Covalent Inhibitors of Monoacylglycerol Lipase Induce Conformational Changes and Proteasomal Degradation

Jordan A. Pham, Thanawat Thaingtamtanha, William McLeish, David Lefebvre, Spencer M. Uguccioni, Roxana Filip, Francesco Gentile, and John Paul Pezacki

J. Am. Chem. Soc. 2026

https://doi.org/10.1021/jacs.5c22859

Monoacylglycerol lipase (MGLL) is a key serine hydrolase that regulates 2-arachidonoylglycerol (2-AG) and eicosanoid signaling. Inhibition of MGLL blocks the conversion of 2-AG into arachidonic acid (AA) with broad therapeutic implications in inflammation, cancer, and viral infection. Carbamate/urea inhibitors such as MJN110, JZL184, and SAR629 are widely used to irreversibly inhibit MGLL through covalent modification of the catalytic serine residue. Here, we demonstrate that this inhibitor class also induces proteasome-dependent degradation of MGLL, functioning as monovalent degraders. We show that loss of MGLL following covalent inhibitor treatment is dependent on the 26S proteasome, while detailed simulations of MGLL dynamics following inhibitor binding suggest that these inhibitors do not destabilize their protein target but instead induce conformational changes that likely facilitate polyubiquitination by exposing two lysine residues. Taken together, these findings establish carbamate/urea inhibitors as dual functional molecules with the propensity to both covalently inhibit their target and act as structural degraders, potentially functioning as monovalent molecular glues, highlighting the need to evaluate the degradation potential alongside inhibitory potency in future screening and drug discovery.

Development and Structural Characterization of UTE-156, a Covalent Inhibitor of the VCP/p97 AAA+ ATPase

Tamayo-Jaramillo D, Hegde S, Jia X, Coffman K, Vankayalapati H, Bearss D, Jones KB, Stark AW, Shen PS. 

Adv Sci. 2026; 13(25):e20545. 

https://doi.org/10.1002/advs.202520545

The AAA+ ATPase valosin-containing protein (VCP/p97) is a central regulator of protein homeostasis that is well characterized for its role in extracting and remodeling ubiquitinated substrates. Dysregulation of VCP activity contributes to the pathogenesis of neurodegenerative diseases and cancer, making it an important therapeutic target. Here, we report the development and characterization of UTE-156, a novel covalent small-molecule inhibitor that modifies Cys522 within the D2 ATPase domain of VCP. UTE-156 potently inhibits VCP ATPase activity, while losing activity against a C522A mutant, supporting a covalent mechanism of action. High-resolution cryo-electron microscopy (cryo-EM) structures reveal that UTE-156 occupies the D2 nucleotide-binding site, sterically blocking ATP binding and inducing conformational remodeling of the pocket. Biochemical and cell-based assays demonstrate strong inhibitory potency but limited solubility and rapid metabolic turnover. These pharmacochemical limitations preclude immediate therapeutic use but underscore its value as a chemical probe. Together, these findings establish UTE-156 as a powerful tool for dissecting VCP function and provide a framework for future optimization of covalent modulators of protein homeostasis.

Nitro-Diphenyl Ethers as Emerging Cysteine-Targeting Covalent Warheads Enable Identification of Novel Target LDLRAP1 for Anticoronaviral Activity

Zeyue Huang, Xiuqi Hu, Zheng Liu, Hongxuan Cao, Yunjie Xiang, Jian Wan, Ivailo Slavchev, Li Rao, Ivanka Nikolova, Petar Grozdanov, Nadya Nikolova, Georgi M. Dobrikov, and Yanliang Ren

Journal of Medicinal Chemistry 2026

DOI: 10.1021/acs.jmedchem.5c03394

Targeted covalent inhibitors (TCIs) are powerful tools in drug discovery, but the high intrinsic reactivity of conventional warheads often compromises selectivity and increases the off-target liability. Here, we reported nitrodiphenyl-ether compounds as a novel irreversible and released-type covalent warhead with exceptionally low reactivity that potently inhibits coronavirus HCoV-OC43 infection. To identify their molecular targets, we designed a panel of active and inactive alkyne-tagged probes and performed chemical proteomic profiling in human host cells. An integrated approach combining activity- and inactivity-based proteome profiling (AIBPP), competitive ABPP, LC–MS/MS, and fluorescence polarization (FP) assays identified low-density lipoprotein receptor adapter protein 1 (LDLRAP1) as the primary target, modified selectively at C119, thereby disrupting the LDLR–LDLRAP1 interaction. Inhibition of this interaction strongly correlated with antiviral efficacy, confirming LDLRAP1 as the functional target. Collectively, this study establishes LDLRAP1 as an unexploited host antiviral target and expands the repertoire of cysteine-targeted covalent warheads for host-directed therapy.

