Azetidines Kill Multidrug-Resistant Mycobacterium tuberculosis without Detectable Resistance by Blocking Mycolate AssemblyClick to copy article linkArticle link copied!
- Yixin CuiYixin CuiSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Yixin Cui
- Alice LanneAlice LanneInstitute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Alice Lanne
- Xudan PengXudan PengState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Xudan Peng
- Edward BrowneEdward BrowneSygnature Discovery, The Discovery Building, BioCity, Pennyfoot Street, Nottingham NG1 1GR, U.K.More by Edward Browne
- Apoorva BhattApoorva BhattInstitute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Apoorva Bhatt
- Nicholas J. ColtmanNicholas J. ColtmanSchool of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Nicholas J. Coltman
- Philip CravenPhilip CravenSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Philip Craven
- Liam R. CoxLiam R. CoxSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Liam R. Cox
- Nicholas J. CundyNicholas J. CundySchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Nicholas J. Cundy
- Katie DaleKatie DaleInstitute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Katie Dale
- Antonio FeulaAntonio FeulaSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Antonio Feula
- Jon FramptonJon FramptonCollege of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Jon Frampton
- Martin FungMartin FungCentre for Regenerative Medicine and Health, Hong Kong Institute of Science & Innovation, Chinese Academy of Sciences, 15 Science Park West Avenue NT, Hong Kong SARMore by Martin Fung
- Michael MortonMichael MortonApconiX Ltd, BIOHUB at Alderly Park, Nether Alderly, Cheshire SK10 4TG, U.K.More by Michael Morton
- Aaron GoffAaron GoffDepartment of Global Health and Infection, Brighton and Sussex Medical School, University of Sussex, Falmer BN1 9PX, U.K.More by Aaron Goff
- Mariwan SalihMariwan SalihSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Mariwan Salih
- Xingfen LangXingfen LangState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Xingfen Lang
- Xingjian LiXingjian LiSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.State Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Xingjian Li
- Chris MoonChris MoonTB Research Group, National Infection Service, Public Health England (UKHSA), Manor Farm Road, Porton, Salisbury SP4 0JG, U.K.More by Chris Moon
- Jordan PascoeJordan PascoeTB Research Group, National Infection Service, Public Health England (UKHSA), Manor Farm Road, Porton, Salisbury SP4 0JG, U.K.More by Jordan Pascoe
- Vanessa PortmanVanessa PortmanSygnature Discovery, The Discovery Building, BioCity, Pennyfoot Street, Nottingham NG1 1GR, U.K.More by Vanessa Portman
- Cara PressCara PressInstitute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Cara Press
- Timothy Schulz-UtermoehlTimothy Schulz-UtermoehlSygnature Discovery, The Discovery Building, BioCity, Pennyfoot Street, Nottingham NG1 1GR, U.K.More by Timothy Schulz-Utermoehl
- Suki LeeSuki LeeCentre for Regenerative Medicine and Health, Hong Kong Institute of Science & Innovation, Chinese Academy of Sciences, 15 Science Park West Avenue NT, Hong Kong SARMore by Suki Lee
- Micky D. TortorellaMicky D. TortorellaState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaCentre for Regenerative Medicine and Health, Hong Kong Institute of Science & Innovation, Chinese Academy of Sciences, 15 Science Park West Avenue NT, Hong Kong SARMore by Micky D. Tortorella
- Zhengchao TuZhengchao TuState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Zhengchao Tu
- Zoe E. UnderwoodZoe E. UnderwoodTB Research Group, National Infection Service, Public Health England (UKHSA), Manor Farm Road, Porton, Salisbury SP4 0JG, U.K.More by Zoe E. Underwood
- Changwei WangChangwei WangState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Changwei Wang
- Akina YoshizawaAkina YoshizawaSchool of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Akina Yoshizawa
- Tianyu ZhangTianyu ZhangState Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaMore by Tianyu Zhang
- Simon J. WaddellSimon J. WaddellDepartment of Global Health and Infection, Brighton and Sussex Medical School, University of Sussex, Falmer BN1 9PX, U.K.More by Simon J. Waddell
- Joanna BaconJoanna BaconTB Research Group, National Infection Service, Public Health England (UKHSA), Manor Farm Road, Porton, Salisbury SP4 0JG, U.K.More by Joanna Bacon
- Luke Alderwick*Luke Alderwick*Email: [email protected]Institute of Microbiology and Infection, School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.Discovery Sciences, Charles River Laboratories, Chesterford Research Park, Saffron Walden CB10 1XL, U.K.More by Luke Alderwick
- John S. Fossey*John S. Fossey*Email: [email protected]School of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by John S. Fossey
- Cleopatra Neagoie*Cleopatra Neagoie*Email: [email protected]State Key Laboratory of Respiratory Disease, China-New Zealand Joint Laboratory on Biomedicine and Health, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science, 190 Kai Yuan Avenue, Science Park, Guangzhou 510530, ChinaCentre for Regenerative Medicine and Health, Hong Kong Institute of Science & Innovation, Chinese Academy of Sciences, 15 Science Park West Avenue NT, Hong Kong SARVisiting Scientist, School of Chemistry, University of Birmingham, Edgbaston, Birmingham, West Midlands B15 2TT, U.K.More by Cleopatra Neagoie
Abstract
Tuberculosis (TB) is the leading cause of global morbidity and mortality resulting from infectious disease, with over 10.6 million new cases and 1.4 million deaths in 2021. This global emergency is exacerbated by the emergence of multidrug-resistant MDR-TB and extensively drug-resistant XDR-TB; therefore, new drugs and new drug targets are urgently required. From a whole cell phenotypic screen, a series of azetidines derivatives termed BGAz, which elicit potent bactericidal activity with MIC99 values <10 μM against drug-sensitive Mycobacterium tuberculosis and MDR-TB, were identified. These compounds demonstrate no detectable drug resistance. The mode of action and target deconvolution studies suggest that these compounds inhibit mycobacterial growth by interfering with cell envelope biogenesis, specifically late-stage mycolic acid biosynthesis. Transcriptomic analysis demonstrates that the BGAz compounds tested display a mode of action distinct from the existing mycobacterial cell wall inhibitors. In addition, the compounds tested exhibit toxicological and PK/PD profiles that pave the way for their development as antitubercular chemotherapies.
This publication is licensed under
License Summary*
You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below:
Creative Commons (CC): This is a Creative Commons license.
Attribution (BY): Credit must be given to the creator.
*Disclaimer
This summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials.
License Summary*
You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below:
Creative Commons (CC): This is a Creative Commons license.
Attribution (BY): Credit must be given to the creator.
*Disclaimer
This summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials.
License Summary*
You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below:
Creative Commons (CC): This is a Creative Commons license.
Attribution (BY): Credit must be given to the creator.
*Disclaimer
This summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials.
Introduction
Results
Identification and Development of Azetidine Derivatives with Antimycobacterial Activity

MIC99 as determined by a modified Gompertz function.
MIC values were determined from three biological replicates using a resazurin end point assay.
Figure 1
Figure 1. BGAz derivatives synthesized.

The M. tb H37Ra::pTYOK is an auto-luminescent strain of mycobacteria. (24) The MIC99 values of BGAz-002–BGAz-005 against M. smegmatis and M. bovis BCG, drug-sensitive M. tuberculosis H37Rv (reference strain) and M. tuberculosis 1192/015 (clinical isolate), and multi-drug-resistant M. tuberculosis 08/00483E (clinical isolate resistant to INH, RIF, PZA and EMB). MIC values were determined from three biological replicates using a resazurin end point assay. The minimum bactericidal concentration (MBC) was determined for BGAz-002–BGAz-005 against M. bovis BCG.
Physiochemical and Toxicological Properties of BGAz-002–BGAz-005

Measures maximum peak serum concentrations of the drug.
The time of first occurrence of Cmax.
The terminal half-life of the drug.
The area under the plasma drug concentration–time curve to infinite time.
As a result of cassette dosing of BGAz-001–005; see the PK Data file in the Supporting Information.
Multiple daily dosing 4 × 30 mg/kg; see the PK Data file in the Supporting Information.
Multiple daily dosing 3 × 30 mg/kg; see the PK Data file in the Supporting Information.
BGAz Compounds Kill M. tuberculosis with Bactericidal Activity
Figure 2
Figure 2. Assessment of bactericidal activity of BGAz-004 and BGAz-005 against M. tuberculosis H37Rv. Average total viable counts (CFU mL–1) of M. tuberculosis cultures exposed to either BGAz-004 (Panel A) or BGAz-005 (Panel B) at concentrations: 0 μM (0.1% DMSO) (circle, closed), 3, 6, 12, 24, 48, and 96 μM or isoniazid (Panel C) at concentrations 0 μM (0.1% DMSO), 0.9, 1.8, 3.7, 7.3, 14.6, and 29.2 μM over a 14-day time-course. Samples were taken after 0, 2, 6, 10, and 14 days of antibiotic exposure, serially diluted, and plated by the method of Miles et al. (27) Statistical comparisons were performed at 6, 10, and 14 days of antibiotic exposure at 96 μM BGAz-004 and BGAz-005 using factorial ANOVA and posthoc Tukey’s honestly significant difference test (Panel D). Data represent three biological repeats ± standard deviation.
Figure 3
Figure 3. Assessment of bactericidal activity of BGAz-004 and BGAz-005 against M. tuberculosis H37Rv. Quantitation of Calcien-Violet-AM (CV-AM) and Sytox-green (SG) fluorescence of M. tuberculosis H37Rv, using flow cytometry, after exposure to BGAz-004 (column A) and BGAz-005 (column B) at concentrations: 0 μM (0.1% DMSO), 3, 6, 12, 24, 48, and 96 μM or (column C) isoniazid at concentrations 0 μM (0.1% DMSO), 0.9, 1.8, 3.7, 7.3, 14.6, and 29.2 μM over a 14-day time-course. The percentages of the population that are unstained or stained with each dye (or both dyes) are represented in four gates (rows P1–P4). Row P1: unstained population (CV-AM–/SG–); row P2: CV-stained population (CV-AM+/SG–); row P3: dual-stained population (CV-AM+/SG+); and row P4: SG-stained population (CV-AM–/SG+). Data represent three biological repeats ± standard deviation. Statistical comparisons were made using factorial ANOVA and posthoc Tukey’s honestly significant difference test.
BGAz-004 and BGAz-005 Inhibit the Incorporation of Mycobacterial Cell Wall Precursors and Display No Detectable Resistance
Figure 4
Figure 4. Effect of BGAz-005 on the incorporation of radiolabeled precursors into the major cellular macromolecules of M. smegmatis. The incorporation of (A) [methyl-3H]thymidine (for DNA), (B) [5,6-3H]uridine (for RNA), (C) l-[4,5-3H]leucine (for protein), (D) [3H]meso-diaminopimelic acid (for peptidoglycan), and (E) [14C]acetic acid (for lipids) was measured over a period of 36 h. The percentage of incorporation measured at 36 h is represented in panel F. Each plot and error bars represent the average of three independent experiments.
BGAz-005 Dysregulates the Expression of Cell Envelope Biosynthetic Genes
Figure 5
Figure 5. Transcriptional response to BGAz-005 exposure demonstrating inhibition of mycobacterial cell envelope biosynthesis. (A) Cluster diagram of all genes showing similarity of biological replicates and separation of drug-treated from carrier control samples. (B) Volcano plot of M. bovis BCG response to BGAz-005, highlighting genes significantly differentially expressed. (C) Heatmap of 286 gene BGAz-005 signature relative to carrier control. Conditions as columns, genes as rows; red coloring highlighting induced genes and blue representing repressed genes. The BGAz-004 signature is clustered alongside, indicating a similar mode of drug action.
BGAz-004 and BGAz-005 Significantly Alter Mycobacterial Cell Envelope Composition
Figure 6
Figure 6. BCG cell envelope lipid analysis upon exposure to BGAz-005. BCG were cultured in 7H9 broth and exposed to increasing concentrations of BGAz-005. Lipids were selectively labeled with [14C]-acetic acid for 12 h, and cell envelope lipids were selectively removed by solvent extraction, separated by TLC (chloroform/methanol/water, 80:20:2, v/v/v), and visualized by autoradiography. (A) Equal volumes of lipids loaded adjusted for BCG growth; (B) equal counts of lipids (25,000 cpm) loaded; (C) mycolic acid methyl ester (MAME) analysis of cell-wall bound mycolates released by 5% TBAH and separated by TLC (petroleum ether/acetone, 95:5, v/v); (D) quantification of BCG lipids from panels A–C by densitometry. M. smegmatis cell envelope lipid analysis upon exposure to BGAz-005. (E) M. smegmatis were cultured in 7H9 broth, exposed to increasing concentrations of BGAz-005 for 6 h and the cell envelope lipids selectively removed by solvent extraction. Equal volumes of lipid adjusted by bacterial growth were separated by TLC (chloroform/methanol/water, 80:20:2, v/v/v) and stained with MPA or (F) alpha-naphthol. (G) Equal volumes of lipid adjusted by bacterial growth were separated by TLC (hexane/diethyl ether/acetic acid), 70:30:1, v/v/v and stained with MPA. (H) Equal volumes of lipid adjusted by bacterial growth were separated by 2D-TLC (direction 1 chloroform/methanol 96:4, v/v, direction 2 toluene/acetone 80:20, v/v) and stained with MPA.
BGAz-004 and BGAz-005 Target Late-Stage Mycolic Acid Biosynthesis Enzymes
Figure 7
Figure 7. Assessing the MIC shift of BGAz-004 and BGAz-005 against the AG85 complex. MIC values of BGAz-004, BGAz-005, and Ebselen were determined against BCG harboring overexpression vectors and compared to empty vector controls (pTIC6) in order to identify a shift in MIC against fbpA (A), fbpB (B), and fbpC (C). Fold change in MIC shift (D). The MIC99 was calculated using an end point resazurin assay and the Gomperz equation for MIC determination (GraphPad Prism). Data are of triplicate repeats.
Discussion and Conclusions
Experimental Section
Chemistry at GIBH
Synthesis of BGAz-001–BGAz-005
Bacterial Strains and Growth Conditions
Determination of MIC and MBC
Determination of MIClux50 against Autoluminescent M.tb H37Ra
Determination of MIC99 for Clinical M.tb Strains
Physiochemical and Toxicological Analysis
Kinetic Solubility
Mouse PPB
Mouse Microsomal Clearance
Mouse Hepatocyte Clearance
Caco-2 and Efflux
Pharmacokinetic Studies
Cytochrome P450 Activities
HepG2 Mitochondrial Dysfunction
hERG Cardiotoxicity Function
Mycobacterial Time Kill Experiments
Bacterial Staining and Flow Cytometry Analyses
Inhibition of Macromolecular Synthesis
Transcriptomic Profiling by RNA-seq Analysis
Radioisotope Labeling of Lipids and Analysis
Target Gene Overexpression Studies
Supporting Information
The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.jmedchem.3c01643.
Supplementary PK data (PDF).
A detailed description of author contributions, general methods, chemical and biological general procedures, synthetic chemistry protocols for the synthesis of BGAz001–BGAz0016, NMR spectra, details of mass spectrometry analysis, additional corresponding references, (82,83) and molecular formula strings (ZIP).
Terms & Conditions
Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.
Acknowledgments
We dedicate this study to the memory of Professor John Fossey, an exceptional synthetic chemist who passed away on April 15, 2022. Professor Fossey’s passion for chirality and stereoselective synthesis marked him as a distinguished scientific leader in the field of chemistry. His legacy endures through his scientific integrity, innovative thinking, and his role as a remarkable scientific mentor and collaborator. Professor John Fossey will be profoundly missed by colleagues, family, and friends, and we extend our deepest sympathies and sincere condolences to his loved ones. We are grateful for the support underpinning much of this study from MRC Confidence in Concepts and EPSRC follow-on fund schemes. The University of Birmingham is acknowledged for support, including travel funds permitting A.Y., X.L., and Y.C. to undertake training placements at GIBH. J.S.F. is grateful to the Royal Society for the training provided because of a previous Industrial Fellowship and the EPSRC for previous funding (EP/J003220/1). Funding for part of this study was received from Public Health England. This work was supported by the National Key R&D Program of China (2021 YFA1300900) and by the Chinese Academy of Sciences Grant (YJKYYQ20210026, 154144KYSB20190005). S.J.W. and A.G. thank the National Centre for the Replacement, Refinement. and Reduction of Animals in Research (NC3Rs) for grant support (NC/R001669/1). Qiong Pan (GIBH), Jingfang Xiong (GIBH). and Miaoqin She (GIBH) are acknowledged for conducting aspects of the PK/PD studies of this report. Dr. Chi Tsang (UoB), Dr. Peter Ashton (UoB), and Jiajia Wei (GIBH) are acknowledged for their helpful discussions and practical support with aspects of mass spectrometry. Dr. Cécile S. Le Duff (UoB) and Dr. Neil Spencer (UoB) gave advice on aspects of NMR spectroscopy underpinning the preliminary or previously reported, findings. Yingxue Liu (GIBH) is acknowledged for help with the purification of final products by HPLC where required.
AG | Arabinogalactan |
Ag85 | antigen 85 |
BGAz | azetidine derivate |
MmpL3 | mycobacterial membrane protein Large 3 |
PKs13 | polyketide synthase 13 |
TDM | trehalose dimycolate |
TMM | trehalose monomycolate |
WGS | whole genome sequencing. |
References
This article references 85 other publications.
- 1www.who.int/gho/tb/en/. World Health Organisation: Global Health Observatory (GHO) data www.who.int/gho/tb/en/.Google ScholarThere is no corresponding record for this reference.
- 2Global tuberculosis report 2022; World Health Organization: Geneva, 2022, Licence: CC BY-NC-SA 3.0 IGO.Google ScholarThere is no corresponding record for this reference.
- 3Combs, D. L.; O’Brien, R. J.; Geiter, L. J. USPHS Tuberculosis Short-Course Chemotherapy Trial 21: Effectiveness, Toxicity, and Acceptability: The Report of Final Results. Ann. Int. Med. 1990, 112, 397– 406, DOI: 10.7326/0003-4819-76-3-112-6-397Google Scholar3https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADyaK3c7nvVyqtg%253D%253D&md5=2df74050189541e5d5b92bbb5acde939USPHS Tuberculosis Short-Course Chemotherapy Trial 21: effectiveness, toxicity, and acceptability. The report of final resultsCombs D L; O'Brien R J; Geiter L JAnnals of internal medicine (1990), 112 (6), 397-406 ISSN:0003-4819.STUDY OBJECTIVE: To determine the effectiveness, toxicity, and acceptability of a 6-month antituberculous regimen compared with a 9-month regimen. DESIGN: A nonblinded, unbalanced, randomized, multicenter clinical trial. SETTING: Twenty-two tuberculosis clinics in public health departments and hospitals in the United States. PATIENTS: Patients were eligible if Mycobacterium tuberculosis, isolated from sputum cultures, was susceptible to study drugs. Of 1451 patients enrolled, 75% (617 of 823) assigned to the 6-month regimen and 71% (445 of 628) assigned to the 9-month regimen were eligible. INTERVENTIONS: Patients took self-administered isoniazid and rifampin daily for 24 weeks (6-month regimen) or 36 weeks (9-month regimen). In addition, patients assigned to the 6-month regimen took self-administered pyrazinamide daily during the first 8 weeks. RESULTS: Patients on the 6-month regimen converted more rapidly than patients on the 9-month regimen (94.6% compared with 89.9% after 16 weeks of therapy, with a difference of 4.7% [95% CI, 0.7% to 8.7%]); had similar rates of adverse drug reactions (7.7% compared with 6.4%, with a difference of 1.3% [95% CI, 0.0% to 4.6%]); had lower noncompliance rates (16.8% compared with 29.2%, with a difference of 12.4% [95% CI, 6.8% to 18.0%]); and had similar relapse rates 96 weeks after completing therapy (3.5% compared with 2.8%, with a difference of 0.7% [95% CI, 0.0% to 3.9%]). A significantly greater proportion of patients assigned to the 6-month regimen successfully completed therapy (61.4% compared with 50.6%; chi 2 = 11.976). CONCLUSIONS: Our results suggest that this 6-month regimen is similar in effectiveness, toxicity, and acceptability to the 9-month regimen for treating pulmonary tuberculosis.
- 4Shah, N. S.; Wright, A.; Bai, G. H.; Barrera, L.; Boulahbal, F.; Martin-Casabona, N.; Drobniewski, F.; Gilpin, C.; Havelkova, M.; Lepe, R.; Lumb, R.; Metchock, B.; Portaels, F.; Rodrigues, M. F.; Rusch-Gerdes, S.; Van Deun, A.; Vincent, V.; Laserson, K.; Wells, C.; Cegielski, J. P. Worldwide emergence of extensively drug-resistant tuberculosis. Emerg. Infect. Dis. 2007, 13, 380– 7, DOI: 10.3201/eid1303.061400Google Scholar4https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXjvVOksbs%253D&md5=dd37e95733ca0384716c0874924d151dWorldwide emergence of extensively drug-resistant tuberculosisShah, N. Sarita; Wright, Abigail; Bai, Gill-Han; Barrera, Lucia; Boulahbal, Fadila; Martin-Casabona, Nuria; Drobniewski, Francis; Gilpin, Chris; Havelkova, Marta; Lepe, Rosario; Lumb, Richard; Metchock, Beverly; Portaels, Francoise; Rodrigues, Maria Filomena; Rusch-Gerdes, Sabine; Van Deun, Armand; Vincent, Veronique; Laserson, Kayla; Wells, Charles; Cegielski, J. PeterEmerging Infectious Diseases (2007), 13 (3), 380-387CODEN: EIDIFA; ISSN:1080-6040. (National Center for Infectious Diseases, Centers for Disease Control and Prevention)Mycobacterium tuberculosis strains that are resistant to an increasing no. of second-line drugs used to treat multidrug-resistant tuberculosis (MDR TB) are becoming a threat to public health worldwide. We surveyed the Network of Supranational Ref. Labs. for M. tuberculosis isolates that were resistant to second-line anti-TB drugs during 2000-2004. We defined extensively drug-resistant TB (XDR TB) as MDR TB with further resistance to ≥3 of the 6 classes of second-line drugs. Of 23 eligible labs., 14 (61%) contributed data on 17,690 isolates, which reflected drug susceptibility results from 48 countries. Of 3,520 (19.9%) MDR TB isolates, 347 (9.9%) met criteria for XDR TB. Further investigation of population-based trends and expanded efforts to prevent drug resistance and effectively treat patients with MDR TB are crucial for protection of public health and control of TB.
- 5Sharma, S. K.; Mohan, A. Multidrug-Resistant Tuberculosis: A Menace That Threatens To Destabilize Tuberculosis Control. Chest 2006, 130, 261– 272, DOI: 10.1016/S0012-3692(15)50981-1Google Scholar5https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28Xot1KjtLc%253D&md5=f61cec249a7ad13d4ff64b444510a740Multidrug-resistant tuberculosis: a menace that threatens to destabilize tuberculosis controlSharma, Surendra K.; Mohan, AlladiChest (2006), 130 (1), 261-272CODEN: CHETBF; ISSN:0012-3692. (American College of Chest Physicians)A review. Multidrug-resistant tuberculosis (MDR-TB), caused by Mycobacterium tuberculosis that is resistant to both isoniazid and rifampicin with or without resistance to other drugs, is a phenomenon that is threatening to destabilize global tuberculosis (TB) control. MDR-TB is a worldwide problem, being present virtually in all countries that were surveyed. According to current World Health Organization and the International Union Against Tuberculosis and Lung Disease ests., the median prevalence of MDR-TB has been 1.1% in newly diagnosed patients. The proportion, however, is considerably higher (median prevalence, 7%) in patients who have previously received anti-TB treatment. While host genetic factors may contribute to the development of MDR-TB, incomplete and inadequate treatment is the most important factor leading to its development, suggesting that it is often a man made tragedy. Efficiently run TB control programs based on a policy of directly obsd. treatment, short course (DOTS), are essential for preventing the emergence of MDR-TB. The management of MDR-TB is a challenge that should be undertaken by experienced clinicians at centers equipped with reliable lab. services for mycobacterial cultures and in vitro sensitivity testing as it the requires prolonged use of costly second-line drugs with a significant potential for toxicity. The judicious use of drugs; supervised standardized treatment; focused clin., radiol., and bacteriol. follow-up; and surgery at the appropriate juncture are key factors in the successful management of these patients. With newer effective anti-TB drugs still a distant dream, innovative approaches such as DOTS-Plus are showing promise for the management of patients with MDR-TB under program conditions and appear to be a hope for future.
- 6Zumla, A.; Nahid, P.; Cole, S. T. Advances in the development of new tuberculosis drugs and treatment regimens. Nat. Rev. Drug Discovery 2013, 12, 388– 404, DOI: 10.1038/nrd4001Google Scholar6https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXms1Kmsrk%253D&md5=fbf12f867a19ffff4d7fb74a98699682Advances in the development of new tuberculosis drugs and treatment regimensZumla, Alimuddin; Nahid, Payam; Cole, Stewart T.Nature Reviews Drug Discovery (2013), 12 (5), 388-404CODEN: NRDDAG; ISSN:1474-1776. (Nature Publishing Group)A review. Despite the introduction 40 years ago of the inexpensive and effective four-drug (isoniazid, rifampicin, pyrazinamide and ethambutol) treatment regimen, tuberculosis (TB) continues to cause considerable morbidity and mortality worldwide. For the first time since the 1960s, new and novel drugs and regimens for all forms of TB are emerging. Such regimens are likely to utilize both repurposed drugs and new chem. entities, and several of these regimens are now progressing through clin. trials. This article covers current concepts and recent advances in TB drug discovery and development, including an update of ongoing TB treatment trials, newer clin. trial designs, TB biomarkers and adjunct host-directed therapies.
- 7Ma, Z.; Lienhardt, C.; McIlleron, H.; Nunn, A. J.; Wang, X. Global tuberculosis drug development pipeline: the need and the reality. Lancet 2010, 375, 2100– 2109, DOI: 10.1016/S0140-6736(10)60359-9Google Scholar7https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3cvpsl2msA%253D%253D&md5=893d7a51ed206dca3cfb1f7a2c3de77fGlobal tuberculosis drug development pipeline: the need and the realityMa Zhenkun; Lienhardt Christian; McIlleron Helen; Nunn Andrew J; Wang XiexiuLancet (London, England) (2010), 375 (9731), 2100-9 ISSN:.Drugs for tuberculosis are inadequate to address the many inherent and emerging challenges of treatment. In the past decade, ten compounds have progressed into the clinical development pipeline, including six new compounds specifically developed for tuberculosis. Despite this progress, the global drug pipeline for tuberculosis is still insufficient to address the unmet needs of treatment. Additional and sustainable efforts, and funding are needed to further improve the pipeline. The key challenges in the development of new treatments are the needs for novel drug combinations, new trial designs, studies in paediatric populations, increased clinical trial capacity, clear regulatory guidelines, and biomarkers for prediction of long-term outcome. Despite substantial progress in efforts to control tuberculosis, the global burden of this disease remains high. To eliminate tuberculosis as a public health concern by 2050, all responsible parties need to work together to strengthen the global antituberculosis drug pipeline and support the development of new antituberculosis drug regimens.
- 8Sharma, A.; De Rosa, M.; Singla, N.; Singh, G.; Barnwal, R. P.; Pandey, A. Tuberculosis: An Overview of the Immunogenic Response, Disease Progression, and Medicinal Chemistry Efforts in the Last Decade toward the Development of Potential Drugs for Extensively Drug-Resistant Tuberculosis Strains. J. Med. Chem. 2021, 64, 4359– 4395, DOI: 10.1021/acs.jmedchem.0c01833Google Scholar8https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3MXotlCrurw%253D&md5=6f8bf51a6e6ab45614f311d81379a616Tuberculosis: An Overview of the Immunogenic Response, Disease Progression, and Medicinal Chemistry Efforts in the Last Decade toward the Development of Potential Drugs for Extensively Drug-Resistant Tuberculosis StrainsSharma, Akanksha; De Rosa, Maria; Singla, Neha; Singh, Gurpal; Barnwal, Ravi P.; Pandey, AnkurJournal of Medicinal Chemistry (2021), 64 (8), 4359-4395CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)A review. Tuberculosis (TB) is a slow growing, potentially debilitating disease that has plagued humanity for centuries and has claimed numerous lives across the globe. Concerted efforts by researchers have culminated in the development of various strategies to combat this malady. This review aims to raise awareness of the rapidly increasing incidences of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis, highlighting the significant modifications that were introduced in the TB treatment regimen over the past decade. A description of the role of pathogen-host immune mechanisms together with strategies for prevention of the disease is discussed. The struggle to develop novel drug therapies has continued in an effort to reduce the treatment duration, improve patient compliance and outcomes, and circumvent TB resistance mechanisms. Herein, we give an overview of the extensive medicinal chem. efforts made during the past decade toward the discovery of new chemotypes, which are potentially active against TB-resistant strains.
- 9Raymer, B.; Bhattacharya, S. K. Lead-like Drugs: A Perspective. J. Med. Chem. 2018, 61, 10375– 10384, DOI: 10.1021/acs.jmedchem.8b00407Google Scholar9https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhtl2is73O&md5=e9fc17d69c7d671459fdaf77d6721829Lead-like Drugs: A PerspectiveRaymer, Brian; Bhattacharya, Samit K.Journal of Medicinal Chemistry (2018), 61 (23), 10375-10384CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)Lead-like drugs, or drugs below mol. wt. 300, are an important and sometimes overlooked component of the current pharmacopeia and contemporary medicinal chem. practice. To examine the recent state-of-the-art in lead-like drug discovery, we surveyed recent drug approvals from 2011 to 2017 and top 200 prescribed medications, as well as provide case studies on recently approved lead-like drugs. Many of these recent drugs are close analogs of previously known drugs or natural substrates, with a key focus of their medicinal chem. optimization being the choice of a low mol. wt. starting point and maintaining low mol. wt. during the optimization. However, the identification of low mol. wt. starting points may be limited by the availability of suitable low mol. wt. screening sets. To increase the discovery rate of lead-like drugs, we suggest an increased focus on inclusion and prosecution of lead-like starting points in screening libraries.
- 10Lovering, F. Escape from Flatland 2: complexity and promiscuity. MedChemComm 2013, 4, 515– 519, DOI: 10.1039/c2md20347bGoogle Scholar10https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXjtFShtL0%253D&md5=c03d3b99da22a684cbbbb00fb1633342Escape from Flatland 2: complexity and promiscuityLovering, FrankMedChemComm (2013), 4 (3), 515-519CODEN: MCCEAY; ISSN:2040-2503. (Royal Society of Chemistry)Toxicity plays a major role in attrition in the clinic and promiscuity has been linked to toxicity. A no. of mol. descriptors have been identified that contribute to promiscuity including ionization and logP. In this study we report on the relationship between complexity, as measured by two descriptors [fraction sp3 (Fsp3) where Fsp3 = (no. of sp3 hybridized carbons/total carbon count) and chiral carbon count], and promiscuity as well as Cyp450 inhibition. We find that increasing complexity reduces promiscuity and Cyp450 inhibition. As an understanding of key property descriptors has helped the pharmaceutical industry to address some of the deficiencies of compds. as pertains to bioavailability, awareness of the descriptors that impact promiscuity should allow us to better address toxicity in the clinic.
- 11Lovering, F.; Bikker, J.; Humblet, C. Escape from Flatland: Increasing Saturation as an Approach to Improving Clinical Success. J. Med. Chem. 2009, 52, 6752– 6756, DOI: 10.1021/jm901241eGoogle Scholar11https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXht1KjtLvN&md5=4ca92c30c17c53d77ad376719bad951eEscape from Flatland: Increasing Saturation as an Approach to Improving Clinical SuccessLovering, Frank; Bikker, Jack; Humblet, ChristineJournal of Medicinal Chemistry (2009), 52 (21), 6752-6756CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)The medicinal chem. community has become increasingly aware of the value of tracking calcd. phys. properties such as mol. wt., topol. polar surface area, rotatable bonds, and hydrogen bond donors and acceptors. The authors hypothesized that the shift to high-throughput synthetic practices over the past decade may be another factor that may predispose mols. to fail by steering discovery efforts toward achiral, arom. compds. The authors have proposed two simple and interpretable measures of the complexity of mols. prepd. as potential drug candidates. The first is carbon bond satn. as defined by fraction Sp3 (Fsp3) where Fsp3 = (no. of Sp3 hybridized carbons/total carbon count). The second is simply whether a chiral carbon exists in the mol. The authors demonstrate that both complexity (as measured by Fsp3) and the presence of chiral centers correlate with success as compds. transition from discovery, through clin. testing, to drugs. To explain these observations, the authors further demonstrate that satn. correlates with soly., an exptl. phys. property important to success in the drug discovery setting.
- 12Yang, Y.; Chen, H.; Nilsson, I.; Muresan, S.; Engkvist, O. Investigation of the Relationship between Topology and Selectivity for Druglike Molecules. J. Med. Chem. 2010, 53, 7709– 7714, DOI: 10.1021/jm1008456Google Scholar12https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXht1Oqtb3L&md5=46f7582bfa567734f0391512e2580717Investigation of the Relationship between Topology and Selectivity for Drug-like MoleculesYang, Yidong; Chen, Hongming; Nilsson, Ingemar; Muresan, Sorel; Engkvist, OlaJournal of Medicinal Chemistry (2010), 53 (21), 7709-7714CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)There is a strong interest in drug discovery and development to advance the understanding of pharmacol. promiscuity. Improved understanding of how a mol. structure is related to promiscuity could help to reduce the attrition of compds. in the drug discovery process. For this purpose, a descriptor is introduced that describes the structural complexity of a compd. based on the size of its mol. framework (MF) in relation to its overall size. It is defined as the fraction of the size of the mol. framework vs. the size of the whole mol. (fMF). It is demonstrated that promiscuity correlates with fMF for large fMF values. The obsd. correlation is not due to lipophilicity. To provide further explanation of this observation, it was found that the no. of terminal ring systems in a compd. is correlated with promiscuity. The anal. presented here might help medicinal chemists to improve the selectivity for compds. in drug discovery projects.
- 13Baell, J. B.; Holloway, G. A. New Substructure Filters for Removal of Pan Assay Interference Compounds (PAINS) from Screening Libraries and for Their Exclusion in Bioassays. J. Med. Chem. 2010, 53, 2719– 2740, DOI: 10.1021/jm901137jGoogle Scholar13https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXhsF2qsLw%253D&md5=fbf397aa4910753c550425708c866fd2New Substructure Filters for Removal of Pan Assay Interference Compounds (PAINS) from Screening Libraries and for Their Exclusion in BioassaysBaell, Jonathan B.; Holloway, Georgina A.Journal of Medicinal Chemistry (2010), 53 (7), 2719-2740CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)This report describes a no. of substructural features which can help to identify compds. that appear as frequent hitters (promiscuous compds.) in many biochem. high throughput screens. The compds. identified by such substructural features are not recognized by filters commonly used to identify reactive compds. Even though these substructural features were identified using only one assay detection technol., such compds. have been reported to be active from many different assays. In fact, these compds. are increasingly prevalent in the literature as potential starting points for further exploration, whereas they may not be.
- 14Baell, J. B.; Nissink, J. W. M. Seven Year Itch: Pan-Assay Interference Compounds (PAINS) in 2017─Utility and Limitations. ACS Chem. Biol. 2018, 13, 36– 44, DOI: 10.1021/acschembio.7b00903Google Scholar14https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhvFSlu7bK&md5=051115704137d6a91b8271702c619682Seven Year Itch: Pan-Assay Interference Compounds (PAINS) in 2017-Utility and LimitationsBaell, Jonathan B.; Nissink, J. Willem M.ACS Chemical Biology (2018), 13 (1), 36-44CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)A review on all necessary considerations for the appropriate use of PAINs filters formulated to process hundreds and thousands of compds. in seconds, and identify PAINS in order to exclude them from further anal.
- 15Colomer, I.; Empson, C. J.; Craven, P.; Owen, Z.; Doveston, R. G.; Churcher, I.; Marsden, S. P.; Nelson, A. A divergent synthetic approach to diverse molecular scaffolds: assessment of lead-likeness using LLAMA, an open-access computational tool. Chem. Commun. 2016, 52, 7209– 7212, DOI: 10.1039/C6CC03244CGoogle Scholar15https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XnsVWqtrs%253D&md5=6f16f9da33bfc182d98c56065e8d3951A divergent synthetic approach to diverse molecular scaffolds: assessment of lead-likeness using LLAMA, an open-access computational toolColomer, Ignacio; Empson, Christopher J.; Craven, Philip; Owen, Zachary; Doveston, Richard G.; Churcher, Ian; Marsden, Stephen P.; Nelson, AdamChemical Communications (Cambridge, United Kingdom) (2016), 52 (45), 7209-7212CODEN: CHCOFS; ISSN:1359-7345. (Royal Society of Chemistry)Complementary cyclization reactions of hex-2-ene-1,6-diamine derivs. were exploited in the synthesis of alternative pyrrolidine, oxazolidinone, pyrimidinone and oxazinone mol. scaffolds. The value of the synthetic approach was analyzed using LLAMA, an open-access computational tool for assessing the lead-likeness and novelty of mol. scaffolds.
- 16
Synthetic chemistry methodology projects within the School of Chemistry at the University of Birmingham initially provided ∼ 200 compounds suitable for biological screening.
There is no corresponding record for this reference. - 17Feula, A.; Dhillon, S. S.; Byravan, R.; Sangha, M.; Ebanks, R.; Hama Salih, M. A.; Spencer, N.; Male, L.; Magyary, I.; Deng, W.-P.; Müller, F.; Fossey, J. S. Synthesis of azetidines and pyrrolidines via iodocyclisation of homoallyl amines and exploration of activity in a zebrafish embryo assay. Org. Biomol. Chem. 2013, 11, 5083– 5093, DOI: 10.1039/c3ob41007bGoogle Scholar17https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhtFaltb%252FE&md5=3538313a5c37148cc75e5039a1c34540Synthesis of azetidines and pyrrolidines via iodocyclisation of homoallyl amines and exploration of activity in a zebrafish embryo assayFeula, Antonio; Dhillon, Sundeep S.; Byravan, Rama; Sangha, Mandeep; Ebanks, Ronald; Hama Salih, Mariwan A.; Spencer, Neil; Male, Louise; Magyary, Istvan; Deng, Wei-Ping; Mueller, Ferenc; Fossey, John S.Organic & Biomolecular Chemistry (2013), 11 (31), 5083-5093CODEN: OBCRAK; ISSN:1477-0520. (Royal Society of Chemistry)Room temp. iodocyclization of homoallyl amines stereoselectively delivers functionalized 2-(iodomethyl)azetidine derivs. in high yield. Increasing the reaction temp. from 20° to 50° switches the reaction outcome to realize the stereoselective formation of functionalized 3-iodopyrrolidine derivs. It was shown that these pyrrolidines are formed via thermal isomerization of the aforementioned azetidines. Primary and secondary amines could be reacted with iodomethyl azetidine derivs. to deliver stable aminomethyl azetidine derivs. With subtle changes to the reaction sequences homoallyl amines could be stereoselectively converted to either cis- or trans-substituted 3-amino pyrrolidine derivs. at will. The stereochem. divergent synthesis of cis and trans substituted pyrrolidines supports an ion pair, aziridinium, isomerization pathway for azetidine to pyrrolidine isomerization. Six azetidine derivs. were probed in a zebrafish embryo developmental assay to detect potential biol. effects through the anal. of morphol. and motility behavior phenotypes. The range of effects across the probed mols. demonstrates the suitability of this assay for screening azetidine derivs. One of the probed mols., rac-(((cis)-1-benzyl-4-phenylazetidin-2-yl)methyl)piperidine, exhibited particularly interesting effects in the developmental assay presenting with hypopigmentation and reduced circulation amongst others. This shows that the zebrafish embryo provides a fast, sensitive and effective way to screen new compds. and in the future in combination with existing in vivo and in vitro assays it will become an integral part in drug discovery and development.
