Preclinical characterization of EGT710, an oral non-peptidomimetic reversible covalent SARS-CoV-2 main protease inhibitor

Stephanie A. Moquin, Suresh B. Lakshminarayana, Kamal Kumar Balavenkatraman, Hilmar Schiller, Allison Claas, Barun Bhhatarai, Ioannis Loisios-Konstantinidis, Katarina Vulic, Chaitanya Kurhade, Birte K. Kalveram, John Yun-Chung Chen, Jing Zou, Xuping Xie, Laura Tandeske, Dustin Dovala, Elizabeth Ornelas, Mark S. Knapp, Daniel Fuller, Zachary Nguyen, David T. Barkan, Lidiya Bebrevska, S. Kirk Wright, Scott A. Busby, Johanne Blais, Pei-Yong Shi, Suzanne Gaudet, Renee Bergeron, Hannah Yu, Julia Zack, Christopher Sarko, Feng Gu, James E. Bradner, John A. Tallarico, Thierry T. Diagana & Julien P. N. Papillon 

npj Drug Discov. 2, 28 (2025). 

https://doi.org/10.1038/s44386-025-00030-5

EGT710 is an orally bioavailable non-peptidomimetic reversible covalent coronavirus main protease (Mpro) inhibitor with low nM cellular activity against SARS-CoV-2. Twice daily dosing of 10 mg/kg of EGT710 decreased lung viral load in a mouse model of SARS-CoV-2 infection to below the limit of detection. Resistance selection resulted in the emergence of several Mpro mutations, with recombinant viruses containing L50F + E166A substitutions showing the largest shift in potency. Development of a viral kinetics model using viremia data from clinical trials, along with a human physiologically based pharmacokinetic model, predicted efficacy in humans with once daily oral doses of >360 mg. EGT710 displays favorable pharmacokinetic properties and an acceptable in vitro and in vivo safety profile, with human exposures at the recommended clinical dose of 600 mg predicted to be below the no adverse effect level in preclinical toxicology studies. Together, EGT710 has a promising preclinical profile and has completed a Phase I study.

Fast and Site-Specific Covalent Targeting of Proteins by Arylfluorosulfate-Modified Aptamers

 Kaining Zhang, Juan Li, Yang Shi, Sining Hou, Zichen Qin, Wenhao Shi, Yiying Zhu, Jingjing Zhang, Aijun Tong, and Yu Xiang

Journal of the American Chemical Society 2025

DOI: 10.1021/jacs.5c14374

Sulfur fluoride exchange (SuFEx) is a versatile click chemistry platform with emerging applications in covalent inhibitor development, among which arylfluorosulfate (AFS) derivatives have shown great promise because of their exclusive selectivity to target proteins. Nevertheless, due to the inherently mild electrophilicity of AFS, the high target specificity often comes at the expense of target reactivity. Here, we challenge this conventional view and show that in vitro selected AFS-modified covalent aptamers (AFS-Aps) can be both highly target selective and reactive. One such AFS-Ap is able to site-specifically modify the extracellular domain (ECD) of human epidermal growth factor receptor 3 (HER3) with a “click” reaction half-life (t1/2) as short as 1 min and covalently inhibit the protein–protein interaction (PPI) of HER3/EGFRs. Another AFS-Ap targeting human vascular endothelial growth factor (VEGF) can also rapidly cross-link VEGF165 and block the PPI of VEGF/VEGFR. The inactivation rate constants (kinact) of these AFS-Aps are one to two orders of magnitude higher than many reported AFS-based covalent molecules. This unexpectedly fast and site-specific covalent targeting requires a precisely organized aptamer–protein binding interface, where the AFS structure, rather than electrophilicity, is the decisive factor. We believe the target reactivity potential of AFS derivatives may have long been underestimated in covalent inhibitor development. AFS-adapted combinatorial techniques are therefore invaluable for discovering covalent inhibitors with both high target reactivity and selectivity.