Shifting the PPARγ conformational ensemble toward a transcriptionally repressive state improves covalent inhibitor efficacy

Liudmyla Arifova, Brian S MacTavish, Zane Laughlin, Mithun Nag, Karadi Giridhar, Jinsai Shang, Min-Hsuan Li, Xiaoyu Yu, Di Zhu, Theodore M Kamenecka, Douglas J Kojetin,

eLife 2025 14:RP106697

https://doi.org/10.7554/eLife.106697.2

The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARγ) regulates transcription in response to ligand binding at an orthosteric pocket within the ligand-binding domain (LBD). We previously showed that two covalent ligands, T0070907 and GW9662—extensively used as PPARγ inhibitors to assess off-target activity—weaken but do not completely block ligand binding via an allosteric mechanism associated with pharmacological inverse agonism (Shang et al., 2024). These covalent inhibitors shift the LBD towards a repressive conformation, where the activation function-2 (AF-2) helix 12 occupies the orthosteric pocket, competing with orthosteric ligand binding. Here, we provide additional support for this allosteric mechanism using two covalent inverse agonists, SR33065 and SR36708, which better stabilize the repressive LBD conformation and are more effective inhibitors of—but also do not completely inhibit—ligand cobinding. Furthermore, we show that ligand cobinding can occur with a previously reported PPARγ dual-site covalent inhibitor, SR16832, which appears to weaken ligand binding through a direct mechanism independent of the allosteric mechanism. These findings underscore the complex nature of the PPARγ LBD conformational ensemble and highlight the need to develop alternative methods for designing more effective covalent inhibitors.

Don’t Lose Your (War)head: Structure–Activity Relationships of Covalent Warheads as Substrates for GST-Catalyzed Glutathione Conjugation

Lavleen K. Mader, Jessica E. Borean, and Jeffrey W. Keillor

Journal of Medicinal Chemistry 2026

DOI: https://doi.org/10.1021/acs.jmedchem.6c00826

The resurgence of targeted covalent inhibitors (TCIs) has expanded the diversity of electrophilic warheads used in drug discovery. TCIs must balance efficient target engagement with resistance to rapid metabolic clearance. In drug development campaigns, intrinsic reactivity toward glutathione (GSH) is commonly used to estimate metabolic liability; however, in vivo GSH conjugation is primarily catalyzed by glutathione S-transferases (GSTs), a phase II metabolic pathway that is not captured by intrinsic reactivity measurements. Here, we establish a quantitative assay to determine GST kcat and KM values across a panel of structurally diverse warheads. We show that their intrinsic reactivities correlate poorly with GST-catalyzed conjugation rates, which are instead governed by warhead- and scaffold-dependent enzyme–substrate interactions. In contrast, GST kcat/KM values correlate closely with compound half-lives in human liver cytosol. Together, these findings establish GST susceptibility as a structurally tunable determinant of metabolic GSH conjugation and provide new principles for the optimization of TCIs.

Efficacy and safety of branebrutinib (BMS-986195), an irreversible Bruton's tyrosine kinase inhibitor, for the treatment of rheumatoid arthritis: a phase 2a, randomised, double-blind, placebo-controlled study

 Østergaard M, Haavardsholm E, Nowak M et al.

The Lancet Rheumatology, 2026

https://doi.org/10.1016/S2665-9913(25)00374-1

1. Background

   Branebrutinib, an oral, highly selective, and irreversible Bruton's tyrosine kinase inhibitor, is a potential candidate for rheumatoid arthritis treatment as Bruton's tyrosine kinase has a role in B-cell activation, autoantibody production, and proinflammatory cytokine release, all of which are implicated in rheumatoid arthritis disease activity and progression. This study assessed the efficacy and safety of branebrutinib in patients with rheumatoid arthritis and an inadequate response to methotrexate.