- 18Feula, A.; Male, L.; Fossey, J. S. Diastereoselective preparation of azetidines and pyrrolidines. Org. Lett. 2010, 12, 5044– 5047, DOI: 10.1021/ol102215eGoogle Scholar18https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXht1ajtL7M&md5=fc6db1a4635d20b732e90500bf24f52eDiastereoselective Preparation of Azetidines and PyrrolidinesFeula, Antonio; Male, Louise; Fossey, John S.Organic Letters (2010), 12 (21), 5044-5047CODEN: ORLEF7; ISSN:1523-7060. (American Chemical Society)Iodine-mediated cyclization of homoallyl amines at room temp. delivered cis-2,4-azetidine through a 4-exo trig cyclization. Isomerization of iodo-azetidines to cis-pyrrolidines could be achieved by heating, with complete stereocontrol. The relative stereochem. of the iodo-azetidines and pyrrolidines was confirmed by NMR spectroscopy and x-ray crystallog. Further functionalization was achieved through nucleophilic displacement of iodine to deliver substituted azetidines and pyrrolidines. 1,2,3-Triazole-appended azetidines and pyrrolidines were also prepd.
- 19Yoshizawa, A.; Feula, A.; Leach, A. G.; Male, L.; Fossey, J. S. Palladium and Platinum 2,4-cis-amino Azetidine and Related Complexes. Front. Chem. 2018, 6, 211, DOI: 10.3389/fchem.2018.00211Google Scholar19https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXisFaisbvN&md5=239ee55e21a937601449564813779a14Palladium and platinum 2,4-cis-amino azetidine and related complexesYoshizawa, Akina; Feula, Antonio; Leach, Andrew G.; Male, Louise; Fossey, John S.Frontiers in Chemistry (Lausanne, Switzerland) (2018), 6 (), 211/1-211/9CODEN: FCLSAA; ISSN:2296-2646. (Frontiers Media S.A.)Seven N,N'-palladium(II) chloride complexes, one N,N'-palladium(II) acetate complex of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. Two platinum(II) chloride N,N'-complexes of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. Computational anal. and detn. of the %Vbur was examd. conducted. A CNN' metallocyclic complex was prepd. by oxidative addn. of palladium(0) to an ortho bromo 2,4-cis-disubstituted azetidine and its crystal structure displays a slightly pyramidalized metal-ligand orientation. Seven N,N'-palladium(II) chloride complexes, one N,N'-palladium(II) acetate complex of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. The racemic ligands adopted a single diastereoisomer form upon coordination to palladium the same chirality at nitrogen. In the palladium(II) acetate complex a coordinated water mol. and H-bonding acetates formed the identified complex. Two platinum(II) chloride N,N'-complexes of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD, and two diastereoisomers were generated upon amine coordination to platinum (under different prepn. conditions). Computational anal. revealed which diastereoisomer was more stable and provided a rationale for why this is the case, and the %Vbur described by the diastereomeric coordination geometry was examd. A CNN' metallocyclic complex was prepd. by oxidative addn. of palladium(0) to an ortho bromo 2,4-cis-disubstituted azetidine and its crystal structure displays a slightly pyramidalised metal-ligand orientation. Ligand salts were not suitable for the synthesis of azetidine complexes, instead leading to N,N' complexation of stereospecifically ring-opened congeners. A minor, pyrrolidine, impurity in an azetidine ligand sample led, initially, to the formation of a highly cryst. complex, identified by single crystal XRD, as well as from the same sample the expected azetidine complex. The isomeric azetidine and pyrrolidine complexes, characterized by single crystal XRD, were studied using the SambVca 2.0 online tool and their steric parameters contrasted revealing the potential for both azetidine and pyrrolidine ligands in future catalytic applications.
- 20Yoshizawa, A.; Feula, A.; Male, L.; Leach, A. G.; Fossey, J. S. Rigid and concave, 2,4-cis-substituted azetidine derivatives: A platform for asymmetric catalysis. Sci. Rep. 2018, 8, 6541, DOI: 10.1038/s41598-018-24784-3Google Scholar20https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1MjmslektA%253D%253D&md5=ae3566571728d61bbcc7fa8d4c7fd426Rigid and concave, 2,4-cis-substituted azetidine derivatives: A platform for asymmetric catalysisYoshizawa Akina; Feula Antonio; Fossey John S; Male Louise; Leach Andrew GScientific reports (2018), 8 (1), 6541 ISSN:.A series of single enantiomer, 2,4-cis-disubstituted amino azetidines were synthesised and used as ligands for copper-catalysed Henry reactions of aldehydes with nitromethane. Optimisation of ligand substituents and the reaction conditions was conducted. The enantiomeric excess of the formed products was highest when alkyl aldehydes were employed in the reaction (>99% e.e.). The absolute stereochemistry of one representative azetidine derivative salt was determined by analysis of the Flack parameter of an XRD single crystal structure. The origin of selectivity in catalysis was investigated computationally, revealing the importance of the amino-substituent in determining the stereochemical outcome. A racemic platinum complex of a cis-disubstituted azetidine is examined by XRD single crystal structure analysis with reference to its steric parameters, and analogies to the computationally determined copper complex catalyst are drawn. A preliminary example of the use of a cis-disubstituted azetidine scaffold in thiourea H-bonding catalyst is noted in the supporting information.
- 21www.birmingham.ac.uk/facilities/bddf. Birmingham Drug Discovery Facility. www.birmingham.ac.uk/facilities/bddf.Google ScholarThere is no corresponding record for this reference.
- 22
The synthesis and preliminary screening data of ∼ 100 azetidine derivatives that were less active, and not investigated further, will be reported elsewhere.
There is no corresponding record for this reference. - 23Palomino, J.-C.; Martin, A.; Camacho, M.; Guerra, H.; Swings, J.; Portaels, F. Resazurin Microtiter Assay Plate: Simple and Inexpensive Method for Detection of Drug Resistance in Mycobacterium tuberculosis. Antimicrob. Agents Chemother. 2002, 46, 2720, DOI: 10.1128/AAC.46.8.2720-2722.2002Google Scholar23https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XlsFGiu7w%253D&md5=555848714bd1528e6a8eb913b5971c9aResazurin microtiter assay plate: simple and inexpensive method for detection of drug resistance in Mycobacterium tuberculosisPalomino, Juan-Carlos; Martin, Anandi; Camacho, Mirtha; Guerra, Humberto; Swings, Jean; Portaels, FrancoiseAntimicrobial Agents and Chemotherapy (2002), 46 (8), 2720-2722CODEN: AMACCQ; ISSN:0066-4804. (American Society for Microbiology)A method for detecting multidrug-resistant Mycobacterium tuberculosis by using a redn. of resazurin is described. Eighty clin. isolates were evaluated against isoniazid and rifampin; results at 7 days were compared with those of the proportion method. Specificity and sensitivity were excellent. The method is simple, inexpensive, and rapid and might be used with other antituberculosis drugs.
- 24Yang, F.; Njire, M. M.; Liu, J.; Wu, T.; Wang, B.; Liu, T.; Cao, Y.; Liu, Z.; Wan, J.; Tu, Z.; Tan, Y.; Tan, S.; Zhang, T. Engineering more stable, selectable marker-free autoluminescent mycobacteria by one step. PLoS One 2015, 10, e0119341 DOI: 10.1371/journal.pone.0119341Google Scholar24https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhs1WhtbjL&md5=c63cbbd4bac4eb72c3fcf387faefd5afEngineering more stable, selectable marker- free autoluminescent mycobacteria by one stepYang, Feng; Njire, Moses M.; Liu, Jia; Wu, Tian; Wang, Bangxing; Liu, Tianzhou; Cao, Yuanyuan; Liu, Zhiyong; Wan, Junting; Tu, Zhengchao; Tan, Yaoju; Tan, Shouyong; Zhang, TianyuPLoS One (2015), 10 (3), e0119341/1-e0119341/13CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)In our previous study, we demonstrated that the use of the autoluminescent Mycobacterium tuberculosis as a reporter strain had the potential to drastically reduce the time, effort, animals and costs consumed in evaluation of the activities of drugs and vaccines in live mice. However, the strains were relatively unstable and lost reporter with time without selection. The kanamycin selection marker used wasn't the best choice as it provides resistance to amino glycosides which are an important class of second line drugs used in tuberculosis treatment. In addn., the marker could limit utility of the strains for screening of new potential drugs or evaluating drug combinations for tuberculosis treatment. Limited selection marker genes for mycobacterial genetic manipulation is a major drawback for such a marker- contg. strain in many research fields. Therefore, selectable marker-free, more stable autoluminescent mycobacteria are highly needed. After trying several strategies, we created such mycobacterial strains successfully by using an integrative vector and removing both the resistance maker and integrase genes by Xer site-specific recombination in one step. The corresponding plasmid vectors developed in this study could be very convenient in constructing other selectable marker-free, more stable reporter mycobacteria with diverse applications.
- 25Qin, L.; Wang, J.; Lu, J.; Yang, H.; Zheng, R.; Liu, Z.; Huang, X.; Feng, Y.; Hu, Z.; Ge, B. A deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosis. BMC Biol. 2019, 17, 7, DOI: 10.1186/s12915-019-0628-6Google Scholar25https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3cjltl2rsA%253D%253D&md5=ff8dac93d32fd72ff22f55c65a72d5bbA deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosisQin Lianhua; Wang Jie; Lu Junmei; Yang Hua; Zheng Ruijuan; Liu Zhonghua; Huang Xiaochen; Feng Yonghong; Hu Zhongyi; Ge BaoxueBMC biology (2019), 17 (1), 7 ISSN:.BACKGROUND: The emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb), especially those that are multidrug resistant poses a serious threat to global tuberculosis control. However, the mechanism underlying the occurrence of drug resistance against more than one drug is poorly understood. Given that the Beijing/W strains are associated with outbreaks and multidrug resistance, they may harbor a genetic advantage and provide useful insight into the disease. One marker found in all Beijing/W Mtb strains is a deletion of RD105 region that results in a gene fusion, Rv0071/74, with a variable number (3-9 m) of VDP (V: Val, D: Asp; P: Pro) repeats (coded by gtggacccg repeat sequences) at the N-terminal. Here, we report that this variable number of VDP repeats in Rv0071/74 regulates the development of multidrug resistance. RESULTS: We collected and analyzed 1255 Beijing/W clinical strains. The results showed that the number of VDP repeats in Rv0071/74 was related to the development of multidrug resistance, and the deletion of Rv0071/74-9 m from Beijing/W clinical strain restored drug susceptibility. Rv0071/74-9 m also increased resistance to multiple drugs when transferred to different mycobacterial strains. Cell-free assays indicate that the domain carrying 4-9 VDP repeats (4-9 m) showed a variable binding affinity with peptidoglycan and Rv0071/74 cleaves peptidoglycan. Furthermore, Rv0071/74-9 m increased cell wall thickness and reduced the intracellular concentration of antibiotics. CONCLUSIONS: These findings not only identify Rv0071/74 with VDP repeats as a newly identified multidrug resistance gene but also provide a new model for the development of multiple drug resistance.
- 26Hoagland, D. T.; Liu, J.; Lee, R. B.; Lee, R. E. New agents for the treatment of drug-resistant Mycobacterium tuberculosis. Adv. Drug Delivery Rev. 2016, 102, 55– 72, DOI: 10.1016/j.addr.2016.04.026Google Scholar26https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XntlSns7c%253D&md5=7c8ca15b9b493cceaf3d51d93b9afcecNew agents for the treatment of drug-resistant Mycobacterium tuberculosisHoagland, Daniel T.; Liu, Jiuyu; Lee, Robin B.; Lee, Richard E.Advanced Drug Delivery Reviews (2016), 102 (), 55-72CODEN: ADDREP; ISSN:0169-409X. (Elsevier B.V.)Inadequate dosing and incomplete treatment regimens, coupled with the ability of the tuberculosis bacilli to cause latent infections that are tolerant of currently used drugs, have fueled the rise of multidrug-resistant tuberculosis (MDR-TB). Treatment of MDR-TB infections is a major clin. challenge that has few viable or effective solns.; therefore patients face a poor prognosis and years of treatment. This review focuses on emerging drug classes that have the potential for treating MDR-TB and highlights their particular strengths as leads including their mode of action, in vivo efficacy, and key medicinal chem. properties. Examples include the newly approved drugs bedaquiline and delaminid, and other agents in clin. and late preclin. development pipeline for the treatment of MDR-TB. Herein, we discuss the challenges to developing drugs to treat tuberculosis and how the field has adapted to these difficulties, with an emphasis on drug discovery approaches that might produce more effective agents and treatment regimens.
- 27Miles, A. A.; Misra, S. S.; Irwin, J. O. The estimation of the bactericidal power of the blood. Epidemiol. Infect. 1938, 38, 732– 749, DOI: 10.1017/S002217240001158XGoogle ScholarThere is no corresponding record for this reference.
- 28Hendon-Dunn, C. L.; Doris, K. S.; Thomas, S. R.; Allnutt, J. C.; Marriott, A. A. N.; Hatch, K. A.; Watson, R. J.; Bottley, G.; Marsh, P. D.; Taylor, S. C.; Bacon, J. A Flow Cytometry Method for Rapidly Assessing Mycobacterium tuberculosis Responses to Antibiotics with Different Modes of Action. Antimicrob. Agents Chemother. 2016, 60, 3869, DOI: 10.1128/AAC.02712-15Google Scholar28https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFSjtb7P&md5=5670956cd3b30b675b72209e8f5f92d1A flow cytometry method for rapidly assessing Mycobacterium tuberculosis responses to antibiotics with different modes of actionHendon-Dunn, Charlotte Louise; Doris, Kathryn Sarah; Thomas, Stephen Richard; Allnutt, Jonathan Charles; Marriott, Alice Ann Neville; Hatch, Kim Alexandra; Watson, Robert James; Bottley, Graham; Marsh, Philip David; Taylor, Stephen Charles; Bacon, JoannaAntimicrobial Agents and Chemotherapy (2016), 60 (7), 3869-3883CODEN: AMACCQ; ISSN:1098-6596. (American Society for Microbiology)Current methods for assessing the drug susceptibility of Mycobacterium tuberculosis are lengthy and do not capture information about viable organisms that are not immediately culturable under std. lab. conditions as a result of antibiotic exposure. We have developed a rapid dual-fluorescence flow cytometry method using markers for cell viability and death. We show that the fluorescent marker calcein violet with an acetoxy-Me ester group (CV-AM) can differentiate between populations of M. tuberculosis growing at different rates, while Sytox green (SG) can differentiate between live and dead mycobacteria. M. tuberculosis was exposed to isoniazid or rifampin at different concns. over time and either dual stained with CV-AM and SG and analyzed by flow cytometry or plated to det. the viability of the cells. Although similar trends in the loss of viability were obsd. when the results of flow cytometry and the plate counting methods were compared, there was a lack of correlation between these two approaches, as the flow cytometry anal. potentially captured information about cell populations that were unable to grow under std. conditions. The flow cytometry approach had an addnl. advantage in that it could provide insights into the mode of action of the drug: antibiotics targeting the cell wall gave a flow cytometry profile distinct from those inhibiting intracellular processes. This rapid drug susceptibility testing method could identify more effective antimycobacterials, provide information about their potential mode of action, and accelerate their progress to the clinic.
- 29Abrahams, K. A.; Cox, J. A. G.; Spivey, V. L.; Loman, N. J.; Pallen, M. J.; Constantinidou, C.; Fernandez, R.; Alemparte, C.; Remuinan, M. J.; Barros, D.; Ballell, L.; Besra, G. S. Identification of Novel Imidazo[1,2-a]pyridine Inhibitors Targeting M. tuberculosis QcrB. PLoS One 2012, 7, e52951 DOI: 10.1371/journal.pone.0052951Google Scholar29https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptVSgsQ%253D%253D&md5=4c00cd8ffb101ce9d40da5c0c4f51c0eIdentification of novel imidazo[1,2-a]pyridine inhibitors targeting M. tuberculosis QcrBAbrahams, Katherine A.; Cox, Jonathan A. G.; Spivey, Vickey L.; Loman, Nicholas J.; Pallen, Mark J.; Constantinidou, Chrystala; Fernandez, Raquel; Alemparte, Carlos; Remuinan, Modesto J.; Barros, David; Ballell, Lluis; Besra, Gurdyal S.PLoS One (2012), 7 (12), e52951CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Mycobacterium tuberculosis is a major human pathogen and the causative agent for the pulmonary disease, tuberculosis (TB). Current treatment programs to combat TB are under threat due to the emergence of multi-drug and extensively-drug resistant TB. Through the use of high throughput whole cell screening of an extensive compd. library a no. of imidazo[1,2-a]pyridine (IP) compds. were obtained as potent lead mols. active against M. tuberculosis and Mycobacterium bovis BCG. The IP inhibitors (1-4) demonstrated min. inhibitory concns. (MICs) in the range of 0.03 to 5 μM against a panel of M. tuberculosis strains. M. bovis BCG spontaneous resistant mutants were generated against IP 1, 3 and 4 at 5 × MIC and subsequent whole genome sequencing identified a single nucleotide polymorphism 937ACC>937GCC (T313A) in qcrB, which encodes the b subunit of the electron transport ubiquinol cytochrome C reductase. This mutation also conferred cross-resistance against IP 1, 3 and 4 demonstrating a common target. Gene dosage expts. confirmed M. bovis BCG QcrB as the target where over-expression in M. bovis BCG led to an increase in MIC from 0.5 to >8 μM for IP 3. An acute murine model of TB infection established bacteriostatic activity of the IP series, which await further detailed characterization.
ARTN
- 30Abrahams, K. A.; Chung, C.-W.; Ghidelli-Disse, S.; Rullas, J.; Rebollo-López, M. J.; Gurcha, S. S.; Cox, J. A. G.; Mendoza, A.; Jiménez-Navarro, E.; Martínez-Martínez, M. S.; Neu, M.; Shillings, A.; Homes, P.; Argyrou, A.; Casanueva, R.; Loman, N. J.; Moynihan, P. J.; Lelièvre, J.; Selenski, C.; Axtman, M.; Kremer, L.; Bantscheff, M.; Angulo-Barturen, I.; Izquierdo, M. C.; Cammack, N. C.; Drewes, G.; Ballell, L.; Barros, D.; Besra, G. S.; Bates, R. H. Identification of KasA as the cellular target of an anti-tubercular scaffold. Nat. Commun. 2016, 7, 12581, DOI: 10.1038/ncomms12581Google Scholar30https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhsVOru7%252FI&md5=a1206008e181383854e026f36ad27489Identification of KasA as the cellular target of an anti-tubercular scaffoldAbrahams, Katherine A.; Chung, Chun-wa; Ghidelli-Disse, Sonja; Rullas, Joaquin; Rebollo-Lopez, Maria Jose; Gurcha, Sudagar S.; Cox, Jonathan A. G.; Mendoza, Alfonso; Jimenez-Navarro, Elena; Martinez-Martinez, Maria Santos; Neu, Margarete; Shillings, Anthony; Homes, Paul; Argyrou, Argyrides; Casanueva, Ruth; Loman, Nicholas J.; Moynihan, Patrick J.; Lelievre, Joel; Selenski, Carolyn; Axtman, Matthew; Kremer, Laurent; Bantscheff, Marcus; Angulo-Barturen, Inigo; Izquierdo, Monica Cacho; Cammack, Nicholas C.; Drewes, Gerard; Ballell, Lluis; Barros, David; Besra, Gurdyal S.; Bates, Robert H.Nature Communications (2016), 7 (), 12581CODEN: NCAOBW; ISSN:2041-1723. (Nature Publishing Group)Phenotypic screens for bactericidal compds. are starting to yield promising hits against tuberculosis. In this regard, whole-genome sequencing of spontaneous resistant mutants generated against an indazole sulfonamide (GSK3011724A) identifies several specific single-nucleotide polymorphisms in the essential Mycobacterium tuberculosis β-ketoacyl synthase (kas) A gene. Here, this genomic-based target assignment is confirmed by biochem. assays, chem. proteomics and structural resoln. of a KasA-GSK3011724A complex by X-ray crystallog. Finally, M. tuberculosis GSK3011724A-resistant mutants increase the in vitro min. inhibitory concn. and the in vivo 99% ED in mice, establishing in vitro and in vivo target engagement. Surprisingly, the lack of target engagement of the related β-ketoacyl synthases (FabH and KasB) suggests a different mode of inhibition when compared with other Kas inhibitors of fatty acid biosynthesis in bacteria. These results clearly identify KasA as the biol. target of GSK3011724A and validate this enzyme for further drug discovery efforts against tuberculosis.
- 31Andries, K.; Verhasselt, P.; Guillemont, J.; Göhlmann, H. W. H.; Neefs, J.-M.; Winkler, H.; Van Gestel, J.; Timmerman, P.; Zhu, M.; Lee, E.; Williams, P.; de Chaffoy, D.; Huitric, E.; Hoffner, S.; Cambau, E.; Truffot-Pernot, C.; Lounis, N.; Jarlier, V. A Diarylquinoline Drug Active on the ATP Synthase of Mycobacterium tuberculosis. Science 2005, 307, 223, DOI: 10.1126/science.1106753Google Scholar31https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXisVKkug%253D%253D&md5=099b643f173bb77e70cf1aa6e9871ffeA Diarylquinoline Drug Active on the ATP Synthase of Mycobacterium tuberculosisAndries, Koen; Verhasselt, Peter; Guillemont, Jerome; Goehlmann, Hinrich W. H.; Neefs, Jean-Marc; Winkler, Hans; Van Gestel, Jef; Timmerman, Philip; Zhu, Min; Lee, Ennis; Williams, Peter; de Chaffoy, Didier; Huitric, Emma; Hoffner, Sven; Cambau, Emmanuelle; Truffot-Pernot, Chantal; Lounis, Nacer; Jarlier, VincentScience (Washington, DC, United States) (2005), 307 (5707), 223-227CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)The incidence of tuberculosis has been increasing substantially on a worldwide basis over the past decade, but no tuberculosis-specific drugs have been discovered in 40 years. We identified a diarylquinoline, R207910, that potently inhibits both drug-sensitive and drug-resistant Mycobacterium tuberculosis in vitro (min. inhibitory concn. 0.06 μg/mL). In mice, R207910 exceeded the bactericidal activities of isoniazid and rifampin by at least 1 log unit. Substitution of drugs included in the World Health Organization's first-line tuberculosis treatment regimen (rifampin, isoniazid, and pyrazinamide) with R207910 accelerated bactericidal activity, leading to complete culture conversion after 2 mo of treatment in some combinations. A single dose of R207910 inhibited mycobacterial growth for 1 wk. Plasma levels assocd. with efficacy in mice were well tolerated in healthy human volunteers. Mutants selected in vitro suggest that the drug targets the proton pump of ATP (ATP) synthase.
- 32Batt, S. M.; Cacho Izquierdo, M.; Castro Pichel, J.; Stubbs, C. J.; Vela-Glez Del Peral, L.; Pérez-Herrán, E.; Dhar, N.; Mouzon, B.; Rees, M.; Hutchinson, J. P.; Young, R. J.; McKinney, J. D.; Barros Aguirre, D.; Ballell, L.; Besra, G. S.; Argyrou, A. Whole Cell Target Engagement Identifies Novel Inhibitors of Mycobacterium tuberculosis Decaprenylphosphoryl-β-d-ribose Oxidase. ACS Infec. Dis. 2015, 1, 615– 626, DOI: 10.1021/acsinfecdis.5b00065Google Scholar32https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtlemtbnE&md5=1e6d6ef29fb37d3e8535328368af2472Whole Cell Target Engagement Identifies Novel Inhibitors of Mycobacterium tuberculosis Decaprenylphosphoryl-β-D-ribose OxidaseBatt, Sarah M.; Cacho Izquierdo, Monica; Castro Pichel, Julia; Stubbs, Christopher J.; Vela-Glez Del Peral, Laura; Perez-Herran, Esther; Dhar, Neeraj; Mouzon, Bernadette; Rees, Mike; Hutchinson, Jonathan P.; Young, Robert J.; McKinney, John D.; Barros Aguirre, David; Ballell, Lluis; Besra, Gurdyal S.; Argyrou, ArgyridesACS Infectious Diseases (2015), 1 (12), 615-626CODEN: AIDCBC; ISSN:2373-8227. (American Chemical Society)We have targeted the Mycobacterium tuberculosis decaprenylphosphoryl-β-D-ribose oxidase (Mt-DprE1) for potential chemotherapeutic intervention of tuberculosis. A multicopy suppression strategy that overexpressed Mt-DprE1 in M. bovis BCG was used to profile the publically available GlaxoSmithKline antimycobacterial compd. set, and one compd. (GSK710) was identified that showed an 8-fold higher min. inhibitory concn. relative to the control strain. Analogs of GSK710 show a clear relationship between whole cell potency and in vitro activity using an enzymic assay employing recombinant Mt-DprE1, with binding affinity measured by fluorescence quenching of the flavin cofactor of the enzyme. M. bovis BCG spontaneous resistant mutants to GSK710 and a closely related analog were isolated and sequencing of ten such mutants revealed a single point mutation at two sites, E221Q or G248S within DprE1, providing further evidence that DprE1 is the main target of these compds. Finally, time-lapse microscopy expts. showed that exposure of M. tuberculosis to a compd. of this series arrests bacterial growth rapidly followed by a slower cytolysis phase.
- 33Manganelli, R.; Voskuil, M. I.; Schoolnik, G. K.; Smith, I. The Mycobacterium tuberculosis ECF sigma factor sigmaE: role in global gene expression and survival in macrophages. Mol. Microbiol. 2001, 41, 423– 37, DOI: 10.1046/j.1365-2958.2001.02525.xGoogle Scholar33https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3MXlvVClur0%253D&md5=ec7927a14ea6841428d0a7e70df82551The Mycobacterium tuberculosis ECF sigma factor σE: role in global gene expression and survival in macrophagesManganelli, Riccardo; Voskuil, Martin I.; Schoolnik, Gary K.; Smith, IssarMolecular Microbiology (2001), 41 (2), 423-437CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Science Ltd.)In previously published work, we identified three Mycobacterium tuberculosis sigma (σ) factor genes responding to heat shock (sigB, sigE and sigH). Two of them (sigB and sigE) also responded to SDS exposure. As these responses to stress suggested that the σ factors encoded by these genes could be involved in pathogenicity, we are studying their role in physiol. and virulence. In this work, we characterize a sigE mutant of M. tuberculosis H37Rv. The sigE mutant strain was more sensitive than the wild-type strain to heat shock, SDS and various oxidative stresses. It was also defective in the ability to grow inside both human and murine unactivated macrophages and was more sensitive than the wild-type strain to the killing activity of activated murine macrophages. Using microarray technol. and quant. reverse transcription-polymerase chain reaction (RT-PCR), we started to define the σE regulon of M. tuberculosis and its involvement in the global regulation of the stress induced by SDS. We showed the requirement for a functional sigE gene for full expression of sigB and for its induction after SDS exposure but not after heat shock. We also identified several genes that are no longer induced when σE is absent. These genes encode proteins belonging to different classes including transcriptional regulators, enzymes involved in fatty acid degrdn. and classical heat shock proteins.
- 34Balazsi, G.; Heath, A. P.; Shi, L.; Gennaro, M. L. The temporal response of the Mycobacterium tuberculosis gene regulatory network during growth arrest. Mol. Syst. Biol. 2008, 4, 225, DOI: 10.1038/msb.2008.63Google Scholar34https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD1cjhvV2gsg%253D%253D&md5=27c4701b97cae4a1f215832cdaf75481The temporal response of the Mycobacterium tuberculosis gene regulatory network during growth arrestBalazsi Gabor; Heath Allison P; Shi Lanbo; Gennaro Maria LMolecular systems biology (2008), 4 (), 225 ISSN:.The virulence of Mycobacterium tuberculosis depends on the ability of the bacilli to switch between replicative (growth) and non-replicative (dormancy) states in response to host immunity. However, the gene regulatory events associated with transition to dormancy are largely unknown. To address this question, we have assembled the largest M. tuberculosis transcriptional-regulatory network to date, and characterized the temporal response of this network during adaptation to stationary phase and hypoxia, using published microarray data. Distinct sets of transcriptional subnetworks (origons) were responsive at various stages of adaptation, showing a gradual progression of network response under both conditions. Most of the responsive origons were in common between the two conditions and may help define a general transcriptional signature of M. tuberculosis growth arrest. These results open the door for a systems-level understanding of transition to non-replicative persistence, a phenotypic state that prevents sterilization of infection by the host immune response and promotes the establishment of latent M. tuberculosis infection, a condition found in two billion people worldwide.
- 35Karp, P. D.; Billington, R.; Caspi, R.; Fulcher, C. A.; Latendresse, M.; Kothari, A.; Keseler, I. M.; Krummenacker, M.; Midford, P. E.; Ong, Q.; Ong, W. K.; Paley, S. M.; Subhraveti, P. The BioCyc collection of microbial genomes and metabolic pathways. Brief. Bioinform. 2019, 20, 1085– 1093, DOI: 10.1093/bib/bbx085Google Scholar35https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3cXht1KqtLvE&md5=53160f313a2b5c6a9891cb442f4b2dd0The BioCyc collection of microbial genomes and metabolic pathwaysKarp, Peter D.; Billington, Richard; Caspi, Ron; Fulcher, Carol A.; Latendresse, Mario; Kothari, Anamika; Keseler, Ingrid M.; Krummenacker, Markus; Midford, Peter E.; Ong, Quang; Ong, Wai Kit; Paley, Suzanne M.; Subhraveti, PallaviBriefings in Bioinformatics (2019), 20 (4), 1085-1093CODEN: BBIMFX; ISSN:1477-4054. (Oxford University Press)BioCyc.org is a microbial genome Web portal that combines thousands of genomes with addnl. information inferred by computer programs, imported from other databases and curated from the biomedical literature by biologist curators. BioCyc also provides an extensive range of query tools, visualization services and anal. software. Recent advances in BioCyc include an expansion in the content of BioCyc in terms of both the no. of genomes and the types of information available for each genome; an expansion in the amt. of curated content within BioCyc; and new developments in the BioCyc software tools including redesigned gene/protein pages and metabolite pages; new search tools; a new sequence-alignment tool; a new tool for visualizing groups of related metabolic pathways; and a facility called SmartTables, which enables biologists to perform analyses that previously would have required a programmer's assistance.
- 36Caspi, R.; Billington, R.; Ferrer, L.; Foerster, H.; Fulcher, C. A.; Keseler, I. M.; Kothari, A.; Krummenacker, M.; Latendresse, M.; Mueller, L. A.; Ong, Q.; Paley, S.; Subhraveti, P.; Weaver, D. S.; Karp, P. D. The MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databases. Nucleic Acids Res. 2016, 44, D471– 80, DOI: 10.1093/nar/gkv1164Google Scholar36https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtV2nsrnJ&md5=7d8dbff74ddb4e6050783c8ba3d68e2bThe MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databasesCaspi, Ron; Billington, Richard; Ferrer, Luciana; Foerster, Hartmut; Fulcher, Carol A.; Keseler, Ingrid M.; Kothari, Anamika; Krummenacker, Markus; Latendresse, Mario; Mueller, Lukas A.; Ong, Quang; Paley, Suzanne; Subhraveti, Pallavi; Weaver, Daniel S.; Karp, Peter D.Nucleic Acids Research (2016), 44 (D1), D471-D480CODEN: NARHAD; ISSN:0305-1048. (Oxford University Press)A review. The MetaCyc database is a freely accessible comprehensive database describing metabolic pathways and enzymes from all domains of life. The majority of MetaCyc pathways are smallmol. metabolic pathways that have been exptl. detd. MetaCyc contains more than 2400 pathways derived from >46 000 publications, and is the largest curated collection of metabolic pathways. BioCyc is a collection of 5700 organism-specific Pathway/Genome Databases (PGDBs), each contg. the full genome and predicted metabolic network of one organism, including metabolites, enzymes, reactions, metabolic pathways, predicted operons, transport systems, and pathway-hole fillers. The BioCyc website offers a variety of tools for querying and analyzing PGDBs, including Omics Viewers and tools for comparative anal. This article provides an update of new developments in MetaCyc and BioCyc during the last two years, including addn. of Gibbs free energy values for compds. and reactions; redesign of the primary gene/protein page; addn. of a tool for creating diagrams contg. multiple linked pathways; several new search capabilities, including searching for genes based on sequence patterns, searching for databases based on an organism's phenotypes, and a cross-organism search; and a metabolite identifier translation service.
- 37Chang, Y.; Fox, B. G. Identification of Rv3230c as the NADPH oxidoreductase of a two-protein DesA3 acyl-CoA desaturase in Mycobacterium tuberculosis H37Rv. Biochemistry 2006, 45, 13476– 86, DOI: 10.1021/bi0615285Google Scholar37https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28XhtFSrs73M&md5=970108deb08558d4f7b1ff6f349111b6Identification of Rv3230c as the NADPH Oxidoreductase of a Two-Protein DesA3 Acyl-CoA Desaturase in Mycobacterium tuberculosis H37RvChang, Yong; Fox, Brian G.Biochemistry (2006), 45 (45), 13476-13486CODEN: BICHAW; ISSN:0006-2960. (American Chemical Society)DesA3 is a membrane-bound stearoyl-CoA Δ9-desaturase that produces oleic acid, a precursor of mycobacterial membrane phospholipids and triglycerides. The sequence of DesA3 is homologous with those of other membrane desaturases, including the presence of the eight-His motif proposed to bind the diiron center active site. This family of desaturases function as multicomponent complexes and thus require electron transfer proteins for efficient catalytic turnover. Here we present evidence that Rv3230c from Mycobacterium tuberculosis H37Rv is a biol. relevant electron transfer partner for DesA3 from the same pathogen. For these studies, Rv3230c was expressed as a partially sol. protein in Escherichia coli; recombinant DesA3 was expressed in Mycobacterium smegmatis as a catalytically active membrane protein. The addn. of E. coli lysates contg. Rv3230c to lysates of M. smegmatis expressing DesA3 gave strong conversion of [1-14C]-18:0-CoA to [1-14C]-cis-Δ9-18:1-CoA and of [1-14C]-16:0-CoA to [1-14C]-cis-Δ9-16:1-CoA. Both M. tuberculosis proteins were required for reconstitution of activity, as various combinations of control lysates lacking either Rv3230c or DesA3 gave minimal or no activity. Furthermore, the specificity of interaction between Rv3230c and DesA3 was implied by the inability of other related redox systems to substitute for Rv3230c. The reconstituted activity was dependent upon the presence of NADPH, could be satd. by increasing the amt. of Rv3230c added, and was also sensitive to the salt concn. in the buffer. The results are consistent with the formation of a protein-protein complex, possibly with electrostatic character. This work defines a multiprotein, acyl-CoA desaturase complex from M. tuberculosis H37Rv to minimally consist of a sol. Rv3230c reductase and integral membrane DesA3 desaturase. Further implications of this finding relative to the properties of other multiprotein iron-enzyme complexes are discussed.
- 38McGillivray, A.; Golden, N. A.; Gautam, U. S.; Mehra, S.; Kaushal, D. The Mycobacterium tuberculosis Rv2745c plays an important role in responding to redox stress. PLoS One 2014, 9, e93604 DOI: 10.1371/journal.pone.0093604Google Scholar38https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhs1ajurjK&md5=33aa9199d6d28f482d53610ed08fd3f2The Mycobacterium tuberculosis Rv2745c plays an important role in responding to redox stressMcGillivray, Amanda; Golden, Nadia Abrahams; Gautam, Uma Shankar; Mehra, Smriti; Kaushal, DeepakPLoS One (2014), 9 (4), e93604/1-e93604/22, 22 pp.CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is the leading cause of death from an infectious disease worldwide. Over the course of its life cycle in vivo, Mtb is exposed to a plethora of environmental stress conditions. Temporal regulation of genes involved in sensing and responding to such conditions is therefore crucial for Mtb to establish an infection. The Rv2745c (clgR) gene encodes a Clp protease gene regulator that is induced in response to a variety of stress conditions and potentially plays a role in Mtb pathogenesis. The isogenic mutant Mtb:ΔRv2745c is significantly more sensitive to in vitro redox stress generated by diamide, relative to wild-type Mtb as well as to a complemented strain. Together with the fact that the expression of Rv2745c is strongly induced in response to redox stress, these results strongly implicate a role for ClgR in the management of intraphagosomal redox stress. Addnl., we obsd. that redox stress led to the dysregulation of the expression of the σH/σE regulon in the isogenic mutant, Mtb:ΔRv2745c. Furthermore, induction of clgR in Mtb and Mtb:ΔRv2745c (comp) did not lead to Clp protease induction, indicating that clgR has addnl. functions that need to be elucidated. Our data indicate that ClgR plays diverse roles in multiple regulatory networks in response to different stress conditions. In addn. to redox stress, the expression of Rv2745c correlates with the expression of genes involved in sulfate assimilation as well as in response to hypoxia and reaeration. Clearly, the Mtb Rv2745c-encoded ClgR performs different functions during stress response and is important for the pathogenicity of Mtb in vivo, regardless of its induction of the Clp proteolytic pathway.
