Anti-tuberculosis Drug Discovery Research Articles

Polypharmacology-Driven Discovery and Design of Highly Selective, Dual and Multitargeting Inhibitors of Mycobacterium tuberculosis - A Review

: The increasing demand for novel antitubercular agents has been the main 'force' of many TB research efforts due to the uncontrolled growing number of drug-resistant strains of M. tuberculosis in the clinical setting. Many strategies have been employed to address the drug-resistant issue, including a trend that is gaining attention, which is the design and discovery of Mtb inhibitors that are either dual- or multitargeting. The multiple-target design concept is not new in medicinal chemistry. With a growing number of newly discovered Mtb proteins, numerous targets are now available for developing new biochemical/cell-based assays and computer-aided drug design (CADD) protocols. To describe the achievements and overarching picture of this field in anti- infective drug discovery, we provide in this review small molecules that exhibit profound inhibitory activity against the tubercle bacilli and are identified to trace two or more Mtb targets. This review also presents emerging design methodologies for developing new anti-TB agents, particularly tailored to structure-based CADD. Dedicated on the special occasion of the 70th birthday of Prof. Dr. Ma. Alicia Aguinaldo, whose scientific efforts elevated antituberculosis drug discovery in the Philippines.

QT-Interval Prolongation in the Treatment of Drug-Resistant Tuberculosis

The review analyzes the research articles, guidelines, and clinical trial results on the effect of fluoroquinolones, bedaquiline, nitroimidazoles, and clofazimine on QTc prolongation. The importance of timely and regular monitoring of cardiotoxicity and the importance of continuing clinical trials for the discovery of anti-tuberculosis drugs with a satisfactory safety profile are presented.

Identification of a Chemical Inhibitor with a Novel Scaffold Targeting Decaprenylphosphoryl-β-D-Ribose Oxidase (DprE1).

Tuberculosis is the second leading cause of death from infectious diseases worldwide. Multidrug-resistant Mycobacterium tuberculosis is spreading throughout the world, creating a crisis. Hence, there is a need to develop anti-tuberculosis drugs with novel structures and versatile mechanisms of action. In this study, we identified antimicrobial compounds with a novel skeleton that inhibits mycobacterium decaprenylphosphoryl-β-D-ribose oxidase (DprE1). A multi-step, in silico, structure-based drug screening identified potential DprE1 inhibitors from a library of 154,118 compounds. We experimentally verified the growth inhibitory effects of the eight selected candidate compounds against Mycobacterium smegmatis. Molecular dynamics simulations were performed to understand the mechanism of molecular interactions between DprE1 and ompound 4. Eight compounds were selected through in silico screening. Compound 4 showed strong growth inhibition against M. smegmatis. Molecular dynamics simulation (50 ns) predicted direct and stable binding of Compound 4 to the active site of DprE1. The structural analysis of the novel scaffold in Compound 4 can pave way for antituberculosis drug development and discovery.

DprE1 Inhibitors: Enduring Aspirations for Future Antituberculosis Drug Discovery.

DprE1 is a crucial enzyme involved in the cell wall synthesis of Mycobacterium tuberculosis and a promising target for antituberculosis drug development. However, its unique structural characteristics for ligand binding and association with DprE2 make developing new clinical compounds challenging. This review provides an in-depth analysis of the structural requirements for both covalent and non-covalent inhibitors, their 2D and 3D binding patterns, as well as their biological activity data in vitro and in vivo, including pharmacokinetic information. We also introduce a protein quality score (PQS) and an active-site map of the DprE1 enzyme to help medicinal chemists better understand DprE1 inhibition and develop new and effective anti-TB drugs. Furthermore, we examine the resistance mechanisms associated with DprE1 inhibitors to understand future developments due to resistance emergence. This comprehensive review offers insight into the DprE1 active site, including protein-binding maps, PQS, and graphical representations of known inhibitors, making it a valuable resource for medicinal chemists working on future antitubercular compounds.