A multicenter, open-label, first-in-human study of TYRA-200 in advanced intrahepatic cholangiocarcinoma and other solid tumors with activating FGFR2 gene alterations (SURF201).

Jordi Rodon Ahnert, Sameek Roychowdhury, Haley Ellis, Fernando F. Blanco, Timothy Burn, Jennifer Michelle Davis, Alex Balcer, Alena Zalutskaya

Journal of Clinical Oncology 2025

Volume 43, Number 4_suppl

https://doi.org/10.1200/JCO.2025.43.4_suppl.TPS646

Background: Approved Fibroblast Growth Factor Receptor (FGFR) inhibitors have demonstrated clinical benefit in locally advanced or metastatic cholangiocarcinoma harboring oncogenic FGFR2 fusions or other rearrangements. However, the emergence of acquired resistance mutations limits the clinical activity and duration of responses to currently available inhibitors. TYRA-200 is an orally bioavailable FGFR1/2/3 inhibitor designed to specifically address these clinically observed acquired resistance alterations in FGFR2. Methods: SURF201 is a single arm, multicenter, open-label, first-in-human, dose escalation and expansion study designed to investigate TYRA-200 in patients (pts) with advanced intrahepatic cholangiocarcinoma and other solid tumors with primary activating FGFR2 gene alterations and on-target acquired known FGFR2 resistance mutations. The study is being conducted in two parts. Part A dose escalation uses an i3+3 design and is evaluating the safety, tolerability, and the pharmacokinetic profile of TYRA-200. Part B dose expansion will further characterize the safety profile and evaluate the preliminary anti-tumor activity of TYRA-200 by RECIST v1.1 in pts with unresectable locally advanced/metastatic intrahepatic cholangiocarcinoma who have previously received an FGFR inhibitor(s) and have developed acquired FGFR2 kinase domain resistance mutations. The study is currently planned for approximately four centers in the US and is actively enrolling (NCT06160752). Clinical trial information: NCT06160752.

Discovery of Fulzerasib (GFH925) for the Treatment of KRAS G12C-Mutated Solid Tumors

Tao Jiang, Chonglan Lin, Siyuan Le, Leitao Zhang, Tao Liang, Lijian Cai, Xiaoling Lan, Mei Ge, Zhubo Liu, Wan He, Ling Peng, Yanhui Zhao, Jinmin Ren, Feng Yan, Qiang Lu, Jiong Lan, and Fusheng Zhou

J. Med. Chem. 2025, 68, 15, 15386–15402

DOI: 10.1021/acs.jmedchem.4c03183

RAS mutations are the most prevalent genetic alterations in human tumors, accounting for 30% of all cases. Among these mutations, KRAS G12C emerged as the first druggable target through covalent attachment, which locks the protein in its inactive state. Employing a structure-based drug design strategy, we identified fulzerasib (GFH925), which features a novel lactam-based tetracyclic naphthyridinone scaffold. This molecule demonstrates high in vitro potency and selectivity, favorable pharmacokinetic profiles across species, and significant in vivo antitumor efficacy in various cancer-related xenograft models, including intracranial tumors. Fulzerasib has recently received accelerated approval in China for adult NSCLC patients with the KRAS G12C mutation after prior systemic therapy.

Discovery of an Internal Alkyne Warhead Scaffold for Irreversible hTG2 Inhibition

Lavleen K. Mader, Namita Maunick, Jessica E. Borean, Jeffrey W. Keillor

RSC Med. Chem., 2025

https://doi.org/10.1039/D5MD00777A

Human tissue transglutaminase (hTG2) is a multifunctional enzyme with both protein cross-linking and G-protein activity. Dysregulation of these functions has been implicated in diseases such as celiac disease and cancer, prompting the development of hTG2 inhibitors, many of which act covalently via a pendant electrophilic warhead. Most small molecule hTG2 inhibitors to date feature terminal, sterically minimal warheads, based on the assumption that bulkier electrophiles impair binding and reactivity. Here, we report structure–activity relationships (SAR) of a novel internal alkynyl warhead scaffold for irreversible inhibition of hTG2. This series includes one of the most potent non-peptidic hTG2 inhibitors reported to date. We demonstrate that this scaffold not only inhibits transamidase activity but also abolishes GTP binding, while exhibiting excellent isozyme selectivity. In addition, we investigate the tunability and stability of this warhead, providing insights into its broader applicability. Through detailed kinetic analysis, this study establishes a new scaffold for irreversible hTG2 inhibition and expands the design principles for covalent warheads beyond traditional terminal systems.

