Antitubercular Triazines: Optimization and Intrabacterial Metabolism

Wang, Xin; Inoyama, Daigo; Russo, Riccardo; Li, Shao Gang; Jadhav, Ravindra D.; Stratton, Thomas P.; Mittal, Nisha; Bilotta, Joseph A.; Singleton, Eric; Kim, Thomas; Paget, Steve D.; Pottorf, Richard S.; Ahn, Yong Mo; Davila-Pagan, Alejandro; Kandasamy, Srinivasan; Grady, Courtney; Hussain, Syed M.; Soteropoulos, Patricia; Zimmerman, Matthew; Ho, Hsin Pin; Park, Steven; Dartois, Véronique; Ekins, Sean; Connell, Nancy; Kumar, Pradeep; Freundlich, Joel S.

doi:10.1016/j.chembiol.2019.10.010

Cited by 24 publications

(36 citation statements)

References 67 publications

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“…These results indicate that Mtb experiences DNA damage when treated with JSF-2019. A detailed analysis of JSF-2019 resistant mutants ( 41 ) uncovered the presence of mutations in rv2983 and rv2623 ( 44 – 46 ). Mutations in rv2983 have previously been found to generate resistance to fluoroquinolones ( 44 ) while overexpression of Rv2623 has been linked to exposure of Mtb to ofloxacin or moxifloxacin ( 45 , 46 ).…”

Section: Resultsmentioning

confidence: 99%

“…We unblinded the compound to learn if our conclusions were corroborated with previous mechanistic studies performed. Unknown compound 3 is JSF-2019, a triazine that resembles pretomanid in both its F420-dependent production of NO• and its ability to inhibit mycolic acid synthesis, albeit at a different step in the pathway (41). The mechanistic similarity of JSF-2019 and pretomanid validated the MorphEUS prediction of JSF-2019 acting like pretomanid at low dose but did not provide insight into the MorphEUS prediction of DNA targeting activity at high dose.…”

Section: Morpheus Correctly Classifies Cellular Targets Of Unknown Drmentioning

confidence: 97%

“…We hypothesized that the production of NO• by JSF-2019 at high doses induces DNA damage through DNA alkylation (42) in addition to its known cell wall-targeting activity (41). To test if JSF-2019 perturbs DNA processing pathways, we evaluated transcriptional profiles for ofloxacin-and JSF-2019-treated Mtb and found enrichment of coregulated genes involved in DNA damage (43) as well as nucleotide metabolism and biosynthesis (SI Appendix, Fig.…”

Section: Morpheus Correctly Classifies Cellular Targets Of Unknown Drmentioning

confidence: 99%

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Morphological profiling of tubercle bacilli identifies drug pathways of action

Smith

Pullen

Olson

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Morphological profiling is a method to classify target pathways of antibacterials based on how bacteria respond to treatment through changes to cellular shape and spatial organization. Here we utilized the cell-to-cell variation in morphological features of Mycobacterium tuberculosis bacilli to develop a rapid profiling platform called Morphological Evaluation and Understanding of Stress (MorphEUS). MorphEUS classified 94% of tested drugs correctly into broad categories according to modes of action previously identified in the literature. In the other 6%, MorphEUS pointed to key off-target activities. We observed cell wall damage induced by bedaquiline and moxifloxacin through secondary effects downstream from their main target pathways. We implemented MorphEUS to correctly classify three compounds in a blinded study and identified an off-target effect for one compound that was not readily apparent in previous studies. We anticipate that the ability of MorphEUS to rapidly identify pathways of drug action and the proximal cause of cellular damage in tubercle bacilli will make it applicable to other pathogens and cell types where morphological responses are subtle and heterogeneous.

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Section: Resultsmentioning

confidence: 99%

Section: Morpheus Correctly Classifies Cellular Targets Of Unknown Drmentioning

confidence: 97%

Section: Morpheus Correctly Classifies Cellular Targets Of Unknown Drmentioning

confidence: 99%

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Morphological profiling of tubercle bacilli identifies drug pathways of action

Smith

Pullen

Olson

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…3A, white star and 3B). Components of the mycobacterial cell-wall, in particular peptidoglycan (PG) and arabinogalactan (AG), are linked to energy production in the cell with components of glycolysis feeding directly into the synthesis of PG and AG (25, 26). In standard laboratory nutrient-replete medium, the presence of sugars allows Mtb to generate ATP from both glycolysis and TCA cycle through substrate-level phosphorylation and oxidative phosphorylation via the electron transport chain (6).…”

Section: Resultsmentioning

confidence: 99%

“…The transcriptional profile of JSF-2019, bedaquiline and moxifloxacin were obtained from GSE126718 (26), GSE43749 (6) and GSE71200 (10), respectively. The transcriptional profiles of other compounds were extracted from GSE1642 (11).…”

Section: Methodsmentioning

confidence: 99%

Morphological profiling of tubercule bacilli identifies drug pathways of action

Smith

Pullen

Olson

et al. 2020

Preprint

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34 35 36 Keywords 37 tuberculosis, drug discovery, cell morphology, high throughput 38 39 40 41 42 43 2 Abstract 44Morphological profiling is a method to classify target pathways of antibacterials based on how 45 bacteria respond to treatment through changes to cellular shape and spatial organization. Here, we utilized 46 the cell-to-cell variation in morphological features of Mycobacterium tuberculosis bacilli to develop a rapid 47 profiling platform called Morphological Evaluation and Understanding of Stress (MorphEUS). MorphEUS 48 classified 94% of tested drugs correctly into broad categories according to modes of action previously 49 identified in the literature. In the other 6%, MorphEUS pointed to key off-target or secondary bactericidal 50 activities. We observed cell-wall damaging activity induced by bedaquiline and moxifloxacin through 51 secondary effects downstream from their main target pathways. We implemented MorphEUS to correctly 52 classify three compounds in a blinded study and identified an off-target effect for one compound that was 53 not readily apparent in previous studies. We anticipate that the ability of MorphEUS to rapidly identify 54 pathways of drug action and the proximal cause of bactericidal activity in tubercule bacilli will make it 55 applicable to other pathogens and cell types where morphological responses are subtle and 56 heterogeneous. 58 Significance Statement 59Tuberculosis is a leading cause of death in the world and requires treatment with an arduous 60 multidrug regimen. Many new tuberculosis drugs are in development, and the drug development pipeline 61 would benefit from more rapid methods to learn drug mechanism of action and off-target effects. Here, we 62 describe a high throughput imaging method for rapidly classifying drugs into categories based on the 63 primary and secondary cellular damage called Morphological Evaluation and Understanding of drug-Stress 64 (MorphEUS). We anticipate that MorphEUS will assist in rapidly pinpointing causes of cellular death in 65 response to drug treatment in tuberculosis and other bacterial pathogens. 66 67 68 69 3 Main Text 70 Introduction 71Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), causes more deaths 72 annually than any other infectious agent (1). Tuberculosis treatment is lengthy, lasting between four months 73 to over a year (1). The difficult regimen, rate of relapse, and incidence of drug resistant Mtb has motivated 74 a significant effort to develop new antibacterial compounds that are effective in sterilizing Mtb infection (2). 75Many new drug classes and derivative compounds have been developed (2), but rapidly identifying the 76 primary and secondary pathways of action of each compound is often a protracted process due to the 77 difficulty in generating resistant mutants, and dissecting the broad-reaching metabolic effects of drug 78 treatment leading to death (3). Furthermore, drug action on bacterial cells can elicit dynamic responses in 79 multiple pathways both on-and off-target, s...

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