Abstract
A novel small molecule thiocarbazate (PubChem SID 26681509), a potent inhibitor of human cathepsin L (EC 3.4.22.15) with an IC50 of 56 nM, was developed after a 57,821-compound screen of the National Institutes of Health Molecular Libraries Small Molecule Repository. After a 4-h preincubation with cathepsin L, this compound became even more potent, demonstrating an IC50 of 1.0 nM. The thiocarbazate was determined to be a slow-binding and slowly reversible competitive inhibitor. Through a transient kinetic analysis for single-step reversibility, inhibition rate constants were kon = 24,000 M-1s-1 and koff = 2.2 × 10-5 s-1 (Ki = 0.89 nM). Molecular docking studies were undertaken using the experimentally derived X-ray crystal structure of papain/CLIK-148 (1cvz. pdb). These studies revealed critical hydrogen bonding patterns of the thiocarbazate with key active site residues in papain. The thiocarbazate displayed 7- to 151-fold greater selectivity toward cathepsin L than papain and cathepsins B, K, V, and S with no activity against cathepsin G. The inhibitor demonstrated a lack of toxicity in human aortic endothelial cells and zebrafish. In addition, the thiocarbazate inhibited in vitro propagation of malaria parasite Plasmodium falciparum with an IC50 of 15.4 μM and inhibited Leishmania major with an IC50 of 12.5 μM.
Footnotes
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Financial support for this work was provided by National Institutes of Health grants 5U54-HG003915-02 and 5U54-HG003915-03).
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ABBREVIATIONS: LC-MS, liquid chromatography-mass spectrometry; SID, substance identifier; SID 861540, 2-[[5-[(1S)-1-amino-2-(1H-indol-3-yl)ethyl]-1,3,4-oxadiazol-2-yl]sulfanyl]-N-(2-ethylphenyl)acetamide hydrochloride; SID 26681509, S-[2-[(2-ethylphenyl)amino]-2-oxoethyl] [2-[(2S)-3-(1H-indol-3-yl)-2-[(2-methylpropan-2-yl)oxycarbonylamino]propanoyl]hydrazinyl]methanethioate; AMC, 7-amido-4-methylcoumarin; DMSO, dimethyl sulfoxide; Z-, N-benzyloxycarbonyl-; E-64, 1-[l-N-(trans-epoxysuccinyl)leucyl]amino-4-guanidinobutane.
- Received February 25, 2008.
- Accepted April 10, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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