRs), including sweet (TAS1Rs) and bitter (TAS2Rs) taste receptors, are essential for detecting nutrients and avoiding toxins, influencing dietary behavior, metabolism, and overall health. Beyond their role in taste perception, these receptors are widely expressed in extra-gustatory tissues, including the respiratory and gastrointestinal systems, where they regulate innate immune responses, hormone secretion, and energy balance. Dysfunctions or polymorphisms in TAS1Rs and TAS2Rs have been linked to various diseases such as asthma, type 2 diabetes, obesity, dental caries, periodontitis, and certain cancers. The structural features of these receptors, including their ligand-binding domains and signaling pathways, are central to their diverse functions. Recent studies also highlight their potential as therapeutic targets for managing conditions like metabolic syndrome and immune-related disorders. This review provides a detailed examination of the structural and functional dynamics of TAS1Rs and TAS2Rs, emphasizing their roles in disease mechanisms and exploring therapeutic strategies. While challenges remain in structural resolution and functional characterization, advancements in molecular modeling and pharmacological approaches shed light on their clinical potential. Understanding the tissue-specific roles and molecular mechanisms of taste GPCRs can pave the way for innovative treatments targeting these receptors, offering significant promise in addressing a range of health conditions.

Psychedelics are undergoing a renaissance as potential therapy for psychiatric disorders, with more than 200 clinical trials being studied across several countries1-3. However, the precise mechanisms by which these drugs bring about benefits and the potential clinical risks are not yet fully understood. The serotonin 2A receptor (5-HT2AR) was reported to be a Gq-coupled receptor and the primary interoceptive target of psychedelics4,5. Here we compared psychedelics and their non-hallucinogenic analogues (nHAs) using in vitro and in vivo approaches, finding that 5-HT2AR-mediated non-canonical Gi signalling is essential for hallucinogenic effect. We further presented five cryo-electron microscopy structures of 5-HT2AR-Gi/Gq in complex with psychedelics or nHAs. Structural analysis and pharmacological investigation revealed that a special contact between nHAs with 5-HT2AR mediated the signalling bias. Building on this insight, we identified a 2,5-dimethoxy-4-iodoamphetamine derivative, DOI-NBOMe, which exhibits potent and selective Gq-biased activity, and demonstrates promising therapeutic effects in mouse models without hallucinogenic effect. Our finding uncovers the functional mechanisms underlying the Gi signalling mediated by 5-HT2AR and provides valuable insights for designing psychedelic-based drugs with minimized risk from hallucinogenic effects.

Developing highly selective PARP1 inhibitors is a promising strategy to improve the therapeutic window compared with dual PARP1/2 inhibitors. Through structure-guided design and optimization, we developed (R)-A17, a novel, highly selective PARP1 inhibitor featuring a unique tricyclic scaffold. It demonstrates nanomolar enzymatic potency (PARP1 IC50 = 2.4 nM), exceptional 65.8-fold selectivity over PARP2, and robust antiproliferative activity in BRCA-deficient cells. (R)-A17 exhibits favorable pharmacokinetic properties with complete oral bioavailability (F = 100%) in mice. Mechanistically, it selectively inhibits PARP1-mediated PARylation and induces DNA damage and G2/M phase arrest. In vivo, oral administration of (R)-A17 produced dose-dependent antitumor efficacy in BRCA mutant xenograft models, achieving tumor growth inhibition values of 57.8, 86.3, and 91.3% at doses of 0.3, 1, and 3 mg/kg, respectively, while also synergizing effectively with liposomal irinotecan in an HCT116 xenograft model. These results establish (R)-A17 as a promising candidate and validate the design strategy for the next-generation PARP1-targeted therapy.

Opioid analgesics provide potent pain relief but are limited by severe adverse effects, tolerance, and interindividual genetic variability in response. Poly-pharmacology and allosteric modulation of opioid receptors offer promising strategies to enhance analgesic efficacy while mitigating these limitations. Pan-positive allosteric modulators (pan-PAMs), which simultaneously potentiate multiple opioid receptor subtypes, integrate the advantages of both approaches and represent an emerging therapeutic paradigm for pain management. However, the molecular mechanisms underlying pan-PAM activity at opioid receptors remain poorly understood. Here, we characterize BMS-986187 as a pan-PAM of opioid receptors and report the cryo-electron microscopy (cryo-EM) structures of multiple opioid receptor subtypes bound to this modulator, revealing a previously unidentified allosteric pocket. Structural and functional analyses revealed a conserved binding motif that mediates PAM recognition across the opioid receptor family and revealed the essential contributions of key opioid receptor residues to allosteric modulation by BMS-986187. Functionally, BMS-986187 enhances analgesic efficacy through an opioid-sparing effect, allowing lower opioid doses and reducing side effects, while restoring activity in loss-of-function (LOF) μ-opioid receptor variants. These findings define a previously unrecognized allosteric site in opioid receptors and establish a structural framework for the rational design of safer and more effective opioid therapeutics through allosteric modulation.