- 39Hards, K.; Robson, J. R.; Berney, M.; Shaw, L.; Bald, D.; Koul, A.; Andries, K.; Cook, G. M. Bactericidal mode of action of bedaquiline. J. Antimicrob. Chemother. 2015, 70, 2028– 2037, DOI: 10.1093/jac/dkv054Google Scholar39https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhtFyltr7N&md5=22c8df11e6b904ba3645fa7ac74c5074Bactericidal mode of action of bedaquilineHards, Kiel; Robson, Jennifer R.; Berney, Michael; Shaw, Lisa; Bald, Dirk; Koul, Anil; Andries, Koen; Cook, Gregory M.Journal of Antimicrobial Chemotherapy (2015), 70 (7), 2028-2037CODEN: JACHDX; ISSN:0305-7453. (Oxford University Press)Objectives: It is not fully understood why inhibiting ATP synthesis in Mycobacterium species leads to death in non-replicating cells. We investigated the bactericidal mode of action of the anti-tubercular F1Fo-ATP synthase inhibitor bedaquiline (Sirturo) in order to further understand the lethality of ATP synthase inhibition. Methods:Mycobacterium smegmatis strains were used for all the expts. Growth and survival during a bedaquiline challenge were performed in multiple media types. A time-course microarray was performed during initial bedaquiline challenge in minimal medium. Oxygen consumption and proton-motive force measurements were performed on whole cells and inverted membrane vesicles, resp. Results: A killing of 3 log10 cfu/mL was achieved 4-fold more quickly in minimal medium (a glycerol carbon source) vs. rich medium (LB with Tween 80) during bedaquiline challenge. Assessing the accelerated killing condition, we identified a transcriptional remodelling of metab. that was consistent with respiratory dysfunction but inconsistent with ATP depletion. In glycerol-energized cell suspensions, bedaquiline caused an immediate 2.3-fold increase in oxygen consumption. Bedaquiline collapsed the transmembrane pH gradient, but not the membrane potential, in a dose-dependent manner. Both these effects were dependent on binding to the F1Fo-ATP synthase. Conclusions: Challenge with bedaquiline results in an electroneutral uncoupling of respiration-driven ATP synthesis. This may be a determinant of the bactericidal effects of bedaquiline, while ATP depletion may be a determinant of its delayed onset of killing. We propose that bedaquiline binds to and perturbs the a-c subunit interface of the Fo, leading to futile proton cycling, which is known to be lethal to mycobacteria.
- 40Mishra, S.; Shukla, P.; Bhaskar, A.; Anand, K.; Baloni, P.; Jha, R. K.; Mohan, A.; Rajmani, R. S.; Nagaraja, V.; Chandra, N.; Singh, A. Efficacy of β-lactam/β-lactamase inhibitor combination is linked to WhiB4-mediated changes in redox physiology of Mycobacterium tuberculosis. eLife 2017, 6, e25624 DOI: 10.7554/eLife.25624Google Scholar40https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXitlGhur7M&md5=708a67ddb1108ab51d5b9dee9e017b14Efficacy of β-lactam/β-lactamase inhibitor combination is linked to WhiB4-mediated changes in redox physiology of Mycobacterium tuberculosisMishra, Saurabh; Shukla, Prashant; Bhaskar, Ashima; Anand, Kushi; Baloni, Priyanka; Jha, Rajiv Kumar; Mohan, Abhilash; Rajmani, Raju S.; Nagaraja, Valakunja; Chandra†, Nagasuma; Singh, AmiteLife (2017), 6 (), e25624/1-e25624/30CODEN: ELIFA8; ISSN:2050-084X. (eLife Sciences Publications Ltd.)Mycobacterium tuberculosis (Mtb) expresses a broad-spectrum b-lactamase (BlaC) that mediates resistance to one of the highly effective antibacterials, β-lactams. Nonetheless, b-lactams showed mycobactericidal activity in combination with β-lactamase inhibitor, clavulanate (Clav). However, the mechanistic aspects of how Mtb responds to β-lactams such as Amoxicillin in combination with Clav (referred as Augmentin [AG]) are not clear. Here, we identified cytoplasmic redox potential and intracellular redox sensor, WhiB4, as key determinants of mycobacterial resistance against AG. Using computer-based, biochem., redox-biosensor, and genetic strategies, we uncovered a functional linkage between specific determinants of b-lactam resistance (e.g. β-lactamase) and redox potential in Mtb. We also describe the role of WhiB4 in coordinating the activity of b-lactamase in a redox-dependent manner to tolerate AG. Disruption of WhiB4 enhances AG tolerance, whereas overexpression potentiates AG activity against drug-resistant Mtb. Our findings suggest that AG can be exploited to diminish drug-resistance in Mtb through redox-based interventions.
- 41Waddell, S. J.; Stabler, R. A.; Laing, K.; Kremer, L.; Reynolds, R. C.; Besra, G. S. The use of microarray analysis to determine the gene expression profiles of Mycobacterium tuberculosis in response to anti-bacterial compounds. Tuberculosis 2004, 84, 263– 74, DOI: 10.1016/j.tube.2003.12.005Google Scholar41https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD2czgs1KisA%253D%253D&md5=75c1c424e53be6ec2c94a7231853fb75The use of microarray analysis to determine the gene expression profiles of Mycobacterium tuberculosis in response to anti-bacterial compoundsWaddell Simon J; Stabler Richard A; Laing Ken; Kremer Laurent; Reynolds Robert C; Besra Gurdyal STuberculosis (Edinburgh, Scotland) (2004), 84 (3-4), 263-74 ISSN:1472-9792.The response of Mycobacterium tuberculosis to six anti-microbial agents was determined by microarray analysis in an attempt to define mechanisms of innate resistance in M. tuberculosis. The gene expression profiles of M. tuberculosis after treatment at the minimal inhibitory concentration (MIC) for 4 h with isoniazid, isoxyl, tetrahydrolipstatin, SRI#221, SR1#967 and SR1#9190 were compared to untreated M. tuberculosis. A common response to drug exposure was defined, and this expression profile overlapped with a number of other mycobacterial stress responses recently identified by microarray analysis. Compound-specific responses were also distinguished including a number of putative transcriptional regulators and translocation-related genes. These genes may contribute to the intrinsic resistance of M. tuberculosis to anti-microbial compounds. Further investigation into these mechanisms may elucidate novel pathways contributing to mycobacterial drug resistance and influence anti-mycobacterial drug development strategies.
- 42Boshoff, H. I.; Myers, T. G.; Copp, B. R.; McNeil, M. R.; Wilson, M. A.; Barry, C. E., 3rd The transcriptional responses of Mycobacterium tuberculosis to inhibitors of metabolism: Novel insights into drug mechanisms of action. J. Biol. Chem. 2004, 279, 40174– 84, DOI: 10.1074/jbc.M406796200Google Scholar42https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXnsVKrsLs%253D&md5=677489fe3a9981eba62d990ec037f71aThe Transcriptional Responses of Mycobacterium tuberculosis to Inhibitors of Metabolism: Novel insights into drug mechanisms of actionBoshoff, Helena I. M.; Myers, Timothy G.; Copp, Brent R.; McNeil, Michael R.; Wilson, Michael A.; Barry, Clifton E., IIIJournal of Biological Chemistry (2004), 279 (38), 40174-40184CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)The differential transcriptional response of Mycobacterium tuberculosis to drugs and growth-inhibitory conditions was monitored to generate a data set of 430 microarray profiles. Unbiased grouping of these profiles independently clustered agents of known mechanism of action accurately and was successful at predicting the mechanism of action of several unknown agents. These predictions were validated biochem. for two agents of previously uncategorized mechanism, pyridoacridones and phenothiazines. Anal. of this data set further revealed 150 underlying clusters of coordinately regulated genes offering the first glimpse at the full metabolic potential of this organism. A signature subset of these gene clusters was sufficient to classify all known agents as to mechanism of action. Transcriptional profiling of both crude and purified natural products can provide crit. information on both mechanism and detoxification prior to purifn. that can be used to guide the drug discovery process. Thus, the transcriptional profile generated by a crude marine natural product recapitulated the mechanistic prediction from the pure active component. The underlying gene clusters further provide fundamental insights into the metabolic response of bacteria to drug-induced stress and provide a rational basis for the selection of crit. metabolic targets for screening for new agents with improved activity against this important human pathogen.
- 43Amaral, L.; Viveiros, M. Thioridazine: A Non-Antibiotic Drug Highly Effective, in Combination with First Line Anti-Tuberculosis Drugs, against Any Form of Antibiotic Resistance of Mycobacterium tuberculosis Due to Its Multi-Mechanisms of Action. Antibiotics 2017, 6, 3, DOI: 10.3390/antibiotics6010003Google ScholarThere is no corresponding record for this reference.
- 44Lee, R. E.; Protopopova, M.; Crooks, E.; Slayden, R. A.; Terrot, M.; Barry, C. E., 3rd Combinatorial lead optimization of [1,2]-diamines based on ethambutol as potential antituberculosis preclinical candidates. J. Comb. Chem. 2003, 5, 172– 87, DOI: 10.1021/cc020071pGoogle Scholar44https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXkvFKguw%253D%253D&md5=3005c111ca5aa61044f016dcfafc6d09Combinatorial Lead Optimization of [1,2]-Diamines Based on Ethambutol as Potential Antituberculosis Preclinical CandidatesLee, Richard E.; Protopopova, Marina; Crooks, Emma; Slayden, Richard A.; Terrot, Marianne; Barry, Clifton E., IIIJournal of Combinatorial Chemistry (2003), 5 (2), 172-187CODEN: JCCHFF; ISSN:1520-4766. (American Chemical Society)Despite relatively modest potency, ethambutol (EMB, (S,S)-[N,N-di-2-amino-1-butanol]ethylenediamine) is a mainstay of contemporary chemotherapy for the treatment of tuberculosis. We have developed a solid-phase synthesis of 1,2-diamine analogs of EMB using a novel acylation-redn. sequence that is compatible with high-throughput 96-well format chem. Using this procedure, we have synthesized 63,238 diamine analogs in pools of 10 that are suitable for testing. MIC and a target-based reporter assay were used to direct deconvolution of 2796 individual compds. from these mixts., and the 69 most potent mols. were resynthesized in milligram quantities for hit confirmation. Purifn. of these individual active diamine analogs allowed the identification of 26 compds. with activity equal to or greater than EMB. Amines which occurred most frequently in active compds. included many with large hydrophobic moieties, suggesting that optimization was perhaps selecting for the isoprenoid binding site of the arabinosyltransferase target of EMB. N-Geranyl-N'-(2-adamantyl)ethane-1,2-diamine, the most active of these diamines, displayed a 14-35-fold improvement in activity in vitro against Mycobacterium tuberculosis, as compared to EMB.
- 45Makarov, V.; Manina, G.; Mikusova, K.; Möllmann, U.; Ryabova, O.; Saint-Joanis, B.; Dhar, N.; Pasca, M. R.; Buroni, S.; Lucarelli, A. P.; Milano, A.; De Rossi, E.; Belanova, M.; Bobovska, A.; Dianiskova, P.; Kordulakova, J.; Sala, C.; Fullam, E.; Schneider, P.; McKinney, J. D.; Brodin, P.; Christophe, T.; Waddell, S.; Butcher, P.; Albrethsen, J.; Rosenkrands, I.; Brosch, R.; Nandi, V.; Bharath, S.; Gaonkar, S.; Shandil, R. K.; Balasubramanian, V.; Balganesh, T.; Tyagi, S.; Grosset, J.; Riccardi, G.; Cole, S. T. Benzothiazinones Kill Mycobacterium tuberculosis by Blocking Arabinan Synthesis. Science 2009, 324, 801, DOI: 10.1126/science.1171583Google Scholar45https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXlsVeksrY%253D&md5=1d80f14ba88460ef5c8ea31793dbb592Benzothiazinones Kill Mycobacterium tuberculosis by Blocking Arabinan SynthesisMakarov, Vadim; Manina, Giulia; Mikusova, Katarina; Moellmann, Ute; Ryabova, Olga; Saint-Joanis, Brigitte; Dhar, Neeraj; Pasca, Maria Rosalia; Buroni, Silvia; Lucarelli, Anna Paola; Milano, Anna; De Rossi, Edda; Belanova, Martina; Bobovska, Adela; Dianiskova, Petronela; Kordulakova, Jana; Sala, Claudia; Fullam, Elizabeth; Schneider, Patricia; McKinney, John D.; Brodin, Priscille; Christophe, Thierry; Waddell, Simon; Butcher, Philip; Albrethsen, Jakob; Rosenkrands, Ida; Brosch, Roland; Nandi, Vrinda; Bharath, Sowmya; Gaonkar, Sheshagiri; Shandil, Radha K.; Balasubramanian, Venkataraman; Balganesh, Tanjore; Tyagi, Sandeep; Grosset, Jacques; Riccardi, Giovanna; Cole, Stewart T.Science (Washington, DC, United States) (2009), 324 (5928), 801-804CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)New drugs are required to counter the tuberculosis (TB) pandemic. Here, we describe the synthesis and characterization of 1,3-benzothiazin-4-ones (BTZs), a new class of antimycobacterial agents that kill Mycobacterium tuberculosis in vitro, ex vivo, and in mouse models of TB. Using genetics and biochem., we identified the enzyme decaprenylphosphoryl-β-D-ribose 2'-epimerase as a major BTZ target. Inhibition of this enzymic activity abolishes the formation of decaprenylphosphoryl arabinose, a key precursor that is required for the synthesis of the cell-wall arabinans, thus provoking cell lysis and bacterial death. The most advanced compd., BTZ043, is a candidate for inclusion in combination therapies for both drug-sensitive and extensively drug-resistant TB.
- 46Xu, Z.; Meshcheryakov, V. A.; Poce, G.; Chng, S.-S. MmpL3 is the flippase for mycolic acids in mycobacteria. Proc. Natl. Acad. Sci. U.S.A. 2017, 114, 7993, DOI: 10.1073/pnas.1700062114Google Scholar46https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtFCrtL7O&md5=1c8a806ca2a1eaebefe7c1e6bf3bcab9MmpL3 is the flippase for mycolic acids in mycobacteriaXu, Zhujun; Meshcheryakov, Vladimir A.; Poce, Giovanna; Chng, Shu-SinProceedings of the National Academy of Sciences of the United States of America (2017), 114 (30), 7993-7998CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)The defining feature of the mycobacterial outer membrane (OM) is the presence of mycolic acids (MAs), which, in part, render the bilayer extremely hydrophobic and impermeable to external insults, including many antibiotics. Although the biosynthetic pathway of MAs is well studied, the mechanism(s) by which these lipids are transported across the cell envelope is(are) much less known. Mycobacterial membrane protein Large 3 (MmpL3), an essential inner membrane (IM) protein, is implicated in MA transport, but its exact function has not been elucidated. It is believed to be the cellular target of several antimycobacterial compds.; however, evidence for direct inhibition of MmpL3 activity is also lacking. Here, we establish that MmpL3 is the MA flippase at the IM of mycobacteria and is the mol. target of BM212, a 1,5-diarylpyrrole compd. We develop assays that selectively access mycolates on the surface of Mycobacterium smegmatis spheroplasts, allowing us to monitor flipping of MAs across the IM. Using these assays, we establish the mechanism of action of BM212 as a potent MmpL3 inhibitor, and use it as a mol. probe to demonstrate the requirement for functional MmpL3 in the transport of MAs across the IM. Finally, we show that BM212 binds MmpL3 directly and inhibits its activity. Our work provides fundamental insights into OM biogenesis and MA transport in mycobacteria. Furthermore, our assays serve as an important platform for accelerating the validation of small mols. that target MmpL3, and their development as future antituberculosis drugs.
- 47Portevin, D.; de Sousa, D.; Auria, C.; Houssin, C.; Grimaldi, C.; Chami, M.; Daffé, M.; Guilhot, C. A polyketide synthase catalyzes the last condensation step of mycolic acid biosynthesis in mycobacteria and related organisms. Proc. Natl. Acad. Sci. U. S. A. 2004, 101, 314, DOI: 10.1073/pnas.0305439101Google Scholar47https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXjvFamuw%253D%253D&md5=dbbe12644994eba3860c2733d429f4b1A polyketide synthase catalyzes the last condensation step of mycolic acid biosynthesis in mycobacteria and related organismsPortevin, Damien; de Sousa-d'Auria, Celia; Houssin, Christine; Grimaldi, Christine; Chami, Mohamed; Daffe, Mamadou; Guilhot, ChristopheProceedings of the National Academy of Sciences of the United States of America (2004), 101 (1), 314-319CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Mycolic acids are major and specific constituents of the cell envelope of Corynebacterineae, a suborder of bacterial species including several important human pathogens such as Mycobacterium tuberculosis, Mycobacterium leprae, or Corynebacterium diphtheriae. These long-chain fatty acids are involved in the unusual architecture and impermeability of the cell envelope of these bacteria. The condensase, the enzyme responsible for the final condensation step in mycolic acid biosynthesis, has remained an enigma for decades. By in silico anal. of various mycobacterial genomes, we identified a candidate enzyme, Pks13, that contains the four catalytic domains required for the condensation reaction. Orthologs of this enzyme were found in other Corynebacterineae species. A Corynebacterium glutamicum strain with a deletion in the pks13 gene was shown to be deficient in mycolic acid prodn. whereas it was able to produce the fatty acids precursors. This mutant strain displayed an altered cell envelope structure. We showed that the pks13 gene was essential for the survival of Mycobacterium smegmatis. A conditional M. smegmatis mutant carrying its only copy of pks13 on a thermosensitive plasmid exhibited mycolic acid biosynthesis defect if grown at nonpermissive temp. These results indicate that Pks13 is the condensase, a promising target for the development of new antimicrobial drugs against Corynebacterineae.
- 48Gavalda, S.; Bardou, F.; Laval, F.; Bon, C.; Malaga, W.; Chalut, C.; Guilhot, C.; Mourey, L.; Daffé, M.; Quémard, A. The Polyketide Synthase Pks13 Catalyzes a Novel Mechanism of Lipid Transfer in Mycobacteria. Chem. Biol. 2014, 21, 1660– 1669, DOI: 10.1016/j.chembiol.2014.10.011Google Scholar48https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhvF2gs7%252FO&md5=aa1ce63f8e0bec476d5e9fb91ef7c63cThe Polyketide Synthase Pks13 Catalyzes a Novel Mechanism of Lipid Transfer in MycobacteriaGavalda, Sabine; Bardou, Fabienne; Laval, Francoise; Bon, Cecile; Malaga, Wladimir; Chalut, Christian; Guilhot, Christophe; Mourey, Lionel; Daffe, Mamadou; Quemard, AnnaikChemistry & Biology (Oxford, United Kingdom) (2014), 21 (12), 1660-1669CODEN: CBOLE2; ISSN:1074-5521. (Elsevier Ltd.)Mycolate-contg. compds. constitute major strategic elements of the protective coat surrounding the tubercle bacillus. We have previously shown that FAAL32-Pks13 polyketide synthase catalyzes the condensation reaction, which produces α-alkyl β-ketoacids, direct precursors of mycolic acids. In contrast to the current biosynthesis model, we show here that Pks13 catalyzes itself the release of the neosynthesized products and demonstrate that this function is carried by its thioesterase-like domain. Most importantly, in agreement with the prediction of a trehalose-binding pocket in its catalytic site, this domain exhibits an acyltransferase activity and transfers Pks13's products onto an acceptor mol., mainly trehalose, leading to the formation of the trehalose monomycolate precursor. Thus, this work allows elucidation of the hinge step of the mycolate-contg. compd. biosynthesis pathway. Above all, it highlights a unique mechanism of transfer of polyketide synthase products in mycobacteria, which is distinct from the conventional intervention of the discrete polyketide-assocd. protein (Pap)-type acyltransferases.
- 49Su, C.-C.; Klenotic, P. A.; Bolla, J. R.; Purdy, G. E.; Robinson, C. V.; Yu, E. W. MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine. Proc. Natl. Acad. Sci. U.S.A. 2019, 116, 11241, DOI: 10.1073/pnas.1901346116Google Scholar49https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXhtV2itLrK&md5=36f2807f7a55ec997fb211206054eef7MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamineSu, Chih-Chia; Klenotic, Philip A.; Bolla, Jani Reddy; Purdy, Georgiana E.; Robinson, Carol V.; Yu, Edward W.Proceedings of the National Academy of Sciences of the United States of America (2019), 116 (23), 11241-11246CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)The cell envelope of Mycobacterium tuberculosis is notable for the abundance of mycolic acids (MAs), essential to mycobacterial viability, and of other species-specific lipids. The mycobacterial cell envelope is extremely hydrophobic, which contributes to virulence and antibiotic resistance. However, exactly how fatty acids and lipidic elements are transported across the cell envelope for cell-wall biosynthesis is unclear. Mycobacterial membrane protein Large 3 (MmpL3) is essential and required for transport of trehalose monomycolates (TMMs), precursors of MA-contg. trehalose dimycolates (TDM) and mycolyl arabinogalactan peptidoglycan, but the exact function of MmpL3 remains elusive. Here, we report a crystal structure of Mycobacterium smegmatis MmpL3 at a resoln. of 2.59 Å, revealing a monomeric mol. that is structurally distinct from all known bacterial membrane proteins. A previously unknown MmpL3 ligand, phosphatidylethanolamine (PE), was discovered inside this transporter. We also show, via native mass spectrometry, that MmpL3 specifically binds both TMM and PE, but not TDM, in the micromolar range. These observations provide insight into the function of MmpL3 and suggest a possible role for this protein in shuttling a variety of lipids to strengthen the mycobacterial cell wall.
- 50Cox, J. A. G.; Abrahams, K. A.; Alemparte, C.; Ghidelli-Disse, S.; Rullas, J.; Angulo-Barturen, I.; Singh, A.; Gurcha, S. S.; Nataraj, V.; Bethell, S.; Remuiñán, M. J.; Encinas, L.; Jervis, P. J.; Cammack, N. C.; Bhatt, A.; Kruse, U.; Bantscheff, M.; Fütterer, K.; Barros, D.; Ballell, L.; Drewes, G.; Besra, G. S. THPP target assignment reveals EchA6 as an essential fatty acid shuttle in mycobacteria. Nat. Microbiol. 2016, 1, 15006, DOI: 10.1038/nmicrobiol.2015.6Google Scholar50https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXkvFyqtbk%253D&md5=d1704f5bc0a939f1213078f71cdcbf49THPP target assignment reveals EchA6 as an essential fatty acid shuttle in MycobacteriaCox, Jonathan A. G.; Abrahams, Katherine A.; Alemparte, Carlos; Ghidelli-Disse, Sonja; Rullas, Joaquin; Angulo-Barturen, Inigo; Singh, Albel; Gurcha, Sudagar S.; Nataraj, Vijayashankar; Bethell, Stephen; Remuinan, Modesto J.; Encinas, Lourdes; Jervis, Peter J.; Cammack, Nicholas C.; Bhatt, Apoorva; Kruse, Ulrich; Bantscheff, Marcus; Futterer, Klaus; Barros, David; Ballell, Lluis; Drewes, Gerard; Besra, Gurdyal S.Nature Microbiology (2016), 1 (2), 15006CODEN: NMAICH; ISSN:2058-5276. (Nature Publishing Group)Phenotypic screens for bactericidal compds. against drug-resistant tuberculosis are beginning to yield novel inhibitors. However, reliable target identification remains challenging. Here, we show that tetrahydropyrazo[1,5-a]pyrimidine-3-carboxamide (THPP) selectively pulls down EchA6 in a stereospecific manner, instead of the previously assigned target Mycobacterium tuberculosis MmpL3. While homologous to mammalian enoyl-CoA hydratases, EchA6 is non-catalytic yet essential and binds long-chain acyl-CoAs. THPP inhibitors compete with CoA-binding, suppress mycolic acid synthesis, and are bactericidal in a mouse model of chronic tuberculosis infection. A point mutation, W133A, abrogated THPP-binding and increased both the in vitro min. inhibitory concn. and the in vivo ED 99 in mice. Surprisingly, EchA6 interacts with selected enzymes of fatty acid synthase II (FAS-II) in bacterial two-hybrid assays, suggesting essentiality may be linked to feeding long-chain fatty acids to FAS-II. Finally, our data show that spontaneous resistance-conferring mutations can potentially obscure the actual target or alternative targets of small mol. inhibitors.
- 51Belisle, J. T.; Vissa, V. D.; Sievert, T.; Takayama, K.; Brennan, P. J.; Besra, G. S. Role of the Major Antigen of Mycobacterium tuberculosis in Cell Wall Biogenesis. Science 1997, 276, 1420, DOI: 10.1126/science.276.5317.1420Google Scholar51https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2sXjsFOjtLg%253D&md5=b810258a42304fcb17cd57b2308aee85Role of the major antigen of Mycobacterium tuberculosis in cell wall biogenesisBelisle, John T.; Vissa, Varalakshmi D.; Sievert, Todd; Takayama, Kuni; Brennan, Patrick J.; Besra, Gurdyal S.Science (Washington, D. C.) (1997), 276 (5317), 1420-1422CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)The dominant exported proteins and protective antigens of Mycobacterium tuberculosis are a triad of related gene products called the antigen 85 (Ag85) complex. Each has also been implicated in disease pathogenesis through its fibronectin-binding capacities. A carboxylesterase domain was found within the amino acid sequences of Ag85A, B, and C, and each protein acted as a mycolyltransferase involved in the final stages of mycobacterial cell wall assembly, as shown by direct enzyme assay and site-directed mutagenesis. Furthermore, the use of an antagonist (6-azido-6-deoxy-α,α'-trehalose) of this activity demonstrates that these proteins are essential and potential targets for new antimycobacterial drugs.
- 52Favrot, L.; Grzegorzewicz, A. E.; Lajiness, D. H.; Marvin, R. K.; Boucau, J.; Isailovic, D.; Jackson, M.; Ronning, D. R. Mechanism of inhibition of Mycobacterium tuberculosis antigen 85 by ebselen. Nat. Commun. 2013, 4, 2748, DOI: 10.1038/ncomms3748Google Scholar52https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2c7jtlarsA%253D%253D&md5=0e00cdc6af699b2ee93fce10c7cbfc0cMechanism of inhibition of Mycobacterium tuberculosis antigen 85 by ebselenFavrot Lorenza; Grzegorzewicz Anna E; Lajiness Daniel H; Marvin Rachel K; Boucau Julie; Isailovic Dragan; Jackson Mary; Ronning Donald RNature communications (2013), 4 (), 2748 ISSN:.The increasing prevalence of drug-resistant tuberculosis highlights the need for identifying new antitubercular drugs that can treat these infections. The antigen 85 (Ag85) complex has emerged as an intriguing mycobacterial drug target due to its central role in synthesizing major components of the inner and outer leaflets of the mycobacterial outer membrane. Here we identify ebselen (EBS) as a potent inhibitor of the Mycobacterium tuberculosis Ag85 complex. Mass spectrometry data show that EBS binds covalently to a cysteine residue (C209) located near the Ag85C active site. The crystal structure of Ag85C in the presence of EBS shows that C209 modification restructures the active site, thereby disrupting the hydrogen-bonded network within the active site that is essential for enzymatic activity. C209 mutations display marked decreases in enzymatic activity. These data suggest that compounds using this mechanism of action will strongly inhibit the Ag85 complex and minimize the selection of drug resistance.
- 53Mdluli, K.; Kaneko, T.; Upton, A. The tuberculosis drug discovery and development pipeline and emerging drug targets. Cold Spring Harb. Perspect. Med. 2015, 5, a021154, DOI: 10.1101/cshperspect.a021154Google ScholarThere is no corresponding record for this reference.
- 54Ling, L. L.; Schneider, T.; Peoples, A. J.; Spoering, A. L.; Engels, I.; Conlon, B. P.; Mueller, A.; Schäberle, T. F.; Hughes, D. E.; Epstein, S.; Jones, M.; Lazarides, L.; Steadman, V. A.; Cohen, D. R.; Felix, C. R.; Fetterman, K. A.; Millett, W. P.; Nitti, A. G.; Zullo, A. M.; Chen, C.; Lewis, K. A new antibiotic kills pathogens without detectable resistance. Nature 2015, 517, 455– 459, DOI: 10.1038/nature14098Google Scholar54https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFOju7w%253D&md5=27e302d7d44a549a91aa52113c3b4ad8A new antibiotic kills pathogens without detectable resistanceLing, Losee L.; Schneider, Tanja; Peoples, Aaron J.; Spoering, Amy L.; Engels, Ina; Conlon, Brian P.; Mueller, Anna; Schaberle, Till F.; Hughes, Dallas E.; Epstein, Slava; Jones, Michael; Lazarides, Linos; Steadman, Victoria A.; Cohen, Douglas R.; Felix, Cintia R.; Fetterman, K. Ashley; Millett, William P.; Nitti, Anthony G.; Zullo, Ashley M.; Chen, Chao; Lewis, KimNature (London, United Kingdom) (2015), 517 (7535), 455-459CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)Antibiotic resistance is spreading faster than the introduction of new compds. into clin. practice, causing a public health crisis. Most antibiotics were produced by screening soil microorganisms, but this limited resource of cultivable bacteria was overmined by the 1960s. Synthetic approaches to produce antibiotics have been unable to replace this platform. Uncultured bacteria make up approx. 99% of all species in external environments, and are an untapped source of new antibiotics. We developed several methods to grow uncultured organisms by cultivation in situ or by using specific growth factors. Here we report a new antibiotic that we term teixobactin, discovered in a screen of uncultured bacteria. Teixobactin inhibits cell wall synthesis by binding to a highly conserved motif of lipid II (precursor of peptidoglycan) and lipid III (precursor of cell wall teichoic acid). We did not obtain any mutants of Staphylococcus aureus or Mycobacterium tuberculosis resistant to teixobactin. The properties of this compd. suggest a path towards developing antibiotics that are likely to avoid development of resistance.
- 55
Isoniazid was included as a positive control (0.9–29.2 μM), and comparison, with it being an antibiotic that targets the cell wall and has rapid bactericidal activity.
There is no corresponding record for this reference. - 56Hendon-Dunn, C. L.; Pertinez, H.; Marriott, A. A. N.; Hatch, K. A.; Allnutt, J. C.; Davies, G.; Bacon, J. Regrowth of Mycobacterium tuberculosis Populations Exposed to Antibiotic Combinations Is Due to the Presence of Isoniazid and Not Bacterial Growth Rate. Antimicrob. Agents Chemother. 2019, 63, e00570-19 DOI: 10.1128/AAC.00570-19Google ScholarThere is no corresponding record for this reference.
- 57Banerjee, A.; Dubnau, E.; Quemard, A.; Balasubramanian, V.; Um, K. S.; Wilson, T.; Collins, D.; de Lisle, G.; Jacobs, W. R. inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosis. Science 1994, 263, 227, DOI: 10.1126/science.8284673Google Scholar57https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2cXhsFylt7c%253D&md5=12f3eabbca02009180332c1e65db8c15inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosisBanerjee, Asesh; Dubnau, Eugenie; Quemard, Annaik; Balasubramanian, V.; Um, Kyung Sun; Wilson, Theresa; Collins, Des; de Lisle, Geoffrey; Jacobs, William, R., Jr.Science (Washington, DC, United States) (1994), 263 (5144), 227-30CODEN: SCIEAS; ISSN:0036-8075.Isoniazid (isonicotinic acid hydrazide, INH) is one of the most widely used antituberculosis drugs, yet its precise target of action on Mycobacterium tuberculosis is unknown. A missense mutation within the mycobacterial inhA gene was shown to confer resistance to both INH and ethionamide (ETH) in M. smegmatis and in M. bovis. The wild-type inhA gene also conferred INH and ETH resistance when transferred on a multicopy plasmid vector to M. smegmatis and M. bovis BCG. The InhA protein shows a significant sequence conservation with the Escherichia coli enzyme EnvM, and cell-free assays indicate that it may be involved in mycolic acid biosynthesis. These results suggest that InhA is likely a primary target of action for INH and ETH.
- 58Takayama, K.; Kilburn, J. O. Inhibition of synthesis of arabinogalactan by ethambutol in Mycobacterium smegmatis. Antimicrob. Agents Chemother. 1989, 33, 1493, DOI: 10.1128/AAC.33.9.1493Google Scholar58https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL1MXmt1yrsbg%253D&md5=568b1756e3a57a9ebd1a871e4df22e5dInhibition of synthesis of arabinogalactan by ethambutol in Mycobacterium smegmatisTakayama, Kuni; Kilburn, James O.Antimicrobial Agents and Chemotherapy (1989), 33 (9), 1493-9CODEN: AMACCQ; ISSN:0066-4804.Ethambutol at 3.0 μg/mL inhibited the transfer of label from D-[14C]glucose into the D-arabinose residue of arabinogalactan in whole cells of a drug-susceptible strain of M. smegmatis. This inhibition began almost immediately after exposure of the cells to the drug. When drug-resistant M. smegmatis was used in a similar expt., no such drug inhibition was detected. A much higher concn. of ethambutol (>50 μg/mL) was required to show this inhibition. The drug also inhibited synthesis of arabinose-contg. oligosaccharides when a cell-free enzyme system was used. These results suggest that the site of action of ethambutol is somewhere on the pathway between the conversion of D-glucose to D-arabinose and the transfer of arabinose into arabinogalactan. The primary mode of action of ethambutol appears to be inhibition of arabinogalactan synthesis.
- 59Tahlan, K.; Wilson, R.; Kastrinsky, D. B.; Arora, K.; Nair, V.; Fischer, E.; Barnes, S. W.; Walker, J. R.; Alland, D.; Barry, C. E.; Boshoff, H. I. SQ109 Targets MmpL3, a Membrane Transporter of Trehalose Monomycolate Involved in Mycolic Acid Donation to the Cell Wall Core of Mycobacterium tuberculosis. Antimicrob. Agents Chemother. 2012, 56, 1797, DOI: 10.1128/AAC.05708-11Google Scholar59https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XltV2ru7k%253D&md5=6309318d9e4d2f0c75a9fed20b8108eaSQ109 targets MmpL3, a membrane transporter of trehalose monomycolate involved in mycolic acid donation to the cell wall core of Mycobacterium tuberculosisTahlan, Kapil; Wilson, Regina; Kastrinsky, David B.; Arora, Kriti; Nair, Vinod; Fischer, Elizabeth; Barnes, S. Whitney; Walker, John R.; Alland, David; Barry, Clifton E., III; Boshoff, Helena I.Antimicrobial Agents and Chemotherapy (2012), 56 (4), 1797-1809CODEN: AMACCQ; ISSN:0066-4804. (American Society for Microbiology)SQ109, a 1,2-diamine related to ethambutol, is currently in clin. trials for the treatment of tuberculosis, but its mode of action remains unclear. Here, we demonstrate that SQ109 disrupts cell wall assembly, as evidenced by macromol. incorporation assays and ultrastructural analyses. SQ109 interferes with the assembly of mycolic acids into the cell wall core of Mycobacterium tuberculosis, as bacilli exposed to SQ109 show immediate inhibition of trehalose dimycolate (TDM) prodn. and fail to attach mycolates to the cell wall arabinogalactan. These effects were not due to inhibition of mycolate synthesis, since total mycolate levels were unaffected, but instead resulted in the accumulation of trehalose monomycolate (TMM), the precursor of TDM and cell wall mycolates. In vitro assays using purified enzymes showed that this was not due to inhibition of the secreted Ag85 mycolyltransferases. We were unable to achieve spontaneous generation of SQ109-resistant mutants; however, analogs of this compd. that resulted in similar shutdown of TDM synthesis with concomitant TMM accumulation were used to spontaneously generate resistant mutants that were also cross-resistant to SQ109. Whole-genome sequencing of these mutants showed that these all had mutations in the essential mmpL3 gene, which encodes a transmembrane transporter. Our results suggest that MmpL3 is the target of SQ109 and that MmpL3 is a transporter of mycobacterial TMM.
- 60Prosser, G. A.; de Carvalho, L. P. S. Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserine. FEBS Journal 2013, 280, 1150– 1166, DOI: 10.1111/febs.12108Google Scholar60https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXis1Wmtbw%253D&md5=01551bd01443b8a028e6dde0ea3e03f8Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserineProsser, Gareth A.; de Carvalho, Luiz Pedro S.FEBS Journal (2013), 280 (4), 1150-1166CODEN: FJEOAC; ISSN:1742-464X. (Wiley-Blackwell)D-Cycloserine (DCS) is an antibiotic that is currently used in second-line treatment of tuberculosis. DCS is a structural analog of D-alanine, and targets two enzymes involved in the cytosolic stages of peptidoglycan synthesis: alanine racemase (Alr) and D-alanine:D-alanine ligase (Ddl). The mechanisms of inhibition of DCS have been well-assessed using Alr and Ddl enzymes from various bacterial species, but little is known regarding the interactions of DCS with the mycobacterial orthologues of these enzymes. We have over-expressed and purified recombinant Mycobacterium tuberculosis Ddl (MtDdl; Rv2981c), and report a kinetic examn. of the enzyme with both its native substrate and DCS. MtDdl is activated by K+, follows an ordered ter ter mechanism and displays distinct affinities for D-Ala at each D-Ala binding site (Km,D-Ala1 = 0.075 mm,Km,D-Ala2 = 3.6 mm). ATP is the first substrate to bind and is necessary for subsequent binding of D-alanine or DCS. The pH dependence of MtDdl kinetic parameters indicate that general base chem. is involved in the catalytic step. DCS was found to competitively inhibit D-Ala binding at both MtDdl D-Ala sites with equal affinity (Ki,DCS1 = 14 μm,Ki,DCS2 = 25 μm); however, each enzyme active site can only accommodate a single DCS mol. at a given time. The pH dependence of Ki,DCS2 revealed a loss of DCS binding affinity at high pH (pKa = 7.5), suggesting that DCS binds optimally in the zwitterionic form. The results of this study may assist in the design and development of novel Ddl-specific inhibitors for use as anti-mycobacterial agents.
- 61Briffotaux, J.; Liu, S.; Gicquel, B. Genome-Wide Transcriptional Responses of Mycobacterium to Antibiotics. Front. Microbiol. 2019, 10, 249, DOI: 10.3389/fmicb.2019.00249Google Scholar61https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3cbhtlSkug%253D%253D&md5=aff38e10263f3b6e7c854010022fbee1Genome-Wide Transcriptional Responses of Mycobacterium to AntibioticsBriffotaux Julien; Liu Shengyuan; Gicquel Brigitte; Briffotaux Julien; Gicquel Brigitte; Gicquel BrigitteFrontiers in microbiology (2019), 10 (), 249 ISSN:1664-302X.Antibiotics can stimulate or depress gene expression in bacteria. The analysis of transcriptional responses of Mycobacterium to antimycobacterial compounds has improved our understanding of the mode of action of various drug classes and the efficacy and effect of such compounds on the global metabolism of Mycobacterium. This approach can provide new insights for known antibiotics, for example those currently used for tuberculosis treatment, as well as help to identify the mode of action and predict the targets of new compounds identified by whole-cell screening assays. In addition, changes in gene expression profiles after antimycobacterial treatment can provide information about the adaptive ability of bacteria to escape the effects of antibiotics and allow monitoring of the physiology of the bacteria during treatment. Genome-wide expression profiling also makes it possible to pinpoint genes differentially expressed between drug sensitive Mycobacterium and multidrug-resistant clinical isolates. Finally, genes involved in adaptive responses and drug tolerance could become new targets for improving the efficacy of existing antibiotics.
