Enzymatic, structural, and biophysical characterization of a single-domain antibody (VHH) selectively and tightly inhibiting neutrophil elastase and exhibiting favorable developability properties.

Autor: Redeghieri P; Nano-Antibodies to Explore Protein Structure and Functions (NEPTUNS), Centre for Protein Engineering, InBios, Department of Life Sciences, University of Liège, Liège, Belgium., Moray J; Nano-Antibodies to Explore Protein Structure and Functions (NEPTUNS), Centre for Protein Engineering, InBios, Department of Life Sciences, University of Liège, Liège, Belgium., Kerff F; Biological Macromolecule Crystallography, Centre for Protein Engineering, InBios, Department of Life Sciences, University of Liège, Liège, Belgium., Gohy S; Department of Pneumology, ENT and Dermatology, Institute of Experimental and Clinical Research (IREC), Université Catholique de Louvain, Brussels, Belgium.; Cystic Fibrosis Reference Centre, Cliniques Universitaires Saint-Luc, Brussels, Belgium., Leal T; Louvain Center for Toxicology and Applied Pharmacology (LTAP), Institute of Experimental and Clinical Research (IREC), Université Catholique de Louvain, Brussels, Belgium., Muyldermans S; Laboratory of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium., Vanbever R; Louvain Drug Research Institute (LDRI), Université Catholique de Louvain, Brussels, Belgium., Morales-Yánez FJ; Nano-Antibodies to Explore Protein Structure and Functions (NEPTUNS), Centre for Protein Engineering, InBios, Department of Life Sciences, University of Liège, Liège, Belgium., Dumoulin M; Nano-Antibodies to Explore Protein Structure and Functions (NEPTUNS), Centre for Protein Engineering, InBios, Department of Life Sciences, University of Liège, Liège, Belgium.
Jazyk: angličtina
Zdroj: Protein science : a publication of the Protein Society [Protein Sci] 2024 Dec; Vol. 33 (12), pp. e5227.
DOI: 10.1002/pro.5227
Abstrakt: Human neutrophil elastase (hNE), a serine protease released by neutrophils during inflammation, plays a major role in the pathophysiology of several conditions especially in inflammatory lung diseases. Its inhibition constitutes, therefore, a promising therapeutic strategy to combat these diseases. In this work, we characterized the in vitro properties of a VHH (i.e., the antigen binding domain of camelid heavy chain-only antibodies), referred to as NbE201. This VHH is able to inhibit tightly, selectively and competitively both human and murine elastases with the inhibition constants (K i ) of 4.1 ± 0.9 nM and 36.8 ± 3.9 nM, respectively. The IC 50 for the inhibition of the hydrolysis of elastin is in the same range to that of alpha-1 antitrypsin (i.e., the main endogenous inhibitor of hNE also used in the clinic) and 14 times better than that of Sivelestat (i.e., the 2nd clinically approved hNE inhibitor). The X-ray crystal structure of the NbE201-hNE complex reveals that the Complementarity Determining Regions CDR1 and CDR3 of the VHH bind into the substrate binding pocket of hNE and prevent the access to small or macromolecular substrates. They do not, however, bind deep enough into the pocket to be hydrolyzed. NbE201 is highly stable towards oxidation, deamidation, and chemical or thermal denaturation. NbE201 is therefore likely to tolerate manufacturing processes during drug development. These results highlight the high potential of NbE201 as a (pre)clinical tool to diagnose and treat diseases associated with excessive hNE activity, and for fundamental research to better understand the role of hNE in these conditions.
(© 2024 The Protein Society.)
Databáze: MEDLINE
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