Coumarin derivatives were recently shown to constitute a totally new class of inhibitors of the zinc metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1), being hydrolyzed within the CA active site to 2-hydroxycinnamic acids. We explore here a new series of variously substituted coumarins and a thiocoumarin for their interaction with 13 mammalian CA isoforms, detecting low nanomolar and isoform selective inhibitors. The mechanism of action of this class of inhibitors is delineated in detail by resolving the X-ray crystal structure of CA II in complex with trans-2-hydroxy-cinnamic acid, the in situ hydrolysis product of simple coumarin. Thiocoumarins also act as efficient CAIs, similarly to coumarins. The versatility of the (thio)coumarin chemistry, the cis-trans isomerization evidenced here, and easy derivatization of the (thio)coumarin rings, coupled with the nanomolar inhibition range of several isozymes, afford isoform-selective CAIs with various biomedical applications, which render these classes of compounds superior to the clinically used sulfonamides.

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A series of esters and amides of 6-(chloromethyl)-2-oxo-2H-1-benzopyran-3-carboxylic acid were synthesized and evaluated in vitro for their inhibitory activity toward bovine alpha-chymotrypsin and human leukocyte elastase. Both series behaved as time-dependent inhibitors of alpha-chymotrypsin, but ester-type coumarins were clearly more efficient than the corresponding amides in inactivating the serine proteinase. The best inactivations were observed with "aromatic" esters, in particular with meta-substituted phenyl esters such as m-chlorophenyl 6-(chloromethyl)-2-oxo-2H-1-benzopyran-3-carboxylate, which appears to be one of the most powerful inactivators of alpha-chymotrypsin yet reported (kinact/KI = 760,000 M-1 S-1 at pH 7.5 and 25 degrees C). Usually, the coumarin derivatives failed to inhibit significantly human leukocyte elastase. As a result, the reported series of aromatic coumarinic esters behaves as a new chemical family of selective alpha-chymotrypsin inhibitors.

Tryptophan catabolism mediated by indoleamine 2,3-dioxygenase (IDO) is an important mechanism of peripheral immune tolerance contributing to tumoral immune resistance. IDO inhibition is thus an active area of research in drug development. Recently, our group has shown that tryptophan 2,3-dioxygenase (TDO), an unrelated hepatic enzyme also catalyzing the first step of tryptophan degradation, is also expressed in many tumors and that this expression prevents tumor rejection by locally depleting tryptophan. Herein, we report a structure-activity study on a series of 3-(2-(pyridyl)ethenyl)indoles. More than 70 novel derivatives were synthesized, and their TDO inhibitory potency was evaluated. The rationalization of the structure-activity relationships (SARs) revealed essential features to attain high TDO inhibition and notably a dense H-bond network mainly involving His(55) and Thr(254) residues. Our study led to the identification of a very promising compound (58) displaying good TDO inhibition (K(i) = 5.5 μM), high selectivity, and good oral bioavailability. Indeed, 58 was chosen for preclinical evaluation.

To overcome the intrinsic resistance of cancer cells to apoptotic stimuli, we designed and synthesized approximately 50 novel β-carbolines structurally related to harmine. Harmine is known for its anticancer properties and is a DYRK1A inhibitor. Of the synthesized compounds, the most active in terms of growth inhibition of five cancer cell lines are cytostatic and approximately 100 times more potent than harmine but demonstrated no DYRK1A inhibitory activity. These novel β-carbolines display similar growth inhibitory activity in cancer cells that are sensitive and resistant to apoptotic stimuli. Using ChemGPS-NP, we found that the more active β-carbolines are all more lipophilic and larger than the less active compounds. Lastly, on the basis of the NCI human tumor cell line anticancer drug screen and the NCI COMPARE algorithm, it appears that some of these compounds, including 5a and 5k, seem to act as protein synthesis inhibitors.

Pyridyl esters of 6-substituted 2-oxo-2H-1-benzopyran-3-carboxylic acid were designed as mechanism-based inhibitors of human leukocyte elastase. Compounds of series 4 specifically inhibited this enzyme. Several of the tested compounds (series 2 and 3) acted as powerful time-dependent inhibitors of both human leukocyte elastase and alpha-chymotrypsin; some compounds of these series inhibited thrombin. Trypsin was not inhibited. A transient inactivation was observed for human leukocyte elastase (k(i)/K(I) = 107 000 M(-1). s(-1) for 4c) and thrombin (k(i)/K(I) = 7 200 M(-1).s(-1) for 3b) as demonstrated by spontaneous or hydroxylamine-accelerated reactivation, irrespective of the nature of the substituent at the 6-position. Conversely, alpha-chymotrypsin was irreversibly inhibited by 6-chloromethyl derivatives (k(i)/K(I) = 107 400 M(-1). s(-1) for 3b). The presence of a latent alkylating function at the 6-position (chloromethyl group) was required for leading to this inactivation. In the absence of such an alkylating function (series 4), human leukocyte elastase was specifically inhibited suggesting that this new series of human leukocyte elastase inhibitors may be of potential therapeutic interest in degradative and degenerative processes involving this enzyme.

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AbstractÐNovel coumarinic derivatives were synthesized and tested for their inhibitory potency toward a-CT and HLE. Cycloalkyl esters and amides were found to be essentially inactive on both enzymes. On the opposite, aromatic esters strongly inactivated a-CT whereas HLE was less e ciently inhibited with dichlorophenyl ester derivatives (k inact /K I =4000 M À1 s À1 for 36). Representative examples of amide, ester, thioester and ketone derivatives were prepared in order to evaluate the in¯uence of the link between the coumarinic ring and the phenyl side chain. The irreversible inactivation of a-CT by 6-chloromethyl derivatives should be due to alkylation of a histidine residue as suggested by the amino acid analysis of the modi®ed chymotrypsin. Conversely the inhibition of HLE was transient. Intrinsic reactivity of coumarins has been calculated using a model of a nucleophilic reaction between the ligand and the couple methanol±water. From this calculation, it appears that di erences in the inhibitory potency expressed by these molecules cannot only be explained by di erences in the reactivity of the lactonic carbonyl group toward the nucleophilic attack. #