- 62Johnsson, K.; King, D. S.; Schultz, P. G. Studies on the Mechanism of Action of Isoniazid and Ethionamide in the Chemotherapy of Tuberculosis. J. Am. Chem. Soc. 1995, 117, 5009– 5010, DOI: 10.1021/ja00122a038Google Scholar62https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2MXlt1Srt7Y%253D&md5=3492989087bfdd266d0d8c7b10037225Studies on the Mechanism of Action of Isoniazid and Ethionamide in the Chemotherapy of TuberculosisJohnsson, Kai; King, David S.; Schultz, Peter G.Journal of the American Chemical Society (1995), 117 (17), 5009-10CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)The inactivation of the enoyl-reductase InhA from Mycobacterium tuberculosis by reactive intermediates formed during the oxidn. of isoniazid and ethionamide was studied. Both drugs can generate electrophilic intermediates capable of reacting with a nucleophilic group of InhA, leading to its inactivation. After inactivation of InhA by isoniazid, one mol. of isoniazid per InhA is covalently bound to the enzyme. Mapping studies suggest that Cys243 is the residue modified in the course of the inactivation.
- 63Milanes, C. L.; Pernalete, N.; Starosta, R.; Perez-Gonzalez, M.; Paz-Martinez, V.; Bellorin-Font, E. Altered response of adenylate cyclase to parathyroid hormone during compensatory renal growth. Kidney Int. 1989, 36, 802– 9, DOI: 10.1038/ki.1989.265Google Scholar63https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK3cXjsVWjtw%253D%253D&md5=91a1c3931d282d6703be188411f389ecAltered response of adenylate cyclase to parathyroid hormone during compensatory renal growthMilanes, Carmen L.; Pernalete, Nidia; Starosta, Rebeca; Perez-Gonzalez, Margarita; Paz-Martinez, Virgilio; Bellorin-Font, EzequielKidney International (1989), 36 (5), 802-9CODEN: KDYIA5; ISSN:0085-2538.The loss of renal mass is assocd. with functional adaptations in the remaining nephrons to maintain homeostasis. Although parathyroid hormone (PTH) is important in the adaptations to phosphate, the mechanisms are not completely defined. In the present study, the response of the adenylate cyclase system to PTH was examd. in renal cortical membranes of rat kidneys ten days after unilateral nephrectomy. The kidneys obtained at the time of the initial nephrectomy were used as controls. Unilateral nephrectomy resulted in contralateral compensatory renal growth, as demonstrated by a 24% increase in wt. in the remaining kidney. Glomerular filtration rate (GFR) after unilateral nephrectomy was 62% of the control, whereas basal fractional phosphate excretion was higher in rats with unilateral nephrectomy (7.7% vs. 2.9%). PTH infusion resulted in a similar increase of fractional phosphate excretion and urinary cAMP in both groups. In the absence of added guanine nucleotides, PTH-dependent adenylate cyclase activity in cortical membranes from kidneys with compensatory growth was decreased as compared to controls (Vmax 807.5 pmol cAMP/mg protein/30 min vs. 1384.8, resp.). The apparent affinity for PTH stimulation of adenylate cyclase (Kact) was unchanged. Magnesium-dependent adenylate cyclase activity was also decreased in the membranes from kidneys with compensatory growth. However, the kinetics of adenylate cyclase for the substrates ATP-Mg or ATP-Mn were similar. The addn. of Gpp(NH)p resulted in a similar maximal response to PTH in the two groups, indicating an increased response of the enzyme to PTH in the presence of the guanine nucleotide. Cholera toxin-dependent ADP-ribosylation of the stimulatory guanine nucleotide binding protein (Gs) showed a marked decrease in the apparent content of the alpha subunit in membranes from kidneys with compensatory growth compared to controls. On the contrary, pertussis toxin-dependent ADP-ribosylation of the inhibitory guanine nucleotide binding protein (Gi) did not show differences in the content of the alpha subunit in both groups of membranes. Since the transduction of the hormone signal from the receptor is mediated by G proteins, the results suggest that during compensatory renal growth a decrease in the alpha subunit of Gs could account for the impaired response of adenylate cyclase to PTH in vitro, which could be overcome by high concns. of guanine nucleotides.
- 64Bacon, J.; Alderwick, L. J.; Allnutt, J. A.; Gabasova, E.; Watson, R.; Hatch, K. A.; Clark, S. O.; Jeeves, R. E.; Marriott, A.; Rayner, E.; Tolley, H.; Pearson, G.; Hall, G.; Besra, G. S.; Wernisch, L.; Williams, A.; Marsh, P. D. Non-replicating Mycobacterium tuberculosis elicits a reduced infectivity profile with corresponding modifications to the cell wall and extracellular matrix. PLoS One 2014, 9, e87329 DOI: 10.1371/journal.pone.0087329Google Scholar64https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhsVGrtrnJ&md5=4d7773618b4185c2ee98f32dd97bd15bNon-replicating Mycobacterium tuberculosis elicits a reduced infectivity profile with corresponding modifications to the cell wall and extracellular matrixBacon, Joanna; Alderwick, Luke J.; Allnutt, Jon A.; Gabasova, Evelina; Watson, Robert; Hatch, Kim A.; Clark, Simon O.; Jeeves, Rose E.; Marriott, Alice; Rayner, Emma; Tolley, Howard; Pearson, Geoff; Hall, Graham; Besra, Gurdyal S.; Wernisch, Lorenz; Williams, Ann; Marsh, Philip D.PLoS One (2014), 9 (2), e87329/1-e87329/17, 17 pp.CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)A key feature of Mycobacterium tuberculosis is its ability to become dormant in the host. Little is known of the mechanisms by which these bacilli are able to persist in this state. Therefore, the focus of this study was to emulate environmental conditions encountered by M. tuberculosis in the granuloma, and det. the effect of such conditions on the physiol. and infectivity of the organism. Non-replicating persistent (NRP) M. tuberculosis was established by the gradual depletion of nutrients in an oxygen-replete and controlled environment. In contrast to rapidly dividing bacilli, NRP bacteria exhibited a distinct phenotype by accumulating an extracellular matrix rich in free mycolate and lipoglycans, with increased arabinosylation. Microarray studies demonstrated a substantial down-regulation of genes involved in energy metab. in NRP bacteria. Despite this redn. in metabolic activity, cells were still able to infect guinea pigs, but with a delay in the development of disease when compared to exponential phase bacilli. Using these approaches to investigate the interplay between the changing environment of the host and altered physiol. of NRP bacteria, this study sheds new light on the conditions that are pertinent to M. tuberculosis dormancy and how this organism could be establishing latent disease.
- 65Rose, J. D.; Maddry, J. A.; Comber, R. N.; Suling, W. J.; Wilson, L. N.; Reynolds, R. C. Synthesis and biological evaluation of trehalose analogs as potential inhibitors of mycobacterial cell wall biosynthesis. Carbohydr. Res. 2002, 337, 105– 120, DOI: 10.1016/S0008-6215(01)00288-9Google Scholar65https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XptlCjsg%253D%253D&md5=84944b2ab25eb9763377b537c6515a29Synthesis and biological evaluation of trehalose analogs as potential inhibitors of mycobacterial cell wall biosynthesisRose, Jerry D.; Maddry, Joseph A.; Comber, Robert N.; Suling, William J.; Wilson, Larry N.; Reynolds, Robert C.Carbohydrate Research (2002), 337 (2), 105-120CODEN: CRBRAT; ISSN:0008-6215. (Elsevier Science Ltd.)Analogs of trehalose are reported that were designed to interfere with mycolylation pathways in the mycobacterial cell wall. Several derivs. of 6,6'-dideoxytrehalose, including N,N'-dialkylamino and 6,6'-bis(sulfonamido) analogs, were prepd. and evaluated for antimycobacterial activity against Mycobacterium tuberculosis H37Ra and a panel of clin. isolates of Mycobacterium avium. 6,6'-Diaminotrehalose and its diazido precursor were both inactive, but significant activity apparently related to aliph. chain length was found among the sulfonamides, N-alkylamines, and one of the amidines.
- 66Barry, C. S.; Backus, K. M.; Barry, C. E.; Davis, B. G. ESI-MS Assay of M. tuberculosis Cell Wall Antigen 85 Enzymes Permits Substrate Profiling and Design of a Mechanism-Based Inhibitor. J. Am. Chem. Soc. 2011, 133, 13232– 13235, DOI: 10.1021/ja204249pGoogle Scholar66https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXpvFSnsbc%253D&md5=8d09f08160f78d634a08e87a0b516babESI-MS Assay of M. tuberculosis Cell Wall Antigen 85 Enzymes Permits Substrate Profiling and Design of a Mechanism-Based InhibitorBarry, Conor S.; Backus, Keriann M.; Barry, Clifton E., III; Davis, Benjamin G.Journal of the American Chemical Society (2011), 133 (34), 13232-13235CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)Mycobacterium tuberculosis Antigen 85 enzymes are vital to the integrity of the highly impermeable cell envelope and are potential therapeutic targets. Kinetic anal. using a label-free assay revealed both mechanistic details and a substrate profile that allowed the design and construction of a selective in vitro mechanism-based inhibitor.
- 67Viljoen, A.; Richard, M.; Nguyen, P. C.; Fourquet, P.; Camoin, L.; Paudal, R. R.; Gnawali, G. R.; Spilling, C. D.; Cavalier, J.-F.; Canaan, S.; Blaise, M.; Kremer, L. Cyclipostins and cyclophostin analogs inhibit the antigen 85C from Mycobacterium tuberculosis both in vitro and in vivo. J. Biol. Chem. 2018, 293, 2755– 2769, DOI: 10.1074/jbc.RA117.000760Google Scholar67https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXjt1Shsro%253D&md5=50d705aacc7e65e5b0fb448305c4e160Cyclipostins and cyclophostin analogs inhibit the antigen 85C from Mycobacterium tuberculosis both in vitro and in vivoViljoen, Albertus; Richard, Matthias; Nguyen, Phuong Chi; Fourquet, Patrick; Camoin, Luc; Paudal, Rishi R.; Gnawali, Giri R.; Spilling, Christopher D.; Cavalier, Jean-Francois; Canaan, Stephane; Blaise, Mickael; Kremer, LaurentJournal of Biological Chemistry (2018), 293 (8), 2755-2769CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)An increasing prevalence of cases of drug-resistant tuberculosis requires the development of more efficacious chemotherapies. The authors previously reported the discovery of a new class of cyclipostins and cyclophostin (CyC) analogs exhibiting potent activity against Mycobacterium tuberculosis both in vitro and in infected macrophages. Competitive labeling/enrichment assays combined with MS have identified several serine or cysteine enzymes in lipid and cell wall metab. as putative targets of these CyC compds. These targets included members of the antigen 85 (Ag85) complex (i.e. Ag85A, Ag85B, and Ag85C), responsible for biosynthesis of trehalose dimycolate and mycolylation of arabinogalactan. Herein, the authors used biochem. and structural approaches to validate the Ag85 complex as a pharmacol. target of the CyC analogs. The authors found that CyC7β, CyC8β, and CyC17 bind covalently to the catalytic Ser124 residue in Ag85C; inhibit mycolyltransferase activity (i.e. the transfer of a fatty acid mol. onto trehalose); and reduce triacylglycerol synthase activity, a property previously attributed to Ag85A. Supporting these results, an x-ray structure of Ag85C in complex with CyC8β disclosed that this inhibitor occupies Ag85C's substrate-binding pocket. Importantly, metabolic labeling of M. tuberculosis cultures revealed that the CyC compds. impair both trehalose dimycolate synthesis and mycolylation of arabinogalactan. Overall, the study provides compelling evidence that CyC analogs can inhibit the activity of the Ag85 complex in vitro and in mycobacteria, opening the door to a new strategy for inhibiting Ag85. The high-resoln. crystal structure obtained will further guide the rational optimization of new CyC scaffolds with greater specificity and potency against M. tuberculosis.
- 68Crespo, M.; Martinez, M.; de Pablo, E. Activation volumes for intramolecular oxidative C–X (X = H, F, Cl or Br) addition to platinum(II) imine complexes as a proof of the intimate mechanism. J. Chem. Soc., Dalton Trans. 1997, 1231– 1236, DOI: 10.1039/a606872cGoogle Scholar68https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2sXivVCqsLc%253D&md5=6580b106746a62d72545615d45e26c27Activation volumes for intramolecular oxidative C-X (X = H, F, Cl or Br) addition to platinum(II) imine complexes as a proof of the intimate mechanismCrespo, Margarita; Martinez, Manuel; de Pablo, EstherJournal of the Chemical Society, Dalton Transactions: Inorganic Chemistry (1997), (7), 1231-1235CODEN: JCDTBI; ISSN:0300-9246. (Royal Society of Chemistry)The kinetics of C-X (X = H, F, Cl or Br) bond activation of ring-substituted, PhCH:NCH2Ph, type imines via intramol. oxidative addn. to Pt(II) complexes was studied in acetone and toluene soln. at different temps. and pressures. Although the activation parameters detd. are within the range expected, the latter is extremely lage (ΔH⧧ from 25 to 70 kJ mol-1, ΔS⧧ from -220 to -45 J K-1 mol-1, ΔV⧧ from -31.2 to -9.5 cm3 mol-1). No differences were found for the reactions carried out in acetone or toluene, indicating that no polar transition state is formed during the reaction and that a common highly ordered three-centered C-Pt-X interaction is present for all the imines used. A good correlation was also obtained for the ΔS⧧ and ΔV⧧ values, independently of the solvent used, confirming the nonpolarity of the transition state. A deviation from this pattern is obsd. only for fluorinated imines both in acetone and toluene solns.; this result is interpreted by considering an earlier transition state for the oxidative addn. of C-F that has not yet produced an important vol. contraction of the Pt center despite the important spatial organization of the ligand, as shown by the very neg. values of ΔS⧧.
- 69Clark, P. W.; Dyke, S. F.; Smith, G.; Kennard, C. H. L. The cyclopalladation of benzylidenebenzylamines. J. Organomet. Chem. 1987, 330, 447– 460, DOI: 10.1016/S0022-328X(00)99057-0Google Scholar69https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL1cXksFCqsLc%253D&md5=587bb5c956cc1c516de4625448348c12The cyclopalladation of benzylidenebenzylaminesClark, P. W.; Dyke, S. F.; Smith, G.; Kennard, C. H. L.Journal of Organometallic Chemistry (1987), 330 (3), 447-60CODEN: JORCAI; ISSN:0022-328X.Cyclopalladation of benzylidenebenzylamines I (R = H; R1, R2, R3 = H, OMe; R4 = H, Me, Ph) with 1 equiv Li2PdCl4 gave dimers II (same R1-R4, X = Cl). Reaction of I (R = Br, same R1-R4) with bis(dibenzylideneacetone)palladium (III) gave II (X = Br, same R1-R4). The cyclopalladated products were characterized by 1H and 13C NMR spectroscopy and as their products derived from further reaction with acetylacetonate, PPh3, or NaOAc. Reinvestigation of the reaction of I (R - R4 = H) with Pd(OAc)2 gave the acetyl-substituted dimer IV, which reacted with CO to give phthalimidine V. Cyclopalladation of 2-BrC6H4CH2N:CR5Ph (R5 = H, Me, Ph) with III gave dimeric palladocycles which reacted with acetylacetonate to give monomers VI (same R5). The x-ray crystal structures of VI (R5 = H) and a monomeric acetylacetonate deriv. of II (R1-R4 = H) were detd. Competitive cyclopalladations of several N-6-bromobenzyl-6'-bromobenzalimines with III gave dimers VII (R6, R7 = H, OMe).
- 70Anderson, C. M.; Crespo, M.; Kfoury, N.; Weinstein, M. A.; Tanski, J. M. Regioselective C–H Activation Preceded by Csp2–Csp3 Reductive Elimination from Cyclometalated Platinum(IV) Complexes. Organometallics 2013, 32, 4199– 4207, DOI: 10.1021/om400398gGoogle Scholar70https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhtFOgtr%252FE&md5=9068798fd8cbe7aff054746d16192b96Regioselective C-H Activation Preceded by Csp2-Csp3 Reductive Elimination from Cyclometalated Platinum(IV) ComplexesAnderson, Craig M.; Crespo, Margarita; Kfoury, Nicole; Weinstein, Michael A.; Tanski, Joseph M.Organometallics (2013), 32 (15), 4199-4207CODEN: ORGND7; ISSN:0276-7333. (American Chemical Society)Reductive elimination reactions of the cyclometalated Pt(IV) compds. [PtMe2Cl{C6H4CH:NCH2(4-ClC6H4)}L] and [PtMe2Br{C6H4CH:NCH2Ph}L] (L = SMe2, PPh3) to form Csp3-Csp2 bonds, followed by either exclusive Csp2-H bond activation (L = SMe2) or competition between Csp2-H and Csp3-H bond activation (L = PPh3), are reported. Isomerization to give endo products instead of the expected exo complex was obsd. for the ligand C6H4CH:NCH2(2-BrC6H5), and formation of an endo six-membered platinacycle occurs for the ligand 2,4,6-Me3C6H2CH:NCH2(2-BrC6H4).
- 71Zhang, T.; Li, S.-Y.; Nuermberger, E. L. Autoluminescent Mycobacterium tuberculosis for Rapid, Real-Time, Non-Invasive Assessment of Drug and Vaccine Efficacy. PLoS One 2012, 7, e29774 DOI: 10.1371/journal.pone.0029774Google Scholar71https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsVCqsLY%253D&md5=9576cdfd3c6a058a826fd9736dca8b40Autoluminescent Mycobacterium tuberculosis for rapid, real-time, non-invasive assessment of drug and vaccine efficacyZhang, Tianyu; Li, Si-Yang; Nuermberger, Eric L.PLoS One (2012), 7 (1), e29774CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Preclin. efforts to discover and develop new drugs and vaccines for tuberculosis are hampered by the reliance on colony-forming unit (CFU) counts as primary outcomes for in vivo efficacy studies and the slow growth of Mycobacterium tuberculosis. The utility of bioluminescent M. tuberculosis reporter strains for real-time in vitro and ex vivo assessment of drug and vaccine activity has been demonstrated but a simple, non-invasive, real-time surrogate marker to replace CFU counts for real-time evaluation of drug and vaccine efficacy in vivo has not been described. We describe the development of a fully virulent and stable autoluminescent strain of M. tuberculosis and proof-of-concept expts. demonstrating its utility for in vivo bioluminescence imaging to assess the efficacy of new drugs and vaccines for tuberculosis in a mouse model. Relative light unit (RLU) counts paralleled CFU counts during the active phase of bacterial growth, with a lower limit of detection of approx. 106 CFU in live, anesthetized mice. Expts. distinguishing active from inactive anti-tuberculosis drugs and bacteriostatic drug effects from bactericidal effects were completed in less than 5 days. The ability of a recombinant BCG vaccine to limit bacterial growth was demonstrated within 3 wk. Use of this autoluminescent reporter strain has the potential to drastically reduce the time, effort, animals and costs consumed in the evaluation of drug activity in vitro and the in vivo assessment of drug and vaccine efficacy.
- 72Liu, Y.; Gao, Y.; Liu, J.; Tan, Y.; Liu, Z.; Chhotaray, C.; Jiang, H.; Lu, Z.; Chiwala, G.; Wang, S.; Makafe, G.; Islam, M. M.; Hameed, H. M. A.; Cai, X.; Wang, C.; Li, X.; Tan, S.; Zhang, T. The compound TB47 is highly bactericidal against Mycobacterium ulcerans in a Buruli ulcer mouse model. Nat. Commun. 2019, 10, 524, DOI: 10.1038/s41467-019-08464-yGoogle Scholar72https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXnt1Wks74%253D&md5=f1a01c719038fbbd9f9ce8972e33be27The compound TB47 is highly bactericidal against Mycobacterium ulcerans in a Buruli ulcer mouse modelLiu, Yang; Gao, Yamin; Liu, Jianxiong; Tan, Yaoju; Liu, Zhiyong; Chhotaray, Chiranjibi; Jiang, Huofeng; Lu, Zhili; Chiwala, Gift; Wang, Shuai; Makafe, Gaelle; Islam, Md. Mahmudul; Hameed, H. M. Adnan; Cai, Xingshan; Wang, Changwei; Li, Xinjie; Tan, Shouyong; Zhang, TianyuNature Communications (2019), 10 (1), 524CODEN: NCAOBW; ISSN:2041-1723. (Nature Research)Buruli ulcer (BU) is an emerging infectious disease that causes disfiguring skin ulcers. The causative agent, Mycobacterium ulcerans, secretes toxin called mycolactone that triggers inflammation and immunopathol. Existing treatments are lengthy and consist of drugs developed for tuberculosis. Here, we report that a pyrazolo[1,5-a]pyridine-3-carboxamide, TB47, is highly bactericidal against M. ulcerans both in vitro and in vivo. In the validated mouse model of BU, TB47 alone reduces M. ulcerans burden in mouse footpads by more than 2.5 log10 CFU compared to the std. BU treatment regimen recommended by the WHO. We show that mutations of ubiquinol-cytochrome C reductase cytochrome subunit B confer resistance to TB47 and the dissimilarity of CydABs from different mycobacteria may account for their differences in susceptibility to TB47. TB47 is highly potent against M. ulcerans and possesses desirable pharmacol. attributes and low toxicity that warrant further assessment of this agent for treatment of BU.
- 73Nateche, F.; Martin, A.; Baraka, S.; Palomino, J. C.; Khaled, S.; Portaels, F. Application of the resazurin microtitre assay for detection of multidrug resistance in Mycobacterium tuberculosis in Algiers. J. Med. Microbiol. 2006, 55, 857– 860, DOI: 10.1099/jmm.0.46513-0Google Scholar73https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD28zlvVCgsA%253D%253D&md5=152ee2d0bd954157d5c27338cdddf3e5Application of the resazurin microtitre assay for detection of multidrug resistance in Mycobacterium tuberculosis in AlgiersNateche Farida; Baraka Saliha; Khaled Safia; Nateche Farida; Martin Anandi; Palomino Juan Carlos; Portaels FrancoiseJournal of medical microbiology (2006), 55 (Pt 7), 857-860 ISSN:0022-2615.This study assessed the performance of a rapid, low-cost, colorimetric method, the resazurin microtitre assay (REMA) plate method, for the detection of resistance to isoniazid and rifampicin in 136 clinical isolates of Mycobacterium tuberculosis from two hospitals in Algiers. MICs were determined and the results were compared with those obtained with the conventional proportion method on Lowenstein-Jensen medium. Excellent results were obtained for the REMA plate method, with a sensitivity of 100 % for both isoniazid and rifampicin and a specificity of 98.3 and 99.2 %, respectively. The REMA plate method appears to be a reliable method for the rapid determination of multidrug-resistant tuberculosis and is a good alternative for use in resource-limited countries such as Algeria.
- 74Mosaei, H.; Molodtsov, V.; Kepplinger, B.; Harbottle, J.; Moon, C. W.; Jeeves, R. E.; Ceccaroni, L.; Shin, Y.; Morton-Laing, S.; Marrs, E. C. L.; Wills, C.; Clegg, W.; Yuzenkova, Y.; Perry, J. D.; Bacon, J.; Errington, J.; Allenby, N. E. E.; Hall, M. J.; Murakami, K. S.; Zenkin, N. Mode of Action of Kanglemycin A, an Ansamycin Natural Product that Is Active against Rifampicin-Resistant Mycobacterium tuberculosis. Mol. Cell 2018, 72, 263– 274.e5, DOI: 10.1016/j.molcel.2018.08.028Google Scholar74https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhslKhsr%252FF&md5=63369346e69244e37dd37f851734a0b8Mode of Action of Kanglemycin A, an Ansamycin Natural Product that Is Active against Rifampicin-Resistant Mycobacterium tuberculosisMosaei, Hamed; Molodtsov, Vadim; Kepplinger, Bernhard; Harbottle, John; Moon, Christopher William; Jeeves, Rose Elizabeth; Ceccaroni, Lucia; Shin, Yeonoh; Morton-Laing, Stephanie; Marrs, Emma Claire Louise; Wills, Corinne; Clegg, William; Yuzenkova, Yulia; Perry, John David; Bacon, Joanna; Errington, Jeff; Allenby, Nicholas Edward Ellis; Hall, Michael John; Murakami, Katsuhiko S.; Zenkin, NikolayMolecular Cell (2018), 72 (2), 263-274.e5CODEN: MOCEFL; ISSN:1097-2765. (Elsevier Inc.)Antibiotic-resistant bacterial pathogens pose an urgent healthcare threat, prompting a demand for new medicines. We report the mode of action of the natural ansamycin antibiotic kanglemycin A (KglA). KglA binds bacterial RNA polymerase at the rifampicin-binding pocket but maintains potency against RNA polymerases contg. rifampicin-resistant mutations. KglA has antibiotic activity against rifampicin-resistant Gram-pos. bacteria and multidrug-resistant Mycobacterium tuberculosis (MDR-M. tuberculosis). The X-ray crystal structures of KglA with the Escherichia coli RNA polymerase holoenzyme and Thermus thermophilus RNA polymerase-promoter complex reveal an altered-compared with rifampicin-conformation of KglA within the rifampicin-binding pocket. Unique deoxysugar and succinate ansa bridge substituents make addnl. contacts with a sep., hydrophobic pocket of RNA polymerase and preclude the formation of initial dinucleotides, resp. Previous ansa-chain modifications in the rifamycin series have proven unsuccessful. Thus, KglA represents a key starting point for the development of a new class of ansa-chain derivatized ansamycins to tackle rifampicin resistance.
- 75James, B. W.; Williams, A.; Marsh, P. D. The physiology and pathogenicity of Mycobacterium tuberculosis grown under controlled conditions in a defined medium. J. Appl. Microbiol. 2000, 88, 669– 77, DOI: 10.1046/j.1365-2672.2000.01020.xGoogle Scholar75https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXjtl2jurc%253D&md5=8f9a6b179536fbfdeca405fd9373acecThe physiology and pathogenicity of Mycobacterium tuberculosis grown under controlled conditions in a defined mediumJames, B. W.; Williams, A.; Marsh, P. D.Journal of Applied Microbiology (2000), 88 (4), 669-677CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)A chem.-defined culture medium was developed which supported batch growth of Mycobacterium tuberculosis, strain H37Rv, at a min. doubling time of 14.7 h. This medium also facilitated chemostat culture of M. tuberculosis at a const. doubling time of 24 h. Chemostat growth was optimized at a dissolved oxygen tension of 20% (vol./vol.) and 0.2% (vol./vol.) Tween-80. Chemostat cultures were dispersed suspensions of single bacilli (1.5-3 μm long), or small aggregates, at a mean d. of log10 8.3 cfu ml-1. A limited no. of amino acids was utilized (alanine, asparagine, aspartate and serine were depleted by > 50%; glycine, arginine, isoleucine, leucine and phenylalanine, by approx. 40%). Chemostat-grown cells were pathogenic in aerosol-infected guinea pigs, producing disseminated infection similar to that caused by plate-grown cells. Cells from chemostat culture were significantly more invasive for J774A.1 mouse macrophages than agar- or batch-grown cells. This study demonstrates the suitability of chemostat culture for the growth of pathogenic mycobacteria in a defined physiol. state with potential applications for the controlled prodn. of mycobacterial components for therapeutic and vaccine applications.
- 76Lambert, R. J. W. Susceptibility testing: inoculum size dependency of inhibition using the Colworth MIC technique. J. Appl. Microbiol. 2000, 89, 275– 279, DOI: 10.1046/j.1365-2672.2000.01105.xGoogle Scholar76https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXmvVCntLw%253D&md5=3390c45f9763d95dd0f0efffaa4f54eeSusceptibility testing: Inoculum size dependency of inhibition using the Colworth MIC techniqueLambert, R. J. W.Journal of Applied Microbiology (2000), 89 (2), 275-279CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)The min. inhibitory concn., MIC, is an accepted and well used criterion for measuring the susceptibility of organisms to inhibitors. Many factors influence the MIC value obtained, including temp., inoculum size and type of organism. A modification of the method developed in this lab. to obtain inhibition profiles of antimicrobials was used to examine the effect of inoculum size on the degree of inhibition obsd. with respect to inhibitor concn. The data obtained enabled the prodn. of an empirical model of inhibition, based on a Gompertz function, relating the level of growth obsd. to both the inoculum size and concn. of the inhibitor. The inoculum size dependencies of phenethyl alc., phenoxyethanol, p-chloro-m-cresol, trichloro-phenol, thymol and dodecyltrimethylammonium bromide against Staphylococcus aureus were obtained.
- 77Lambert, R. J.; Pearson, J. Susceptibility testing: accurate and reproducible minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) values. J. Appl. Microbiol. 2000, 88, 784– 90, DOI: 10.1046/j.1365-2672.2000.01017.xGoogle Scholar77https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXktVOhsr8%253D&md5=04b3d3382ed17f7f409388e2e4702309Susceptibility testing: accurate and reproducible minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) valuesLambert, R. J. W.; Pearson, J.Journal of Applied Microbiology (2000), 88 (5), 784-790CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)Measuring the min. inhibitory concn. (MIC) of a substance by current methods is straightforward, whereas obtaining useful comparative information from the tests can be more difficult. A simple technique and a method of data anal. are reported which give the experimentalist more useful information from susceptibility testing. This method makes use of a 100-well microtiter plate and the anal. uses all the growth information, obtained by turbidometry, from each and every well of the microtiter plate. A modified Gompertz function is used to fit the data, from which a more exact value can be obtained for the MIC. The technique also showed that at certain concns. of inhibitor, there was no effect on growth relative to a control well (zero inhibitor). Above a threshold value, which has been termed the non-inhibitory concn. or NIC, growth becomes limiting until it reaches the MIC, where no growth relative to the control is obsd.
- 78Luo, S.; Pal, D.; Shah, S. J.; Kwatra, D.; Paturi, K. D.; Mitra, A. K. Effect of HEPES Buffer on the Uptake and Transport of P-Glycoprotein Substrates and Large Neutral Amino Acids. Mol. Pharmaceutics 2010, 7, 412– 420, DOI: 10.1021/mp900193eGoogle Scholar78https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXisFGmtrw%253D&md5=5271b78dd7cf1b391a5d80eb9ac9cc91Effect of HEPES buffer on uptake and transport of P-glycoprotein substrates and large neutral amino acidsLuo, Shuanghui; Pal, Dhananjay; Shah, Sujay J.; Kwatra, Deep; Paturi, Kalyani D.; Mitra, Ashim K.Molecular Pharmaceutics (2010), 7 (2), 412-420CODEN: MPOHBP; ISSN:1543-8384. (American Chemical Society)HEPES has been widely employed as an org. buffer agent in cell culture medium as well as uptake and transport expts. in vitro. However, concns. of HEPES used in such studies vary from one lab. to another. In this study, we investigated the effect of HEPES on the uptake and bidirectional transport of P-gp substrates employing both Caco-2 and MDCK-MDR1 cells. ATP-dependent uptake of glutamic acid was also examd. ATP prodn. was further quantified applying ATP Detn. Kit. An addn. of HEPES to the growth and incubation media significantly altered the uptake and transport of P-gp substrates in both Caco-2 and MDCK-MDR1 cells. Uptake of P-gp substrates substantially diminished as the HEPES concn. was raised to 25 mM. Bidirectional (A-B and B-A) transport studies revealed that permeability ratio of PappB-A to PappA-B in the presence of 25 mM HEPES was significantly higher than control. The uptake of phenylalanine is an ATP-independent process, whereas the accumulation of glutamic acid is ATP-dependent. While phenylalanine uptake remained unchanged, glutamic acid uptake was elevated with the addn. of HEPES. Verapamil is an inhibitor of P-gp mediated uptake; elevation of cyclosporine uptake in the presence of 5 μM verapamil was compromised by the presence of 25 mM HEPES. The results of ATP assay indicated that HEPES stimulated the prodn. of ATP. This study suggests that the addn. of HEPES in the medium modulated the energy dependent efflux and uptake processes. The effect of HEPES on P-gp mediated drug efflux and transport may provide some mechanistic insight into possible reasons for inconsistencies in the results reported from various labs.
- 79Borgers, K.; Ou, J. Y.; Zheng, P. X.; Tiels, P.; Van Hecke, A.; Plets, E.; Michielsen, G.; Festjens, N.; Callewaert, N.; Lin, Y. C. Reference genome and comparative genome analysis for the WHO reference strain for Mycobacterium bovis BCG Danish, the present tuberculosis vaccine. BMC Genomics 2019, 20, 561, DOI: 10.1186/s12864-019-5909-5Google Scholar79https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3MzlsV2isw%253D%253D&md5=26ef669e165146e848a0916417d8532cReference genome and comparative genome analysis for the WHO reference strain for Mycobacterium bovis BCG Danish, the present tuberculosis vaccineBorgers Katlyn; Tiels Petra; Van Hecke Annelies; Plets Evelyn; Michielsen Gitte; Festjens Nele; Callewaert Nico; Borgers Katlyn; Tiels Petra; Van Hecke Annelies; Plets Evelyn; Michielsen Gitte; Festjens Nele; Callewaert Nico; Ou Jheng-Yang; Zheng Po-Xing; Lin Yao-Cheng; Ou Jheng-Yang; Zheng Po-Xing; Lin Yao-ChengBMC genomics (2019), 20 (1), 561 ISSN:.BACKGROUND: Mycobacterium bovis bacillus Calmette-Guerin (M. bovis BCG) is the only vaccine available against tuberculosis (TB). In an effort to standardize the vaccine production, three substrains, i.e. BCG Danish 1331, Tokyo 172-1 and Russia BCG-1 were established as the WHO reference strains. Both for BCG Tokyo 172-1 as Russia BCG-1, reference genomes exist, not for BCG Danish. In this study, we set out to determine the completely assembled genome sequence for BCG Danish and to establish a workflow for genome characterization of engineering-derived vaccine candidate strains. RESULTS: By combining second (Illumina) and third (PacBio) generation sequencing in an integrated genome analysis workflow for BCG, we could construct the completely assembled genome sequence of BCG Danish 1331 (07/270) (and an engineered derivative that is studied as an improved vaccine candidate, a SapM KO), including the resolution of the analytically challenging long duplication regions. We report the presence of a DU1-like duplication in BCG Danish 1331, while this tandem duplication was previously thought to be exclusively restricted to BCG Pasteur. Furthermore, comparative genome analyses of publicly available data for BCG substrains showed the absence of a DU1 in certain BCG Pasteur substrains and the presence of a DU1-like duplication in some BCG China substrains. By integrating publicly available data, we provide an update to the genome features of the commonly used BCG strains. CONCLUSIONS: We demonstrate how this analysis workflow enables the resolution of genome duplications and of the genome of engineered derivatives of the BCG Danish vaccine strain. The BCG Danish WHO reference genome will serve as a reference for future engineered strains and the established workflow can be used to enhance BCG vaccine standardization.
- 80Kapopoulou, A.; Lew, J. M.; Cole, S. T. The MycoBrowser portal: A comprehensive and manually annotated resource for mycobacterial genomes. Tuberculosis 2011, 91, 8– 13, DOI: 10.1016/j.tube.2010.09.006Google Scholar80https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXht1ylsLg%253D&md5=1e5f6644776728d23e5d00c313ae4b9bThe MycoBrowser portal: A comprehensive and manually annotated resource for mycobacterial genomesKapopoulou, Adamandia; Lew, Jocelyne M.; Cole, Stewart T.Tuberculosis (Oxford, United Kingdom) (2011), 91 (1), 8-13CODEN: TUBECU; ISSN:1472-9792. (Elsevier Ltd.)A review. Summary: In this paper, we present the MycoBrowser portal (http://mycobrowser.epfl.ch/), a resource that provides both in silico generated and manually reviewed information within databases dedicated to the complete genomes of Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium marinum and Mycobacterium smegmatis. A central component of MycoBrowser is TubercuList (http://tuberculist.epfl.ch), which has recently benefited from a new data management system and web interface. These improvements were extended to all MycoBrowser databases. We provide an overview of the functionalities available and the different ways of interrogating the data then discuss how both the new information and the latest features are helping the mycobacterial research communities.
- 81Abrahams, K. A.; Cox, J. A. G.; Fütterer, K.; Rullas, J.; Ortega-Muro, F.; Loman, N. J.; Moynihan, P. J.; Pérez-Herrán, E.; Jiménez, E.; Esquivias, J.; Barros, D.; Ballell, L.; Alemparte, C.; Besra, G. S. Inhibiting mycobacterial tryptophan synthase by targeting the inter-subunit interface. Sci. Rep. 2017, 7, 9430, DOI: 10.1038/s41598-017-09642-yGoogle Scholar81https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1cbhvVSqtw%253D%253D&md5=d5256bcc84a9c06221984116b25aee2eInhibiting mycobacterial tryptophan synthase by targeting the inter-subunit interfaceAbrahams Katherine A; Futterer Klaus; Loman Nicholas J; Moynihan Patrick J; Besra Gurdyal S; Cox Jonathan A G; Rullas Joaquin; Ortega-Muro Fatima; Perez-Herran Esther; Jimenez Elena; Esquivias Jorge; Barros David; Ballell Lluis; Alemparte CarlosScientific reports (2017), 7 (1), 9430 ISSN:.Drug discovery efforts against the pathogen Mycobacterium tuberculosis (Mtb) have been advanced through phenotypic screens of extensive compound libraries. Such a screen revealed sulfolane 1 and indoline-5-sulfonamides 2 and 3 as potent inhibitors of mycobacterial growth. Optimization in the sulfolane series led to compound 4, which has proven activity in an in vivo murine model of Mtb infection. Here we identify the target and mode of inhibition of these compounds based on whole genome sequencing of spontaneous resistant mutants, which identified mutations locating to the essential α- and β-subunits of tryptophan synthase. Over-expression studies confirmed tryptophan synthase as the biological target. Biochemical techniques probed the mechanism of inhibition, revealing the mutant enzyme complex incurs a fitness cost but does not prevent inhibitor binding. Mapping of the resistance conferring mutations onto a low-resolution crystal structure of Mtb tryptophan synthase showed they locate to the interface between the α- and β-subunits. The discovery of anti-tubercular agents inhibiting tryptophan synthase highlights the therapeutic potential of this enzyme and draws attention to the prospect of other amino acid biosynthetic pathways as future Mtb drug targets.
- 82Briffotaux, J.; Huang, W.; Wang, X.; Gicquel, B. MmpS5/MmpL5 as an efflux pump in Mycobacterium species. Tuberculosis 2017, 107, 13– 19, DOI: 10.1016/j.tube.2017.08.001Google Scholar82https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtlSrtrvP&md5=9d12bd4cd8051abd3cc695099ef6f899MmpS5/MmpL5 as an efflux pump in Mycobacterium speciesBriffotaux, Julien; Huang, Wei; Wang, Xinwei; Gicquel, BrigitteTuberculosis (Oxford, United Kingdom) (2017), 107 (), 13-19CODEN: TUBECU; ISSN:1472-9792. (Elsevier Ltd.)A review. Tuberculosis remains an important cause of morbidity and mortality throughout the world, amplified by the expansion of antibiotic resistance. Increasing active efflux of the antibiotic is one of the several strategies used by bacteria to resist to antibiotics. After showing the importance of the RND superfamily of efflux pumps in drug resistance, this review focuses on the protein MmpL5, a transmembrane transporter of Mycobacterium. These exporters should be involved in the variety of roles in bacterial cells, including expelling various drugs. The mutation in the transcriptional regulator, linked to the upregulation of MmpL5 can lead to resistance of antibiotics. The study of these mechanisms should be considered in order to improve the treatment of tuberculosis.