Target Identification in Anti-Tuberculosis Drug Discovery.

Mycobacterium tuberculosis (Mtb) is the etiological agent of tuberculosis (TB), a disease that, although preventable and curable, remains a global epidemic due to the emergence of resistance and a latent form responsible for a long period of treatment. Drug discovery in TB is a challenging task due to the heterogeneity of the disease, the emergence of resistance, and uncomplete knowledge of the pathophysiology of the disease. The limited permeability of the cell wall and the presence of multiple efflux pumps remain a major barrier to achieve effective intracellular drug accumulation. While the complete genome sequence of Mtb has been determined and several potential protein targets have been validated, the lack of adequate models for in vitro and in vivo studies is a limiting factor in TB drug discovery programs. In current therapeutic regimens, less than 0.5% of bacterial proteins are targeted during the biosynthesis of the cell wall and the energetic metabolism of two of the most important processes exploited for TB chemotherapeutics. This review provides an overview on the current challenges in TB drug discovery and emerging Mtb druggable proteins, and explains how chemical probes for protein profiling enabled the identification of new targets and biomarkers, paving the way to disruptive therapeutic regimens and diagnostic tools.

Recent Advances in Anti-Tuberculosis Drug Discovery Based on Hydrazide-Hydrazone and Thiadiazole Derivatives Targeting InhA.

Tuberculosis is an extremely serious problem of global public health. Its incidence is worsened by the presence of multidrug-resistant (MDR) strains of Mycobacterium tuberculosis. More serious forms of drug resistance have been observed in recent years. Therefore, the discovery and/or synthesis of new potent and less toxic anti-tubercular compounds is very critical, especially having in mind the consequences and the delays in treatment caused by the COVID-19 pandemic. Enoyl-acyl carrier protein reductase (InhA) is an important enzyme involved in the biosynthesis of mycolic acid, a major component of the M. tuberculosis cell wall. At the same time, it is a key enzyme in the development of drug resistance, making it an important target for the discovery of new antimycobacterial agents. Many different chemical scaffolds, including hydrazide hydrazones and thiadiazoles, have been evaluated for their InhA inhibitory activity. The aim of this review is to evaluate recently described hydrazide-hydrazone- and thiadiazole-containing derivatives that inhibit InhA activity, resulting in antimycobacterial effects. In addition, a brief review of the mechanisms of action of currently available anti-tuberculosis drugs is provided, including recently approved agents and molecules in clinical trials.

Role of transcription termination factor Rho in anti-tuberculosis drug discovery.

Mycobacterial infections, including multidrug and extreme drug-resistant (MDR and XDR) infections, are a severe challenge and create a virtual antibiotic-deficient era. Bacterial transcription is an established antimicrobial drug target. In mycobacteria, efficient transcription termination relies on the ATP-dependent RNA helicase factor Rho. Rho factor is essential for Mycobacterium tuberculosis (Mtb) survival, and is a valid antibacterial drug target with no homolog in eukaryotes. Rho maintains genomic stability and virulence and prevents pervasive transcription in Mtb. In this review, we provide an overview of the essentiality of Rho in Mtb, which makes it an attractive drug target for inhibitor discovery.

Bifunctional NadC Homologue PyrZ Catalyzes Nicotinic Acid Formation in Pyridomycin Biosynthesis

Pyridomycin is a potent antimycobacterial natural product by specifically inhibiting InhA, a clinically validated antituberculosis drug discovery target. Pyridyl moieties of pyridomycin play an essential role in inhibiting InhA by occupying the reduced form of the nicotinamide adenine dinucleotide (NADH) cofactor binding site. Herein, we biochemically characterize PyrZ that is a multifunctional NadC homologue and catalyzes the successive formation, dephosphorylation, and ribose hydrolysis of nicotinic acid mononucleotide (NAMN) to generate nicotinic acid (NA), a biosynthetic precursor for the pyridyl moiety of pyridomycin. Crystal structures of PyrZ in complex with substrate quinolinic acid (QA) and the final product NA revealed a specific salt bridge formed between K184 and the C3-carboxyl group of QA. This interaction positions QA for accepting the phosphoribosyl group to generate NAMN, retains NAMN within the active site, and mediates its translocation to nucleophile D296 for dephosphorylation. Combining kinetic and thermodynamic analysis with site-directed mutagenesis, the catalytic mechanism of PyrZ dephosphorylation was proposed. Our study discovered an alternative and concise NA biosynthetic pathway involving a unique multifunctional enzyme.

