A large-scale chemical-genetic strategy to design antimicrobial combination chemotherapy for<i>Mycobacterium tuberculosis</i>

Johnson, Eachan O. D.; Office, Emma; Kawate, Tomohiko; Orzechowski, Marek; Hung, Deborah T.

doi:10.1101/772459

Cited by 4 publications

(4 citation statements)

References 26 publications

(17 reference statements)

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“…We have used M. smegmatis to examine the suppression of the whole-cell efflux pump by mycobactin analogues against mycobacteria. A non-virulent mycobacterial strain, M. smegmatis mc 2 155, is frequently employed as a surrogate model for the screening of recently identified efflux pump inhibitors to avoid the complexity associated with M. tuberculosis in the early drug discovery stage (Johnson et al, 2020). However, pathogenic, non-pathogenic, and NTM categories all-share siderophore biosynthesis and recycling pathways (Shyam et al, 2021;Shyam et al, 2022a).…”

Section: Investigation Of Mycobacterium Smegmatis Efflux-pump Inhibit...mentioning

confidence: 99%

Mycobactin analogue interacting with siderophore efflux-pump protein: insights from molecular dynamics simulations and whole-cell assays

Shyam,

Thakur,

Velez

et al. 2024

Front. Antibiot.

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IntroductionIn response to continued public health emergency of antimicrobial resistance (AMR), a significant key strategy is the discovery of novel mycobacterial efflux-pump inhibitors (EPIs) as potential adjuvants in combination drug therapy. Interest in identifying new chemotypes which could potentially synergize with the existing antibiotics and can be deployed as part of a combination therapy. This strategy could delay the emergence of resistance to existing antibiotics and increase their efficacy against resistant strains of mycobacterial species. In recent decades, notable approaches have been accounted for EPI development and have resulted in the discovery of several EPIs including SQ109 and AU1235. In context, to accelerate newer EPIs with novel mode of action here we have discussed mycobactin analogues and highlighted in silico binding orientation with siderophore efflux-pump proteins MmpL4/5.Methods3-(2-hydroxyphenyl)-5-(aryl)-pyrazoline series was investigated for whole-cell efflux-pump inhibitory activity against Mycobacterium smegmatis and Mycobacterium abscessus. Machine learning and molecular dynamics were performed to construct a MmpL4/5 complex embedded in a lipid bilayer to identify the putative binding site and to predict ligand-protein binding energetics. Furthermore, the identified HIT compound was investigated in synergistic assay with bedaquiline.ResultsCompound Il, 2-(5-(4-fluorophenyl)-4,5-dihydro-1H-pyrazol-3-yl)phenol, was identified as the most potent efflux pump inhibitor against M. smegmatis in whole-cell efflux-pump investigation. Followed HIT Il employed against M. abscessus for efflux-pump inhibition investigations and notable whole-cell efflux-pump inhibitory profile has been observed. The theoretical investigations predicted compound Il to be selective towards MmpL4, with significant hydrogen bonding and π-π stacking interactions effectively blocking a critical Asp-Tyr dyad interaction network necessary for proton translocation. Compound Il with bedaquiline highlighted an additive profile against the M. abscessus pathogen.ConclusionsMD simulations and whole-cell assays are indicating potential development of compound Il as an adjunct to the existing therapeutic regimen against mycobacterial infections.

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Section: Investigation Of Mycobacterium Smegmatis Efflux-pump Inhibit...mentioning

confidence: 99%

Mycobactin analogue interacting with siderophore efflux-pump protein: insights from molecular dynamics simulations and whole-cell assays

Shyam,

Thakur,

Velez

et al. 2024

Front. Antibiot.

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“…Chemicals can also be screened against hypomorph libraries to analyze potential off-target effects and to validate mechanisms of action. For instance, proteolytic degradation systems were recently used in a large chemical-genetic screen to combine small molecules with a pool of Mtb strains depleted in essential genes (Johnson et al, 2020). Because these hypomorphs are hypersensitive, this approach was found to yield more hit compounds of interest than whole-cell chemical screens alone, and their effects on different strains shed light on compound mechanisms of action.…”

Section: Genetic Knockdownsmentioning

confidence: 99%

“…An inhibitor of a new target was also identified, which interfered with the essential Mtb efflux pump EfpA. This inhibitor, which was effective against an EfpA hypomorph, was then chemically modified for increased potency against a wildtype Mtb strain (Johnson et al, 2020). This example shows the utility of combining genetic knockdowns with whole-cell chemical screens to identify small molecules that would not be unearthed by exclusively screening wild-type cells.…”

Section: Genetic Knockdownsmentioning

confidence: 99%

Failing upwards: Genetics-based strategies to improve antibiotic discovery and efficacy in Mycobacterium tuberculosis

Tomasi

Rubín

2022

Front. Cell. Infect. Microbiol.

