Thiazolyl 4-carboxylate ketone as a new warhead for a highly potent SARS-CoV-2 main protease inhibitor

  • Eur J Med Chem. 2026 Feb 5:303:118436. doi: 10.1016/j.ejmech.2025.118436.
Maria A Theodoropoulou  1 ,  Haifa El Kilani  2 ,  Christiana Mantzourani  3 ,  Dirk Jochmans  4 ,  Johan Neyts  5 ,  Kaixuan Zhang  6 ,  Judith Röske  7 ,  Maroula G Kokotou  8 ,  Rolf Hilgenfeld  9 ,  George Kokotos  10
Affiliations
  • 1. Department of Chemistry, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece; Center of Excellence for Drug Design and Discovery, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece. Electronic address: [email protected].
  • 2. Institute of Molecular Medicine, University of Lübeck, Ratzeburger Allee 160, 23562, Lübeck, Germany. Electronic address: [email protected].
  • 3. Department of Chemistry, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece; Center of Excellence for Drug Design and Discovery, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece. Electronic address: [email protected].
  • 4. Virology, Antiviral Drug & Vaccine Research Group, Department of Microbiology, Immunology and Transplantation, Rega Institute, KU Leuven, Leuven, Belgium. Electronic address: [email protected].
  • 5. Virology, Antiviral Drug & Vaccine Research Group, Department of Microbiology, Immunology and Transplantation, Rega Institute, KU Leuven, Leuven, Belgium. Electronic address: [email protected].
  • 6. Institute of Molecular Medicine, University of Lübeck, Ratzeburger Allee 160, 23562, Lübeck, Germany. Electronic address: [email protected].
  • 7. Institute of Molecular Medicine, University of Lübeck, Ratzeburger Allee 160, 23562, Lübeck, Germany. Electronic address: [email protected].
  • 8. Laboratory of Chemistry, Department of Food Science and Human Nutrition, Agricultural University of Athens, Iera Odos 75, 11855, Athens, Greece. Electronic address: [email protected].
  • 9. Institute of Molecular Medicine, University of Lübeck, Ratzeburger Allee 160, 23562, Lübeck, Germany. Electronic address: [email protected].
  • 10. Department of Chemistry, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece; Center of Excellence for Drug Design and Discovery, National and Kapodistrian University of Athens, Panepistimiopolis, 15771, Athens, Greece. Electronic address: [email protected].
PMID: 41344111 DOI: 10.1016/j.ejmech.2025.118436
Abstract

The SARS-CoV-2 main protease (Mpro), an enzyme essential for viral replication and lacking a human homologue, has emerged as a highly attractive target for the development of novel Antiviral agents. Although several Mpro inhibitors have been developed - some receiving regulatory approval - their use is sometimes limited by drug-drug interactions. In this study, we designed and synthesized peptidomimetic SARS-CoV-2 Mpro inhibitors incorporating a novel thiazolyl 4-carboxylate ketone warhead, previously employed by our group in the development of cytosolic Phospholipase A2 inhibitors. The synthesized compounds were evaluated for their in vitro inhibitory potency against SARS-CoV-2 Mpro, and a highly potent Mpro inhibitor (GK730) was identified (IC50 5.75 nM). The melting temperature of the Mpro-GK730 complex revealed high stability, consistent with the high inhibitory potency. The X-ray crystal structures of inhibitors GK729 and GK730 bound to Mpro were determined, providing insights into the binding interactions and mechanism of action. Studies on the host cell proteases Cathepsin B and L showed that GK730 did not inhibit Cathepsin B, while exhibited weak inhibition of Cathepsin L. Furthermore, GK730 demonstrated an EC50 value of 5.70 μM against a wild-type SARS-CoV-2 strain in Vero E6 cells and minimal cytotoxicity (CC50 value greater than 100 μM).

Keywords
Inhibitors; Main protease; SARS-CoV-2; Thiazolyl 4-carboxylate ketone warhead; X-ray crystal structure.
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