In vitro and ex vivo activity of new derivatives of acetohydroxyacid synthase inhibitors against Mycobacterium tuberculosis and non-tuberculous mycobacteria

Sohn, Honglae; Lee, Kil-Soo; Ko, Young-Kwan; Ryu, Jae-Wook; Woo, J. H.; Koo, Dong-Wan; Shin, Seung-Ho; Ahn, SeJin; Shin, Anna; Song, Chang‐Hwa; Jo, Eun-Kyeong; Park, Jeong‐Kyu; Kim, Hwa Jung

doi:10.1016/j.ijantimicag.2008.01.016

Cited by 32 publications

(27 citation statements)

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“…The in vivo and ex vivo growth inhibition of M. tuberculosis by sulfometuron methyl and its derivatives, shown in earlier studies (Grandoni et al, 1998;Sohn et al, 2008), may possibly have resulted from inhibition of all of the in vivo-active isozymes of AHAS. In vitro inhibition studies with other AHAS isozymes of M. tuberculosis will provide the necessary evidence for this contention.…”

Section: Discussionmentioning

confidence: 87%

Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice

et al. 2009

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Acetohydroxyacid synthase (AHAS) is the first enzyme in the branched-chain amino acid biosynthesis pathway in bacteria. Bioinformatics analysis revealed that the Mycobacterium tuberculosis genome contains four genes (ilvB1, ilvB2, ilvG and ilvX) coding for the large catalytic subunit of AHAS, whereas only one gene (ilvN or ilvH) coding for the smaller regulatory subunit of this enzyme was found. In order to understand the physiological role of AHAS in survival of the organism in vitro and in vivo, we inactivated the ilvB1 gene of M. tuberculosis. The mutant strain was found to be auxotrophic for all of the three branched-chain amino acids (isoleucine, leucine and valine), when grown with either C 6 or C 2 carbon sources, suggesting that the ilvB1 gene product is the major AHAS in M. tuberculosis. Depletion of these branched chain amino acids in the medium led to loss of viability of the DilvB1 strain in vitro, resulting in a 4-log reduction in colony-forming units after 10 days. Survival kinetics of the mutant strain cultured in macrophages maintained with sub-optimal concentrations of the branched-chain amino acids did not show any loss of viability, indicating either that the intracellular environment was rich in these amino acids or that the other AHAS catalytic subunits were functional under these conditions. Furthermore, the growth kinetics of the DilvB1 strain in mice indicated that although this mutant strain showed defective growth in vivo, it could persist in the infected mice for a long time, and therefore could be a potential vaccine candidate.

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Section: Discussionmentioning

confidence: 87%

Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice

et al. 2009

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“…Since both classes of compounds inhibit AHAS enzymes, the difference is likely to be in uptake of the compound into bacterial cells. Sulfonylureas have also been shown to inhibit growth of Mycobacterium tuberculosis both in vitro and in a mouse model (25,44) and of Brucella suis in macrophages (5). These results indicate that AHAS inhibitors have excellent potential for development as antimicrobials against infections of the respiratory tract or other "clean" body sites, although their utility against fulminant A. pleuropneumoniae infection may be limited in vivo due to the effect of the Apx toxins.…”

Section: Discussionmentioning

confidence: 99%

Branched-Chain Amino Acids Are Required for the Survival and Virulence ofActinobacillus pleuropneumoniaein Swine

et al. 2009

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In Actinobacillus pleuropneumoniae, which causes porcine pleuropneumonia, ilvI was identified as an in vivo-induced (ivi) gene and encodes the enzyme acetohydroxyacid synthase (AHAS) required for branchedchain amino acid (BCAA) biosynthesis. ilvI and 7 of 32 additional ivi promoters were upregulated in vitro when grown in chemically defined medium (CDM) lacking BCAA. Based on these observations, we hypothesized that BCAA would be found at limiting concentrations in pulmonary secretions and that A. pleuropneumoniae mutants unable to synthesize BCAA would be attenuated in a porcine infection model. Quantitation of free amino acids in porcine pulmonary epithelial lining fluid showed concentrations of BCAA ranging from 8 to 30 mol/liter, which is 10 to 17% of the concentration in plasma. The expression of both ilvI and lrp, a global regulator that is required for ilvI expression, was strongly upregulated in CDM containing concentrations of BCAA similar to those found in pulmonary secretions. Deletion-disruption mutants of ilvI and lrp were both auxotrophic for BCAA in CDM and attenuated compared to wild-type A. pleuropneumoniae in competitive index experiments in a pig infection model. Wild-type A. pleuropneumoniae grew in CDM؉BCAA but not in CDM؊BCAA in the presence of sulfonylurea AHAS inhibitors. These results clearly demonstrate that BCAA availability is limited in the lungs and support the hypothesis that A. pleuropneumoniae, and potentially other pulmonary pathogens, uses limitation of BCAA as a cue to regulate the expression of genes required for survival and virulence. These results further suggest a potential role for AHAS inhibitors as antimicrobial agents against pulmonary pathogens.Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia, a disease of significant economic importance throughout the swine-raising areas of the world (6, 48). This pathogen possesses several well-studied virulence factors, including Apx toxins (20), capsular polysaccharides (57, 58), lipopolysaccharide (1, 17, 41), fimbriae (63), and ironscavenging proteins (13, 50), which aid in the pathogenesis of acute pleuropneumonia marked by edema, hemorrhage, and necrosis (6, 26). In a search for additional virulence factors of this pathogen, we developed an in vivo expression technology (IVET) system and used this genetic tool to identify A. pleuropneumoniae gene promoters that are upregulated in vivo in the swine lung during infection compared to growth on laboratory media (22, 55).One of the A. pleuropneumoniae in vivo-induced (ivi) promoters that we identified drives the ilvIH operon, which encodes both large and small subunits of acetohydroxy acid synthase isozyme III (AHAS) (55). AHAS enzymes catalyze pivotal steps in the biosynthesis of the branched-chain amino acids (BCAA) isoleucine, leucine, and valine (31). In a survey of IVET, signature-tagged mutagenesis, and microarray studies of other pathogens, we observed that genes involved in BCAA biosynthesis were frequently identified in studies of pathogen...

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“…To combat these MDR/XDR-TB infections, new and effective pharmaceuticals are urgently needed [5,6]. Recently, acetohydroxyacid synthase (AHAS; EC 2.2.1.6) has been identified as an attractive target for designing a new generation of anti-TB agents [7][8][9]. AHAS is a key enzyme in the biosynthesis of branched-chain amino acids (leucine, isoleucine and valine) by higher plants, algae, fungi and bacteria, and no homologous enzyme has been observed in humans or animals [10].…”

Section: Introductionmentioning

confidence: 99%

Evaluation of the in vitro and intracellular efficacy of new monosubstituted sulfonylureas against extensively drug-resistant tuberculosis

Wang

Cao

et al. 2012

International Journal of Antimicrobial Agents

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In vitro and ex vivo activity of new derivatives of acetohydroxyacid synthase inhibitors against Mycobacterium tuberculosis and non-tuberculous mycobacteria

Cited by 32 publications

References 7 publications

Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice

Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice

Branched-Chain Amino Acids Are Required for the Survival and Virulence ofActinobacillus pleuropneumoniaein Swine

Evaluation of the in vitro and intracellular efficacy of new monosubstituted sulfonylureas against extensively drug-resistant tuberculosis

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