A Systematic Review on Antituberculosis Drug Discovery and Antimycobacterial Potential of Biologically Synthesized Silver Nanoparticles: Overview and Future Perspectives

Abstract Rapid emergence and quick evolution of drug-resistant and aggressive mycobacterial strains have resulted in the present antimycobacterial drug crisis and the persistence of tuberculosis as a major public health problem. Green/biological nanotechnologies constitute an interesting area of research for discovering antimycobacterial agents. This review focused on the biological (green) synthesis of silver nanoparticles (AgNPs) as an alternative source of antimycobacterial agents. Data for this study were searched and screened from three electronic databases (Google Scholar, PubMed and ScienceDirect) following the Preferred Reporting Items for Systematic Reviews and Meta-analyses flowchart. Data from in total 17 eligible studies were reported in this systematic review. Twelve of the 17 studies used plants to fabricate AgNPs, whereas the remaining five studies used microorganisms (bacteria and/or fungi). Silver as part of silver nitrate (AgNO3) was the metal precursor reported for the synthesis of AgNPs in these studies. Silver nanoparticles were mostly spherical, with sizes ranging from 12 to 140 nm. Results based on minimum inhibitory concentrations varied between studies and were divided into three groups: (i) those more effective than the antibiotic (controls), (ii) those more effective than plant extracts, and (iii) those less effective than the antibiotic controls. In addition, little or no cytotoxicity effects were reported. Silver nanoparticles were also shown to be highly specific or selective toward mycobacterial strains. This systematic review highlights the antimycobacterial potential of biologically synthesized AgNPs, underscoring the possibility of discovering/developing new antimycobacterial agents using biological synthesis approaches with less toxicity and high selectivity.

Identification of novel inhibitors for mycobacterial polyketide synthase 13 via in silico drug screening assisted by the parallel compound screening with genetic algorithm-based programs.

Identifying small compounds capable of inhibiting Mycobacterium tuberculosis polyketide synthase 13 (Pks13), in charge of final step of mycolic acid biosynthesis, could lead to the development of a novel antituberculosis drug. This study screened for lead compounds capable of targeting M. tuberculosis Pks13 from a chemical library comprising 154,118 compounds through multiple in silico docking simulations. The parallel compound screening (PCS), conducted via two genetic algorithm-based programs was applied in the screening strategy. Out of seven experimentally validated compounds, four compounds showed inhibitory effects on the growth of the model mycobacteria (Mycobacterium smegmatis). Subsequent docking simulation of analogs of the promising leads with the assistance of PCS resulted in the identification of three additional compounds with potent antimycobacterial effects (compounds A1, A2, and A5). Further, molecular dynamics simulation predicted stable interaction between M. tuberculosis Pks13 active site and compound A2, which showed potent antimycobacterial activity comparable to that of isoniazid. The present study demonstrated the efficacy of in silico structure-based drug screening through PCS in antituberculosis drug discovery.

Single-Fluorescence ATP Sensor Based on Fluorescence Resonance Energy Transfer Reveals Role of Antibiotic-Induced ATP Perturbation in Mycobacterial Killing.