- 83Abrahams, K. A.; Cox, J. A. G.; Spivey, V. L.; Loman, N. J.; Pallen, M. J.; Constantinidou, C.; Fernández, R.; Alemparte, C.; Remuiñán, M. J.; Barros, D.; Ballell, L.; Besra, G. S. Identification of Novel Imidazo[1,2-a]pyridine Inhibitors Targeting M. tuberculosis QcrB. PLoS One 2012, 7, e52951 DOI: 10.1371/journal.pone.0052951Google Scholar83https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptVSgsQ%253D%253D&md5=4c00cd8ffb101ce9d40da5c0c4f51c0eIdentification of novel imidazo[1,2-a]pyridine inhibitors targeting M. tuberculosis QcrBAbrahams, Katherine A.; Cox, Jonathan A. G.; Spivey, Vickey L.; Loman, Nicholas J.; Pallen, Mark J.; Constantinidou, Chrystala; Fernandez, Raquel; Alemparte, Carlos; Remuinan, Modesto J.; Barros, David; Ballell, Lluis; Besra, Gurdyal S.PLoS One (2012), 7 (12), e52951CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Mycobacterium tuberculosis is a major human pathogen and the causative agent for the pulmonary disease, tuberculosis (TB). Current treatment programs to combat TB are under threat due to the emergence of multi-drug and extensively-drug resistant TB. Through the use of high throughput whole cell screening of an extensive compd. library a no. of imidazo[1,2-a]pyridine (IP) compds. were obtained as potent lead mols. active against M. tuberculosis and Mycobacterium bovis BCG. The IP inhibitors (1-4) demonstrated min. inhibitory concns. (MICs) in the range of 0.03 to 5 μM against a panel of M. tuberculosis strains. M. bovis BCG spontaneous resistant mutants were generated against IP 1, 3 and 4 at 5 × MIC and subsequent whole genome sequencing identified a single nucleotide polymorphism 937ACC>937GCC (T313A) in qcrB, which encodes the b subunit of the electron transport ubiquinol cytochrome C reductase. This mutation also conferred cross-resistance against IP 1, 3 and 4 demonstrating a common target. Gene dosage expts. confirmed M. bovis BCG QcrB as the target where over-expression in M. bovis BCG led to an increase in MIC from 0.5 to >8 μM for IP 3. An acute murine model of TB infection established bacteriostatic activity of the IP series, which await further detailed characterization.
- 84Gupta, R. S.; Lo, B.; Son, J. Phylogenomics and Comparative Genomic Studies Robustly Support Division of the Genus Mycobacterium into an Emended Genus Mycobacterium and Four Novel Genera. Front. Microbiol. 2018, 9, 67, DOI: 10.3389/fmicb.2018.00067Google Scholar84https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1MrmvFKmtA%253D%253D&md5=bd2862ca3414afb67e794631a0f1434aPhylogenomics and Comparative Genomic Studies Robustly Support Division of the Genus Mycobacterium into an Emended Genus Mycobacterium and Four Novel GeneraGupta Radhey S; Lo Brian; Son JeenFrontiers in microbiology (2018), 9 (), 67 ISSN:1664-302X.The genus Mycobacterium contains 188 species including several major human pathogens as well as numerous other environmental species. We report here comprehensive phylogenomics and comparative genomic analyses on 150 genomes of Mycobacterium species to understand their interrelationships. Phylogenetic trees were constructed for the 150 species based on 1941 core proteins for the genus Mycobacterium, 136 core proteins for the phylum Actinobacteria and 8 other conserved proteins. Additionally, the overall genome similarity amongst the Mycobacterium species was determined based on average amino acid identity of the conserved protein families. The results from these analyses consistently support the existence of five distinct monophyletic groups within the genus Mycobacterium at the highest level, which are designated as the "Tuberculosis-Simiae," "Terrae," "Triviale," "Fortuitum-Vaccae," and "Abscessus-Chelonae" clades. Some of these clades have also been observed in earlier phylogenetic studies. Of these clades, the "Abscessus-Chelonae" clade forms the deepest branching lineage and does not form a monophyletic grouping with the "Fortuitum-Vaccae" clade of fast-growing species. In parallel, our comparative analyses of proteins from mycobacterial genomes have identified 172 molecular signatures in the form of conserved signature indels and conserved signature proteins, which are uniquely shared by either all Mycobacterium species or by members of the five identified clades. The identified molecular signatures (or synapomorphies) provide strong independent evidence for the monophyly of the genus Mycobacterium and the five described clades and they provide reliable means for the demarcation of these clades and for their diagnostics. Based on the results of our comprehensive phylogenomic analyses and numerous identified molecular signatures, which consistently and strongly support the division of known mycobacterial species into the five described clades, we propose here division of the genus Mycobacterium into an emended genus Mycobacterium encompassing the "Tuberculosis-Simiae" clade, which includes all of the major human pathogens, and four novel genera viz. Mycolicibacterium gen. nov., Mycolicibacter gen. nov., Mycolicibacillus gen. nov. and Mycobacteroides gen. nov. corresponding to the "Fortuitum-Vaccae," "Terrae," "Triviale," and "Abscessus-Chelonae" clades, respectively. With the division of mycobacterial species into these five distinct groups, attention can now be focused on unique genetic and molecular characteristics that differentiate members of these groups.
- 85Neagoie, C. D.; Peng, X.; Tortorella, M. D.; Fossey, J. S.; Alderwick, L. J.; Feula, A.; Yoshizawa, A.. Preparation of antibacterial compounds: WO2020206594 A1 2020, 10– 15.Google ScholarThere is no corresponding record for this reference.
Cited By
This article has not yet been cited by other publications.
Article Views
Altmetric
Citations
Article Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.
Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.
The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated.
Recommended Articles
Abstract
Figure 1
Figure 1. BGAz derivatives synthesized.
Figure 2
Figure 2. Assessment of bactericidal activity of BGAz-004 and BGAz-005 against M. tuberculosis H37Rv. Average total viable counts (CFU mL–1) of M. tuberculosis cultures exposed to either BGAz-004 (Panel A) or BGAz-005 (Panel B) at concentrations: 0 μM (0.1% DMSO) (circle, closed), 3, 6, 12, 24, 48, and 96 μM or isoniazid (Panel C) at concentrations 0 μM (0.1% DMSO), 0.9, 1.8, 3.7, 7.3, 14.6, and 29.2 μM over a 14-day time-course. Samples were taken after 0, 2, 6, 10, and 14 days of antibiotic exposure, serially diluted, and plated by the method of Miles et al. (27) Statistical comparisons were performed at 6, 10, and 14 days of antibiotic exposure at 96 μM BGAz-004 and BGAz-005 using factorial ANOVA and posthoc Tukey’s honestly significant difference test (Panel D). Data represent three biological repeats ± standard deviation.
Figure 3
Figure 3. Assessment of bactericidal activity of BGAz-004 and BGAz-005 against M. tuberculosis H37Rv. Quantitation of Calcien-Violet-AM (CV-AM) and Sytox-green (SG) fluorescence of M. tuberculosis H37Rv, using flow cytometry, after exposure to BGAz-004 (column A) and BGAz-005 (column B) at concentrations: 0 μM (0.1% DMSO), 3, 6, 12, 24, 48, and 96 μM or (column C) isoniazid at concentrations 0 μM (0.1% DMSO), 0.9, 1.8, 3.7, 7.3, 14.6, and 29.2 μM over a 14-day time-course. The percentages of the population that are unstained or stained with each dye (or both dyes) are represented in four gates (rows P1–P4). Row P1: unstained population (CV-AM–/SG–); row P2: CV-stained population (CV-AM+/SG–); row P3: dual-stained population (CV-AM+/SG+); and row P4: SG-stained population (CV-AM–/SG+). Data represent three biological repeats ± standard deviation. Statistical comparisons were made using factorial ANOVA and posthoc Tukey’s honestly significant difference test.
Figure 4
Figure 4. Effect of BGAz-005 on the incorporation of radiolabeled precursors into the major cellular macromolecules of M. smegmatis. The incorporation of (A) [methyl-3H]thymidine (for DNA), (B) [5,6-3H]uridine (for RNA), (C) l-[4,5-3H]leucine (for protein), (D) [3H]meso-diaminopimelic acid (for peptidoglycan), and (E) [14C]acetic acid (for lipids) was measured over a period of 36 h. The percentage of incorporation measured at 36 h is represented in panel F. Each plot and error bars represent the average of three independent experiments.
Figure 5
Figure 5. Transcriptional response to BGAz-005 exposure demonstrating inhibition of mycobacterial cell envelope biosynthesis. (A) Cluster diagram of all genes showing similarity of biological replicates and separation of drug-treated from carrier control samples. (B) Volcano plot of M. bovis BCG response to BGAz-005, highlighting genes significantly differentially expressed. (C) Heatmap of 286 gene BGAz-005 signature relative to carrier control. Conditions as columns, genes as rows; red coloring highlighting induced genes and blue representing repressed genes. The BGAz-004 signature is clustered alongside, indicating a similar mode of drug action.
Figure 6
Figure 6. BCG cell envelope lipid analysis upon exposure to BGAz-005. BCG were cultured in 7H9 broth and exposed to increasing concentrations of BGAz-005. Lipids were selectively labeled with [14C]-acetic acid for 12 h, and cell envelope lipids were selectively removed by solvent extraction, separated by TLC (chloroform/methanol/water, 80:20:2, v/v/v), and visualized by autoradiography. (A) Equal volumes of lipids loaded adjusted for BCG growth; (B) equal counts of lipids (25,000 cpm) loaded; (C) mycolic acid methyl ester (MAME) analysis of cell-wall bound mycolates released by 5% TBAH and separated by TLC (petroleum ether/acetone, 95:5, v/v); (D) quantification of BCG lipids from panels A–C by densitometry. M. smegmatis cell envelope lipid analysis upon exposure to BGAz-005. (E) M. smegmatis were cultured in 7H9 broth, exposed to increasing concentrations of BGAz-005 for 6 h and the cell envelope lipids selectively removed by solvent extraction. Equal volumes of lipid adjusted by bacterial growth were separated by TLC (chloroform/methanol/water, 80:20:2, v/v/v) and stained with MPA or (F) alpha-naphthol. (G) Equal volumes of lipid adjusted by bacterial growth were separated by TLC (hexane/diethyl ether/acetic acid), 70:30:1, v/v/v and stained with MPA. (H) Equal volumes of lipid adjusted by bacterial growth were separated by 2D-TLC (direction 1 chloroform/methanol 96:4, v/v, direction 2 toluene/acetone 80:20, v/v) and stained with MPA.
Figure 7
Figure 7. Assessing the MIC shift of BGAz-004 and BGAz-005 against the AG85 complex. MIC values of BGAz-004, BGAz-005, and Ebselen were determined against BCG harboring overexpression vectors and compared to empty vector controls (pTIC6) in order to identify a shift in MIC against fbpA (A), fbpB (B), and fbpC (C). Fold change in MIC shift (D). The MIC99 was calculated using an end point resazurin assay and the Gomperz equation for MIC determination (GraphPad Prism). Data are of triplicate repeats.
References
This article references 85 other publications.
- 1www.who.int/gho/tb/en/. World Health Organisation: Global Health Observatory (GHO) data www.who.int/gho/tb/en/.There is no corresponding record for this reference.
- 2Global tuberculosis report 2022; World Health Organization: Geneva, 2022, Licence: CC BY-NC-SA 3.0 IGO.There is no corresponding record for this reference.
- 3Combs, D. L.; O’Brien, R. J.; Geiter, L. J. USPHS Tuberculosis Short-Course Chemotherapy Trial 21: Effectiveness, Toxicity, and Acceptability: The Report of Final Results. Ann. Int. Med. 1990, 112, 397– 406, DOI: 10.7326/0003-4819-76-3-112-6-3973https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADyaK3c7nvVyqtg%253D%253D&md5=2df74050189541e5d5b92bbb5acde939USPHS Tuberculosis Short-Course Chemotherapy Trial 21: effectiveness, toxicity, and acceptability. The report of final resultsCombs D L; O'Brien R J; Geiter L JAnnals of internal medicine (1990), 112 (6), 397-406 ISSN:0003-4819.STUDY OBJECTIVE: To determine the effectiveness, toxicity, and acceptability of a 6-month antituberculous regimen compared with a 9-month regimen. DESIGN: A nonblinded, unbalanced, randomized, multicenter clinical trial. SETTING: Twenty-two tuberculosis clinics in public health departments and hospitals in the United States. PATIENTS: Patients were eligible if Mycobacterium tuberculosis, isolated from sputum cultures, was susceptible to study drugs. Of 1451 patients enrolled, 75% (617 of 823) assigned to the 6-month regimen and 71% (445 of 628) assigned to the 9-month regimen were eligible. INTERVENTIONS: Patients took self-administered isoniazid and rifampin daily for 24 weeks (6-month regimen) or 36 weeks (9-month regimen). In addition, patients assigned to the 6-month regimen took self-administered pyrazinamide daily during the first 8 weeks. RESULTS: Patients on the 6-month regimen converted more rapidly than patients on the 9-month regimen (94.6% compared with 89.9% after 16 weeks of therapy, with a difference of 4.7% [95% CI, 0.7% to 8.7%]); had similar rates of adverse drug reactions (7.7% compared with 6.4%, with a difference of 1.3% [95% CI, 0.0% to 4.6%]); had lower noncompliance rates (16.8% compared with 29.2%, with a difference of 12.4% [95% CI, 6.8% to 18.0%]); and had similar relapse rates 96 weeks after completing therapy (3.5% compared with 2.8%, with a difference of 0.7% [95% CI, 0.0% to 3.9%]). A significantly greater proportion of patients assigned to the 6-month regimen successfully completed therapy (61.4% compared with 50.6%; chi 2 = 11.976). CONCLUSIONS: Our results suggest that this 6-month regimen is similar in effectiveness, toxicity, and acceptability to the 9-month regimen for treating pulmonary tuberculosis.
- 4Shah, N. S.; Wright, A.; Bai, G. H.; Barrera, L.; Boulahbal, F.; Martin-Casabona, N.; Drobniewski, F.; Gilpin, C.; Havelkova, M.; Lepe, R.; Lumb, R.; Metchock, B.; Portaels, F.; Rodrigues, M. F.; Rusch-Gerdes, S.; Van Deun, A.; Vincent, V.; Laserson, K.; Wells, C.; Cegielski, J. P. Worldwide emergence of extensively drug-resistant tuberculosis. Emerg. Infect. Dis. 2007, 13, 380– 7, DOI: 10.3201/eid1303.0614004https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXjvVOksbs%253D&md5=dd37e95733ca0384716c0874924d151dWorldwide emergence of extensively drug-resistant tuberculosisShah, N. Sarita; Wright, Abigail; Bai, Gill-Han; Barrera, Lucia; Boulahbal, Fadila; Martin-Casabona, Nuria; Drobniewski, Francis; Gilpin, Chris; Havelkova, Marta; Lepe, Rosario; Lumb, Richard; Metchock, Beverly; Portaels, Francoise; Rodrigues, Maria Filomena; Rusch-Gerdes, Sabine; Van Deun, Armand; Vincent, Veronique; Laserson, Kayla; Wells, Charles; Cegielski, J. PeterEmerging Infectious Diseases (2007), 13 (3), 380-387CODEN: EIDIFA; ISSN:1080-6040. (National Center for Infectious Diseases, Centers for Disease Control and Prevention)Mycobacterium tuberculosis strains that are resistant to an increasing no. of second-line drugs used to treat multidrug-resistant tuberculosis (MDR TB) are becoming a threat to public health worldwide. We surveyed the Network of Supranational Ref. Labs. for M. tuberculosis isolates that were resistant to second-line anti-TB drugs during 2000-2004. We defined extensively drug-resistant TB (XDR TB) as MDR TB with further resistance to ≥3 of the 6 classes of second-line drugs. Of 23 eligible labs., 14 (61%) contributed data on 17,690 isolates, which reflected drug susceptibility results from 48 countries. Of 3,520 (19.9%) MDR TB isolates, 347 (9.9%) met criteria for XDR TB. Further investigation of population-based trends and expanded efforts to prevent drug resistance and effectively treat patients with MDR TB are crucial for protection of public health and control of TB.
- 5Sharma, S. K.; Mohan, A. Multidrug-Resistant Tuberculosis: A Menace That Threatens To Destabilize Tuberculosis Control. Chest 2006, 130, 261– 272, DOI: 10.1016/S0012-3692(15)50981-15https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28Xot1KjtLc%253D&md5=f61cec249a7ad13d4ff64b444510a740Multidrug-resistant tuberculosis: a menace that threatens to destabilize tuberculosis controlSharma, Surendra K.; Mohan, AlladiChest (2006), 130 (1), 261-272CODEN: CHETBF; ISSN:0012-3692. (American College of Chest Physicians)A review. Multidrug-resistant tuberculosis (MDR-TB), caused by Mycobacterium tuberculosis that is resistant to both isoniazid and rifampicin with or without resistance to other drugs, is a phenomenon that is threatening to destabilize global tuberculosis (TB) control. MDR-TB is a worldwide problem, being present virtually in all countries that were surveyed. According to current World Health Organization and the International Union Against Tuberculosis and Lung Disease ests., the median prevalence of MDR-TB has been 1.1% in newly diagnosed patients. The proportion, however, is considerably higher (median prevalence, 7%) in patients who have previously received anti-TB treatment. While host genetic factors may contribute to the development of MDR-TB, incomplete and inadequate treatment is the most important factor leading to its development, suggesting that it is often a man made tragedy. Efficiently run TB control programs based on a policy of directly obsd. treatment, short course (DOTS), are essential for preventing the emergence of MDR-TB. The management of MDR-TB is a challenge that should be undertaken by experienced clinicians at centers equipped with reliable lab. services for mycobacterial cultures and in vitro sensitivity testing as it the requires prolonged use of costly second-line drugs with a significant potential for toxicity. The judicious use of drugs; supervised standardized treatment; focused clin., radiol., and bacteriol. follow-up; and surgery at the appropriate juncture are key factors in the successful management of these patients. With newer effective anti-TB drugs still a distant dream, innovative approaches such as DOTS-Plus are showing promise for the management of patients with MDR-TB under program conditions and appear to be a hope for future.
- 6Zumla, A.; Nahid, P.; Cole, S. T. Advances in the development of new tuberculosis drugs and treatment regimens. Nat. Rev. Drug Discovery 2013, 12, 388– 404, DOI: 10.1038/nrd40016https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXms1Kmsrk%253D&md5=fbf12f867a19ffff4d7fb74a98699682Advances in the development of new tuberculosis drugs and treatment regimensZumla, Alimuddin; Nahid, Payam; Cole, Stewart T.Nature Reviews Drug Discovery (2013), 12 (5), 388-404CODEN: NRDDAG; ISSN:1474-1776. (Nature Publishing Group)A review. Despite the introduction 40 years ago of the inexpensive and effective four-drug (isoniazid, rifampicin, pyrazinamide and ethambutol) treatment regimen, tuberculosis (TB) continues to cause considerable morbidity and mortality worldwide. For the first time since the 1960s, new and novel drugs and regimens for all forms of TB are emerging. Such regimens are likely to utilize both repurposed drugs and new chem. entities, and several of these regimens are now progressing through clin. trials. This article covers current concepts and recent advances in TB drug discovery and development, including an update of ongoing TB treatment trials, newer clin. trial designs, TB biomarkers and adjunct host-directed therapies.
- 7Ma, Z.; Lienhardt, C.; McIlleron, H.; Nunn, A. J.; Wang, X. Global tuberculosis drug development pipeline: the need and the reality. Lancet 2010, 375, 2100– 2109, DOI: 10.1016/S0140-6736(10)60359-97https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3cvpsl2msA%253D%253D&md5=893d7a51ed206dca3cfb1f7a2c3de77fGlobal tuberculosis drug development pipeline: the need and the realityMa Zhenkun; Lienhardt Christian; McIlleron Helen; Nunn Andrew J; Wang XiexiuLancet (London, England) (2010), 375 (9731), 2100-9 ISSN:.Drugs for tuberculosis are inadequate to address the many inherent and emerging challenges of treatment. In the past decade, ten compounds have progressed into the clinical development pipeline, including six new compounds specifically developed for tuberculosis. Despite this progress, the global drug pipeline for tuberculosis is still insufficient to address the unmet needs of treatment. Additional and sustainable efforts, and funding are needed to further improve the pipeline. The key challenges in the development of new treatments are the needs for novel drug combinations, new trial designs, studies in paediatric populations, increased clinical trial capacity, clear regulatory guidelines, and biomarkers for prediction of long-term outcome. Despite substantial progress in efforts to control tuberculosis, the global burden of this disease remains high. To eliminate tuberculosis as a public health concern by 2050, all responsible parties need to work together to strengthen the global antituberculosis drug pipeline and support the development of new antituberculosis drug regimens.
- 8Sharma, A.; De Rosa, M.; Singla, N.; Singh, G.; Barnwal, R. P.; Pandey, A. Tuberculosis: An Overview of the Immunogenic Response, Disease Progression, and Medicinal Chemistry Efforts in the Last Decade toward the Development of Potential Drugs for Extensively Drug-Resistant Tuberculosis Strains. J. Med. Chem. 2021, 64, 4359– 4395, DOI: 10.1021/acs.jmedchem.0c018338https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3MXotlCrurw%253D&md5=6f8bf51a6e6ab45614f311d81379a616Tuberculosis: An Overview of the Immunogenic Response, Disease Progression, and Medicinal Chemistry Efforts in the Last Decade toward the Development of Potential Drugs for Extensively Drug-Resistant Tuberculosis StrainsSharma, Akanksha; De Rosa, Maria; Singla, Neha; Singh, Gurpal; Barnwal, Ravi P.; Pandey, AnkurJournal of Medicinal Chemistry (2021), 64 (8), 4359-4395CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)A review. Tuberculosis (TB) is a slow growing, potentially debilitating disease that has plagued humanity for centuries and has claimed numerous lives across the globe. Concerted efforts by researchers have culminated in the development of various strategies to combat this malady. This review aims to raise awareness of the rapidly increasing incidences of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis, highlighting the significant modifications that were introduced in the TB treatment regimen over the past decade. A description of the role of pathogen-host immune mechanisms together with strategies for prevention of the disease is discussed. The struggle to develop novel drug therapies has continued in an effort to reduce the treatment duration, improve patient compliance and outcomes, and circumvent TB resistance mechanisms. Herein, we give an overview of the extensive medicinal chem. efforts made during the past decade toward the discovery of new chemotypes, which are potentially active against TB-resistant strains.
- 9Raymer, B.; Bhattacharya, S. K. Lead-like Drugs: A Perspective. J. Med. Chem. 2018, 61, 10375– 10384, DOI: 10.1021/acs.jmedchem.8b004079https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhtl2is73O&md5=e9fc17d69c7d671459fdaf77d6721829Lead-like Drugs: A PerspectiveRaymer, Brian; Bhattacharya, Samit K.Journal of Medicinal Chemistry (2018), 61 (23), 10375-10384CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)Lead-like drugs, or drugs below mol. wt. 300, are an important and sometimes overlooked component of the current pharmacopeia and contemporary medicinal chem. practice. To examine the recent state-of-the-art in lead-like drug discovery, we surveyed recent drug approvals from 2011 to 2017 and top 200 prescribed medications, as well as provide case studies on recently approved lead-like drugs. Many of these recent drugs are close analogs of previously known drugs or natural substrates, with a key focus of their medicinal chem. optimization being the choice of a low mol. wt. starting point and maintaining low mol. wt. during the optimization. However, the identification of low mol. wt. starting points may be limited by the availability of suitable low mol. wt. screening sets. To increase the discovery rate of lead-like drugs, we suggest an increased focus on inclusion and prosecution of lead-like starting points in screening libraries.
- 10Lovering, F. Escape from Flatland 2: complexity and promiscuity. MedChemComm 2013, 4, 515– 519, DOI: 10.1039/c2md20347b10https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXjtFShtL0%253D&md5=c03d3b99da22a684cbbbb00fb1633342Escape from Flatland 2: complexity and promiscuityLovering, FrankMedChemComm (2013), 4 (3), 515-519CODEN: MCCEAY; ISSN:2040-2503. (Royal Society of Chemistry)Toxicity plays a major role in attrition in the clinic and promiscuity has been linked to toxicity. A no. of mol. descriptors have been identified that contribute to promiscuity including ionization and logP. In this study we report on the relationship between complexity, as measured by two descriptors [fraction sp3 (Fsp3) where Fsp3 = (no. of sp3 hybridized carbons/total carbon count) and chiral carbon count], and promiscuity as well as Cyp450 inhibition. We find that increasing complexity reduces promiscuity and Cyp450 inhibition. As an understanding of key property descriptors has helped the pharmaceutical industry to address some of the deficiencies of compds. as pertains to bioavailability, awareness of the descriptors that impact promiscuity should allow us to better address toxicity in the clinic.
- 11Lovering, F.; Bikker, J.; Humblet, C. Escape from Flatland: Increasing Saturation as an Approach to Improving Clinical Success. J. Med. Chem. 2009, 52, 6752– 6756, DOI: 10.1021/jm901241e11https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXht1KjtLvN&md5=4ca92c30c17c53d77ad376719bad951eEscape from Flatland: Increasing Saturation as an Approach to Improving Clinical SuccessLovering, Frank; Bikker, Jack; Humblet, ChristineJournal of Medicinal Chemistry (2009), 52 (21), 6752-6756CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)The medicinal chem. community has become increasingly aware of the value of tracking calcd. phys. properties such as mol. wt., topol. polar surface area, rotatable bonds, and hydrogen bond donors and acceptors. The authors hypothesized that the shift to high-throughput synthetic practices over the past decade may be another factor that may predispose mols. to fail by steering discovery efforts toward achiral, arom. compds. The authors have proposed two simple and interpretable measures of the complexity of mols. prepd. as potential drug candidates. The first is carbon bond satn. as defined by fraction Sp3 (Fsp3) where Fsp3 = (no. of Sp3 hybridized carbons/total carbon count). The second is simply whether a chiral carbon exists in the mol. The authors demonstrate that both complexity (as measured by Fsp3) and the presence of chiral centers correlate with success as compds. transition from discovery, through clin. testing, to drugs. To explain these observations, the authors further demonstrate that satn. correlates with soly., an exptl. phys. property important to success in the drug discovery setting.
- 12Yang, Y.; Chen, H.; Nilsson, I.; Muresan, S.; Engkvist, O. Investigation of the Relationship between Topology and Selectivity for Druglike Molecules. J. Med. Chem. 2010, 53, 7709– 7714, DOI: 10.1021/jm100845612https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXht1Oqtb3L&md5=46f7582bfa567734f0391512e2580717Investigation of the Relationship between Topology and Selectivity for Drug-like MoleculesYang, Yidong; Chen, Hongming; Nilsson, Ingemar; Muresan, Sorel; Engkvist, OlaJournal of Medicinal Chemistry (2010), 53 (21), 7709-7714CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)There is a strong interest in drug discovery and development to advance the understanding of pharmacol. promiscuity. Improved understanding of how a mol. structure is related to promiscuity could help to reduce the attrition of compds. in the drug discovery process. For this purpose, a descriptor is introduced that describes the structural complexity of a compd. based on the size of its mol. framework (MF) in relation to its overall size. It is defined as the fraction of the size of the mol. framework vs. the size of the whole mol. (fMF). It is demonstrated that promiscuity correlates with fMF for large fMF values. The obsd. correlation is not due to lipophilicity. To provide further explanation of this observation, it was found that the no. of terminal ring systems in a compd. is correlated with promiscuity. The anal. presented here might help medicinal chemists to improve the selectivity for compds. in drug discovery projects.
- 13Baell, J. B.; Holloway, G. A. New Substructure Filters for Removal of Pan Assay Interference Compounds (PAINS) from Screening Libraries and for Their Exclusion in Bioassays. J. Med. Chem. 2010, 53, 2719– 2740, DOI: 10.1021/jm901137j13https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXhsF2qsLw%253D&md5=fbf397aa4910753c550425708c866fd2New Substructure Filters for Removal of Pan Assay Interference Compounds (PAINS) from Screening Libraries and for Their Exclusion in BioassaysBaell, Jonathan B.; Holloway, Georgina A.Journal of Medicinal Chemistry (2010), 53 (7), 2719-2740CODEN: JMCMAR; ISSN:0022-2623. (American Chemical Society)This report describes a no. of substructural features which can help to identify compds. that appear as frequent hitters (promiscuous compds.) in many biochem. high throughput screens. The compds. identified by such substructural features are not recognized by filters commonly used to identify reactive compds. Even though these substructural features were identified using only one assay detection technol., such compds. have been reported to be active from many different assays. In fact, these compds. are increasingly prevalent in the literature as potential starting points for further exploration, whereas they may not be.
- 14Baell, J. B.; Nissink, J. W. M. Seven Year Itch: Pan-Assay Interference Compounds (PAINS) in 2017─Utility and Limitations. ACS Chem. Biol. 2018, 13, 36– 44, DOI: 10.1021/acschembio.7b0090314https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhvFSlu7bK&md5=051115704137d6a91b8271702c619682Seven Year Itch: Pan-Assay Interference Compounds (PAINS) in 2017-Utility and LimitationsBaell, Jonathan B.; Nissink, J. Willem M.ACS Chemical Biology (2018), 13 (1), 36-44CODEN: ACBCCT; ISSN:1554-8929. (American Chemical Society)A review on all necessary considerations for the appropriate use of PAINs filters formulated to process hundreds and thousands of compds. in seconds, and identify PAINS in order to exclude them from further anal.
- 15Colomer, I.; Empson, C. J.; Craven, P.; Owen, Z.; Doveston, R. G.; Churcher, I.; Marsden, S. P.; Nelson, A. A divergent synthetic approach to diverse molecular scaffolds: assessment of lead-likeness using LLAMA, an open-access computational tool. Chem. Commun. 2016, 52, 7209– 7212, DOI: 10.1039/C6CC03244C15https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XnsVWqtrs%253D&md5=6f16f9da33bfc182d98c56065e8d3951A divergent synthetic approach to diverse molecular scaffolds: assessment of lead-likeness using LLAMA, an open-access computational toolColomer, Ignacio; Empson, Christopher J.; Craven, Philip; Owen, Zachary; Doveston, Richard G.; Churcher, Ian; Marsden, Stephen P.; Nelson, AdamChemical Communications (Cambridge, United Kingdom) (2016), 52 (45), 7209-7212CODEN: CHCOFS; ISSN:1359-7345. (Royal Society of Chemistry)Complementary cyclization reactions of hex-2-ene-1,6-diamine derivs. were exploited in the synthesis of alternative pyrrolidine, oxazolidinone, pyrimidinone and oxazinone mol. scaffolds. The value of the synthetic approach was analyzed using LLAMA, an open-access computational tool for assessing the lead-likeness and novelty of mol. scaffolds.
- 16
Synthetic chemistry methodology projects within the School of Chemistry at the University of Birmingham initially provided ∼ 200 compounds suitable for biological screening.
There is no corresponding record for this reference. - 17Feula, A.; Dhillon, S. S.; Byravan, R.; Sangha, M.; Ebanks, R.; Hama Salih, M. A.; Spencer, N.; Male, L.; Magyary, I.; Deng, W.-P.; Müller, F.; Fossey, J. S. Synthesis of azetidines and pyrrolidines via iodocyclisation of homoallyl amines and exploration of activity in a zebrafish embryo assay. Org. Biomol. Chem. 2013, 11, 5083– 5093, DOI: 10.1039/c3ob41007b17https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhtFaltb%252FE&md5=3538313a5c37148cc75e5039a1c34540Synthesis of azetidines and pyrrolidines via iodocyclisation of homoallyl amines and exploration of activity in a zebrafish embryo assayFeula, Antonio; Dhillon, Sundeep S.; Byravan, Rama; Sangha, Mandeep; Ebanks, Ronald; Hama Salih, Mariwan A.; Spencer, Neil; Male, Louise; Magyary, Istvan; Deng, Wei-Ping; Mueller, Ferenc; Fossey, John S.Organic & Biomolecular Chemistry (2013), 11 (31), 5083-5093CODEN: OBCRAK; ISSN:1477-0520. (Royal Society of Chemistry)Room temp. iodocyclization of homoallyl amines stereoselectively delivers functionalized 2-(iodomethyl)azetidine derivs. in high yield. Increasing the reaction temp. from 20° to 50° switches the reaction outcome to realize the stereoselective formation of functionalized 3-iodopyrrolidine derivs. It was shown that these pyrrolidines are formed via thermal isomerization of the aforementioned azetidines. Primary and secondary amines could be reacted with iodomethyl azetidine derivs. to deliver stable aminomethyl azetidine derivs. With subtle changes to the reaction sequences homoallyl amines could be stereoselectively converted to either cis- or trans-substituted 3-amino pyrrolidine derivs. at will. The stereochem. divergent synthesis of cis and trans substituted pyrrolidines supports an ion pair, aziridinium, isomerization pathway for azetidine to pyrrolidine isomerization. Six azetidine derivs. were probed in a zebrafish embryo developmental assay to detect potential biol. effects through the anal. of morphol. and motility behavior phenotypes. The range of effects across the probed mols. demonstrates the suitability of this assay for screening azetidine derivs. One of the probed mols., rac-(((cis)-1-benzyl-4-phenylazetidin-2-yl)methyl)piperidine, exhibited particularly interesting effects in the developmental assay presenting with hypopigmentation and reduced circulation amongst others. This shows that the zebrafish embryo provides a fast, sensitive and effective way to screen new compds. and in the future in combination with existing in vivo and in vitro assays it will become an integral part in drug discovery and development.
- 18Feula, A.; Male, L.; Fossey, J. S. Diastereoselective preparation of azetidines and pyrrolidines. Org. Lett. 2010, 12, 5044– 5047, DOI: 10.1021/ol102215e18https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXht1ajtL7M&md5=fc6db1a4635d20b732e90500bf24f52eDiastereoselective Preparation of Azetidines and PyrrolidinesFeula, Antonio; Male, Louise; Fossey, John S.Organic Letters (2010), 12 (21), 5044-5047CODEN: ORLEF7; ISSN:1523-7060. (American Chemical Society)Iodine-mediated cyclization of homoallyl amines at room temp. delivered cis-2,4-azetidine through a 4-exo trig cyclization. Isomerization of iodo-azetidines to cis-pyrrolidines could be achieved by heating, with complete stereocontrol. The relative stereochem. of the iodo-azetidines and pyrrolidines was confirmed by NMR spectroscopy and x-ray crystallog. Further functionalization was achieved through nucleophilic displacement of iodine to deliver substituted azetidines and pyrrolidines. 1,2,3-Triazole-appended azetidines and pyrrolidines were also prepd.
- 19Yoshizawa, A.; Feula, A.; Leach, A. G.; Male, L.; Fossey, J. S. Palladium and Platinum 2,4-cis-amino Azetidine and Related Complexes. Front. Chem. 2018, 6, 211, DOI: 10.3389/fchem.2018.0021119https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXisFaisbvN&md5=239ee55e21a937601449564813779a14Palladium and platinum 2,4-cis-amino azetidine and related complexesYoshizawa, Akina; Feula, Antonio; Leach, Andrew G.; Male, Louise; Fossey, John S.Frontiers in Chemistry (Lausanne, Switzerland) (2018), 6 (), 211/1-211/9CODEN: FCLSAA; ISSN:2296-2646. (Frontiers Media S.A.)Seven N,N'-palladium(II) chloride complexes, one N,N'-palladium(II) acetate complex of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. Two platinum(II) chloride N,N'-complexes of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. Computational anal. and detn. of the %Vbur was examd. conducted. A CNN' metallocyclic complex was prepd. by oxidative addn. of palladium(0) to an ortho bromo 2,4-cis-disubstituted azetidine and its crystal structure displays a slightly pyramidalized metal-ligand orientation. Seven N,N'-palladium(II) chloride complexes, one N,N'-palladium(II) acetate complex of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD. The racemic ligands adopted a single diastereoisomer form upon coordination to palladium the same chirality at nitrogen. In the palladium(II) acetate complex a coordinated water mol. and H-bonding acetates formed the identified complex. Two platinum(II) chloride N,N'-complexes of 2,4-cis-azetidines where prepd. and analyzed by single crystal XRD, and two diastereoisomers were generated upon amine coordination to platinum (under different prepn. conditions). Computational anal. revealed which diastereoisomer was more stable and provided a rationale for why this is the case, and the %Vbur described by the diastereomeric coordination geometry was examd. A CNN' metallocyclic complex was prepd. by oxidative addn. of palladium(0) to an ortho bromo 2,4-cis-disubstituted azetidine and its crystal structure displays a slightly pyramidalised metal-ligand orientation. Ligand salts were not suitable for the synthesis of azetidine complexes, instead leading to N,N' complexation of stereospecifically ring-opened congeners. A minor, pyrrolidine, impurity in an azetidine ligand sample led, initially, to the formation of a highly cryst. complex, identified by single crystal XRD, as well as from the same sample the expected azetidine complex. The isomeric azetidine and pyrrolidine complexes, characterized by single crystal XRD, were studied using the SambVca 2.0 online tool and their steric parameters contrasted revealing the potential for both azetidine and pyrrolidine ligands in future catalytic applications.
- 20Yoshizawa, A.; Feula, A.; Male, L.; Leach, A. G.; Fossey, J. S. Rigid and concave, 2,4-cis-substituted azetidine derivatives: A platform for asymmetric catalysis. Sci. Rep. 2018, 8, 6541, DOI: 10.1038/s41598-018-24784-320https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1MjmslektA%253D%253D&md5=ae3566571728d61bbcc7fa8d4c7fd426Rigid and concave, 2,4-cis-substituted azetidine derivatives: A platform for asymmetric catalysisYoshizawa Akina; Feula Antonio; Fossey John S; Male Louise; Leach Andrew GScientific reports (2018), 8 (1), 6541 ISSN:.A series of single enantiomer, 2,4-cis-disubstituted amino azetidines were synthesised and used as ligands for copper-catalysed Henry reactions of aldehydes with nitromethane. Optimisation of ligand substituents and the reaction conditions was conducted. The enantiomeric excess of the formed products was highest when alkyl aldehydes were employed in the reaction (>99% e.e.). The absolute stereochemistry of one representative azetidine derivative salt was determined by analysis of the Flack parameter of an XRD single crystal structure. The origin of selectivity in catalysis was investigated computationally, revealing the importance of the amino-substituent in determining the stereochemical outcome. A racemic platinum complex of a cis-disubstituted azetidine is examined by XRD single crystal structure analysis with reference to its steric parameters, and analogies to the computationally determined copper complex catalyst are drawn. A preliminary example of the use of a cis-disubstituted azetidine scaffold in thiourea H-bonding catalyst is noted in the supporting information.
