Selective Neutral pH Inhibitor of Cathepsin B Designed Based on Cleavage Preferences at Cytosolic and Lysosomal pH Conditions.

Autor:	Yoon MC; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States.; Biomedical Sciences Graduate Program, University of California, San Diego, La Jolla, California 92093, United States., Solania A; Departments of Molecular Medicine and Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, California 92037, United States., Jiang Z; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Christy MP; Scripps Institution of Oceanography, University of California, San Diego, La Jolla, California 92093, United States., Podvin S; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Mosier C; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Lietz CB; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Ito G; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Gerwick WH; Scripps Institution of Oceanography, University of California, San Diego, La Jolla, California 92093, United States., Wolan DW; Departments of Molecular Medicine and Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, California 92037, United States., Hook G; American Life Sciences Pharmaceuticals, Inc., La Jolla, California 92037, United States., O'Donoghue AJ; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States., Hook V; Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, California 92093, United States.; Department of Neurosciences, School of Medicine, University of California, San Diego, La Jolla, California 92037, United States.
Jazyk:	angličtina
Zdroj:	ACS chemical biology [ACS Chem Biol] 2021 Sep 17; Vol. 16 (9), pp. 1628-1643. Date of Electronic Publication: 2021 Aug 20.
DOI:	10.1021/acschembio.1c00138
Abstrakt:	Cathepsin B is a cysteine protease that normally functions within acidic lysosomes for protein degradation, but in numerous human diseases, cathepsin B translocates to the cytosol having neutral pH where the enzyme activates inflammation and cell death. Cathepsin B is active at both the neutral pH 7.2 of the cytosol and the acidic pH 4.6 within lysosomes. We evaluated the hypothesis that cathepsin B may possess pH-dependent cleavage preferences that can be utilized for design of a selective neutral pH inhibitor by (1) analysis of differential cathepsin B cleavage profiles at neutral pH compared to acidic pH using multiplex substrate profiling by mass spectrometry (MSP-MS), (2) design of pH-selective peptide-7-amino-4-methylcoumarin (AMC) substrates, and (3) design and validation of Z-Arg-Lys-acyloxymethyl ketone (AOMK) as a selective neutral pH inhibitor. Cathepsin B displayed preferences for cleaving peptides with Arg in the P2 position at pH 7.2 and Glu in the P2 position at pH 4.6, represented by its primary dipeptidyl carboxypeptidase and modest endopeptidase activity. These properties led to design of the substrate Z-Arg-Lys-AMC having neutral pH selectivity, and its modification with the AOMK warhead to result in the inhibitor Z-Arg-Lys-AOMK. This irreversible inhibitor displays nanomolar potency with 100-fold selectivity for inhibition of cathepsin B at pH 7.2 compared to pH 4.6, shows specificity for cathepsin B over other cysteine cathepsins, and is cell permeable and inhibits intracellular cathepsin B. These findings demonstrate that cathepsin B possesses pH-dependent cleavage properties that can lead to development of a potent, neutral pH inhibitor of this enzyme.
Databáze:	MEDLINE
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