For decades, microtubules—composed of αβ-tubulin dimers—have been primary targets for cancer chemotherapy. While eight binding sites on the tubulin dimer have been structurally characterized, this study reveals a ninth. We found that the tubulin inhibitor Tumabulin-1 (TM1, a BML284 derivative) binds simultaneously to the well-known colchicine site and a previously unknown site, designated as Tumabulin site. This site resides at the interface of α1-tubulin, β1-tubulin, and RB3 within the tubulin–RB3–tubulintyrosine ligase complex. Remarkably, two TM1 molecules bind cooperatively to this relatively large pocket, interacting with all three proteins. Crucially, this binding is dependent on RB3; it is absent when RB3 is missing or the key residue H71 is mutated (H71Q). We further designed and synthesized Tumabulin-2 (TM2) that selectively binds the Tumabulin site, excluding binding the colchicine site. TM2 acts as a molecular glue, strengthening the interaction between RB3 and the tubulin dimer and consequently enhancing RB3’s tubulin-depolymerizing activity. In conclusion, our findings confirm the existence of a ninth tubulin-binding site and offer a promising foundation for developing Tubulin–RB3 molecular glues as a next generation of anticancer therapeutics.

G protein-coupled receptors (GPCRs) are major drug targets, yet genetic variations in these receptors can alter drug responses, leading to significant challenges in healthcare. Despite the prevalence of GPCR-targeting drugs, the effects of these genetic variations on receptor function remain underexplored. This study establishes a framework for allosterically rescuing loss-of-function (LoF) variants in GPCRs, using the dopamine receptor D1 (DRD1) as a model. We characterized 49 DRD1 variants from genetic databases and literature, finding that most variants exhibit reduced membrane expression. Structural analysis indicated that variants within the ligand-binding pocket or near critical activation motifs may impair ligand binding or hinder conformational changes during receptor activation, potentially disrupting orthosteric ligand induced signaling. We categorized the variants into three functional groups: those with enhanced G protein signaling, enhanced β-arrestin recruitment, or complete LoF. Among these, 16 variants disrupt G protein signaling, and 27 impair β-arrestin recruitment in HEK293 cells. Notably, defective G protein signaling caused by LoF variants such as T371.46K and L662.46F were effectively restored using allosteric modulators. These findings highlight the functional impact of DRD1 variants and demonstrate the therapeutic potential of relative selectivity of two signal pathways. This study advances precision medicine by offering strategies to restore receptor function and develop targeted therapies for GPCR-related disorders.

Cholesterol plays a pivotal role in modulating the activity of mechanistic target of rapamycin complex 1 (mTOR1), thereby regulating cell growth and metabolic homeostasis. LYCHOS, a lysosome-localized G-protein-coupled receptor-like protein, emerges as a cholesterol sensor and is capable of transducing the cholesterol signal to affect the mTORC1 function. However, the precise mechanism by which LYCHOS recognizes cholesterol remains unknown. Here, using cryo-electron microscopy, we determined the three-dimensional structural architecture of LYCHOS in complex with cholesterol molecules, revealing a unique arrangement of two sequential structural domains. Through a comprehensive analysis of this structure, we elucidated the specific structural features of these two domains and their collaborative role in the process of cholesterol recognition by LYCHOS. LYCHOS is a lysosome-localized cholesterol sensor involved in mTORC1 activity regulation. Here, the authors report the three-dimensional structure of LYCHOS bound to cholesterols, revealing the recognition mechanism for this molecule.