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Therapeutic advances in the 20th century significantly reduced tuberculosis (TB) mortality. Nonetheless, TB still poses a massive global health challenge with significant annual morbidity and mortality that has been amplified during the COVID-19 pandemic. Unlike most common bacterial infectious diseases, successful TB treatment requires months-long regimens, which complicates the ability to treat all cases quickly and effectively. Improving TB chemotherapy by reducing treatment duration and optimizing combinations of drugs is an important step to reducing relapse. In this review, we outline the limitations of current multidrug regimens against TB and have reviewed the genetic tools available to improve the identification of drug targets. The rational design of regimens that sterilize diverse phenotypic subpopulations will maximize bacterial killing while minimizing both treatment duration and infection relapse. Importantly, the TB field currently has all the necessary genetic and analytical tools to screen for and prioritize drug targets in vitro based on the vulnerability of essential and nonessential genes in the Mtb genome and to translate these findings in in vivo models. Combining genetic methods with chemical screens offers a formidable strategy to redefine the preclinical design of TB therapy by identifying powerful new targets altogether, as well as targets that lend new efficacy to existing drugs.

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“…For example, atpE is the target for only bedaquiline. However, resistance to bedaquiline, clofazimine, and even new tuberculosis inhibitors like BRD-9327 can be conferred by efflux pump regulator mutations in Rv0678 (Johnson et al, 2020).…”

Section: Mechanisms Of Drug Resistancementioning

confidence: 99%

Evolution of Mycobacterium tuberculosis drug resistance in the genomic era

Nimmo

Millard

Faulkner

et al. 2022

Front. Cell. Infect. Microbiol.

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Mycobacterium tuberculosis has acquired drug resistance to all drugs that have been used against it, including those only recently introduced into clinical practice. Compared to other bacteria, it has a well conserved genome due to its role as an obligate human pathogen that has adapted to a niche over five to ten thousand years. These features facilitate reconstruction and dating of M. tuberculosis phylogenies, giving key insights into how resistance has been acquired and spread globally. Resistance to each new drug has occurred within five to ten years of clinical use and has occurred even more rapidly with recently introduced drugs. In most cases, resistance-conferring mutations come with a fitness cost, but this can be overcome by compensatory mutations which restore fitness to that of wild-type bacteria. It is likely that M. tuberculosis acquires drug resistance while maintaining limited genomic variability due the generation of low frequency within-host variation, combined with ongoing purifying selection causing loss of variants without a clear fitness advantage. However, variants that do confer an advantage, such as drug resistance, can increase in prevalence amongst all bacteria within a host and become the dominant clone. These resistant strains can then be transmitted leading to primary drug resistant infection in a new host. As many countries move towards genomic methods for diagnosis of M. tuberculosis infection and drug resistance, it is important to be aware of the implications for the evolution of resistance. Currently, understanding of resistance-conferring mutations is incomplete, and some targeted genetic diagnostics create their own selective pressures. We discuss an example where a rifampicin resistance-conferring mutation which was not routinely covered by standard testing became dominant. Finally, resistance to new drugs such as bedaquiline and delamanid is caused by individually rare mutations occurring across a large mutational genomic target that have been detected over a short time, and do not provide statistical power for genotype-phenotype correlation – in contrast to longer-established drugs that form the backbone of drug-sensitive antituberculosis therapy. Therefore, we need a different approach to identify resistance-conferring mutations of new drugs before their resistance becomes widespread, abrogating their usefulness.

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A large-scale chemical-genetic strategy to design antimicrobial combination chemotherapy forMycobacterium tuberculosis

Cited by 4 publications

References 26 publications

Mycobactin analogue interacting with siderophore efflux-pump protein: insights from molecular dynamics simulations and whole-cell assays

Mycobactin analogue interacting with siderophore efflux-pump protein: insights from molecular dynamics simulations and whole-cell assays

Failing upwards: Genetics-based strategies to improve antibiotic discovery and efficacy in Mycobacterium tuberculosis

Evolution of Mycobacterium tuberculosis drug resistance in the genomic era

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