COUMARIN DERIVATIVE CL167 AS AN IRREVERSIBLE INHIBITOR OF TMPRSS2: BIOCHEMICAL CHARACTERIZATION AND ANTIVIRAL PERSPECTIVES

In the context of the continuous emergence of respiratory virus variants, targeting host proteases involved in viral entry represents a promising strategy for the development of broad-spectrum antivirals. The transmembrane serine protease 2 (TMPRSS2) is a key host factor for cellular entry of several respiratory viruses, including coronaviruses, and thus represents an attractive therapeutic target.1, 2
Here we report the identification of CL167, selected from a series of coumarin derivatives based on phenyl coumarin-3-carboxylate esters substituted with basic functions attached to the phenyl ring. CL167 exhibits submicromolar inhibitory activity against TMPRSS2 (IC₅₀ = 0.31 µM ± 0.02 µM) and displays an improved selectivity profile over thrombin. Its kinetic profile is consistent with irreversible inhibition of TMPRSS2 (k_inact/K_I = 592.4 M⁻¹·s⁻¹), as supported by time-dependent inhibition analysis. In addition, its stability in solution was evaluated by mass spectrometry to monitor compound integrity over time. CL167 was prioritized as a chemical tool due to its irreversible mode of action and selective
inhibition of TMPRSS2. Owing to their distinct mechanism, covalent inhibitors present several advantages over classical non-covalent inhibitors and are increasingly considered as
promising candidates for therapeutic development, particularly in the context of infectious
diseases.
These properties motivated further in-depth mechanistic investigations of its interaction with TMPRSS2, combining mass spectrometry and molecular modeling to complement
experimental enzymatic data with detailed structural insights. Moreover, the antiviral activity
of CL167 is currently being evaluated to assess its therapeutic potential as an inhibitor of viral entry.
Taken together, these findings position CL167 as a relevant chemical tool for studying
TMPRSS2 and highlight the potential of coumarin derivatives in the development of
inhibitors targeting host proteases involved in respiratory viral infections.
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https://doi.org/10.1038/s41392-022-01084-x.
(2) Koyou, H. L.; Salleh, M. N.; Jelemie, C. S.; Badrin, M. J. Q.; Prastiyanto, M. E.;
Ramachandran, V. TMPRSS2: A Key Host Factor in SARS-CoV-2 Infection and Potential
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https://doi.org/10.4274/MMJ.galenos.2025.40460.