Our preliminary studies indicate that cevipabulin concurrently binds to both the vinca site and the gatorbulin site, and promotes tubulin degradation. To improve its antiproliferative activity and investigate the structure-activity relationships (SARs), thirty-eight cevipabulin derivatives were designed and synthesized based on the cevipabulin-tubulin cocrystal structure. Among them, compound 8g exerted optimal antiproliferative activity, with IC50 values ranging from 0.016 to 0.035 μM against three tested tumor cell lines. The cocrystal structure of the 8g-tubulin complex revealed that it simultaneously occupies both the vinca site and the gatorbulin site, while maintaining a binding mode similar to that of cevipabulin. Furthermore, 8g promoted αβ-tubulin degradation and displayed good oral bioavailability. In an HT29 xenograft model, oral administration of 8g at doses of 20 and 40 mg/kg every 3 days resulted in potent in vivo antitumor activity, with tumor growth inhibition (TGI) rates of 41.0 % and 49.5 %, respectively. Moreover, 8g exhibited significantly reduced toxicity and fewer adverse effects compared to cevipabulin, supporting its potential as a promising therapeutic agent for cancer treatment.

Olfaction plays a fundamental role in survival and socialization of human beings, and its dysfunction not only includes deficits in smell but also contributes to the progression of various diseases. This review explores the role of mammalian olfactory receptors in odor perception and signaling within the olfactory system, their expression outside olfactory tissues, and recent structural discoveries. The involvement of olfactory receptors is investigated in diseases such as Alzheimer's and immune disorders, highlighting their potential as therapeutic targets. The olfactory activation mechanisms are summarized, offering a valuable reference for disease research and drug development targeting olfactory receptors, with the goal of advancing treatments for olfactory-related diseases.

BackgroundExtracellular leucine rich repeat and fibronectin type III domain containing 1 (ELFN1), a transmembrane protein implicated in tumorigenesis and therapy resistance, remains mechanistically undefined as a pan-cancer target. In this study, we aimed to elucidate the function and potential mechanism of action of ELFN1 across cancers.MethodsThrough integrative analysis of TCGA and GTEx datasets, we systematically characterized ELFN1 across 33 cancer types, including its expression patterns, prognostic value, mutation landscape, methylation modifications, protein-protein interaction (PPI) networks, and the relationship between ELFN1 expression and immune infiltration. KEGG enrichment analysis was also performed to predict the functions and associated cellular pathways of ELFN1. In addition, the molecular docking tool was used to analyze the affinities between ELFN1 protein and drugs. Finally, we assessed the effect of ELFN1 knockdown on colorectal cancer (CRC) cells using in vitro experiments.ResultsOur study revealed significant dysregulation of ELFN1 across various cancer types, with notable diagnostic and prognostic utility in most cancers analyzed. Mechanistically, ELFN1 expression was associated with DNA methylation, DNA repair, genomic instability, and tumor microenvironment (TME) scores in multiple cancer types. Furthermore, Drug sensitivity profiling linked ELFN1 to ABT-737 susceptibility and benzaldehyde resistance through molecular docking. In CRC cells, ELFN1 knockdown significantly inhibited tumor proliferation, migration, and motility.ConclusionThe expression level of ELFN1 may provide insights into tumor development and progression in multiple cancers, including CRC, highlighting its potential utility as an effective prognostic biomarkers and immunotherapeutic targets.

Acute liver injury (ALI) serves as a critical precursor and major etiological factor in the progression and ultimate manifestation of various hepatic disorders. The prevention and treatment of ALI is still a serious global challenge. Given the limited therapeutic options for ALI, exploring novel targeted therapeutic agents becomes imperative. The potential therapeutic efficacy of inhibiting RIPK2 is highlighted, as it may provide significant benefits by attenuating the MAPK pathway and NF-κB signaling. Herein, we propose a CMD-OPT model, a two-stage molecular optimization tool for the rapid discovery of RIPK2 inhibitors with optimal properties. Compound RP20, which targets the ATP binding site, demonstrated excellent kinase specificity, ideal oral pharmacokinetics, and superior therapeutic effects in a model of APAP-induced ALI, positioning RP20 as a promising preclinical candidate. This marks the first application of RIPK2 inhibitors in ALI treatment, opening a novel therapeutic pathway for clinical applications. These results highlight the efficacy of the CMD-OPT model in producing lead compounds from known active molecules, showcasing its significant potential in drug discovery.

