The role of the anaplerotic node in redox homeostasis and pathogenesis of Mycobacterium tuberculosis and its exploitation as a therapeutic target

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Summary. The role of the anaplerotic node in redox homeostasis and pathogenesis of Mycobacterium tuberculosis and its exploitation as a therapeutic targetWe desperately need new treatments to control the tuberculosis pandemic, which is fuelled by the emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb, the causative agent of tuberculosis) as well as a catastrophic synergy with HIV. Mtb is an unusual bacterial pathogen, which has the remarkable ability to cause both acute life threatening disease and also symptomless latent tuberculosis infections that can persist for the lifetime of the human host. It is estimated that 23% of the world has latent tuberculosis, and in 2017 alone, there were 10.0 million cases of tuberculosis and 1.5 million tuberculosis deaths, making TB once again the leading infectious disease globally. A key to the success of tuberculosis as a human disease is ascribed to the extraordinary metabolic flexibility of Mtb. This promotes survival in the variable and harsh environments within the human host, which include exposure to REDuctive and OXidative (redox) stresses. Mtb is able to monitor these stresses in the human host and co-ordinate an appropriate response in order to survive. Maintenance of redox homeostasis is critical to the ability of Mtb to cause disease however many questions in this area of research remain unanswered. A fundamental question is what is the role of metabolism in maintaining redox balance? We have preliminary evidence that a central metabolic hub (the ANA node) consisting of four enzymes is involved in maintaining this balance and have identified a drug which targets this node and kills Mtb. Our previous work showed that this hub is required for the survival of Mtb within its human host cell and we now want to test our hypothesis that this node is essential for the maintenance of redox balance, the ability to cause disease and therefore represents a drug target. We propose to characterise the function of the node in redox control using our mutant strains of Mtb which lack a complete ANA node. Firstly we will investigate the effect of having an incomplete ANA node on the survival, redox balance and immediate metabolic responses of Mtb during exposure to reductive or oxidative stresses. For this work we will use a bespoke system which reports on the redox status of Mtb. The next question is how does the ANA node reconfigure metabolism during redox stress? To tackle this question we will directly analyse the metabolic pathways involved in mediating redox stress using cutting-edge systems-based metabolomics techniques that we have spearheaded at the University of Surrey. By utilising a human cell and mouse model of tuberculosis infection we will probe the essentiality of the ANA node to maintaining redox homeostasis and virulence in the host environment. Finally we will chemically modify a drug which we have shown in preliminary work targets the ANA node and kills Mtb in order to improve the antimicrobial activity. By exploring the mode of action of these tool compounds in the context of redox homeostasis we will validate the ANA node as a drug target. In this way we will elucidate the role of the ANA node in redox control and virulence of this important pathogen and synthesise compounds which can potentially be developed into future treatments for tuberculosis.