In Search of Selective Inhibitors of Cysteine Protease, Cathepsin K (original) (raw)
Related papers
Design of potent and selective human cathepsin K inhibitors that span the active site
Proceedings of the National Academy of Sciences, 1997
Potent and selective active-site-spanning inhibitors have been designed for cathepsin K, a cysteine protease unique to osteoclasts. They act by mechanisms that involve tight binding intermediates, potentially on a hydrolytic pathway. X-ray crystallographic, MS, NMR spectroscopic, and kinetic studies of the mechanisms of inhibition indicate that different intermediates or transition states are being represented that are dependent on the conditions of measurement and the specific groups flanking the carbonyl in the inhibitor. The species observed crystallographically are most consistent with tetrahedral intermediates that may be close approximations of those that occur during substrate hydrolysis. Initial kinetic studies suggest the possibility of irreversible and reversible active-site modification. Representative inhibitors have demonstrated antiresorptive activity both in vitro and in vivo and therefore are promising leads for therapeutic agents for the treatment of osteoporosis. Expansion of these inhibitor concepts can be envisioned for the many other cysteine proteases implicated for therapeutic intervention.
Potency and Selectivity of the Cathepsin L Propeptide as an Inhibitor of Cysteine Proteases †
Biochemistry, 1996
The cathepsin L propeptide (phcl-2) was expressed in Saccharomyces cereVisiae using a human procathepsin L/R-factor fusion construct containing a stop codon at position -1 (the C-terminal amino acid of the proregion). Since the yield after purification was very low, the cathepsin L propeptide was also obtained by an alternate procedure through controlled processing of an inactive mutant of procathepsin L (Cys25Ser/Thr110Ala) expressed in Pichia pastoris, by small amounts of cathepsin L. The peptide resulting from the cleavage of the proenzyme (phcl-1) was then purified by HPLC. The purified propeptides were characterized by N-terminal sequencing and mass spectrometry and correspond to incomplete forms of the proregion (87 and 81 aa for phcl-1 and phcl-2 respectively, compared to 96 aa for the complete cathepsin L propeptide). The two peptides were found to be potent and selective inhibitors of cathepsin L at pH 5.5, with K i values of 0.088 nM for phcl-1 and 0.66 nM for phcl-2. The K i for inhibition of cathepsin S was much higher (44.6 nM with phcl-1), and no inhibition of cathepsin B or papain could be detected at up to 1 µM of the propeptide. The inhibitory activity was also found to be strongly pHdependent. Two synthetic peptides of 75 and 44 aa corresponding to N-terminal truncated versions of the propeptide were also prepared by solid phase synthesis and displayed K i values of 11 nM and 2900 nM, respectively, against cathepsin L. The data obtained for the 4 propeptide derivatives of various lengths indicate that the first 20 residues in the N-terminal region of the propeptide are more important for inhibition than the C-terminal region which contributes little to the overall inhibitory activity. † NRCC Publication No. 39920. Supported by postdoctoral fellowships from the Conselho Nacional de Desenvolvimento Cientifico e Tecnologico, Brazil (E.C.), as well as NATO and the Ministère
Design and evaluation of inhibitors for dipeptidyl peptidase I (Cathepsin C)
Archives of Biochemistry and Biophysics, 2004
Dipeptidyl peptidase I (DPPI, cathepsin C) is a lysosomal cysteine protease that can activate zymogens of several different serine proteases by one step or sequential removal of dipeptides from the N-termini of the pro-protease protein substrates. To find DPPI inhibitors more suitable for cellular applications than diazomethyl ketones, we synthesized three types of inhibitors: dipeptide acyloxymethyl ketones, fluoromethyl ketones, and vinyl sulfones (VS). The acyloxymethyl ketones inhibited DPPI slowly and are moderate inhibitors of cellular DPPI. The fluoromethyl ketones were potent, but the inhibited DPPI regained activity quickly. The dipeptide vinyl sulfones were effective inhibitors for DPPI, but they also inhibited cathepsins B, H, and L weakly. The best inhibitor, Ala-Hph-VS-Ph, had a k 2 =K I of 2,000,000 M À1 s À1 . The vinyl sulfones also inhibited intracellular DPPI, and for this application the more stable inhibitors exhibit better potency. We conclude that vinyl sulfones are promising inhibitors to study the intracellular functions of DPPI.
Inhibition of a Cathepsin L-Like Cysteine Protease by a Chimeric Propeptide-Derived Inhibitor †
Biochemistry, 2005
Like other papain-related cathepsins, congopain from Trypanosoma congolense is synthesized as a zymogen. We have previously identified a proregion-derived peptide (Pcp27), acting as a weak and reversible inhibitor of congopain. Pcp27 contains a 5-mer YHNGA motif, which is essential for selectivity in the inhibition of its mature form []. In the work presented here, a homology model of procongopain was generated and subsequently used to model a chimeric 50mer peptide (called H3-Pcp27) corresponding to the covalent linkage of an unrelated peptide (H3 helix from Antennapedia) to Pcp27. Molecular simulations suggested that H3-Pcp27 (pI ) 9.99) maintains an N-terminal helical conformation, and establishes more complementary electrostatic interactions (E coul ) -25.77 kcal/mol) than 16N-Pcp27, the 34-mer Pcp27 sequence plus the 16 native residues upstream from the proregion (E coul ) 0.20 kcal/mol), with the acid catalytic domain (pI ) 5.2) of the mature enzyme. In silico results correlated with the significant improvement of congopain inhibition by H3-Pcp27 (K i ) 24 nM), compared to 16N-Pcp27 (K i ) 1 µM). In addition, virtual alanine scanning of H3 and 16N identified the residues contributing most to binding affinity. Both peptides did not inhibit human cathepsins B and L. In conclusion, these data support the notion that the positively charged H3 helix favors binding, without modifying the selectivity of Pcp27 for congopain.