Heart failure with preserved ejection fraction (HFpEF) represents a significant global health burden, yet effective pharmacotherapies remain elusive. The angiotensin-like 1 receptor, also known as the apelin receptor (APLNR), is a promising target for treating HFpEF due to its role in modulating cardiovascular function. Despite the cardioprotective effects of endogenous ligand, apelin, achieving G-protein-biased agonism for therapeutic benefit poses a significant challenge. In this study, we unravel the biased signal transduction pathway mediated by a reported partial Gi-protein-biased APLNR agonist CMF-019 and developed a biased chemical space remodeling approach to identify exclusive G-protein-biased agonists targeting APLNR. These agonists exhibited enhanced Gi-protein-biased function and protective effects in both in vitro and in vivo. Our findings not only enhance comprehension of APLNR-biased agonism but also establish drug design strategies for modifying and reshaping biased chemical landscapes in other G-protein-coupled receptors (GPCRs).

G protein-coupled receptors (GPCRs) are highly dynamic membrane receptors with numerous subtypes and complex signal transduction pathways. Precise regulation of GPCR signaling is closely related to disease treatment but presents significant challenges with classical orthosteric ligands. Allosteric modulators, a class of emerging drug candidates, can selectively bind to the allosteric sites located outside the conserved orthosteric pocket. In particular, biased allosteric modulators (BAMs) can stabilize specific conformations of GPCRs to harness signal transduction with high selectivity and specificity, offering a novel approach to modulate GPCR pharmacology and develop safer therapeutic agents. In recent years, significant progress has been made in the study of GPCR allosteric modulation due to advancements in structural biology. However, knowledge about GPCR-biased allostery is still in its infancy. In this chapter, we present the most recent breakthroughs in the discovery of BAM binding site in GPCRs and provide structural insights into biased allostery of GPCR signaling.

The increase in the number of macrolide-resistant Mycoplasma pneumoniae (MP) poses a threat to human health worldwide. The present research investigates the role of the NLRP3 inflammasome in mycoplasma pneumoniae pneumonia (MPP). MPP patient (n = 40) and healthy control (n = 20)-derived serum samples were collected. MP (strain ATCC15531)-infected C57BL/6 J or Nlrp3-/- mice with or without MCC950 treatment were used to explore the role of the NLRP3 inflammasome. The concentrations of inflammatory cytokines were determined by enzyme-linked immunosorbent assay. Histomorphological changes were determined by hematoxylin-eosin staining. The transcriptional and translational levels of NLRP3 were detected with quantitative PCR and western blot. Serum interleukin (IL)-1β and IL-18 levels were higher in MPP patients, along with elevated NLRP3 mRNA and protein levels. High NLRP3 expression was associated with fever duration, duration of admission, c-reactive protein and lactate dehydrogenase levels, and macrolide resistance. During the progression of MP infection, the NLRP3 inflammasome was progressively activated in mice, accompanied by increasing lung injury and inflammation. However, MP-infected Nlrp3-/- mice showed decreased lung injury and inflammation. Additionally, MCC950 weakened lung injury and inflammation in MPP mice, and the combination of azithromycin and MCC950 exerted a stronger effect than azithromycin or MCC950 alone. The NLRP3 inflammasome activation boosts lung injury, inflammation, and macrolide resistance in MPP, implying that interfering with the NLRP3 inflammasome may be a stratagem for MPP administration.

We report a novel FGFR2::TXLNB fusion in a child with hippocampal PLNTY. This fusion likely drives tumorigenesis via homodimerization and activation of MAPK/PI3K pathways, expanding PLNTY's molecular spectrum and suggesting a potential therapeutic target.

G protein-coupled sphingosine-1-phosphate receptor 1 (S1PR1), a drug target for inflammatory bowel disease (IBD), enables immune cells to egress from lymph nodes, but the treatment increases the risk of immunosuppression. The functional signaling pathway triggered by S1PR1 activation in endothelial cells and its therapeutic application remains unclear. Here, we showed that S1PR1 is highly expressed in endothelial cells of IBD patients and positively correlated with endothelial markers. Gi-biased agonist-SAR247799 activated S1PR1 and reversed pathology in male mouse and organoid IBD models by protecting the integrity of the endothelial barrier without affecting immune cell egress. Cryo-electron microscopy structure of S1PR1-Gi signaling complex bound to SAR247799 with a resolution of 3.47 Å revealed the recognition mode for the biased ligand. With the efficacy of SAR247799 in treating other endothelial dysfunction-associated inflammatory diseases, our study offers mechanistic insights into the Gi-biased S1PR1 agonist and represents a strategy for endothelial dysfunction-associated disease treatment.