Covalent Probes Reveal Small-Molecule Binding Pockets in Structured RNA and Enable Bioactive Compound Design

Sandra Kovachka, Jielei Wang, Amirhossein Taghavi, Yilin Jia, Taro Asaba, Karen C. Wolff, Mason Martin, Xueyi Yang, Samantha M. Meyer, Sabine Ottilie, Mina Heacock, Zhong Cheng, Case W. McNamara, Gurudutt Dubey, Arnab K. Chatterjee, Sumit Chanda, José Gallego, Jessica L. Childs-Disney, and Matthew D. Disney

Journal of the American Chemical Society 2025

DOI: 10.1021/jacs.5c11898

The SARS-CoV-2 frameshift stimulation element (FSE) is a critical RNA structure that is essential for viral replication and represents a promising target for antiviral intervention. Here, Chemical Cross-Linking and Isolation by Pull-down (Chem-CLIP) covalent target validation and binding site mapping was applied, to identify small-molecule binding pockets within the FSE and ultimately develop a ligandability map. These studies employed ∼ 190 Chem-CLIP fragments, including the fluoroquinolone merafloxacin, previously shown to interact with this element. Covalent mapping defined merafloxacin’s binding pocket at a nucleotide-level resolution and revealed interactions that, along with structure-based design, efficient one-pot on-plate synthesis and competitive displacement assays, enabled the development of bioactive compounds with antiviral activity. Complementary chemical probing with dimethyl sulfate (DMS) in the presence of a bioactive ligand, coupled to Deconvolution of RNA Alternative Conformations (DRACO), revealed that compound binding increased the reactivity of specific nucleotides with DMS, indicative of changes in local RNA folding. These results highlight the importance of combining Chem-CLIP and DMS profiling to differentiate direct ligand binding from ligand-induced changes in RNA structure. In addition, in silico pocket analysis of FSE structures derived from cryogenic-electron microscopy (cryo-EM) studies identified four recurring cavities, including the experimentally determined merafloxacin and Chem-CLIP fragments binding pockets. Altogether, the findings advance our understanding of RNA–ligand interactions and support a strategy to design and discover small molecules that bind RNA structures.

AI-assisted delivery of novel covalent WRN inhibitors from a non-covalent fragment screen

Geoffrey M.T. Smith, Laksh Aithani, Charlotte E. Barrett, Alwin O. Bucher, Christopher D.O. Cooper, Sébastien L. Degorce, Andrew S. Doré, Catherine T. Fletcher, Sophie Huber, Rosemary Huckvale, Amanda J. Kennedy,  Abigail A. Mornement, Mark Pickworth, Prakash Rucktooa, Conor C.G. Scully,  Sarah E. Skerratt

Bioorganic & Medicinal Chemistry Letters, 2025

https://doi.org/10.1016/j.bmcl.2025.130421

Werner (WRN) helicase, has emerged as a promising therapeutic target for cancers associated with microsatellite instability (MSI). This letter describes the discovery of small molecule inhibitors from a fragment screen that occupy a cryptic, allosteric site of WRN helicase. Key findings include the identification of benzimidazole and amino-indazole scaffolds, exploiting their proximity to Cys727 via covalent modification. The use of our proprietary co-folding model DragonFold assisted the identification of novel WRN helicase inhibitors. These, together with near-neighbor profiling, offer tools for furthering the understanding of WRN and BLM helicase function, and potential therapeutic avenues for MSI-associated cancers.