- 21www.birmingham.ac.uk/facilities/bddf. Birmingham Drug Discovery Facility. www.birmingham.ac.uk/facilities/bddf.There is no corresponding record for this reference.
- 22
The synthesis and preliminary screening data of ∼ 100 azetidine derivatives that were less active, and not investigated further, will be reported elsewhere.
There is no corresponding record for this reference. - 23Palomino, J.-C.; Martin, A.; Camacho, M.; Guerra, H.; Swings, J.; Portaels, F. Resazurin Microtiter Assay Plate: Simple and Inexpensive Method for Detection of Drug Resistance in Mycobacterium tuberculosis. Antimicrob. Agents Chemother. 2002, 46, 2720, DOI: 10.1128/AAC.46.8.2720-2722.200223https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XlsFGiu7w%253D&md5=555848714bd1528e6a8eb913b5971c9aResazurin microtiter assay plate: simple and inexpensive method for detection of drug resistance in Mycobacterium tuberculosisPalomino, Juan-Carlos; Martin, Anandi; Camacho, Mirtha; Guerra, Humberto; Swings, Jean; Portaels, FrancoiseAntimicrobial Agents and Chemotherapy (2002), 46 (8), 2720-2722CODEN: AMACCQ; ISSN:0066-4804. (American Society for Microbiology)A method for detecting multidrug-resistant Mycobacterium tuberculosis by using a redn. of resazurin is described. Eighty clin. isolates were evaluated against isoniazid and rifampin; results at 7 days were compared with those of the proportion method. Specificity and sensitivity were excellent. The method is simple, inexpensive, and rapid and might be used with other antituberculosis drugs.
- 24Yang, F.; Njire, M. M.; Liu, J.; Wu, T.; Wang, B.; Liu, T.; Cao, Y.; Liu, Z.; Wan, J.; Tu, Z.; Tan, Y.; Tan, S.; Zhang, T. Engineering more stable, selectable marker-free autoluminescent mycobacteria by one step. PLoS One 2015, 10, e0119341 DOI: 10.1371/journal.pone.011934124https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhs1WhtbjL&md5=c63cbbd4bac4eb72c3fcf387faefd5afEngineering more stable, selectable marker- free autoluminescent mycobacteria by one stepYang, Feng; Njire, Moses M.; Liu, Jia; Wu, Tian; Wang, Bangxing; Liu, Tianzhou; Cao, Yuanyuan; Liu, Zhiyong; Wan, Junting; Tu, Zhengchao; Tan, Yaoju; Tan, Shouyong; Zhang, TianyuPLoS One (2015), 10 (3), e0119341/1-e0119341/13CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)In our previous study, we demonstrated that the use of the autoluminescent Mycobacterium tuberculosis as a reporter strain had the potential to drastically reduce the time, effort, animals and costs consumed in evaluation of the activities of drugs and vaccines in live mice. However, the strains were relatively unstable and lost reporter with time without selection. The kanamycin selection marker used wasn't the best choice as it provides resistance to amino glycosides which are an important class of second line drugs used in tuberculosis treatment. In addn., the marker could limit utility of the strains for screening of new potential drugs or evaluating drug combinations for tuberculosis treatment. Limited selection marker genes for mycobacterial genetic manipulation is a major drawback for such a marker- contg. strain in many research fields. Therefore, selectable marker-free, more stable autoluminescent mycobacteria are highly needed. After trying several strategies, we created such mycobacterial strains successfully by using an integrative vector and removing both the resistance maker and integrase genes by Xer site-specific recombination in one step. The corresponding plasmid vectors developed in this study could be very convenient in constructing other selectable marker-free, more stable reporter mycobacteria with diverse applications.
- 25Qin, L.; Wang, J.; Lu, J.; Yang, H.; Zheng, R.; Liu, Z.; Huang, X.; Feng, Y.; Hu, Z.; Ge, B. A deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosis. BMC Biol. 2019, 17, 7, DOI: 10.1186/s12915-019-0628-625https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3cjltl2rsA%253D%253D&md5=ff8dac93d32fd72ff22f55c65a72d5bbA deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosisQin Lianhua; Wang Jie; Lu Junmei; Yang Hua; Zheng Ruijuan; Liu Zhonghua; Huang Xiaochen; Feng Yonghong; Hu Zhongyi; Ge BaoxueBMC biology (2019), 17 (1), 7 ISSN:.BACKGROUND: The emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb), especially those that are multidrug resistant poses a serious threat to global tuberculosis control. However, the mechanism underlying the occurrence of drug resistance against more than one drug is poorly understood. Given that the Beijing/W strains are associated with outbreaks and multidrug resistance, they may harbor a genetic advantage and provide useful insight into the disease. One marker found in all Beijing/W Mtb strains is a deletion of RD105 region that results in a gene fusion, Rv0071/74, with a variable number (3-9 m) of VDP (V: Val, D: Asp; P: Pro) repeats (coded by gtggacccg repeat sequences) at the N-terminal. Here, we report that this variable number of VDP repeats in Rv0071/74 regulates the development of multidrug resistance. RESULTS: We collected and analyzed 1255 Beijing/W clinical strains. The results showed that the number of VDP repeats in Rv0071/74 was related to the development of multidrug resistance, and the deletion of Rv0071/74-9 m from Beijing/W clinical strain restored drug susceptibility. Rv0071/74-9 m also increased resistance to multiple drugs when transferred to different mycobacterial strains. Cell-free assays indicate that the domain carrying 4-9 VDP repeats (4-9 m) showed a variable binding affinity with peptidoglycan and Rv0071/74 cleaves peptidoglycan. Furthermore, Rv0071/74-9 m increased cell wall thickness and reduced the intracellular concentration of antibiotics. CONCLUSIONS: These findings not only identify Rv0071/74 with VDP repeats as a newly identified multidrug resistance gene but also provide a new model for the development of multiple drug resistance.
- 26Hoagland, D. T.; Liu, J.; Lee, R. B.; Lee, R. E. New agents for the treatment of drug-resistant Mycobacterium tuberculosis. Adv. Drug Delivery Rev. 2016, 102, 55– 72, DOI: 10.1016/j.addr.2016.04.02626https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XntlSns7c%253D&md5=7c8ca15b9b493cceaf3d51d93b9afcecNew agents for the treatment of drug-resistant Mycobacterium tuberculosisHoagland, Daniel T.; Liu, Jiuyu; Lee, Robin B.; Lee, Richard E.Advanced Drug Delivery Reviews (2016), 102 (), 55-72CODEN: ADDREP; ISSN:0169-409X. (Elsevier B.V.)Inadequate dosing and incomplete treatment regimens, coupled with the ability of the tuberculosis bacilli to cause latent infections that are tolerant of currently used drugs, have fueled the rise of multidrug-resistant tuberculosis (MDR-TB). Treatment of MDR-TB infections is a major clin. challenge that has few viable or effective solns.; therefore patients face a poor prognosis and years of treatment. This review focuses on emerging drug classes that have the potential for treating MDR-TB and highlights their particular strengths as leads including their mode of action, in vivo efficacy, and key medicinal chem. properties. Examples include the newly approved drugs bedaquiline and delaminid, and other agents in clin. and late preclin. development pipeline for the treatment of MDR-TB. Herein, we discuss the challenges to developing drugs to treat tuberculosis and how the field has adapted to these difficulties, with an emphasis on drug discovery approaches that might produce more effective agents and treatment regimens.
- 27Miles, A. A.; Misra, S. S.; Irwin, J. O. The estimation of the bactericidal power of the blood. Epidemiol. Infect. 1938, 38, 732– 749, DOI: 10.1017/S002217240001158XThere is no corresponding record for this reference.
- 28Hendon-Dunn, C. L.; Doris, K. S.; Thomas, S. R.; Allnutt, J. C.; Marriott, A. A. N.; Hatch, K. A.; Watson, R. J.; Bottley, G.; Marsh, P. D.; Taylor, S. C.; Bacon, J. A Flow Cytometry Method for Rapidly Assessing Mycobacterium tuberculosis Responses to Antibiotics with Different Modes of Action. Antimicrob. Agents Chemother. 2016, 60, 3869, DOI: 10.1128/AAC.02712-1528https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFSjtb7P&md5=5670956cd3b30b675b72209e8f5f92d1A flow cytometry method for rapidly assessing Mycobacterium tuberculosis responses to antibiotics with different modes of actionHendon-Dunn, Charlotte Louise; Doris, Kathryn Sarah; Thomas, Stephen Richard; Allnutt, Jonathan Charles; Marriott, Alice Ann Neville; Hatch, Kim Alexandra; Watson, Robert James; Bottley, Graham; Marsh, Philip David; Taylor, Stephen Charles; Bacon, JoannaAntimicrobial Agents and Chemotherapy (2016), 60 (7), 3869-3883CODEN: AMACCQ; ISSN:1098-6596. (American Society for Microbiology)Current methods for assessing the drug susceptibility of Mycobacterium tuberculosis are lengthy and do not capture information about viable organisms that are not immediately culturable under std. lab. conditions as a result of antibiotic exposure. We have developed a rapid dual-fluorescence flow cytometry method using markers for cell viability and death. We show that the fluorescent marker calcein violet with an acetoxy-Me ester group (CV-AM) can differentiate between populations of M. tuberculosis growing at different rates, while Sytox green (SG) can differentiate between live and dead mycobacteria. M. tuberculosis was exposed to isoniazid or rifampin at different concns. over time and either dual stained with CV-AM and SG and analyzed by flow cytometry or plated to det. the viability of the cells. Although similar trends in the loss of viability were obsd. when the results of flow cytometry and the plate counting methods were compared, there was a lack of correlation between these two approaches, as the flow cytometry anal. potentially captured information about cell populations that were unable to grow under std. conditions. The flow cytometry approach had an addnl. advantage in that it could provide insights into the mode of action of the drug: antibiotics targeting the cell wall gave a flow cytometry profile distinct from those inhibiting intracellular processes. This rapid drug susceptibility testing method could identify more effective antimycobacterials, provide information about their potential mode of action, and accelerate their progress to the clinic.
- 29Abrahams, K. A.; Cox, J. A. G.; Spivey, V. L.; Loman, N. J.; Pallen, M. J.; Constantinidou, C.; Fernandez, R.; Alemparte, C.; Remuinan, M. J.; Barros, D.; Ballell, L.; Besra, G. S. Identification of Novel Imidazo[1,2-a]pyridine Inhibitors Targeting M. tuberculosis QcrB. PLoS One 2012, 7, e52951 DOI: 10.1371/journal.pone.005295129https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptVSgsQ%253D%253D&md5=4c00cd8ffb101ce9d40da5c0c4f51c0eIdentification of novel imidazo[1,2-a]pyridine inhibitors targeting M. tuberculosis QcrBAbrahams, Katherine A.; Cox, Jonathan A. G.; Spivey, Vickey L.; Loman, Nicholas J.; Pallen, Mark J.; Constantinidou, Chrystala; Fernandez, Raquel; Alemparte, Carlos; Remuinan, Modesto J.; Barros, David; Ballell, Lluis; Besra, Gurdyal S.PLoS One (2012), 7 (12), e52951CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Mycobacterium tuberculosis is a major human pathogen and the causative agent for the pulmonary disease, tuberculosis (TB). Current treatment programs to combat TB are under threat due to the emergence of multi-drug and extensively-drug resistant TB. Through the use of high throughput whole cell screening of an extensive compd. library a no. of imidazo[1,2-a]pyridine (IP) compds. were obtained as potent lead mols. active against M. tuberculosis and Mycobacterium bovis BCG. The IP inhibitors (1-4) demonstrated min. inhibitory concns. (MICs) in the range of 0.03 to 5 μM against a panel of M. tuberculosis strains. M. bovis BCG spontaneous resistant mutants were generated against IP 1, 3 and 4 at 5 × MIC and subsequent whole genome sequencing identified a single nucleotide polymorphism 937ACC>937GCC (T313A) in qcrB, which encodes the b subunit of the electron transport ubiquinol cytochrome C reductase. This mutation also conferred cross-resistance against IP 1, 3 and 4 demonstrating a common target. Gene dosage expts. confirmed M. bovis BCG QcrB as the target where over-expression in M. bovis BCG led to an increase in MIC from 0.5 to >8 μM for IP 3. An acute murine model of TB infection established bacteriostatic activity of the IP series, which await further detailed characterization.
ARTN
- 30Abrahams, K. A.; Chung, C.-W.; Ghidelli-Disse, S.; Rullas, J.; Rebollo-López, M. J.; Gurcha, S. S.; Cox, J. A. G.; Mendoza, A.; Jiménez-Navarro, E.; Martínez-Martínez, M. S.; Neu, M.; Shillings, A.; Homes, P.; Argyrou, A.; Casanueva, R.; Loman, N. J.; Moynihan, P. J.; Lelièvre, J.; Selenski, C.; Axtman, M.; Kremer, L.; Bantscheff, M.; Angulo-Barturen, I.; Izquierdo, M. C.; Cammack, N. C.; Drewes, G.; Ballell, L.; Barros, D.; Besra, G. S.; Bates, R. H. Identification of KasA as the cellular target of an anti-tubercular scaffold. Nat. Commun. 2016, 7, 12581, DOI: 10.1038/ncomms1258130https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhsVOru7%252FI&md5=a1206008e181383854e026f36ad27489Identification of KasA as the cellular target of an anti-tubercular scaffoldAbrahams, Katherine A.; Chung, Chun-wa; Ghidelli-Disse, Sonja; Rullas, Joaquin; Rebollo-Lopez, Maria Jose; Gurcha, Sudagar S.; Cox, Jonathan A. G.; Mendoza, Alfonso; Jimenez-Navarro, Elena; Martinez-Martinez, Maria Santos; Neu, Margarete; Shillings, Anthony; Homes, Paul; Argyrou, Argyrides; Casanueva, Ruth; Loman, Nicholas J.; Moynihan, Patrick J.; Lelievre, Joel; Selenski, Carolyn; Axtman, Matthew; Kremer, Laurent; Bantscheff, Marcus; Angulo-Barturen, Inigo; Izquierdo, Monica Cacho; Cammack, Nicholas C.; Drewes, Gerard; Ballell, Lluis; Barros, David; Besra, Gurdyal S.; Bates, Robert H.Nature Communications (2016), 7 (), 12581CODEN: NCAOBW; ISSN:2041-1723. (Nature Publishing Group)Phenotypic screens for bactericidal compds. are starting to yield promising hits against tuberculosis. In this regard, whole-genome sequencing of spontaneous resistant mutants generated against an indazole sulfonamide (GSK3011724A) identifies several specific single-nucleotide polymorphisms in the essential Mycobacterium tuberculosis β-ketoacyl synthase (kas) A gene. Here, this genomic-based target assignment is confirmed by biochem. assays, chem. proteomics and structural resoln. of a KasA-GSK3011724A complex by X-ray crystallog. Finally, M. tuberculosis GSK3011724A-resistant mutants increase the in vitro min. inhibitory concn. and the in vivo 99% ED in mice, establishing in vitro and in vivo target engagement. Surprisingly, the lack of target engagement of the related β-ketoacyl synthases (FabH and KasB) suggests a different mode of inhibition when compared with other Kas inhibitors of fatty acid biosynthesis in bacteria. These results clearly identify KasA as the biol. target of GSK3011724A and validate this enzyme for further drug discovery efforts against tuberculosis.
- 31Andries, K.; Verhasselt, P.; Guillemont, J.; Göhlmann, H. W. H.; Neefs, J.-M.; Winkler, H.; Van Gestel, J.; Timmerman, P.; Zhu, M.; Lee, E.; Williams, P.; de Chaffoy, D.; Huitric, E.; Hoffner, S.; Cambau, E.; Truffot-Pernot, C.; Lounis, N.; Jarlier, V. A Diarylquinoline Drug Active on the ATP Synthase of Mycobacterium tuberculosis. Science 2005, 307, 223, DOI: 10.1126/science.110675331https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2MXisVKkug%253D%253D&md5=099b643f173bb77e70cf1aa6e9871ffeA Diarylquinoline Drug Active on the ATP Synthase of Mycobacterium tuberculosisAndries, Koen; Verhasselt, Peter; Guillemont, Jerome; Goehlmann, Hinrich W. H.; Neefs, Jean-Marc; Winkler, Hans; Van Gestel, Jef; Timmerman, Philip; Zhu, Min; Lee, Ennis; Williams, Peter; de Chaffoy, Didier; Huitric, Emma; Hoffner, Sven; Cambau, Emmanuelle; Truffot-Pernot, Chantal; Lounis, Nacer; Jarlier, VincentScience (Washington, DC, United States) (2005), 307 (5707), 223-227CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)The incidence of tuberculosis has been increasing substantially on a worldwide basis over the past decade, but no tuberculosis-specific drugs have been discovered in 40 years. We identified a diarylquinoline, R207910, that potently inhibits both drug-sensitive and drug-resistant Mycobacterium tuberculosis in vitro (min. inhibitory concn. 0.06 μg/mL). In mice, R207910 exceeded the bactericidal activities of isoniazid and rifampin by at least 1 log unit. Substitution of drugs included in the World Health Organization's first-line tuberculosis treatment regimen (rifampin, isoniazid, and pyrazinamide) with R207910 accelerated bactericidal activity, leading to complete culture conversion after 2 mo of treatment in some combinations. A single dose of R207910 inhibited mycobacterial growth for 1 wk. Plasma levels assocd. with efficacy in mice were well tolerated in healthy human volunteers. Mutants selected in vitro suggest that the drug targets the proton pump of ATP (ATP) synthase.
- 32Batt, S. M.; Cacho Izquierdo, M.; Castro Pichel, J.; Stubbs, C. J.; Vela-Glez Del Peral, L.; Pérez-Herrán, E.; Dhar, N.; Mouzon, B.; Rees, M.; Hutchinson, J. P.; Young, R. J.; McKinney, J. D.; Barros Aguirre, D.; Ballell, L.; Besra, G. S.; Argyrou, A. Whole Cell Target Engagement Identifies Novel Inhibitors of Mycobacterium tuberculosis Decaprenylphosphoryl-β-d-ribose Oxidase. ACS Infec. Dis. 2015, 1, 615– 626, DOI: 10.1021/acsinfecdis.5b0006532https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtlemtbnE&md5=1e6d6ef29fb37d3e8535328368af2472Whole Cell Target Engagement Identifies Novel Inhibitors of Mycobacterium tuberculosis Decaprenylphosphoryl-β-D-ribose OxidaseBatt, Sarah M.; Cacho Izquierdo, Monica; Castro Pichel, Julia; Stubbs, Christopher J.; Vela-Glez Del Peral, Laura; Perez-Herran, Esther; Dhar, Neeraj; Mouzon, Bernadette; Rees, Mike; Hutchinson, Jonathan P.; Young, Robert J.; McKinney, John D.; Barros Aguirre, David; Ballell, Lluis; Besra, Gurdyal S.; Argyrou, ArgyridesACS Infectious Diseases (2015), 1 (12), 615-626CODEN: AIDCBC; ISSN:2373-8227. (American Chemical Society)We have targeted the Mycobacterium tuberculosis decaprenylphosphoryl-β-D-ribose oxidase (Mt-DprE1) for potential chemotherapeutic intervention of tuberculosis. A multicopy suppression strategy that overexpressed Mt-DprE1 in M. bovis BCG was used to profile the publically available GlaxoSmithKline antimycobacterial compd. set, and one compd. (GSK710) was identified that showed an 8-fold higher min. inhibitory concn. relative to the control strain. Analogs of GSK710 show a clear relationship between whole cell potency and in vitro activity using an enzymic assay employing recombinant Mt-DprE1, with binding affinity measured by fluorescence quenching of the flavin cofactor of the enzyme. M. bovis BCG spontaneous resistant mutants to GSK710 and a closely related analog were isolated and sequencing of ten such mutants revealed a single point mutation at two sites, E221Q or G248S within DprE1, providing further evidence that DprE1 is the main target of these compds. Finally, time-lapse microscopy expts. showed that exposure of M. tuberculosis to a compd. of this series arrests bacterial growth rapidly followed by a slower cytolysis phase.
- 33Manganelli, R.; Voskuil, M. I.; Schoolnik, G. K.; Smith, I. The Mycobacterium tuberculosis ECF sigma factor sigmaE: role in global gene expression and survival in macrophages. Mol. Microbiol. 2001, 41, 423– 37, DOI: 10.1046/j.1365-2958.2001.02525.x33https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3MXlvVClur0%253D&md5=ec7927a14ea6841428d0a7e70df82551The Mycobacterium tuberculosis ECF sigma factor σE: role in global gene expression and survival in macrophagesManganelli, Riccardo; Voskuil, Martin I.; Schoolnik, Gary K.; Smith, IssarMolecular Microbiology (2001), 41 (2), 423-437CODEN: MOMIEE; ISSN:0950-382X. (Blackwell Science Ltd.)In previously published work, we identified three Mycobacterium tuberculosis sigma (σ) factor genes responding to heat shock (sigB, sigE and sigH). Two of them (sigB and sigE) also responded to SDS exposure. As these responses to stress suggested that the σ factors encoded by these genes could be involved in pathogenicity, we are studying their role in physiol. and virulence. In this work, we characterize a sigE mutant of M. tuberculosis H37Rv. The sigE mutant strain was more sensitive than the wild-type strain to heat shock, SDS and various oxidative stresses. It was also defective in the ability to grow inside both human and murine unactivated macrophages and was more sensitive than the wild-type strain to the killing activity of activated murine macrophages. Using microarray technol. and quant. reverse transcription-polymerase chain reaction (RT-PCR), we started to define the σE regulon of M. tuberculosis and its involvement in the global regulation of the stress induced by SDS. We showed the requirement for a functional sigE gene for full expression of sigB and for its induction after SDS exposure but not after heat shock. We also identified several genes that are no longer induced when σE is absent. These genes encode proteins belonging to different classes including transcriptional regulators, enzymes involved in fatty acid degrdn. and classical heat shock proteins.
- 34Balazsi, G.; Heath, A. P.; Shi, L.; Gennaro, M. L. The temporal response of the Mycobacterium tuberculosis gene regulatory network during growth arrest. Mol. Syst. Biol. 2008, 4, 225, DOI: 10.1038/msb.2008.6334https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD1cjhvV2gsg%253D%253D&md5=27c4701b97cae4a1f215832cdaf75481The temporal response of the Mycobacterium tuberculosis gene regulatory network during growth arrestBalazsi Gabor; Heath Allison P; Shi Lanbo; Gennaro Maria LMolecular systems biology (2008), 4 (), 225 ISSN:.The virulence of Mycobacterium tuberculosis depends on the ability of the bacilli to switch between replicative (growth) and non-replicative (dormancy) states in response to host immunity. However, the gene regulatory events associated with transition to dormancy are largely unknown. To address this question, we have assembled the largest M. tuberculosis transcriptional-regulatory network to date, and characterized the temporal response of this network during adaptation to stationary phase and hypoxia, using published microarray data. Distinct sets of transcriptional subnetworks (origons) were responsive at various stages of adaptation, showing a gradual progression of network response under both conditions. Most of the responsive origons were in common between the two conditions and may help define a general transcriptional signature of M. tuberculosis growth arrest. These results open the door for a systems-level understanding of transition to non-replicative persistence, a phenotypic state that prevents sterilization of infection by the host immune response and promotes the establishment of latent M. tuberculosis infection, a condition found in two billion people worldwide.
- 35Karp, P. D.; Billington, R.; Caspi, R.; Fulcher, C. A.; Latendresse, M.; Kothari, A.; Keseler, I. M.; Krummenacker, M.; Midford, P. E.; Ong, Q.; Ong, W. K.; Paley, S. M.; Subhraveti, P. The BioCyc collection of microbial genomes and metabolic pathways. Brief. Bioinform. 2019, 20, 1085– 1093, DOI: 10.1093/bib/bbx08535https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BB3cXht1KqtLvE&md5=53160f313a2b5c6a9891cb442f4b2dd0The BioCyc collection of microbial genomes and metabolic pathwaysKarp, Peter D.; Billington, Richard; Caspi, Ron; Fulcher, Carol A.; Latendresse, Mario; Kothari, Anamika; Keseler, Ingrid M.; Krummenacker, Markus; Midford, Peter E.; Ong, Quang; Ong, Wai Kit; Paley, Suzanne M.; Subhraveti, PallaviBriefings in Bioinformatics (2019), 20 (4), 1085-1093CODEN: BBIMFX; ISSN:1477-4054. (Oxford University Press)BioCyc.org is a microbial genome Web portal that combines thousands of genomes with addnl. information inferred by computer programs, imported from other databases and curated from the biomedical literature by biologist curators. BioCyc also provides an extensive range of query tools, visualization services and anal. software. Recent advances in BioCyc include an expansion in the content of BioCyc in terms of both the no. of genomes and the types of information available for each genome; an expansion in the amt. of curated content within BioCyc; and new developments in the BioCyc software tools including redesigned gene/protein pages and metabolite pages; new search tools; a new sequence-alignment tool; a new tool for visualizing groups of related metabolic pathways; and a facility called SmartTables, which enables biologists to perform analyses that previously would have required a programmer's assistance.
- 36Caspi, R.; Billington, R.; Ferrer, L.; Foerster, H.; Fulcher, C. A.; Keseler, I. M.; Kothari, A.; Krummenacker, M.; Latendresse, M.; Mueller, L. A.; Ong, Q.; Paley, S.; Subhraveti, P.; Weaver, D. S.; Karp, P. D. The MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databases. Nucleic Acids Res. 2016, 44, D471– 80, DOI: 10.1093/nar/gkv116436https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtV2nsrnJ&md5=7d8dbff74ddb4e6050783c8ba3d68e2bThe MetaCyc database of metabolic pathways and enzymes and the BioCyc collection of pathway/genome databasesCaspi, Ron; Billington, Richard; Ferrer, Luciana; Foerster, Hartmut; Fulcher, Carol A.; Keseler, Ingrid M.; Kothari, Anamika; Krummenacker, Markus; Latendresse, Mario; Mueller, Lukas A.; Ong, Quang; Paley, Suzanne; Subhraveti, Pallavi; Weaver, Daniel S.; Karp, Peter D.Nucleic Acids Research (2016), 44 (D1), D471-D480CODEN: NARHAD; ISSN:0305-1048. (Oxford University Press)A review. The MetaCyc database is a freely accessible comprehensive database describing metabolic pathways and enzymes from all domains of life. The majority of MetaCyc pathways are smallmol. metabolic pathways that have been exptl. detd. MetaCyc contains more than 2400 pathways derived from >46 000 publications, and is the largest curated collection of metabolic pathways. BioCyc is a collection of 5700 organism-specific Pathway/Genome Databases (PGDBs), each contg. the full genome and predicted metabolic network of one organism, including metabolites, enzymes, reactions, metabolic pathways, predicted operons, transport systems, and pathway-hole fillers. The BioCyc website offers a variety of tools for querying and analyzing PGDBs, including Omics Viewers and tools for comparative anal. This article provides an update of new developments in MetaCyc and BioCyc during the last two years, including addn. of Gibbs free energy values for compds. and reactions; redesign of the primary gene/protein page; addn. of a tool for creating diagrams contg. multiple linked pathways; several new search capabilities, including searching for genes based on sequence patterns, searching for databases based on an organism's phenotypes, and a cross-organism search; and a metabolite identifier translation service.
- 37Chang, Y.; Fox, B. G. Identification of Rv3230c as the NADPH oxidoreductase of a two-protein DesA3 acyl-CoA desaturase in Mycobacterium tuberculosis H37Rv. Biochemistry 2006, 45, 13476– 86, DOI: 10.1021/bi061528537https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28XhtFSrs73M&md5=970108deb08558d4f7b1ff6f349111b6Identification of Rv3230c as the NADPH Oxidoreductase of a Two-Protein DesA3 Acyl-CoA Desaturase in Mycobacterium tuberculosis H37RvChang, Yong; Fox, Brian G.Biochemistry (2006), 45 (45), 13476-13486CODEN: BICHAW; ISSN:0006-2960. (American Chemical Society)DesA3 is a membrane-bound stearoyl-CoA Δ9-desaturase that produces oleic acid, a precursor of mycobacterial membrane phospholipids and triglycerides. The sequence of DesA3 is homologous with those of other membrane desaturases, including the presence of the eight-His motif proposed to bind the diiron center active site. This family of desaturases function as multicomponent complexes and thus require electron transfer proteins for efficient catalytic turnover. Here we present evidence that Rv3230c from Mycobacterium tuberculosis H37Rv is a biol. relevant electron transfer partner for DesA3 from the same pathogen. For these studies, Rv3230c was expressed as a partially sol. protein in Escherichia coli; recombinant DesA3 was expressed in Mycobacterium smegmatis as a catalytically active membrane protein. The addn. of E. coli lysates contg. Rv3230c to lysates of M. smegmatis expressing DesA3 gave strong conversion of [1-14C]-18:0-CoA to [1-14C]-cis-Δ9-18:1-CoA and of [1-14C]-16:0-CoA to [1-14C]-cis-Δ9-16:1-CoA. Both M. tuberculosis proteins were required for reconstitution of activity, as various combinations of control lysates lacking either Rv3230c or DesA3 gave minimal or no activity. Furthermore, the specificity of interaction between Rv3230c and DesA3 was implied by the inability of other related redox systems to substitute for Rv3230c. The reconstituted activity was dependent upon the presence of NADPH, could be satd. by increasing the amt. of Rv3230c added, and was also sensitive to the salt concn. in the buffer. The results are consistent with the formation of a protein-protein complex, possibly with electrostatic character. This work defines a multiprotein, acyl-CoA desaturase complex from M. tuberculosis H37Rv to minimally consist of a sol. Rv3230c reductase and integral membrane DesA3 desaturase. Further implications of this finding relative to the properties of other multiprotein iron-enzyme complexes are discussed.
- 38McGillivray, A.; Golden, N. A.; Gautam, U. S.; Mehra, S.; Kaushal, D. The Mycobacterium tuberculosis Rv2745c plays an important role in responding to redox stress. PLoS One 2014, 9, e93604 DOI: 10.1371/journal.pone.009360438https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhs1ajurjK&md5=33aa9199d6d28f482d53610ed08fd3f2The Mycobacterium tuberculosis Rv2745c plays an important role in responding to redox stressMcGillivray, Amanda; Golden, Nadia Abrahams; Gautam, Uma Shankar; Mehra, Smriti; Kaushal, DeepakPLoS One (2014), 9 (4), e93604/1-e93604/22, 22 pp.CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is the leading cause of death from an infectious disease worldwide. Over the course of its life cycle in vivo, Mtb is exposed to a plethora of environmental stress conditions. Temporal regulation of genes involved in sensing and responding to such conditions is therefore crucial for Mtb to establish an infection. The Rv2745c (clgR) gene encodes a Clp protease gene regulator that is induced in response to a variety of stress conditions and potentially plays a role in Mtb pathogenesis. The isogenic mutant Mtb:ΔRv2745c is significantly more sensitive to in vitro redox stress generated by diamide, relative to wild-type Mtb as well as to a complemented strain. Together with the fact that the expression of Rv2745c is strongly induced in response to redox stress, these results strongly implicate a role for ClgR in the management of intraphagosomal redox stress. Addnl., we obsd. that redox stress led to the dysregulation of the expression of the σH/σE regulon in the isogenic mutant, Mtb:ΔRv2745c. Furthermore, induction of clgR in Mtb and Mtb:ΔRv2745c (comp) did not lead to Clp protease induction, indicating that clgR has addnl. functions that need to be elucidated. Our data indicate that ClgR plays diverse roles in multiple regulatory networks in response to different stress conditions. In addn. to redox stress, the expression of Rv2745c correlates with the expression of genes involved in sulfate assimilation as well as in response to hypoxia and reaeration. Clearly, the Mtb Rv2745c-encoded ClgR performs different functions during stress response and is important for the pathogenicity of Mtb in vivo, regardless of its induction of the Clp proteolytic pathway.
- 39Hards, K.; Robson, J. R.; Berney, M.; Shaw, L.; Bald, D.; Koul, A.; Andries, K.; Cook, G. M. Bactericidal mode of action of bedaquiline. J. Antimicrob. Chemother. 2015, 70, 2028– 2037, DOI: 10.1093/jac/dkv05439https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhtFyltr7N&md5=22c8df11e6b904ba3645fa7ac74c5074Bactericidal mode of action of bedaquilineHards, Kiel; Robson, Jennifer R.; Berney, Michael; Shaw, Lisa; Bald, Dirk; Koul, Anil; Andries, Koen; Cook, Gregory M.Journal of Antimicrobial Chemotherapy (2015), 70 (7), 2028-2037CODEN: JACHDX; ISSN:0305-7453. (Oxford University Press)Objectives: It is not fully understood why inhibiting ATP synthesis in Mycobacterium species leads to death in non-replicating cells. We investigated the bactericidal mode of action of the anti-tubercular F1Fo-ATP synthase inhibitor bedaquiline (Sirturo) in order to further understand the lethality of ATP synthase inhibition. Methods:Mycobacterium smegmatis strains were used for all the expts. Growth and survival during a bedaquiline challenge were performed in multiple media types. A time-course microarray was performed during initial bedaquiline challenge in minimal medium. Oxygen consumption and proton-motive force measurements were performed on whole cells and inverted membrane vesicles, resp. Results: A killing of 3 log10 cfu/mL was achieved 4-fold more quickly in minimal medium (a glycerol carbon source) vs. rich medium (LB with Tween 80) during bedaquiline challenge. Assessing the accelerated killing condition, we identified a transcriptional remodelling of metab. that was consistent with respiratory dysfunction but inconsistent with ATP depletion. In glycerol-energized cell suspensions, bedaquiline caused an immediate 2.3-fold increase in oxygen consumption. Bedaquiline collapsed the transmembrane pH gradient, but not the membrane potential, in a dose-dependent manner. Both these effects were dependent on binding to the F1Fo-ATP synthase. Conclusions: Challenge with bedaquiline results in an electroneutral uncoupling of respiration-driven ATP synthesis. This may be a determinant of the bactericidal effects of bedaquiline, while ATP depletion may be a determinant of its delayed onset of killing. We propose that bedaquiline binds to and perturbs the a-c subunit interface of the Fo, leading to futile proton cycling, which is known to be lethal to mycobacteria.
- 40Mishra, S.; Shukla, P.; Bhaskar, A.; Anand, K.; Baloni, P.; Jha, R. K.; Mohan, A.; Rajmani, R. S.; Nagaraja, V.; Chandra, N.; Singh, A. Efficacy of β-lactam/β-lactamase inhibitor combination is linked to WhiB4-mediated changes in redox physiology of Mycobacterium tuberculosis. eLife 2017, 6, e25624 DOI: 10.7554/eLife.2562440https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXitlGhur7M&md5=708a67ddb1108ab51d5b9dee9e017b14Efficacy of β-lactam/β-lactamase inhibitor combination is linked to WhiB4-mediated changes in redox physiology of Mycobacterium tuberculosisMishra, Saurabh; Shukla, Prashant; Bhaskar, Ashima; Anand, Kushi; Baloni, Priyanka; Jha, Rajiv Kumar; Mohan, Abhilash; Rajmani, Raju S.; Nagaraja, Valakunja; Chandra†, Nagasuma; Singh, AmiteLife (2017), 6 (), e25624/1-e25624/30CODEN: ELIFA8; ISSN:2050-084X. (eLife Sciences Publications Ltd.)Mycobacterium tuberculosis (Mtb) expresses a broad-spectrum b-lactamase (BlaC) that mediates resistance to one of the highly effective antibacterials, β-lactams. Nonetheless, b-lactams showed mycobactericidal activity in combination with β-lactamase inhibitor, clavulanate (Clav). However, the mechanistic aspects of how Mtb responds to β-lactams such as Amoxicillin in combination with Clav (referred as Augmentin [AG]) are not clear. Here, we identified cytoplasmic redox potential and intracellular redox sensor, WhiB4, as key determinants of mycobacterial resistance against AG. Using computer-based, biochem., redox-biosensor, and genetic strategies, we uncovered a functional linkage between specific determinants of b-lactam resistance (e.g. β-lactamase) and redox potential in Mtb. We also describe the role of WhiB4 in coordinating the activity of b-lactamase in a redox-dependent manner to tolerate AG. Disruption of WhiB4 enhances AG tolerance, whereas overexpression potentiates AG activity against drug-resistant Mtb. Our findings suggest that AG can be exploited to diminish drug-resistance in Mtb through redox-based interventions.
- 41Waddell, S. J.; Stabler, R. A.; Laing, K.; Kremer, L.; Reynolds, R. C.; Besra, G. S. The use of microarray analysis to determine the gene expression profiles of Mycobacterium tuberculosis in response to anti-bacterial compounds. Tuberculosis 2004, 84, 263– 74, DOI: 10.1016/j.tube.2003.12.00541https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD2czgs1KisA%253D%253D&md5=75c1c424e53be6ec2c94a7231853fb75The use of microarray analysis to determine the gene expression profiles of Mycobacterium tuberculosis in response to anti-bacterial compoundsWaddell Simon J; Stabler Richard A; Laing Ken; Kremer Laurent; Reynolds Robert C; Besra Gurdyal STuberculosis (Edinburgh, Scotland) (2004), 84 (3-4), 263-74 ISSN:1472-9792.The response of Mycobacterium tuberculosis to six anti-microbial agents was determined by microarray analysis in an attempt to define mechanisms of innate resistance in M. tuberculosis. The gene expression profiles of M. tuberculosis after treatment at the minimal inhibitory concentration (MIC) for 4 h with isoniazid, isoxyl, tetrahydrolipstatin, SRI#221, SR1#967 and SR1#9190 were compared to untreated M. tuberculosis. A common response to drug exposure was defined, and this expression profile overlapped with a number of other mycobacterial stress responses recently identified by microarray analysis. Compound-specific responses were also distinguished including a number of putative transcriptional regulators and translocation-related genes. These genes may contribute to the intrinsic resistance of M. tuberculosis to anti-microbial compounds. Further investigation into these mechanisms may elucidate novel pathways contributing to mycobacterial drug resistance and influence anti-mycobacterial drug development strategies.