   Methods

   This phase 2a, randomised, double-blind, placebo-controlled study was designed to assess the efficacy and safety of branebrutinib in patients with rheumatoid arthritis, systemic lupus erythematosus, or primary Sjögren's disease. Here, we report the results of the rheumatoid arthritis substudy, done in the USA, Poland, and Spain across 24 sites. The study included a 12-week double-blind treatment period followed by an additional 12-week open-label period with abatacept treatment. Only data for the double-blind treatment period are reported here. Eligible patients were aged 18–75 years, met the 2010 American College of Rheumatology (ACR)–European Alliance of Associations for Rheumatology criteria for rheumatoid arthritis, had disease duration less than 4 years, and had inadequate response to methotrexate. Patients were randomly assigned (3:1) to receive branebrutinib 9 mg once daily or placebo for 12 weeks. Randomisation was carried out centrally according to a computer-generated block randomisation scheme using interactive response technology. All parties were masked to treatment allocation. The primary endpoint was the proportion of patients who had 50% improvement in the ACR response criteria (ACR50) at week 12, assessed in all participants randomly assigned to treatment (full analysis set). Safety was assessed in patients who received at least one dose of branebrutinib or placebo. This trial was registered with ClinicalTrials.gov, NCT04186871. Patients with lived experience of rheumatoid arthritis were not involved in the study design.

   Findings

   Between Jan 7, 2020, to Dec 5, 2022, 85 patients were randomly assigned to receive branebrutinib (n=64) or placebo (n=21). 63 (74%) of 85 patients were female, 22 (26%) were male, 80 (94%) were White, and the mean age was 49·1 years (SD 12·0). The primary endpoint of ACR50 response at week 12 was not met; 12 (19%) of 64 patients had an ACR50 response in the branebrutinib group compared with seven (33%) of 21 patients in the placebo group (p=0·16). Adverse events were similar between the two groups (30 [47%] of 64 in the branebrutinib group and 8 [38%] of 21 in the placebo group), with no reported serious adverse events or deaths.

   Interpretation

   There was no significant difference between branebrutinib and placebo for any clinical efficacy measures. The 12-week safety profiles were similar between treatment groups, and branebrutinib was well tolerated with a favourable safety profile.

   Funding

   Bristol Myers Squibb.

A Novel Covalent Inhibitor Fragment for the SARS-CoV-2 Main Protease Identified by Target-Specific Deep Learning

Weijun Zhou, Angel D′Oliviera, Xuhang Dai, Jeffrey S. Mugridge, and Yingkai Zhang

ACS Chemical Biology 2026

DOI: 10.1021/acschembio.6c00120

The SARS-CoV-2 main protease (Mpro, also known as 3CLpro) is an attractive antiviral drug target due to its essential role in viral replication and absence of human homologues. Development of new coronavirus-specific Mpro inhibitors will be important as SARS-CoV-2 continues to evolve. Leveraging the rapidly expanding pool of diverse, experimental Mpro-inhibitor data, we developed a target-specific deep learning workflow to accelerate the discovery of new Mpro inhibitor compounds and fragment-like starting points. This workflow combined a fine-tuned inhibitor prediction model with solubility (logS) and lipophilicity (logP) models, molecular similarity analysis, and literature mining to prioritize novel, drug-like candidates. Applied to a purchasable library of over 500,000 compounds, the approach rapidly identified 24 candidates for experimental testing. Biochemical assays revealed a novel, small covalent inhibitor fragment (A02) with an apparent IC50 of 1.5 μM, prior to any synthetic optimization or derivatization. A 1.76 Å crystal structure of Mpro bound to A02 confirmed covalent modification of the catalytic Mpro cysteine (C145), unique engagement of the underutilized Mpro S3′ pocket, and the potential for derivatives of this scaffold to interact with additional Mpro pockets in future optimization efforts. Together, these results demonstrate the potential for target-specific deep learning approaches to guide the rapid screening and discovery of new inhibitor leads or drug scaffolds.