ChemMedChem, 2006
Description of the enzyme and NMR assays 1 H-NMR spectra of the reaction of 2 with 4-methoxy thiophenol Ratios of reaction products of the reaction of cpd. 2 with 4-methoxy thiophenol Mass spectrum of the reaction of 2 with 4-methoxy thiophenol Detailed results of the computations References S2 Syntheses and analytical data of inhibitor 2 General information HR-ESI mass spectra were recorded on a FT-ICR mass spectrometer APEX II, Bruker. NMR spectra were recorded on an AVANCE 400 MHz spectrometer from Bruker Biospin GmbH, Germany (solvent: CDCl 3 , 1 H-NMR: 400.13 MHz; 13 C-NMR: 100.61 MHz). IR spectra were recorded on a PharmalyzIR FT-IR spectrometer from BioRad, USA. α values were detemined on a 241 polarimeter from PerkinElmer, USA. Column chromatography was performed with silica gel 60 from Merck (0.063-0.2 mm or 70-230 mesh). All solvents were purified and dried prior to use according to standard literature procedures. All reactions were performed in a N 2 atmosphere under strict exclusion of humidity. Cathepsin L (Paramecium tetraurelia) was purchased from Calbiochem. Cbz-Phe-Arg-AMC was purchased from Bachem. (2R,3R)-Diethyl 1-formyl aziridine-2,3-dicarboxylate (2) 72.0 mg (0.55 mmol) formyl pivaloyl anhydride 1 are added at room temperature to a solution of 94.0 mg (0.5 mmol) (2R,3R)-diethyl aziridine-2,3-dicarboxylate (1) 2 in 10 mL CH 2 Cl 2. The solution is stirred for 12 h under nitrogen atmosphere. Then, the solution is diluted with 40 mL CH 2 Cl 2 and extracted with sat. NaHCO 3 solution (2 x 25 mL), and water (1 x 25 mL). The organic layer is dried with Na 2 SO 4 and removed i. vac.. The crude product is purified by column chromatography on silica gel 60, cyclohexane/ethyl acetate 2:1 (R f = 0.28). Yield: 78 mg (72%), colourless crystals, mp 28-29° C; [a] D 25
Journal of Molecular Modeling, 2014
Several cellular disorders have been related to the overexpression of the cysteine protease cathepsin B (CatB), such as rheumatic arthritis, muscular dystrophy, osteoporosis, Alzheimer's disease, and tumor metastasis. Therefore, inhibiting CatB may be a way to control unregulated cellular functions and prevent tissue malformations. The inhibitory action of 1,2,4-thiadiazole (TDZ) derivatives has been associated in the literature with their ability to form disulfide bridges with the catalytic cysteine of CatB. In this work, we present molecular modeling and docking studies of a series of eight 1,2,4-thiadiazole compounds. Substitutions at two positions (3 and 5) on the 1,2,4-thiadiazole ring were analyzed, and the docking scores were correlated to experimental data. A correlation was found with the sequence of scores of four related compounds with different substituents at position 5. No correlation was observed for changes at position 3. In addition, quantum chemistry calculations were performed on smaller molecular models to study the mechanism of inhibition of TDZ at the active site of CatB. All possible protonation states of the ligand and the active site residues were assessed. The tautomeric form in which the proton is located on N2 was identified as the species that has the structural and energetic characteristics that would allow the ring opening of 1,2,4thiadiazole.
Substrate-derived triazolo- and azapeptides as inhibitors of cathepsins K and S
European journal of medicinal chemistry, 2018
Cathepsin (Cat) K is a critical bone-resorbing protease and is a relevant target for the treatment of osteoporosis and bone metastasis, while CatS is an attractive target for drugs in autoimmune diseases (e.g. rheumatoid arthritis), emphysema or neuropathic pain. Despite major achievements, current pharmacological inhibitors are still lacking in safety and may have damaging side effects. A promising strategy for developing safer reversible and competitive inhibitors as new lead compounds could be to insert non-cleavable bonds at the scissile P1-P1' position of selective substrates of CatS and CatK. Accordingly, we introduced a 1,4-disubstituted 1,2,3-triazole heterocycle that mimics most of the features of a trans-amide bond, or we incorporated a semicarbazide bond (azaGly residue) by replacing the α-carbon of the glycyl residue at P1 by a nitrogen atom. AzaGly-containing peptidomimetics inhibited powerfully their respective target proteases in the nM range, while triazolopeptid...