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Functionalized N-aryl azetidinones as novel mechanism-based inhibitors of neutrophil elastase

A functionalized N‐aryl azetidinone has been shown to inactive human leukocyte elastase (HLE) and porcine pancreatic elastase (PPE) by an enzyme‐mediated process. The inactivation is characterized by the following kinetic constants at pH 8.0 and 37°C: k max=0.035 s−1, K 1=1.2 × 10 −4 M for HLE, 0.08 s−1 and 2.7 × 10−4 M for PPE, respectively. Two parent molecules devoid of the latent leaving group failed to inactive HLE and PPE and behaved as substrates of these enzymes. A suicide mechanism involving the formation of an acyl‐enzyme and the simultaneous unmasking of a latent quinonimmonium methide ion which irreversibly reacts with an active site nucleophile. Moreover, the inhibitor is still effective at inhibiting elastase preabsorbed onto elastin.

Inhibition of human leukocyte elastase by functionalized N-aryl azetidin-2-ones: substituent effects at C-3 and benzylic positions

Summary A series of functionalized N -aryl azetidin-2-ones with a latent alkylating group was prepared by a flexible four-step synthesis. They met criteria expected for a suicide-type inactivation of human leukocyte elastase (HLE) and porcine pancreatic elastase (PPE), with no inactivation of trypsin- and chymotrypsin-like proteases. The inhibition potency was dependent on the halogen substituents at C-3 (F, F; Cl, Cl; Br, Br) and the nature and the position relative to nitrogen of the latent benzylic leaving group (F, Cl, Br). Better inactivations of HLE compared with PPE were observed with azetidinones gem -disubstituted by Cl and Br rather than by F. Their protio analogs, which are devoid of the latent quinoniminium methide electrophile, behave as simple substrates of elastases.

Synthesis of N-glyoxylyl peptides and their in vitro evaluation as HIV-1 protease inhibitors

A series of novel synthetic peptides containing an N-terminal glyoxylyl function (CHOCO-) have been tested as inhibitors of HIV-1 protease. The N-glyoxylyl peptide CHOCO-Pro-Ile-Val-NH2, which fulfills the specificity requirements of the MA/CA protease cleavage site together with the criteria of transition state analogue of the catalyzed reaction, was found to be a moderate competitive inhibitor although favorable interactions were visualized between its hydrated form and the catalytic aspartates using molecular modeling. Increasing the length of the peptide sequence led to compounds acting only as substrates.

Inhibition of human leukocyte elastase by functionalized N-aryl-3,3-dihalogenoazetidin-2-ones. Stereospecific synthesis and chiral recognition of dissymmetrically C3-substituted β-lactams

Abstract (3R)- and (3S)-N-(2-chloromethylphenyl)-3-bromo-3-fluoroazetidin-2-ones 2 were synthesized via the separation of diastereoisomeric phenylglycinol derivatives of the starting 2,3-dibromo-2-fluoropropanoic acid. Acidic hydrolysis of the hydroxyamides led to the chiral trihalogenopropanoic acids. Then, an expeditious four step synthesis provided the (3S)- and (3R)-azetidinones 2, both of which behaved as strictly irreversible inhibitors of HLE. The configuration of the bromofluorocarbon was shown to have a significant effect on the partition ratio: kcat/kinact=4.6 and 34.3 for (3S)- and 3R)-2, respectively.

ACYLOXYBENZYL HALIDES, INHIBITORS OF ELASTASES

3-Benzyl-6-chloromethyl-3,4-dihydrocoumarin inhibits human leucocyte elastase (HLE) and porcine pancreatic elastase (PPE) through a mechanism-based process characterized by the following apparent enzyme-inhibitor dissociation constants, Ki, and limiting inactivation rate constants k2: 200 microM (HLE), 69 microM (PPE) and 5.10(-2) s-1 (HLE), 17.7.10(-2) s-1 (PPE) at pH 8.0, 37 degrees C. Bis(4-acyloxyphenyl)methane derivatives with a benzylic halogen as potential leaving group have also been synthesized and studied. They transiently inactivate PPE and HLE through the formation of an acyl-enzyme.

Protection of vascular basement membrane and microcirculation from elastase-induced damage with a fluorinated β-lactam derivative

N-(2-chloromethylphenyl) 3,3-difluoroazetidin-2-one (AA 231-1), a specific suicide-type inhibitor of elastase which is known to suppress the lysis of chromogenic oligopeptides, elastin and elastic fibers, is effective also in preventing the degradation of the vascular basement membrane. The degradation of porcine glomerular basement membrane by purified human leukocyte elastase (HLE), was reduced in proportion of inhibitor dose (8.3 microM for 50% inhibition). It is noteworthy that there was no reduction of the inhibitory effect when the addition of AA 231-1 was delayed for 1 h after the addition of the enzyme to the substrate. In the guinea pig, reduction of the dermal microhemorrhage due to HLE was related to the dose of inhibitor and to its preincubation time with HLE before intradermal injection. The inflammatory hemorrhage associated with the Arthus skin reaction was moderately depressed by AA 231-1 in situ. A part of the vascular permeability induced by HLE also responded to the inhibitor. In spite of the tissular diffusion and the time-dependence parameters which restrict responsiveness of elastase to AA 231-1 in vivo this biochemical compound should be helpful in the study and possibly the cure of vascular injury related to elastase.

Synthesis and inhibition of human leucocyte elastase by functionalized N-aryl azetidin-2-ones: effect of different substituents on the aromatic ring.

N‐aryl‐3,3‐difluoroazetidin‐2‐ones featured by a latent electrophilic methylene quinoniminium function have been synthesized and evaluated as inhibitors of human leucocyte elastase.

Evaluation of the Inhibitory Activity on Serine and Aspartic Proteases of 4-Amino-4H-1,2,4-triazole and 5-Aminothiazole Derivatives Structurally Related to β-Lactam Antibiotics

Abstract— Twenty new derivatives of 4‐amino‐4H‐1,2,4‐triazole and 5‐aminothiazole have been examined for their inhibitory potential towards serine and aspartic proteases. Upon prolonged incubation with enzyme, the phenylacetylaminothiazolium salts exhibit progressive, time‐dependent inhibition of chymotrypsin according to a first‐order process. The formation of a tetrahedral transition state‐like complex by attack of the active‐site serine at the C2‐position of the pseudobase form of the thiazolium may be responsible for the observed effect. Triazolium salts appeared to be simple competitive inhibitors of this enzyme, effective in the Mm range concentration. Poor inhibitions of trypsin and pepsin were also obtained in the triazolium series. In spite of their structural analogy with β‐lactams, the selected derivatives failed to inhibit human leucocyte elastase.