- 42Boshoff, H. I.; Myers, T. G.; Copp, B. R.; McNeil, M. R.; Wilson, M. A.; Barry, C. E., 3rd The transcriptional responses of Mycobacterium tuberculosis to inhibitors of metabolism: Novel insights into drug mechanisms of action. J. Biol. Chem. 2004, 279, 40174– 84, DOI: 10.1074/jbc.M40679620042https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXnsVKrsLs%253D&md5=677489fe3a9981eba62d990ec037f71aThe Transcriptional Responses of Mycobacterium tuberculosis to Inhibitors of Metabolism: Novel insights into drug mechanisms of actionBoshoff, Helena I. M.; Myers, Timothy G.; Copp, Brent R.; McNeil, Michael R.; Wilson, Michael A.; Barry, Clifton E., IIIJournal of Biological Chemistry (2004), 279 (38), 40174-40184CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)The differential transcriptional response of Mycobacterium tuberculosis to drugs and growth-inhibitory conditions was monitored to generate a data set of 430 microarray profiles. Unbiased grouping of these profiles independently clustered agents of known mechanism of action accurately and was successful at predicting the mechanism of action of several unknown agents. These predictions were validated biochem. for two agents of previously uncategorized mechanism, pyridoacridones and phenothiazines. Anal. of this data set further revealed 150 underlying clusters of coordinately regulated genes offering the first glimpse at the full metabolic potential of this organism. A signature subset of these gene clusters was sufficient to classify all known agents as to mechanism of action. Transcriptional profiling of both crude and purified natural products can provide crit. information on both mechanism and detoxification prior to purifn. that can be used to guide the drug discovery process. Thus, the transcriptional profile generated by a crude marine natural product recapitulated the mechanistic prediction from the pure active component. The underlying gene clusters further provide fundamental insights into the metabolic response of bacteria to drug-induced stress and provide a rational basis for the selection of crit. metabolic targets for screening for new agents with improved activity against this important human pathogen.
- 43Amaral, L.; Viveiros, M. Thioridazine: A Non-Antibiotic Drug Highly Effective, in Combination with First Line Anti-Tuberculosis Drugs, against Any Form of Antibiotic Resistance of Mycobacterium tuberculosis Due to Its Multi-Mechanisms of Action. Antibiotics 2017, 6, 3, DOI: 10.3390/antibiotics6010003There is no corresponding record for this reference.
- 44Lee, R. E.; Protopopova, M.; Crooks, E.; Slayden, R. A.; Terrot, M.; Barry, C. E., 3rd Combinatorial lead optimization of [1,2]-diamines based on ethambutol as potential antituberculosis preclinical candidates. J. Comb. Chem. 2003, 5, 172– 87, DOI: 10.1021/cc020071p44https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3sXkvFKguw%253D%253D&md5=3005c111ca5aa61044f016dcfafc6d09Combinatorial Lead Optimization of [1,2]-Diamines Based on Ethambutol as Potential Antituberculosis Preclinical CandidatesLee, Richard E.; Protopopova, Marina; Crooks, Emma; Slayden, Richard A.; Terrot, Marianne; Barry, Clifton E., IIIJournal of Combinatorial Chemistry (2003), 5 (2), 172-187CODEN: JCCHFF; ISSN:1520-4766. (American Chemical Society)Despite relatively modest potency, ethambutol (EMB, (S,S)-[N,N-di-2-amino-1-butanol]ethylenediamine) is a mainstay of contemporary chemotherapy for the treatment of tuberculosis. We have developed a solid-phase synthesis of 1,2-diamine analogs of EMB using a novel acylation-redn. sequence that is compatible with high-throughput 96-well format chem. Using this procedure, we have synthesized 63,238 diamine analogs in pools of 10 that are suitable for testing. MIC and a target-based reporter assay were used to direct deconvolution of 2796 individual compds. from these mixts., and the 69 most potent mols. were resynthesized in milligram quantities for hit confirmation. Purifn. of these individual active diamine analogs allowed the identification of 26 compds. with activity equal to or greater than EMB. Amines which occurred most frequently in active compds. included many with large hydrophobic moieties, suggesting that optimization was perhaps selecting for the isoprenoid binding site of the arabinosyltransferase target of EMB. N-Geranyl-N'-(2-adamantyl)ethane-1,2-diamine, the most active of these diamines, displayed a 14-35-fold improvement in activity in vitro against Mycobacterium tuberculosis, as compared to EMB.
- 45Makarov, V.; Manina, G.; Mikusova, K.; Möllmann, U.; Ryabova, O.; Saint-Joanis, B.; Dhar, N.; Pasca, M. R.; Buroni, S.; Lucarelli, A. P.; Milano, A.; De Rossi, E.; Belanova, M.; Bobovska, A.; Dianiskova, P.; Kordulakova, J.; Sala, C.; Fullam, E.; Schneider, P.; McKinney, J. D.; Brodin, P.; Christophe, T.; Waddell, S.; Butcher, P.; Albrethsen, J.; Rosenkrands, I.; Brosch, R.; Nandi, V.; Bharath, S.; Gaonkar, S.; Shandil, R. K.; Balasubramanian, V.; Balganesh, T.; Tyagi, S.; Grosset, J.; Riccardi, G.; Cole, S. T. Benzothiazinones Kill Mycobacterium tuberculosis by Blocking Arabinan Synthesis. Science 2009, 324, 801, DOI: 10.1126/science.117158345https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1MXlsVeksrY%253D&md5=1d80f14ba88460ef5c8ea31793dbb592Benzothiazinones Kill Mycobacterium tuberculosis by Blocking Arabinan SynthesisMakarov, Vadim; Manina, Giulia; Mikusova, Katarina; Moellmann, Ute; Ryabova, Olga; Saint-Joanis, Brigitte; Dhar, Neeraj; Pasca, Maria Rosalia; Buroni, Silvia; Lucarelli, Anna Paola; Milano, Anna; De Rossi, Edda; Belanova, Martina; Bobovska, Adela; Dianiskova, Petronela; Kordulakova, Jana; Sala, Claudia; Fullam, Elizabeth; Schneider, Patricia; McKinney, John D.; Brodin, Priscille; Christophe, Thierry; Waddell, Simon; Butcher, Philip; Albrethsen, Jakob; Rosenkrands, Ida; Brosch, Roland; Nandi, Vrinda; Bharath, Sowmya; Gaonkar, Sheshagiri; Shandil, Radha K.; Balasubramanian, Venkataraman; Balganesh, Tanjore; Tyagi, Sandeep; Grosset, Jacques; Riccardi, Giovanna; Cole, Stewart T.Science (Washington, DC, United States) (2009), 324 (5928), 801-804CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)New drugs are required to counter the tuberculosis (TB) pandemic. Here, we describe the synthesis and characterization of 1,3-benzothiazin-4-ones (BTZs), a new class of antimycobacterial agents that kill Mycobacterium tuberculosis in vitro, ex vivo, and in mouse models of TB. Using genetics and biochem., we identified the enzyme decaprenylphosphoryl-β-D-ribose 2'-epimerase as a major BTZ target. Inhibition of this enzymic activity abolishes the formation of decaprenylphosphoryl arabinose, a key precursor that is required for the synthesis of the cell-wall arabinans, thus provoking cell lysis and bacterial death. The most advanced compd., BTZ043, is a candidate for inclusion in combination therapies for both drug-sensitive and extensively drug-resistant TB.
- 46Xu, Z.; Meshcheryakov, V. A.; Poce, G.; Chng, S.-S. MmpL3 is the flippase for mycolic acids in mycobacteria. Proc. Natl. Acad. Sci. U.S.A. 2017, 114, 7993, DOI: 10.1073/pnas.170006211446https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtFCrtL7O&md5=1c8a806ca2a1eaebefe7c1e6bf3bcab9MmpL3 is the flippase for mycolic acids in mycobacteriaXu, Zhujun; Meshcheryakov, Vladimir A.; Poce, Giovanna; Chng, Shu-SinProceedings of the National Academy of Sciences of the United States of America (2017), 114 (30), 7993-7998CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)The defining feature of the mycobacterial outer membrane (OM) is the presence of mycolic acids (MAs), which, in part, render the bilayer extremely hydrophobic and impermeable to external insults, including many antibiotics. Although the biosynthetic pathway of MAs is well studied, the mechanism(s) by which these lipids are transported across the cell envelope is(are) much less known. Mycobacterial membrane protein Large 3 (MmpL3), an essential inner membrane (IM) protein, is implicated in MA transport, but its exact function has not been elucidated. It is believed to be the cellular target of several antimycobacterial compds.; however, evidence for direct inhibition of MmpL3 activity is also lacking. Here, we establish that MmpL3 is the MA flippase at the IM of mycobacteria and is the mol. target of BM212, a 1,5-diarylpyrrole compd. We develop assays that selectively access mycolates on the surface of Mycobacterium smegmatis spheroplasts, allowing us to monitor flipping of MAs across the IM. Using these assays, we establish the mechanism of action of BM212 as a potent MmpL3 inhibitor, and use it as a mol. probe to demonstrate the requirement for functional MmpL3 in the transport of MAs across the IM. Finally, we show that BM212 binds MmpL3 directly and inhibits its activity. Our work provides fundamental insights into OM biogenesis and MA transport in mycobacteria. Furthermore, our assays serve as an important platform for accelerating the validation of small mols. that target MmpL3, and their development as future antituberculosis drugs.
- 47Portevin, D.; de Sousa, D.; Auria, C.; Houssin, C.; Grimaldi, C.; Chami, M.; Daffé, M.; Guilhot, C. A polyketide synthase catalyzes the last condensation step of mycolic acid biosynthesis in mycobacteria and related organisms. Proc. Natl. Acad. Sci. U. S. A. 2004, 101, 314, DOI: 10.1073/pnas.030543910147https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2cXjvFamuw%253D%253D&md5=dbbe12644994eba3860c2733d429f4b1A polyketide synthase catalyzes the last condensation step of mycolic acid biosynthesis in mycobacteria and related organismsPortevin, Damien; de Sousa-d'Auria, Celia; Houssin, Christine; Grimaldi, Christine; Chami, Mohamed; Daffe, Mamadou; Guilhot, ChristopheProceedings of the National Academy of Sciences of the United States of America (2004), 101 (1), 314-319CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Mycolic acids are major and specific constituents of the cell envelope of Corynebacterineae, a suborder of bacterial species including several important human pathogens such as Mycobacterium tuberculosis, Mycobacterium leprae, or Corynebacterium diphtheriae. These long-chain fatty acids are involved in the unusual architecture and impermeability of the cell envelope of these bacteria. The condensase, the enzyme responsible for the final condensation step in mycolic acid biosynthesis, has remained an enigma for decades. By in silico anal. of various mycobacterial genomes, we identified a candidate enzyme, Pks13, that contains the four catalytic domains required for the condensation reaction. Orthologs of this enzyme were found in other Corynebacterineae species. A Corynebacterium glutamicum strain with a deletion in the pks13 gene was shown to be deficient in mycolic acid prodn. whereas it was able to produce the fatty acids precursors. This mutant strain displayed an altered cell envelope structure. We showed that the pks13 gene was essential for the survival of Mycobacterium smegmatis. A conditional M. smegmatis mutant carrying its only copy of pks13 on a thermosensitive plasmid exhibited mycolic acid biosynthesis defect if grown at nonpermissive temp. These results indicate that Pks13 is the condensase, a promising target for the development of new antimicrobial drugs against Corynebacterineae.
- 48Gavalda, S.; Bardou, F.; Laval, F.; Bon, C.; Malaga, W.; Chalut, C.; Guilhot, C.; Mourey, L.; Daffé, M.; Quémard, A. The Polyketide Synthase Pks13 Catalyzes a Novel Mechanism of Lipid Transfer in Mycobacteria. Chem. Biol. 2014, 21, 1660– 1669, DOI: 10.1016/j.chembiol.2014.10.01148https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhvF2gs7%252FO&md5=aa1ce63f8e0bec476d5e9fb91ef7c63cThe Polyketide Synthase Pks13 Catalyzes a Novel Mechanism of Lipid Transfer in MycobacteriaGavalda, Sabine; Bardou, Fabienne; Laval, Francoise; Bon, Cecile; Malaga, Wladimir; Chalut, Christian; Guilhot, Christophe; Mourey, Lionel; Daffe, Mamadou; Quemard, AnnaikChemistry & Biology (Oxford, United Kingdom) (2014), 21 (12), 1660-1669CODEN: CBOLE2; ISSN:1074-5521. (Elsevier Ltd.)Mycolate-contg. compds. constitute major strategic elements of the protective coat surrounding the tubercle bacillus. We have previously shown that FAAL32-Pks13 polyketide synthase catalyzes the condensation reaction, which produces α-alkyl β-ketoacids, direct precursors of mycolic acids. In contrast to the current biosynthesis model, we show here that Pks13 catalyzes itself the release of the neosynthesized products and demonstrate that this function is carried by its thioesterase-like domain. Most importantly, in agreement with the prediction of a trehalose-binding pocket in its catalytic site, this domain exhibits an acyltransferase activity and transfers Pks13's products onto an acceptor mol., mainly trehalose, leading to the formation of the trehalose monomycolate precursor. Thus, this work allows elucidation of the hinge step of the mycolate-contg. compd. biosynthesis pathway. Above all, it highlights a unique mechanism of transfer of polyketide synthase products in mycobacteria, which is distinct from the conventional intervention of the discrete polyketide-assocd. protein (Pap)-type acyltransferases.
- 49Su, C.-C.; Klenotic, P. A.; Bolla, J. R.; Purdy, G. E.; Robinson, C. V.; Yu, E. W. MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamine. Proc. Natl. Acad. Sci. U.S.A. 2019, 116, 11241, DOI: 10.1073/pnas.190134611649https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXhtV2itLrK&md5=36f2807f7a55ec997fb211206054eef7MmpL3 is a lipid transporter that binds trehalose monomycolate and phosphatidylethanolamineSu, Chih-Chia; Klenotic, Philip A.; Bolla, Jani Reddy; Purdy, Georgiana E.; Robinson, Carol V.; Yu, Edward W.Proceedings of the National Academy of Sciences of the United States of America (2019), 116 (23), 11241-11246CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)The cell envelope of Mycobacterium tuberculosis is notable for the abundance of mycolic acids (MAs), essential to mycobacterial viability, and of other species-specific lipids. The mycobacterial cell envelope is extremely hydrophobic, which contributes to virulence and antibiotic resistance. However, exactly how fatty acids and lipidic elements are transported across the cell envelope for cell-wall biosynthesis is unclear. Mycobacterial membrane protein Large 3 (MmpL3) is essential and required for transport of trehalose monomycolates (TMMs), precursors of MA-contg. trehalose dimycolates (TDM) and mycolyl arabinogalactan peptidoglycan, but the exact function of MmpL3 remains elusive. Here, we report a crystal structure of Mycobacterium smegmatis MmpL3 at a resoln. of 2.59 Å, revealing a monomeric mol. that is structurally distinct from all known bacterial membrane proteins. A previously unknown MmpL3 ligand, phosphatidylethanolamine (PE), was discovered inside this transporter. We also show, via native mass spectrometry, that MmpL3 specifically binds both TMM and PE, but not TDM, in the micromolar range. These observations provide insight into the function of MmpL3 and suggest a possible role for this protein in shuttling a variety of lipids to strengthen the mycobacterial cell wall.
- 50Cox, J. A. G.; Abrahams, K. A.; Alemparte, C.; Ghidelli-Disse, S.; Rullas, J.; Angulo-Barturen, I.; Singh, A.; Gurcha, S. S.; Nataraj, V.; Bethell, S.; Remuiñán, M. J.; Encinas, L.; Jervis, P. J.; Cammack, N. C.; Bhatt, A.; Kruse, U.; Bantscheff, M.; Fütterer, K.; Barros, D.; Ballell, L.; Drewes, G.; Besra, G. S. THPP target assignment reveals EchA6 as an essential fatty acid shuttle in mycobacteria. Nat. Microbiol. 2016, 1, 15006, DOI: 10.1038/nmicrobiol.2015.650https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXkvFyqtbk%253D&md5=d1704f5bc0a939f1213078f71cdcbf49THPP target assignment reveals EchA6 as an essential fatty acid shuttle in MycobacteriaCox, Jonathan A. G.; Abrahams, Katherine A.; Alemparte, Carlos; Ghidelli-Disse, Sonja; Rullas, Joaquin; Angulo-Barturen, Inigo; Singh, Albel; Gurcha, Sudagar S.; Nataraj, Vijayashankar; Bethell, Stephen; Remuinan, Modesto J.; Encinas, Lourdes; Jervis, Peter J.; Cammack, Nicholas C.; Bhatt, Apoorva; Kruse, Ulrich; Bantscheff, Marcus; Futterer, Klaus; Barros, David; Ballell, Lluis; Drewes, Gerard; Besra, Gurdyal S.Nature Microbiology (2016), 1 (2), 15006CODEN: NMAICH; ISSN:2058-5276. (Nature Publishing Group)Phenotypic screens for bactericidal compds. against drug-resistant tuberculosis are beginning to yield novel inhibitors. However, reliable target identification remains challenging. Here, we show that tetrahydropyrazo[1,5-a]pyrimidine-3-carboxamide (THPP) selectively pulls down EchA6 in a stereospecific manner, instead of the previously assigned target Mycobacterium tuberculosis MmpL3. While homologous to mammalian enoyl-CoA hydratases, EchA6 is non-catalytic yet essential and binds long-chain acyl-CoAs. THPP inhibitors compete with CoA-binding, suppress mycolic acid synthesis, and are bactericidal in a mouse model of chronic tuberculosis infection. A point mutation, W133A, abrogated THPP-binding and increased both the in vitro min. inhibitory concn. and the in vivo ED 99 in mice. Surprisingly, EchA6 interacts with selected enzymes of fatty acid synthase II (FAS-II) in bacterial two-hybrid assays, suggesting essentiality may be linked to feeding long-chain fatty acids to FAS-II. Finally, our data show that spontaneous resistance-conferring mutations can potentially obscure the actual target or alternative targets of small mol. inhibitors.
- 51Belisle, J. T.; Vissa, V. D.; Sievert, T.; Takayama, K.; Brennan, P. J.; Besra, G. S. Role of the Major Antigen of Mycobacterium tuberculosis in Cell Wall Biogenesis. Science 1997, 276, 1420, DOI: 10.1126/science.276.5317.142051https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2sXjsFOjtLg%253D&md5=b810258a42304fcb17cd57b2308aee85Role of the major antigen of Mycobacterium tuberculosis in cell wall biogenesisBelisle, John T.; Vissa, Varalakshmi D.; Sievert, Todd; Takayama, Kuni; Brennan, Patrick J.; Besra, Gurdyal S.Science (Washington, D. C.) (1997), 276 (5317), 1420-1422CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)The dominant exported proteins and protective antigens of Mycobacterium tuberculosis are a triad of related gene products called the antigen 85 (Ag85) complex. Each has also been implicated in disease pathogenesis through its fibronectin-binding capacities. A carboxylesterase domain was found within the amino acid sequences of Ag85A, B, and C, and each protein acted as a mycolyltransferase involved in the final stages of mycobacterial cell wall assembly, as shown by direct enzyme assay and site-directed mutagenesis. Furthermore, the use of an antagonist (6-azido-6-deoxy-α,α'-trehalose) of this activity demonstrates that these proteins are essential and potential targets for new antimycobacterial drugs.
- 52Favrot, L.; Grzegorzewicz, A. E.; Lajiness, D. H.; Marvin, R. K.; Boucau, J.; Isailovic, D.; Jackson, M.; Ronning, D. R. Mechanism of inhibition of Mycobacterium tuberculosis antigen 85 by ebselen. Nat. Commun. 2013, 4, 2748, DOI: 10.1038/ncomms374852https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2c7jtlarsA%253D%253D&md5=0e00cdc6af699b2ee93fce10c7cbfc0cMechanism of inhibition of Mycobacterium tuberculosis antigen 85 by ebselenFavrot Lorenza; Grzegorzewicz Anna E; Lajiness Daniel H; Marvin Rachel K; Boucau Julie; Isailovic Dragan; Jackson Mary; Ronning Donald RNature communications (2013), 4 (), 2748 ISSN:.The increasing prevalence of drug-resistant tuberculosis highlights the need for identifying new antitubercular drugs that can treat these infections. The antigen 85 (Ag85) complex has emerged as an intriguing mycobacterial drug target due to its central role in synthesizing major components of the inner and outer leaflets of the mycobacterial outer membrane. Here we identify ebselen (EBS) as a potent inhibitor of the Mycobacterium tuberculosis Ag85 complex. Mass spectrometry data show that EBS binds covalently to a cysteine residue (C209) located near the Ag85C active site. The crystal structure of Ag85C in the presence of EBS shows that C209 modification restructures the active site, thereby disrupting the hydrogen-bonded network within the active site that is essential for enzymatic activity. C209 mutations display marked decreases in enzymatic activity. These data suggest that compounds using this mechanism of action will strongly inhibit the Ag85 complex and minimize the selection of drug resistance.
- 53Mdluli, K.; Kaneko, T.; Upton, A. The tuberculosis drug discovery and development pipeline and emerging drug targets. Cold Spring Harb. Perspect. Med. 2015, 5, a021154, DOI: 10.1101/cshperspect.a021154There is no corresponding record for this reference.
- 54Ling, L. L.; Schneider, T.; Peoples, A. J.; Spoering, A. L.; Engels, I.; Conlon, B. P.; Mueller, A.; Schäberle, T. F.; Hughes, D. E.; Epstein, S.; Jones, M.; Lazarides, L.; Steadman, V. A.; Cohen, D. R.; Felix, C. R.; Fetterman, K. A.; Millett, W. P.; Nitti, A. G.; Zullo, A. M.; Chen, C.; Lewis, K. A new antibiotic kills pathogens without detectable resistance. Nature 2015, 517, 455– 459, DOI: 10.1038/nature1409854https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFOju7w%253D&md5=27e302d7d44a549a91aa52113c3b4ad8A new antibiotic kills pathogens without detectable resistanceLing, Losee L.; Schneider, Tanja; Peoples, Aaron J.; Spoering, Amy L.; Engels, Ina; Conlon, Brian P.; Mueller, Anna; Schaberle, Till F.; Hughes, Dallas E.; Epstein, Slava; Jones, Michael; Lazarides, Linos; Steadman, Victoria A.; Cohen, Douglas R.; Felix, Cintia R.; Fetterman, K. Ashley; Millett, William P.; Nitti, Anthony G.; Zullo, Ashley M.; Chen, Chao; Lewis, KimNature (London, United Kingdom) (2015), 517 (7535), 455-459CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)Antibiotic resistance is spreading faster than the introduction of new compds. into clin. practice, causing a public health crisis. Most antibiotics were produced by screening soil microorganisms, but this limited resource of cultivable bacteria was overmined by the 1960s. Synthetic approaches to produce antibiotics have been unable to replace this platform. Uncultured bacteria make up approx. 99% of all species in external environments, and are an untapped source of new antibiotics. We developed several methods to grow uncultured organisms by cultivation in situ or by using specific growth factors. Here we report a new antibiotic that we term teixobactin, discovered in a screen of uncultured bacteria. Teixobactin inhibits cell wall synthesis by binding to a highly conserved motif of lipid II (precursor of peptidoglycan) and lipid III (precursor of cell wall teichoic acid). We did not obtain any mutants of Staphylococcus aureus or Mycobacterium tuberculosis resistant to teixobactin. The properties of this compd. suggest a path towards developing antibiotics that are likely to avoid development of resistance.
- 55
Isoniazid was included as a positive control (0.9–29.2 μM), and comparison, with it being an antibiotic that targets the cell wall and has rapid bactericidal activity.
There is no corresponding record for this reference. - 56Hendon-Dunn, C. L.; Pertinez, H.; Marriott, A. A. N.; Hatch, K. A.; Allnutt, J. C.; Davies, G.; Bacon, J. Regrowth of Mycobacterium tuberculosis Populations Exposed to Antibiotic Combinations Is Due to the Presence of Isoniazid and Not Bacterial Growth Rate. Antimicrob. Agents Chemother. 2019, 63, e00570-19 DOI: 10.1128/AAC.00570-19There is no corresponding record for this reference.
- 57Banerjee, A.; Dubnau, E.; Quemard, A.; Balasubramanian, V.; Um, K. S.; Wilson, T.; Collins, D.; de Lisle, G.; Jacobs, W. R. inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosis. Science 1994, 263, 227, DOI: 10.1126/science.828467357https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2cXhsFylt7c%253D&md5=12f3eabbca02009180332c1e65db8c15inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosisBanerjee, Asesh; Dubnau, Eugenie; Quemard, Annaik; Balasubramanian, V.; Um, Kyung Sun; Wilson, Theresa; Collins, Des; de Lisle, Geoffrey; Jacobs, William, R., Jr.Science (Washington, DC, United States) (1994), 263 (5144), 227-30CODEN: SCIEAS; ISSN:0036-8075.Isoniazid (isonicotinic acid hydrazide, INH) is one of the most widely used antituberculosis drugs, yet its precise target of action on Mycobacterium tuberculosis is unknown. A missense mutation within the mycobacterial inhA gene was shown to confer resistance to both INH and ethionamide (ETH) in M. smegmatis and in M. bovis. The wild-type inhA gene also conferred INH and ETH resistance when transferred on a multicopy plasmid vector to M. smegmatis and M. bovis BCG. The InhA protein shows a significant sequence conservation with the Escherichia coli enzyme EnvM, and cell-free assays indicate that it may be involved in mycolic acid biosynthesis. These results suggest that InhA is likely a primary target of action for INH and ETH.
- 58Takayama, K.; Kilburn, J. O. Inhibition of synthesis of arabinogalactan by ethambutol in Mycobacterium smegmatis. Antimicrob. Agents Chemother. 1989, 33, 1493, DOI: 10.1128/AAC.33.9.149358https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL1MXmt1yrsbg%253D&md5=568b1756e3a57a9ebd1a871e4df22e5dInhibition of synthesis of arabinogalactan by ethambutol in Mycobacterium smegmatisTakayama, Kuni; Kilburn, James O.Antimicrobial Agents and Chemotherapy (1989), 33 (9), 1493-9CODEN: AMACCQ; ISSN:0066-4804.Ethambutol at 3.0 μg/mL inhibited the transfer of label from D-[14C]glucose into the D-arabinose residue of arabinogalactan in whole cells of a drug-susceptible strain of M. smegmatis. This inhibition began almost immediately after exposure of the cells to the drug. When drug-resistant M. smegmatis was used in a similar expt., no such drug inhibition was detected. A much higher concn. of ethambutol (>50 μg/mL) was required to show this inhibition. The drug also inhibited synthesis of arabinose-contg. oligosaccharides when a cell-free enzyme system was used. These results suggest that the site of action of ethambutol is somewhere on the pathway between the conversion of D-glucose to D-arabinose and the transfer of arabinose into arabinogalactan. The primary mode of action of ethambutol appears to be inhibition of arabinogalactan synthesis.
- 59Tahlan, K.; Wilson, R.; Kastrinsky, D. B.; Arora, K.; Nair, V.; Fischer, E.; Barnes, S. W.; Walker, J. R.; Alland, D.; Barry, C. E.; Boshoff, H. I. SQ109 Targets MmpL3, a Membrane Transporter of Trehalose Monomycolate Involved in Mycolic Acid Donation to the Cell Wall Core of Mycobacterium tuberculosis. Antimicrob. Agents Chemother. 2012, 56, 1797, DOI: 10.1128/AAC.05708-1159https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XltV2ru7k%253D&md5=6309318d9e4d2f0c75a9fed20b8108eaSQ109 targets MmpL3, a membrane transporter of trehalose monomycolate involved in mycolic acid donation to the cell wall core of Mycobacterium tuberculosisTahlan, Kapil; Wilson, Regina; Kastrinsky, David B.; Arora, Kriti; Nair, Vinod; Fischer, Elizabeth; Barnes, S. Whitney; Walker, John R.; Alland, David; Barry, Clifton E., III; Boshoff, Helena I.Antimicrobial Agents and Chemotherapy (2012), 56 (4), 1797-1809CODEN: AMACCQ; ISSN:0066-4804. (American Society for Microbiology)SQ109, a 1,2-diamine related to ethambutol, is currently in clin. trials for the treatment of tuberculosis, but its mode of action remains unclear. Here, we demonstrate that SQ109 disrupts cell wall assembly, as evidenced by macromol. incorporation assays and ultrastructural analyses. SQ109 interferes with the assembly of mycolic acids into the cell wall core of Mycobacterium tuberculosis, as bacilli exposed to SQ109 show immediate inhibition of trehalose dimycolate (TDM) prodn. and fail to attach mycolates to the cell wall arabinogalactan. These effects were not due to inhibition of mycolate synthesis, since total mycolate levels were unaffected, but instead resulted in the accumulation of trehalose monomycolate (TMM), the precursor of TDM and cell wall mycolates. In vitro assays using purified enzymes showed that this was not due to inhibition of the secreted Ag85 mycolyltransferases. We were unable to achieve spontaneous generation of SQ109-resistant mutants; however, analogs of this compd. that resulted in similar shutdown of TDM synthesis with concomitant TMM accumulation were used to spontaneously generate resistant mutants that were also cross-resistant to SQ109. Whole-genome sequencing of these mutants showed that these all had mutations in the essential mmpL3 gene, which encodes a transmembrane transporter. Our results suggest that MmpL3 is the target of SQ109 and that MmpL3 is a transporter of mycobacterial TMM.
- 60Prosser, G. A.; de Carvalho, L. P. S. Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserine. FEBS Journal 2013, 280, 1150– 1166, DOI: 10.1111/febs.1210860https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXis1Wmtbw%253D&md5=01551bd01443b8a028e6dde0ea3e03f8Kinetic mechanism and inhibition of Mycobacterium tuberculosis D-alanine:D-alanine ligase by the antibiotic D-cycloserineProsser, Gareth A.; de Carvalho, Luiz Pedro S.FEBS Journal (2013), 280 (4), 1150-1166CODEN: FJEOAC; ISSN:1742-464X. (Wiley-Blackwell)D-Cycloserine (DCS) is an antibiotic that is currently used in second-line treatment of tuberculosis. DCS is a structural analog of D-alanine, and targets two enzymes involved in the cytosolic stages of peptidoglycan synthesis: alanine racemase (Alr) and D-alanine:D-alanine ligase (Ddl). The mechanisms of inhibition of DCS have been well-assessed using Alr and Ddl enzymes from various bacterial species, but little is known regarding the interactions of DCS with the mycobacterial orthologues of these enzymes. We have over-expressed and purified recombinant Mycobacterium tuberculosis Ddl (MtDdl; Rv2981c), and report a kinetic examn. of the enzyme with both its native substrate and DCS. MtDdl is activated by K+, follows an ordered ter ter mechanism and displays distinct affinities for D-Ala at each D-Ala binding site (Km,D-Ala1 = 0.075 mm,Km,D-Ala2 = 3.6 mm). ATP is the first substrate to bind and is necessary for subsequent binding of D-alanine or DCS. The pH dependence of MtDdl kinetic parameters indicate that general base chem. is involved in the catalytic step. DCS was found to competitively inhibit D-Ala binding at both MtDdl D-Ala sites with equal affinity (Ki,DCS1 = 14 μm,Ki,DCS2 = 25 μm); however, each enzyme active site can only accommodate a single DCS mol. at a given time. The pH dependence of Ki,DCS2 revealed a loss of DCS binding affinity at high pH (pKa = 7.5), suggesting that DCS binds optimally in the zwitterionic form. The results of this study may assist in the design and development of novel Ddl-specific inhibitors for use as anti-mycobacterial agents.
- 61Briffotaux, J.; Liu, S.; Gicquel, B. Genome-Wide Transcriptional Responses of Mycobacterium to Antibiotics. Front. Microbiol. 2019, 10, 249, DOI: 10.3389/fmicb.2019.0024961https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3cbhtlSkug%253D%253D&md5=aff38e10263f3b6e7c854010022fbee1Genome-Wide Transcriptional Responses of Mycobacterium to AntibioticsBriffotaux Julien; Liu Shengyuan; Gicquel Brigitte; Briffotaux Julien; Gicquel Brigitte; Gicquel BrigitteFrontiers in microbiology (2019), 10 (), 249 ISSN:1664-302X.Antibiotics can stimulate or depress gene expression in bacteria. The analysis of transcriptional responses of Mycobacterium to antimycobacterial compounds has improved our understanding of the mode of action of various drug classes and the efficacy and effect of such compounds on the global metabolism of Mycobacterium. This approach can provide new insights for known antibiotics, for example those currently used for tuberculosis treatment, as well as help to identify the mode of action and predict the targets of new compounds identified by whole-cell screening assays. In addition, changes in gene expression profiles after antimycobacterial treatment can provide information about the adaptive ability of bacteria to escape the effects of antibiotics and allow monitoring of the physiology of the bacteria during treatment. Genome-wide expression profiling also makes it possible to pinpoint genes differentially expressed between drug sensitive Mycobacterium and multidrug-resistant clinical isolates. Finally, genes involved in adaptive responses and drug tolerance could become new targets for improving the efficacy of existing antibiotics.
- 62Johnsson, K.; King, D. S.; Schultz, P. G. Studies on the Mechanism of Action of Isoniazid and Ethionamide in the Chemotherapy of Tuberculosis. J. Am. Chem. Soc. 1995, 117, 5009– 5010, DOI: 10.1021/ja00122a03862https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2MXlt1Srt7Y%253D&md5=3492989087bfdd266d0d8c7b10037225Studies on the Mechanism of Action of Isoniazid and Ethionamide in the Chemotherapy of TuberculosisJohnsson, Kai; King, David S.; Schultz, Peter G.Journal of the American Chemical Society (1995), 117 (17), 5009-10CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)The inactivation of the enoyl-reductase InhA from Mycobacterium tuberculosis by reactive intermediates formed during the oxidn. of isoniazid and ethionamide was studied. Both drugs can generate electrophilic intermediates capable of reacting with a nucleophilic group of InhA, leading to its inactivation. After inactivation of InhA by isoniazid, one mol. of isoniazid per InhA is covalently bound to the enzyme. Mapping studies suggest that Cys243 is the residue modified in the course of the inactivation.
- 63Milanes, C. L.; Pernalete, N.; Starosta, R.; Perez-Gonzalez, M.; Paz-Martinez, V.; Bellorin-Font, E. Altered response of adenylate cyclase to parathyroid hormone during compensatory renal growth. Kidney Int. 1989, 36, 802– 9, DOI: 10.1038/ki.1989.26563https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK3cXjsVWjtw%253D%253D&md5=91a1c3931d282d6703be188411f389ecAltered response of adenylate cyclase to parathyroid hormone during compensatory renal growthMilanes, Carmen L.; Pernalete, Nidia; Starosta, Rebeca; Perez-Gonzalez, Margarita; Paz-Martinez, Virgilio; Bellorin-Font, EzequielKidney International (1989), 36 (5), 802-9CODEN: KDYIA5; ISSN:0085-2538.The loss of renal mass is assocd. with functional adaptations in the remaining nephrons to maintain homeostasis. Although parathyroid hormone (PTH) is important in the adaptations to phosphate, the mechanisms are not completely defined. In the present study, the response of the adenylate cyclase system to PTH was examd. in renal cortical membranes of rat kidneys ten days after unilateral nephrectomy. The kidneys obtained at the time of the initial nephrectomy were used as controls. Unilateral nephrectomy resulted in contralateral compensatory renal growth, as demonstrated by a 24% increase in wt. in the remaining kidney. Glomerular filtration rate (GFR) after unilateral nephrectomy was 62% of the control, whereas basal fractional phosphate excretion was higher in rats with unilateral nephrectomy (7.7% vs. 2.9%). PTH infusion resulted in a similar increase of fractional phosphate excretion and urinary cAMP in both groups. In the absence of added guanine nucleotides, PTH-dependent adenylate cyclase activity in cortical membranes from kidneys with compensatory growth was decreased as compared to controls (Vmax 807.5 pmol cAMP/mg protein/30 min vs. 1384.8, resp.). The apparent affinity for PTH stimulation of adenylate cyclase (Kact) was unchanged. Magnesium-dependent adenylate cyclase activity was also decreased in the membranes from kidneys with compensatory growth. However, the kinetics of adenylate cyclase for the substrates ATP-Mg or ATP-Mn were similar. The addn. of Gpp(NH)p resulted in a similar maximal response to PTH in the two groups, indicating an increased response of the enzyme to PTH in the presence of the guanine nucleotide. Cholera toxin-dependent ADP-ribosylation of the stimulatory guanine nucleotide binding protein (Gs) showed a marked decrease in the apparent content of the alpha subunit in membranes from kidneys with compensatory growth compared to controls. On the contrary, pertussis toxin-dependent ADP-ribosylation of the inhibitory guanine nucleotide binding protein (Gi) did not show differences in the content of the alpha subunit in both groups of membranes. Since the transduction of the hormone signal from the receptor is mediated by G proteins, the results suggest that during compensatory renal growth a decrease in the alpha subunit of Gs could account for the impaired response of adenylate cyclase to PTH in vitro, which could be overcome by high concns. of guanine nucleotides.
- 64Bacon, J.; Alderwick, L. J.; Allnutt, J. A.; Gabasova, E.; Watson, R.; Hatch, K. A.; Clark, S. O.; Jeeves, R. E.; Marriott, A.; Rayner, E.; Tolley, H.; Pearson, G.; Hall, G.; Besra, G. S.; Wernisch, L.; Williams, A.; Marsh, P. D. Non-replicating Mycobacterium tuberculosis elicits a reduced infectivity profile with corresponding modifications to the cell wall and extracellular matrix. PLoS One 2014, 9, e87329 DOI: 10.1371/journal.pone.008732964https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhsVGrtrnJ&md5=4d7773618b4185c2ee98f32dd97bd15bNon-replicating Mycobacterium tuberculosis elicits a reduced infectivity profile with corresponding modifications to the cell wall and extracellular matrixBacon, Joanna; Alderwick, Luke J.; Allnutt, Jon A.; Gabasova, Evelina; Watson, Robert; Hatch, Kim A.; Clark, Simon O.; Jeeves, Rose E.; Marriott, Alice; Rayner, Emma; Tolley, Howard; Pearson, Geoff; Hall, Graham; Besra, Gurdyal S.; Wernisch, Lorenz; Williams, Ann; Marsh, Philip D.PLoS One (2014), 9 (2), e87329/1-e87329/17, 17 pp.CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)A key feature of Mycobacterium tuberculosis is its ability to become dormant in the host. Little is known of the mechanisms by which these bacilli are able to persist in this state. Therefore, the focus of this study was to emulate environmental conditions encountered by M. tuberculosis in the granuloma, and det. the effect of such conditions on the physiol. and infectivity of the organism. Non-replicating persistent (NRP) M. tuberculosis was established by the gradual depletion of nutrients in an oxygen-replete and controlled environment. In contrast to rapidly dividing bacilli, NRP bacteria exhibited a distinct phenotype by accumulating an extracellular matrix rich in free mycolate and lipoglycans, with increased arabinosylation. Microarray studies demonstrated a substantial down-regulation of genes involved in energy metab. in NRP bacteria. Despite this redn. in metabolic activity, cells were still able to infect guinea pigs, but with a delay in the development of disease when compared to exponential phase bacilli. Using these approaches to investigate the interplay between the changing environment of the host and altered physiol. of NRP bacteria, this study sheds new light on the conditions that are pertinent to M. tuberculosis dormancy and how this organism could be establishing latent disease.
