Identifying New Probes of Essential Targets in Mycobacterium tuberculosis

Abstract

Despite the availability of effective treatments, Mycobacterium tuberculosis (Mtb) remains a public health crisis. An estimated 9 million people are infected with Mtb every year, resulting in 1.5 million deaths. Efforts to control the tuberculosis epidemic are stymied by a lengthy treatment period, a large reservoir of latently infected individuals, and the emergence of multi- and extensively drug-resistant strains of Mtb. Drugs with novel mechanisms of action are urgently needed in order to address these challenges, but standard screening methods are failing to identify these types of compounds. While numerous functions have been characterized as essential for the survival of Mtb, very few are inhibited by current drugs. Furthermore, few existing drugs are able to kill the non-replicating, phenotypically drug-resistant state of Mtb thought to contribute to lengthy treatment times and latent infections. This research explores the mechanisms of new small molecule inhibitors against Mtb, thereby expanding our knowledge of the bacteria’s inner-workings and identifying new therapeutic targets. Chapter 2 describes the identification of a novel azetidine, BRD4592, that kills Mtb through allosteric inhibition of tryptophan synthase (TrpAB), a previously untargeted enzyme. Given that Mtb can scavenge tryptophan from its environment, questions of TrpAB’s in vivo essentiality are of top importance. Building on our discovery of BRD4592 and utilizing additional inhibitors of amino acid synthesis, we explore the mycobacterial response to amino acid starvation (Chapter 3). We show TrpAB is required for the in vivo survival of Mtb and although the inhibitors induce a protective response to cell stress, the response does not prevent eventual cell death. Finally, Chapter 4 details the characterization of four compounds active against non-replicating Mtb. One of the compounds binds nickel and cobalt and disrupts metal ion homeostasis. The other three compounds act as prodrugs, requiring modification by Mtb enzymes for activity. Through this work, we discovered that the Mtb enzyme MymA can function as an activating enzyme. MymA is not only responsible for activating one of our newly identified compounds, but also can activate ethionamide, an existing antitubercular drug. Together, the compounds identified in this dissertation provide tools to better understand Mtb biology and, ultimately, to combat the Mtb epidemic.