Pyrrolidine carboxamides as a novel class of inhibitors of enoyl acyl carrier protein reductase from Mycobacterium tuberculosis.

In view of the worldwide spread of multidrug resistance of Mycobacterium tuberculosis, there is an urgent need to discover antituberculosis agent with novel structures. InhA, the enoyl acyl carrier protein reductase (ENR) from M. tuberculosis, is one of the key enzymes involved in the mycobacterial fatty acid elongation cycle and has been validated as an effective antimicrobial target. We report here the discovery, through high-throughput screening, of a series of pyrrolidine carboxamides as a novel class of potent InhA inhibitors. Crystal structures of InhA complexed with three inhibitors have been used to elucidate the inhibitor binding mode. The potency of the lead compound was improved over 160-fold by subsequent optimization through iterative microtiter library synthesis followed by in situ activity screening without purification. Resolution of racemic mixtures of several inhibitors indicate that only one enantiomer is active as an inhibitor of InhA.

[1]  S. Abdel-Meguid,et al.  Molecular basis for triclosan activity involves a flipping loop in the active site , 2008, Protein science : a publication of the Protein Society.

[2]  Chi‐Huey Wong,et al.  A quick diversity-oriented amide-forming reaction to optimize P-subsite residues of HIV protease inhibitors. , 2002, Chemistry & biology.

[3]  Z. Otwinowski,et al.  [20] Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[4]  C. Chung,et al.  Effect of detergent on "promiscuous" inhibitors. , 2003, Journal of medicinal chemistry.

[5]  G. Besra,et al.  Pathway to Synthesis and Processing of Mycolic Acids in Mycobacterium tuberculosis , 2005, Clinical Microbiology Reviews.

[6]  W. Jacobs,et al.  inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosis. , 1994, Science.

[7]  Antoni R. Slabas,et al.  Molecular basis of triclosan activity , 1999, Nature.

[8]  J. Musser,et al.  Genotypic characterization of drug-resistant Mycobacterium tuberculosis isolates from Peru. , 1998, Tubercle and lung disease : the official journal of the International Union against Tuberculosis and Lung Disease.

[9]  R. Heath,et al.  Fatty acid biosynthesis as a target for novel antibacterials. , 2004, Current opinion in investigational drugs.

[10]  Tom Alber,et al.  Automated protein crystal structure determination using ELVES. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[11]  D W Rice,et al.  Inhibitor Binding Studies on Enoyl Reductase Reveal Conformational Changes Related to Substrate Recognition* , 1999, The Journal of Biological Chemistry.

[12]  S. Parikh,et al.  Roles of tyrosine 158 and lysine 165 in the catalytic mechanism of InhA, the enoyl-ACP reductase from Mycobacterium tuberculosis. , 1999, Biochemistry.

[13]  Böge Kp.  Neuroleptic activity and dopamine-uptake inhibition in 1-piperazino-3-phenylindans. , 1983 .

[14]  T. A. Jones,et al.  Databases in protein crystallography. , 1998, Acta crystallographica. Section D, Biological crystallography.

[15]  B. Shoichet,et al.  A common mechanism underlying promiscuous inhibitors from virtual and high-throughput screening. , 2002, Journal of medicinal chemistry.

[16]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[17]  D. Rice,et al.  Crystallographic analysis of triclosan bound to enoyl reductase. , 1999, Journal of molecular biology.

[18]  S. Parikh,et al.  Structural basis and mechanism of enoyl reductase inhibition by triclosan. , 1999, Journal of molecular biology.

[19]  I. Taylor,et al.  Kinetic and structural characteristics of the inhibition of enoyl (acyl carrier protein) reductase by triclosan. , 1999, Biochemistry.

[20]  Jie J. Zheng,et al.  The structural biology of type II fatty acid biosynthesis. , 2005, Annual review of biochemistry.

[21]  J C Sacchettini,et al.  Enzymatic characterization of the target for isoniazid in Mycobacterium tuberculosis. , 1995, Biochemistry.

[22]  B. Hammock,et al.  Optimization of amide-based inhibitors of soluble epoxide hydrolase with improved water solubility. , 2005, Journal of medicinal chemistry.

[23]  J. W. Campbell,et al.  Bacterial fatty acid biosynthesis: targets for antibacterial drug discovery. , 2001, Annual review of microbiology.

[24]  C. Rock,et al.  The reductase steps of the type II fatty acid synthase as antimicrobial targets , 2004, Lipids.

[25]  J. Sacchettini,et al.  Crystal structure and function of the isoniazid target of Mycobacterium tuberculosis , 1995, Science.

[26]  David Alland,et al.  Targeting Tuberculosis and Malaria through Inhibition of Enoyl Reductase , 2003, Journal of Biological Chemistry.

[27]  R. Heath,et al.  Broad Spectrum Antimicrobial Biocides Target the FabI Component of Fatty Acid Synthesis* , 1998, The Journal of Biological Chemistry.

[28]  Chi‐Huey Wong,et al.  Rapid diversity-oriented synthesis in microtiter plates for in situ screening: discovery of potent and selective alpha-fucosidase inhibitors. , 2003, Angewandte Chemie.

[29]  Chi‐Huey Wong,et al.  A Potent and Highly Selective Inhibitor of Human α-1,3-Fucosyltransferase via Click Chemistry , 2003 .

[30]  J. Gathe,et al.  The Reaction of Itaconic Acid with Primary Amines , 1950 .

[31]  S. Cole,et al.  The catalase—peroxidase gene and isoniazid resistance of Mycobacterium tuberculosis , 1992, Nature.

[32]  L. Collins,et al.  Microplate alamar blue assay versus BACTEC 460 system for high-throughput screening of compounds against Mycobacterium tuberculosis and Mycobacterium avium , 1997, Antimicrobial agents and chemotherapy.