- 65Rose, J. D.; Maddry, J. A.; Comber, R. N.; Suling, W. J.; Wilson, L. N.; Reynolds, R. C. Synthesis and biological evaluation of trehalose analogs as potential inhibitors of mycobacterial cell wall biosynthesis. Carbohydr. Res. 2002, 337, 105– 120, DOI: 10.1016/S0008-6215(01)00288-965https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD38XptlCjsg%253D%253D&md5=84944b2ab25eb9763377b537c6515a29Synthesis and biological evaluation of trehalose analogs as potential inhibitors of mycobacterial cell wall biosynthesisRose, Jerry D.; Maddry, Joseph A.; Comber, Robert N.; Suling, William J.; Wilson, Larry N.; Reynolds, Robert C.Carbohydrate Research (2002), 337 (2), 105-120CODEN: CRBRAT; ISSN:0008-6215. (Elsevier Science Ltd.)Analogs of trehalose are reported that were designed to interfere with mycolylation pathways in the mycobacterial cell wall. Several derivs. of 6,6'-dideoxytrehalose, including N,N'-dialkylamino and 6,6'-bis(sulfonamido) analogs, were prepd. and evaluated for antimycobacterial activity against Mycobacterium tuberculosis H37Ra and a panel of clin. isolates of Mycobacterium avium. 6,6'-Diaminotrehalose and its diazido precursor were both inactive, but significant activity apparently related to aliph. chain length was found among the sulfonamides, N-alkylamines, and one of the amidines.
- 66Barry, C. S.; Backus, K. M.; Barry, C. E.; Davis, B. G. ESI-MS Assay of M. tuberculosis Cell Wall Antigen 85 Enzymes Permits Substrate Profiling and Design of a Mechanism-Based Inhibitor. J. Am. Chem. Soc. 2011, 133, 13232– 13235, DOI: 10.1021/ja204249p66https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXpvFSnsbc%253D&md5=8d09f08160f78d634a08e87a0b516babESI-MS Assay of M. tuberculosis Cell Wall Antigen 85 Enzymes Permits Substrate Profiling and Design of a Mechanism-Based InhibitorBarry, Conor S.; Backus, Keriann M.; Barry, Clifton E., III; Davis, Benjamin G.Journal of the American Chemical Society (2011), 133 (34), 13232-13235CODEN: JACSAT; ISSN:0002-7863. (American Chemical Society)Mycobacterium tuberculosis Antigen 85 enzymes are vital to the integrity of the highly impermeable cell envelope and are potential therapeutic targets. Kinetic anal. using a label-free assay revealed both mechanistic details and a substrate profile that allowed the design and construction of a selective in vitro mechanism-based inhibitor.
- 67Viljoen, A.; Richard, M.; Nguyen, P. C.; Fourquet, P.; Camoin, L.; Paudal, R. R.; Gnawali, G. R.; Spilling, C. D.; Cavalier, J.-F.; Canaan, S.; Blaise, M.; Kremer, L. Cyclipostins and cyclophostin analogs inhibit the antigen 85C from Mycobacterium tuberculosis both in vitro and in vivo. J. Biol. Chem. 2018, 293, 2755– 2769, DOI: 10.1074/jbc.RA117.00076067https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXjt1Shsro%253D&md5=50d705aacc7e65e5b0fb448305c4e160Cyclipostins and cyclophostin analogs inhibit the antigen 85C from Mycobacterium tuberculosis both in vitro and in vivoViljoen, Albertus; Richard, Matthias; Nguyen, Phuong Chi; Fourquet, Patrick; Camoin, Luc; Paudal, Rishi R.; Gnawali, Giri R.; Spilling, Christopher D.; Cavalier, Jean-Francois; Canaan, Stephane; Blaise, Mickael; Kremer, LaurentJournal of Biological Chemistry (2018), 293 (8), 2755-2769CODEN: JBCHA3; ISSN:0021-9258. (American Society for Biochemistry and Molecular Biology)An increasing prevalence of cases of drug-resistant tuberculosis requires the development of more efficacious chemotherapies. The authors previously reported the discovery of a new class of cyclipostins and cyclophostin (CyC) analogs exhibiting potent activity against Mycobacterium tuberculosis both in vitro and in infected macrophages. Competitive labeling/enrichment assays combined with MS have identified several serine or cysteine enzymes in lipid and cell wall metab. as putative targets of these CyC compds. These targets included members of the antigen 85 (Ag85) complex (i.e. Ag85A, Ag85B, and Ag85C), responsible for biosynthesis of trehalose dimycolate and mycolylation of arabinogalactan. Herein, the authors used biochem. and structural approaches to validate the Ag85 complex as a pharmacol. target of the CyC analogs. The authors found that CyC7β, CyC8β, and CyC17 bind covalently to the catalytic Ser124 residue in Ag85C; inhibit mycolyltransferase activity (i.e. the transfer of a fatty acid mol. onto trehalose); and reduce triacylglycerol synthase activity, a property previously attributed to Ag85A. Supporting these results, an x-ray structure of Ag85C in complex with CyC8β disclosed that this inhibitor occupies Ag85C's substrate-binding pocket. Importantly, metabolic labeling of M. tuberculosis cultures revealed that the CyC compds. impair both trehalose dimycolate synthesis and mycolylation of arabinogalactan. Overall, the study provides compelling evidence that CyC analogs can inhibit the activity of the Ag85 complex in vitro and in mycobacteria, opening the door to a new strategy for inhibiting Ag85. The high-resoln. crystal structure obtained will further guide the rational optimization of new CyC scaffolds with greater specificity and potency against M. tuberculosis.
- 68Crespo, M.; Martinez, M.; de Pablo, E. Activation volumes for intramolecular oxidative C–X (X = H, F, Cl or Br) addition to platinum(II) imine complexes as a proof of the intimate mechanism. J. Chem. Soc., Dalton Trans. 1997, 1231– 1236, DOI: 10.1039/a606872c68https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK2sXivVCqsLc%253D&md5=6580b106746a62d72545615d45e26c27Activation volumes for intramolecular oxidative C-X (X = H, F, Cl or Br) addition to platinum(II) imine complexes as a proof of the intimate mechanismCrespo, Margarita; Martinez, Manuel; de Pablo, EstherJournal of the Chemical Society, Dalton Transactions: Inorganic Chemistry (1997), (7), 1231-1235CODEN: JCDTBI; ISSN:0300-9246. (Royal Society of Chemistry)The kinetics of C-X (X = H, F, Cl or Br) bond activation of ring-substituted, PhCH:NCH2Ph, type imines via intramol. oxidative addn. to Pt(II) complexes was studied in acetone and toluene soln. at different temps. and pressures. Although the activation parameters detd. are within the range expected, the latter is extremely lage (ΔH⧧ from 25 to 70 kJ mol-1, ΔS⧧ from -220 to -45 J K-1 mol-1, ΔV⧧ from -31.2 to -9.5 cm3 mol-1). No differences were found for the reactions carried out in acetone or toluene, indicating that no polar transition state is formed during the reaction and that a common highly ordered three-centered C-Pt-X interaction is present for all the imines used. A good correlation was also obtained for the ΔS⧧ and ΔV⧧ values, independently of the solvent used, confirming the nonpolarity of the transition state. A deviation from this pattern is obsd. only for fluorinated imines both in acetone and toluene solns.; this result is interpreted by considering an earlier transition state for the oxidative addn. of C-F that has not yet produced an important vol. contraction of the Pt center despite the important spatial organization of the ligand, as shown by the very neg. values of ΔS⧧.
- 69Clark, P. W.; Dyke, S. F.; Smith, G.; Kennard, C. H. L. The cyclopalladation of benzylidenebenzylamines. J. Organomet. Chem. 1987, 330, 447– 460, DOI: 10.1016/S0022-328X(00)99057-069https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaL1cXksFCqsLc%253D&md5=587bb5c956cc1c516de4625448348c12The cyclopalladation of benzylidenebenzylaminesClark, P. W.; Dyke, S. F.; Smith, G.; Kennard, C. H. L.Journal of Organometallic Chemistry (1987), 330 (3), 447-60CODEN: JORCAI; ISSN:0022-328X.Cyclopalladation of benzylidenebenzylamines I (R = H; R1, R2, R3 = H, OMe; R4 = H, Me, Ph) with 1 equiv Li2PdCl4 gave dimers II (same R1-R4, X = Cl). Reaction of I (R = Br, same R1-R4) with bis(dibenzylideneacetone)palladium (III) gave II (X = Br, same R1-R4). The cyclopalladated products were characterized by 1H and 13C NMR spectroscopy and as their products derived from further reaction with acetylacetonate, PPh3, or NaOAc. Reinvestigation of the reaction of I (R - R4 = H) with Pd(OAc)2 gave the acetyl-substituted dimer IV, which reacted with CO to give phthalimidine V. Cyclopalladation of 2-BrC6H4CH2N:CR5Ph (R5 = H, Me, Ph) with III gave dimeric palladocycles which reacted with acetylacetonate to give monomers VI (same R5). The x-ray crystal structures of VI (R5 = H) and a monomeric acetylacetonate deriv. of II (R1-R4 = H) were detd. Competitive cyclopalladations of several N-6-bromobenzyl-6'-bromobenzalimines with III gave dimers VII (R6, R7 = H, OMe).
- 70Anderson, C. M.; Crespo, M.; Kfoury, N.; Weinstein, M. A.; Tanski, J. M. Regioselective C–H Activation Preceded by Csp2–Csp3 Reductive Elimination from Cyclometalated Platinum(IV) Complexes. Organometallics 2013, 32, 4199– 4207, DOI: 10.1021/om400398g70https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhtFOgtr%252FE&md5=9068798fd8cbe7aff054746d16192b96Regioselective C-H Activation Preceded by Csp2-Csp3 Reductive Elimination from Cyclometalated Platinum(IV) ComplexesAnderson, Craig M.; Crespo, Margarita; Kfoury, Nicole; Weinstein, Michael A.; Tanski, Joseph M.Organometallics (2013), 32 (15), 4199-4207CODEN: ORGND7; ISSN:0276-7333. (American Chemical Society)Reductive elimination reactions of the cyclometalated Pt(IV) compds. [PtMe2Cl{C6H4CH:NCH2(4-ClC6H4)}L] and [PtMe2Br{C6H4CH:NCH2Ph}L] (L = SMe2, PPh3) to form Csp3-Csp2 bonds, followed by either exclusive Csp2-H bond activation (L = SMe2) or competition between Csp2-H and Csp3-H bond activation (L = PPh3), are reported. Isomerization to give endo products instead of the expected exo complex was obsd. for the ligand C6H4CH:NCH2(2-BrC6H5), and formation of an endo six-membered platinacycle occurs for the ligand 2,4,6-Me3C6H2CH:NCH2(2-BrC6H4).
- 71Zhang, T.; Li, S.-Y.; Nuermberger, E. L. Autoluminescent Mycobacterium tuberculosis for Rapid, Real-Time, Non-Invasive Assessment of Drug and Vaccine Efficacy. PLoS One 2012, 7, e29774 DOI: 10.1371/journal.pone.002977471https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsVCqsLY%253D&md5=9576cdfd3c6a058a826fd9736dca8b40Autoluminescent Mycobacterium tuberculosis for rapid, real-time, non-invasive assessment of drug and vaccine efficacyZhang, Tianyu; Li, Si-Yang; Nuermberger, Eric L.PLoS One (2012), 7 (1), e29774CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Preclin. efforts to discover and develop new drugs and vaccines for tuberculosis are hampered by the reliance on colony-forming unit (CFU) counts as primary outcomes for in vivo efficacy studies and the slow growth of Mycobacterium tuberculosis. The utility of bioluminescent M. tuberculosis reporter strains for real-time in vitro and ex vivo assessment of drug and vaccine activity has been demonstrated but a simple, non-invasive, real-time surrogate marker to replace CFU counts for real-time evaluation of drug and vaccine efficacy in vivo has not been described. We describe the development of a fully virulent and stable autoluminescent strain of M. tuberculosis and proof-of-concept expts. demonstrating its utility for in vivo bioluminescence imaging to assess the efficacy of new drugs and vaccines for tuberculosis in a mouse model. Relative light unit (RLU) counts paralleled CFU counts during the active phase of bacterial growth, with a lower limit of detection of approx. 106 CFU in live, anesthetized mice. Expts. distinguishing active from inactive anti-tuberculosis drugs and bacteriostatic drug effects from bactericidal effects were completed in less than 5 days. The ability of a recombinant BCG vaccine to limit bacterial growth was demonstrated within 3 wk. Use of this autoluminescent reporter strain has the potential to drastically reduce the time, effort, animals and costs consumed in the evaluation of drug activity in vitro and the in vivo assessment of drug and vaccine efficacy.
- 72Liu, Y.; Gao, Y.; Liu, J.; Tan, Y.; Liu, Z.; Chhotaray, C.; Jiang, H.; Lu, Z.; Chiwala, G.; Wang, S.; Makafe, G.; Islam, M. M.; Hameed, H. M. A.; Cai, X.; Wang, C.; Li, X.; Tan, S.; Zhang, T. The compound TB47 is highly bactericidal against Mycobacterium ulcerans in a Buruli ulcer mouse model. Nat. Commun. 2019, 10, 524, DOI: 10.1038/s41467-019-08464-y72https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXnt1Wks74%253D&md5=f1a01c719038fbbd9f9ce8972e33be27The compound TB47 is highly bactericidal against Mycobacterium ulcerans in a Buruli ulcer mouse modelLiu, Yang; Gao, Yamin; Liu, Jianxiong; Tan, Yaoju; Liu, Zhiyong; Chhotaray, Chiranjibi; Jiang, Huofeng; Lu, Zhili; Chiwala, Gift; Wang, Shuai; Makafe, Gaelle; Islam, Md. Mahmudul; Hameed, H. M. Adnan; Cai, Xingshan; Wang, Changwei; Li, Xinjie; Tan, Shouyong; Zhang, TianyuNature Communications (2019), 10 (1), 524CODEN: NCAOBW; ISSN:2041-1723. (Nature Research)Buruli ulcer (BU) is an emerging infectious disease that causes disfiguring skin ulcers. The causative agent, Mycobacterium ulcerans, secretes toxin called mycolactone that triggers inflammation and immunopathol. Existing treatments are lengthy and consist of drugs developed for tuberculosis. Here, we report that a pyrazolo[1,5-a]pyridine-3-carboxamide, TB47, is highly bactericidal against M. ulcerans both in vitro and in vivo. In the validated mouse model of BU, TB47 alone reduces M. ulcerans burden in mouse footpads by more than 2.5 log10 CFU compared to the std. BU treatment regimen recommended by the WHO. We show that mutations of ubiquinol-cytochrome C reductase cytochrome subunit B confer resistance to TB47 and the dissimilarity of CydABs from different mycobacteria may account for their differences in susceptibility to TB47. TB47 is highly potent against M. ulcerans and possesses desirable pharmacol. attributes and low toxicity that warrant further assessment of this agent for treatment of BU.
- 73Nateche, F.; Martin, A.; Baraka, S.; Palomino, J. C.; Khaled, S.; Portaels, F. Application of the resazurin microtitre assay for detection of multidrug resistance in Mycobacterium tuberculosis in Algiers. J. Med. Microbiol. 2006, 55, 857– 860, DOI: 10.1099/jmm.0.46513-073https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BD28zlvVCgsA%253D%253D&md5=152ee2d0bd954157d5c27338cdddf3e5Application of the resazurin microtitre assay for detection of multidrug resistance in Mycobacterium tuberculosis in AlgiersNateche Farida; Baraka Saliha; Khaled Safia; Nateche Farida; Martin Anandi; Palomino Juan Carlos; Portaels FrancoiseJournal of medical microbiology (2006), 55 (Pt 7), 857-860 ISSN:0022-2615.This study assessed the performance of a rapid, low-cost, colorimetric method, the resazurin microtitre assay (REMA) plate method, for the detection of resistance to isoniazid and rifampicin in 136 clinical isolates of Mycobacterium tuberculosis from two hospitals in Algiers. MICs were determined and the results were compared with those obtained with the conventional proportion method on Lowenstein-Jensen medium. Excellent results were obtained for the REMA plate method, with a sensitivity of 100 % for both isoniazid and rifampicin and a specificity of 98.3 and 99.2 %, respectively. The REMA plate method appears to be a reliable method for the rapid determination of multidrug-resistant tuberculosis and is a good alternative for use in resource-limited countries such as Algeria.
- 74Mosaei, H.; Molodtsov, V.; Kepplinger, B.; Harbottle, J.; Moon, C. W.; Jeeves, R. E.; Ceccaroni, L.; Shin, Y.; Morton-Laing, S.; Marrs, E. C. L.; Wills, C.; Clegg, W.; Yuzenkova, Y.; Perry, J. D.; Bacon, J.; Errington, J.; Allenby, N. E. E.; Hall, M. J.; Murakami, K. S.; Zenkin, N. Mode of Action of Kanglemycin A, an Ansamycin Natural Product that Is Active against Rifampicin-Resistant Mycobacterium tuberculosis. Mol. Cell 2018, 72, 263– 274.e5, DOI: 10.1016/j.molcel.2018.08.02874https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhslKhsr%252FF&md5=63369346e69244e37dd37f851734a0b8Mode of Action of Kanglemycin A, an Ansamycin Natural Product that Is Active against Rifampicin-Resistant Mycobacterium tuberculosisMosaei, Hamed; Molodtsov, Vadim; Kepplinger, Bernhard; Harbottle, John; Moon, Christopher William; Jeeves, Rose Elizabeth; Ceccaroni, Lucia; Shin, Yeonoh; Morton-Laing, Stephanie; Marrs, Emma Claire Louise; Wills, Corinne; Clegg, William; Yuzenkova, Yulia; Perry, John David; Bacon, Joanna; Errington, Jeff; Allenby, Nicholas Edward Ellis; Hall, Michael John; Murakami, Katsuhiko S.; Zenkin, NikolayMolecular Cell (2018), 72 (2), 263-274.e5CODEN: MOCEFL; ISSN:1097-2765. (Elsevier Inc.)Antibiotic-resistant bacterial pathogens pose an urgent healthcare threat, prompting a demand for new medicines. We report the mode of action of the natural ansamycin antibiotic kanglemycin A (KglA). KglA binds bacterial RNA polymerase at the rifampicin-binding pocket but maintains potency against RNA polymerases contg. rifampicin-resistant mutations. KglA has antibiotic activity against rifampicin-resistant Gram-pos. bacteria and multidrug-resistant Mycobacterium tuberculosis (MDR-M. tuberculosis). The X-ray crystal structures of KglA with the Escherichia coli RNA polymerase holoenzyme and Thermus thermophilus RNA polymerase-promoter complex reveal an altered-compared with rifampicin-conformation of KglA within the rifampicin-binding pocket. Unique deoxysugar and succinate ansa bridge substituents make addnl. contacts with a sep., hydrophobic pocket of RNA polymerase and preclude the formation of initial dinucleotides, resp. Previous ansa-chain modifications in the rifamycin series have proven unsuccessful. Thus, KglA represents a key starting point for the development of a new class of ansa-chain derivatized ansamycins to tackle rifampicin resistance.
- 75James, B. W.; Williams, A.; Marsh, P. D. The physiology and pathogenicity of Mycobacterium tuberculosis grown under controlled conditions in a defined medium. J. Appl. Microbiol. 2000, 88, 669– 77, DOI: 10.1046/j.1365-2672.2000.01020.x75https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXjtl2jurc%253D&md5=8f9a6b179536fbfdeca405fd9373acecThe physiology and pathogenicity of Mycobacterium tuberculosis grown under controlled conditions in a defined mediumJames, B. W.; Williams, A.; Marsh, P. D.Journal of Applied Microbiology (2000), 88 (4), 669-677CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)A chem.-defined culture medium was developed which supported batch growth of Mycobacterium tuberculosis, strain H37Rv, at a min. doubling time of 14.7 h. This medium also facilitated chemostat culture of M. tuberculosis at a const. doubling time of 24 h. Chemostat growth was optimized at a dissolved oxygen tension of 20% (vol./vol.) and 0.2% (vol./vol.) Tween-80. Chemostat cultures were dispersed suspensions of single bacilli (1.5-3 μm long), or small aggregates, at a mean d. of log10 8.3 cfu ml-1. A limited no. of amino acids was utilized (alanine, asparagine, aspartate and serine were depleted by > 50%; glycine, arginine, isoleucine, leucine and phenylalanine, by approx. 40%). Chemostat-grown cells were pathogenic in aerosol-infected guinea pigs, producing disseminated infection similar to that caused by plate-grown cells. Cells from chemostat culture were significantly more invasive for J774A.1 mouse macrophages than agar- or batch-grown cells. This study demonstrates the suitability of chemostat culture for the growth of pathogenic mycobacteria in a defined physiol. state with potential applications for the controlled prodn. of mycobacterial components for therapeutic and vaccine applications.
- 76Lambert, R. J. W. Susceptibility testing: inoculum size dependency of inhibition using the Colworth MIC technique. J. Appl. Microbiol. 2000, 89, 275– 279, DOI: 10.1046/j.1365-2672.2000.01105.x76https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXmvVCntLw%253D&md5=3390c45f9763d95dd0f0efffaa4f54eeSusceptibility testing: Inoculum size dependency of inhibition using the Colworth MIC techniqueLambert, R. J. W.Journal of Applied Microbiology (2000), 89 (2), 275-279CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)The min. inhibitory concn., MIC, is an accepted and well used criterion for measuring the susceptibility of organisms to inhibitors. Many factors influence the MIC value obtained, including temp., inoculum size and type of organism. A modification of the method developed in this lab. to obtain inhibition profiles of antimicrobials was used to examine the effect of inoculum size on the degree of inhibition obsd. with respect to inhibitor concn. The data obtained enabled the prodn. of an empirical model of inhibition, based on a Gompertz function, relating the level of growth obsd. to both the inoculum size and concn. of the inhibitor. The inoculum size dependencies of phenethyl alc., phenoxyethanol, p-chloro-m-cresol, trichloro-phenol, thymol and dodecyltrimethylammonium bromide against Staphylococcus aureus were obtained.
- 77Lambert, R. J.; Pearson, J. Susceptibility testing: accurate and reproducible minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) values. J. Appl. Microbiol. 2000, 88, 784– 90, DOI: 10.1046/j.1365-2672.2000.01017.x77https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD3cXktVOhsr8%253D&md5=04b3d3382ed17f7f409388e2e4702309Susceptibility testing: accurate and reproducible minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) valuesLambert, R. J. W.; Pearson, J.Journal of Applied Microbiology (2000), 88 (5), 784-790CODEN: JAMIFK; ISSN:1364-5072. (Blackwell Science Ltd.)Measuring the min. inhibitory concn. (MIC) of a substance by current methods is straightforward, whereas obtaining useful comparative information from the tests can be more difficult. A simple technique and a method of data anal. are reported which give the experimentalist more useful information from susceptibility testing. This method makes use of a 100-well microtiter plate and the anal. uses all the growth information, obtained by turbidometry, from each and every well of the microtiter plate. A modified Gompertz function is used to fit the data, from which a more exact value can be obtained for the MIC. The technique also showed that at certain concns. of inhibitor, there was no effect on growth relative to a control well (zero inhibitor). Above a threshold value, which has been termed the non-inhibitory concn. or NIC, growth becomes limiting until it reaches the MIC, where no growth relative to the control is obsd.
- 78Luo, S.; Pal, D.; Shah, S. J.; Kwatra, D.; Paturi, K. D.; Mitra, A. K. Effect of HEPES Buffer on the Uptake and Transport of P-Glycoprotein Substrates and Large Neutral Amino Acids. Mol. Pharmaceutics 2010, 7, 412– 420, DOI: 10.1021/mp900193e78https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXisFGmtrw%253D&md5=5271b78dd7cf1b391a5d80eb9ac9cc91Effect of HEPES buffer on uptake and transport of P-glycoprotein substrates and large neutral amino acidsLuo, Shuanghui; Pal, Dhananjay; Shah, Sujay J.; Kwatra, Deep; Paturi, Kalyani D.; Mitra, Ashim K.Molecular Pharmaceutics (2010), 7 (2), 412-420CODEN: MPOHBP; ISSN:1543-8384. (American Chemical Society)HEPES has been widely employed as an org. buffer agent in cell culture medium as well as uptake and transport expts. in vitro. However, concns. of HEPES used in such studies vary from one lab. to another. In this study, we investigated the effect of HEPES on the uptake and bidirectional transport of P-gp substrates employing both Caco-2 and MDCK-MDR1 cells. ATP-dependent uptake of glutamic acid was also examd. ATP prodn. was further quantified applying ATP Detn. Kit. An addn. of HEPES to the growth and incubation media significantly altered the uptake and transport of P-gp substrates in both Caco-2 and MDCK-MDR1 cells. Uptake of P-gp substrates substantially diminished as the HEPES concn. was raised to 25 mM. Bidirectional (A-B and B-A) transport studies revealed that permeability ratio of PappB-A to PappA-B in the presence of 25 mM HEPES was significantly higher than control. The uptake of phenylalanine is an ATP-independent process, whereas the accumulation of glutamic acid is ATP-dependent. While phenylalanine uptake remained unchanged, glutamic acid uptake was elevated with the addn. of HEPES. Verapamil is an inhibitor of P-gp mediated uptake; elevation of cyclosporine uptake in the presence of 5 μM verapamil was compromised by the presence of 25 mM HEPES. The results of ATP assay indicated that HEPES stimulated the prodn. of ATP. This study suggests that the addn. of HEPES in the medium modulated the energy dependent efflux and uptake processes. The effect of HEPES on P-gp mediated drug efflux and transport may provide some mechanistic insight into possible reasons for inconsistencies in the results reported from various labs.
- 79Borgers, K.; Ou, J. Y.; Zheng, P. X.; Tiels, P.; Van Hecke, A.; Plets, E.; Michielsen, G.; Festjens, N.; Callewaert, N.; Lin, Y. C. Reference genome and comparative genome analysis for the WHO reference strain for Mycobacterium bovis BCG Danish, the present tuberculosis vaccine. BMC Genomics 2019, 20, 561, DOI: 10.1186/s12864-019-5909-579https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BB3MzlsV2isw%253D%253D&md5=26ef669e165146e848a0916417d8532cReference genome and comparative genome analysis for the WHO reference strain for Mycobacterium bovis BCG Danish, the present tuberculosis vaccineBorgers Katlyn; Tiels Petra; Van Hecke Annelies; Plets Evelyn; Michielsen Gitte; Festjens Nele; Callewaert Nico; Borgers Katlyn; Tiels Petra; Van Hecke Annelies; Plets Evelyn; Michielsen Gitte; Festjens Nele; Callewaert Nico; Ou Jheng-Yang; Zheng Po-Xing; Lin Yao-Cheng; Ou Jheng-Yang; Zheng Po-Xing; Lin Yao-ChengBMC genomics (2019), 20 (1), 561 ISSN:.BACKGROUND: Mycobacterium bovis bacillus Calmette-Guerin (M. bovis BCG) is the only vaccine available against tuberculosis (TB). In an effort to standardize the vaccine production, three substrains, i.e. BCG Danish 1331, Tokyo 172-1 and Russia BCG-1 were established as the WHO reference strains. Both for BCG Tokyo 172-1 as Russia BCG-1, reference genomes exist, not for BCG Danish. In this study, we set out to determine the completely assembled genome sequence for BCG Danish and to establish a workflow for genome characterization of engineering-derived vaccine candidate strains. RESULTS: By combining second (Illumina) and third (PacBio) generation sequencing in an integrated genome analysis workflow for BCG, we could construct the completely assembled genome sequence of BCG Danish 1331 (07/270) (and an engineered derivative that is studied as an improved vaccine candidate, a SapM KO), including the resolution of the analytically challenging long duplication regions. We report the presence of a DU1-like duplication in BCG Danish 1331, while this tandem duplication was previously thought to be exclusively restricted to BCG Pasteur. Furthermore, comparative genome analyses of publicly available data for BCG substrains showed the absence of a DU1 in certain BCG Pasteur substrains and the presence of a DU1-like duplication in some BCG China substrains. By integrating publicly available data, we provide an update to the genome features of the commonly used BCG strains. CONCLUSIONS: We demonstrate how this analysis workflow enables the resolution of genome duplications and of the genome of engineered derivatives of the BCG Danish vaccine strain. The BCG Danish WHO reference genome will serve as a reference for future engineered strains and the established workflow can be used to enhance BCG vaccine standardization.
- 80Kapopoulou, A.; Lew, J. M.; Cole, S. T. The MycoBrowser portal: A comprehensive and manually annotated resource for mycobacterial genomes. Tuberculosis 2011, 91, 8– 13, DOI: 10.1016/j.tube.2010.09.00680https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXht1ylsLg%253D&md5=1e5f6644776728d23e5d00c313ae4b9bThe MycoBrowser portal: A comprehensive and manually annotated resource for mycobacterial genomesKapopoulou, Adamandia; Lew, Jocelyne M.; Cole, Stewart T.Tuberculosis (Oxford, United Kingdom) (2011), 91 (1), 8-13CODEN: TUBECU; ISSN:1472-9792. (Elsevier Ltd.)A review. Summary: In this paper, we present the MycoBrowser portal (http://mycobrowser.epfl.ch/), a resource that provides both in silico generated and manually reviewed information within databases dedicated to the complete genomes of Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium marinum and Mycobacterium smegmatis. A central component of MycoBrowser is TubercuList (http://tuberculist.epfl.ch), which has recently benefited from a new data management system and web interface. These improvements were extended to all MycoBrowser databases. We provide an overview of the functionalities available and the different ways of interrogating the data then discuss how both the new information and the latest features are helping the mycobacterial research communities.
- 81Abrahams, K. A.; Cox, J. A. G.; Fütterer, K.; Rullas, J.; Ortega-Muro, F.; Loman, N. J.; Moynihan, P. J.; Pérez-Herrán, E.; Jiménez, E.; Esquivias, J.; Barros, D.; Ballell, L.; Alemparte, C.; Besra, G. S. Inhibiting mycobacterial tryptophan synthase by targeting the inter-subunit interface. Sci. Rep. 2017, 7, 9430, DOI: 10.1038/s41598-017-09642-y81https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1cbhvVSqtw%253D%253D&md5=d5256bcc84a9c06221984116b25aee2eInhibiting mycobacterial tryptophan synthase by targeting the inter-subunit interfaceAbrahams Katherine A; Futterer Klaus; Loman Nicholas J; Moynihan Patrick J; Besra Gurdyal S; Cox Jonathan A G; Rullas Joaquin; Ortega-Muro Fatima; Perez-Herran Esther; Jimenez Elena; Esquivias Jorge; Barros David; Ballell Lluis; Alemparte CarlosScientific reports (2017), 7 (1), 9430 ISSN:.Drug discovery efforts against the pathogen Mycobacterium tuberculosis (Mtb) have been advanced through phenotypic screens of extensive compound libraries. Such a screen revealed sulfolane 1 and indoline-5-sulfonamides 2 and 3 as potent inhibitors of mycobacterial growth. Optimization in the sulfolane series led to compound 4, which has proven activity in an in vivo murine model of Mtb infection. Here we identify the target and mode of inhibition of these compounds based on whole genome sequencing of spontaneous resistant mutants, which identified mutations locating to the essential α- and β-subunits of tryptophan synthase. Over-expression studies confirmed tryptophan synthase as the biological target. Biochemical techniques probed the mechanism of inhibition, revealing the mutant enzyme complex incurs a fitness cost but does not prevent inhibitor binding. Mapping of the resistance conferring mutations onto a low-resolution crystal structure of Mtb tryptophan synthase showed they locate to the interface between the α- and β-subunits. The discovery of anti-tubercular agents inhibiting tryptophan synthase highlights the therapeutic potential of this enzyme and draws attention to the prospect of other amino acid biosynthetic pathways as future Mtb drug targets.
- 82Briffotaux, J.; Huang, W.; Wang, X.; Gicquel, B. MmpS5/MmpL5 as an efflux pump in Mycobacterium species. Tuberculosis 2017, 107, 13– 19, DOI: 10.1016/j.tube.2017.08.00182https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXhtlSrtrvP&md5=9d12bd4cd8051abd3cc695099ef6f899MmpS5/MmpL5 as an efflux pump in Mycobacterium speciesBriffotaux, Julien; Huang, Wei; Wang, Xinwei; Gicquel, BrigitteTuberculosis (Oxford, United Kingdom) (2017), 107 (), 13-19CODEN: TUBECU; ISSN:1472-9792. (Elsevier Ltd.)A review. Tuberculosis remains an important cause of morbidity and mortality throughout the world, amplified by the expansion of antibiotic resistance. Increasing active efflux of the antibiotic is one of the several strategies used by bacteria to resist to antibiotics. After showing the importance of the RND superfamily of efflux pumps in drug resistance, this review focuses on the protein MmpL5, a transmembrane transporter of Mycobacterium. These exporters should be involved in the variety of roles in bacterial cells, including expelling various drugs. The mutation in the transcriptional regulator, linked to the upregulation of MmpL5 can lead to resistance of antibiotics. The study of these mechanisms should be considered in order to improve the treatment of tuberculosis.
- 83Abrahams, K. A.; Cox, J. A. G.; Spivey, V. L.; Loman, N. J.; Pallen, M. J.; Constantinidou, C.; Fernández, R.; Alemparte, C.; Remuiñán, M. J.; Barros, D.; Ballell, L.; Besra, G. S. Identification of Novel Imidazo[1,2-a]pyridine Inhibitors Targeting M. tuberculosis QcrB. PLoS One 2012, 7, e52951 DOI: 10.1371/journal.pone.005295183https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptVSgsQ%253D%253D&md5=4c00cd8ffb101ce9d40da5c0c4f51c0eIdentification of novel imidazo[1,2-a]pyridine inhibitors targeting M. tuberculosis QcrBAbrahams, Katherine A.; Cox, Jonathan A. G.; Spivey, Vickey L.; Loman, Nicholas J.; Pallen, Mark J.; Constantinidou, Chrystala; Fernandez, Raquel; Alemparte, Carlos; Remuinan, Modesto J.; Barros, David; Ballell, Lluis; Besra, Gurdyal S.PLoS One (2012), 7 (12), e52951CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Mycobacterium tuberculosis is a major human pathogen and the causative agent for the pulmonary disease, tuberculosis (TB). Current treatment programs to combat TB are under threat due to the emergence of multi-drug and extensively-drug resistant TB. Through the use of high throughput whole cell screening of an extensive compd. library a no. of imidazo[1,2-a]pyridine (IP) compds. were obtained as potent lead mols. active against M. tuberculosis and Mycobacterium bovis BCG. The IP inhibitors (1-4) demonstrated min. inhibitory concns. (MICs) in the range of 0.03 to 5 μM against a panel of M. tuberculosis strains. M. bovis BCG spontaneous resistant mutants were generated against IP 1, 3 and 4 at 5 × MIC and subsequent whole genome sequencing identified a single nucleotide polymorphism 937ACC>937GCC (T313A) in qcrB, which encodes the b subunit of the electron transport ubiquinol cytochrome C reductase. This mutation also conferred cross-resistance against IP 1, 3 and 4 demonstrating a common target. Gene dosage expts. confirmed M. bovis BCG QcrB as the target where over-expression in M. bovis BCG led to an increase in MIC from 0.5 to >8 μM for IP 3. An acute murine model of TB infection established bacteriostatic activity of the IP series, which await further detailed characterization.
- 84Gupta, R. S.; Lo, B.; Son, J. Phylogenomics and Comparative Genomic Studies Robustly Support Division of the Genus Mycobacterium into an Emended Genus Mycobacterium and Four Novel Genera. Front. Microbiol. 2018, 9, 67, DOI: 10.3389/fmicb.2018.0006784https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1MrmvFKmtA%253D%253D&md5=bd2862ca3414afb67e794631a0f1434aPhylogenomics and Comparative Genomic Studies Robustly Support Division of the Genus Mycobacterium into an Emended Genus Mycobacterium and Four Novel GeneraGupta Radhey S; Lo Brian; Son JeenFrontiers in microbiology (2018), 9 (), 67 ISSN:1664-302X.The genus Mycobacterium contains 188 species including several major human pathogens as well as numerous other environmental species. We report here comprehensive phylogenomics and comparative genomic analyses on 150 genomes of Mycobacterium species to understand their interrelationships. Phylogenetic trees were constructed for the 150 species based on 1941 core proteins for the genus Mycobacterium, 136 core proteins for the phylum Actinobacteria and 8 other conserved proteins. Additionally, the overall genome similarity amongst the Mycobacterium species was determined based on average amino acid identity of the conserved protein families. The results from these analyses consistently support the existence of five distinct monophyletic groups within the genus Mycobacterium at the highest level, which are designated as the "Tuberculosis-Simiae," "Terrae," "Triviale," "Fortuitum-Vaccae," and "Abscessus-Chelonae" clades. Some of these clades have also been observed in earlier phylogenetic studies. Of these clades, the "Abscessus-Chelonae" clade forms the deepest branching lineage and does not form a monophyletic grouping with the "Fortuitum-Vaccae" clade of fast-growing species. In parallel, our comparative analyses of proteins from mycobacterial genomes have identified 172 molecular signatures in the form of conserved signature indels and conserved signature proteins, which are uniquely shared by either all Mycobacterium species or by members of the five identified clades. The identified molecular signatures (or synapomorphies) provide strong independent evidence for the monophyly of the genus Mycobacterium and the five described clades and they provide reliable means for the demarcation of these clades and for their diagnostics. Based on the results of our comprehensive phylogenomic analyses and numerous identified molecular signatures, which consistently and strongly support the division of known mycobacterial species into the five described clades, we propose here division of the genus Mycobacterium into an emended genus Mycobacterium encompassing the "Tuberculosis-Simiae" clade, which includes all of the major human pathogens, and four novel genera viz. Mycolicibacterium gen. nov., Mycolicibacter gen. nov., Mycolicibacillus gen. nov. and Mycobacteroides gen. nov. corresponding to the "Fortuitum-Vaccae," "Terrae," "Triviale," and "Abscessus-Chelonae" clades, respectively. With the division of mycobacterial species into these five distinct groups, attention can now be focused on unique genetic and molecular characteristics that differentiate members of these groups.
- 85Neagoie, C. D.; Peng, X.; Tortorella, M. D.; Fossey, J. S.; Alderwick, L. J.; Feula, A.; Yoshizawa, A.. Preparation of antibacterial compounds: WO2020206594 A1 2020, 10– 15.There is no corresponding record for this reference.
Supporting Information
Supporting Information
The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.jmedchem.3c01643.
Supplementary PK data (PDF).
A detailed description of author contributions, general methods, chemical and biological general procedures, synthetic chemistry protocols for the synthesis of BGAz001–BGAz0016, NMR spectra, details of mass spectrometry analysis, additional corresponding references, (82,83) and molecular formula strings (ZIP).
Terms & Conditions
Most electronic Supporting Information files are available without a subscription to ACS Web Editions. Such files may be downloaded by article for research use (if there is a public use license linked to the relevant article, that license may permit other uses). Permission may be obtained from ACS for other uses through requests via the RightsLink permission system: http://pubs.acs.org/page/copyright/permissions.html.