inhibition of mycobacterium tuberculosis reca · responsible for approximately 1.6 million deaths...

RESEARCH POSTER PRESENTATION DESIGN © 2015 www.PosterPresentations.com • Mycobacterium tuberculosis (Mtb) infects more than 10 million new individuals and is responsible for approximately 1.6 million deaths per year globally 1 . • Tuberculosis is curable, but drug regimens are long, 6-20 months, and tuberculosis often persists in communities where medical care is largely inaccessible 2 . • Drug resistance in Mtb is rising, nullifying the effects of first and second-line antibiotics and creating a need for new drugs 3 . • Under stress, persistent bacteria can become more physiologically tolerant to drugs, although no mutations have been acquired, by activating protein signaling pathways that initiate an SOS response that leads to resistant-like phenotypes - giving the bacteria more time to acquire resistance- conferring mutations 4 . • Choudhary et. al. found that RecA/LexA expression is increased in response to fluoroquinolones in Mtb, which leads to the activation of an SOS response and increased drug tolerance 5 . • Inhibition of RecA sensitized the persisters to antibiotics. • Although there is an available inhibitor, suramin, it binds at 50 μM affinity, which is considered too high for use in the clinic. • The development of high affinity inhibitory ligands against RecA could be used to sensitize Mtb populations tolerant to first- line drugs, extending the usability of these drugs and relieving pressure to come up with newer drugs to overcome antimicrobial resistance. Inhibition of Mycobacterium tuberculosis RecA 2 Atomwise, 717 Market Street, Suite 800, San Francisco, CA 94103 Christopher Beaudoin 1* , Terry O’Brien 2 , Tom Blundell 1 ACKNOWLEDGMENTS Antibiotic Research UK Marcin Skwark BACKGROUND METHODS FUTURE • Validate compounds with ATP-binding affinity functional assays • Work with Atomwise chemists to potentially synthesize derivatives of the successful compounds with higher affinity • Test compounds in vivo using Mtb cell culture 1 80 Tennis Court Road, Department of Biochemistry, University of Cambridge CB2 1GA, *[email protected] REFERENCES 1) World Health Organization: Global Tuberculosis Report (2018) 2) World Health Organization: Guidelines for treatment of drug-susceptible tuberculosis and patient care (2017) 3) Al-Saeedi M & Al-Hajoj S (2017) Infect. Drug Resist. 10: 333–342 4) Hemsley CM, Luo JX, Andreae CA, Butler CS, Soyer OS & Titball RW (2014) Antimicrob. Agents Chemother. 58: 5775– 5783 5) Choudhary E, Sharma R, Kumar Y, Agarwal N (2019). Frontiers in Cellular and Infection Microbiology. 9:70. 6) Worth CL, Preissner R, Blundell TL (2011). Nucleic acids research. 39:W215-22. 7) biotium.com/product/glomelt-thermal- shift-protein-stability-kit/ 8) Bai, N., Roder, H., Dickson, A., & Karanicolas, J. (2019). Scientific reports, 9(1), 2650. 9)www.creativebiomart.net/resource/princi ple-protocol-x-ray-crystallography Crystal structure of Mtb RecA (PDB: 4PSV) Mutation-structure analysis using SDM 6 Example of Atomwise ligand predictions Differential Scanning Fluorimetry will be used to screen which compounds bind to Mtb RecA 7 Isothermal titration calorimetry will be used to measure binding affinity 8 X-Ray crystallography will be used to determine structure of compound-RecA complex 9 Surface Plasmon Resonance will be used to measure binding affinity 8 Example plasmid that will be used to produce Mtb RecA in E. coli Compounds predicted by Atomwise to inhibit the ATP binding spot of Mtb RecA (example compounds)

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Page 1: Inhibition of Mycobacterium tuberculosis RecA · responsible for approximately 1.6 million deaths per year globally1. • Tuberculosis is curable, but drug regimens are long, 6-20

www.PosterPresentations.com

• Mycobacterium tuberculosis (Mtb) infects more than 10 million new individuals and is responsible for approximately 1.6 million deaths per year globally1.

• Tuberculosis is curable, but drug regimens are long, 6-20 months, and tuberculosis often persists in communities where medical care is largely inaccessible2.

• Drug resistance in Mtb is rising, nullifying the effects of first and second-line antibiotics and creating a need for new drugs3.

• Under stress, persistent bacteria can become more physiologically tolerant to drugs, although no mutations have been acquired, by activating protein signaling pathways that initiate an SOS response that leads to resistant-like phenotypes - giving the bacteria more time to acquire resistance-conferring mutations4.

• Choudhary et. al. found that RecA/LexAexpression is increased in response to fluoroquinolones in Mtb, which leads to the activation of an SOS response and increased drug tolerance5.

• Inhibition of RecA sensitized the persistersto antibiotics.

• Although there is an available inhibitor, suramin, it binds at 50 μM affinity, which is considered too high for use in the clinic.

• The development of high affinity inhibitory ligands against RecA could be used to sensitize Mtb populations tolerant to first-line drugs, extending the usability of these drugs and relieving pressure to come up with newer drugs to overcome antimicrobial resistance.

Inhibition of Mycobacterium tuberculosis RecA

2Atomwise, 717 Market Street, Suite 800, San Francisco, CA 94103

Christopher Beaudoin1*, Terry O’Brien2, Tom Blundell1

ACKNOWLEDGMENTS

Antibiotic Research UK

Marcin Skwark

BACKGROUND METHODS FUTURE• Validate compounds with ATP-binding affinity

functional assays

• Work with Atomwise chemists to potentially synthesize derivatives of the successful compounds with higher affinity

• Test compounds in vivo using Mtb cell culture

180 Tennis Court Road, Department of Biochemistry, University of Cambridge CB2 1GA, *[email protected]

REFERENCES