ABSTRACTThe rapid emergence of multidrug-resistant/extensively drug-resistant tuberculosis (TB) is responsible for treatment failure in patients with TB and significantly endangers global public health. Recently, bioenergetics has become a new paradigm for anti-TB drug discovery and is based on the link between bacterial ATP levels and drug efficacy. A better understanding of the role of ATP fluctuations during antibiotic treatment may provide insight into antibiotic-mediated killing of mycobacteria. Here, we employed an advanced single-fluorescence FRET (fluorescence resonance energy transfer)-based ATP biosensor, ATPser, for the stable and convenient detection of intracellular ATP fluctuations in mycobacteria. This strategy correlated closely with the results obtained from conventional luminescence ATP assays, indicating the reliability of the system for bioenergetics analysis in mycobacteria. Moreover, the reporter strains expressing ATPser displayed obvious ATP changes when subjected to different stresses, such as starvation and ATP depletion. Interestingly, we observed that different antibiotics induced fluctuations in cellular ATP levels in individual cells of various magnitudes, revealing a strong connection between ATP fluctuations and drug efficacy. Furthermore, drug combinations accelerated ATP perturbation, resulting in increased cell death. We concluded that ATPser enabled real-time measurement of ATP at the single-cell level in mycobacteria, and monitoring ATP dynamics in drug-treated bacteria may shed light on novel treatment strategies.IMPORTANCE Bioenergetics has emerged as a new paradigm for antituberculosis (anti-TB) drug discovery, and the cellular ATP level is the core indicator reflecting bacterial metabolic homeostasis. Although several bulk assays have been designed for the measurement of cellular ATP content, a more convenient strategy is required for real-time ATP measurement of single viable cells. In this study, by combining the ε-subunit of Bacillus subtilis FoF1-ATP synthase with a circularly permuted green fluorescent protein [(cp)GFP], we constructed a FRET-based single-fluorescence ATP sensor, ATPser, for real-time single-cell ATP detection among a mycobacterial population. Using the ATPser, we designed different drug combinations containing components that have similar/opposite effects on ATP alternation. Our results demonstrated that increased cellular ATP fluctuations were associated with depletion of mycobacterial viability, while counteracting ATP fluctuations weakened the killing effect of the drug regime. Thus, potentially efficient drug combinations can be considered based on their similar effects on mycobacterial ATP levels, and ATPser may be a useful tool to study mycobacterial bioenergetics and to guide drug regime design.

A triterpene ketone from an anti-mycobacterial tuberculosis chromatography eluate from the n-hexane fraction of the fruits of Harungana madagascariensis Lam. Ex Poiret (Hypericaceae)

Clinical cases of drug resistant tuberculosis (TB) are a global threat because of the cost and extended duration of treatment regimen. This underscores the continuous drive to discover new therapeutic agents that are faster, simpler and affordable often by recourse to natural products. This report highlights the potentials of the Nigerian medicinal plant Harungana madagascariensis Lam. Ex Poiret (Hypericaceae) as a source of lead agents for the discovery and development of anti-tuberculosis drugs. The fruit of H. madagascariensis was extracted with 70% aqueous ethanol by cold maceration. The crude 70 % aqueous ethanol extract was partitioned with n-hexane to give n-hexane (NHF) soluble portions. The NHF was fractionated further on a column packed with normal phase silica gel (200-400 Mesh size) to afford the chromatography fractions used in this study. The structure of isolated compound was elucidated using spectroscopic (NMR, IR, and MS) techniques. Anti-Mycobacteria tuberculosis susceptibility screening of the chromatography fractions was done using Lowenstein Jensen method with rifampicin, ethambutol, isoniazid and dihydrostreptomycin as reference anti-TB drugs for comparison. Four fractions NHF1-4 were obtained from the NHF. The chromatography fractions: NHF2 and NHF3 inhibited the growth of the Mycobacteria tuberculosis. From the NHF2 was isolated the known bioactive triterpene ketone friedelan-3-one. This preliminary screening underscores the potentials of this ethno medicinal plant as a source of lead compounds for the development of anti-TB drugs.

Dissecting the Mycobacterium bovis BCG Response to Macrophage Infection to Help Prioritize Targets for Anti-Tuberculosis Drug and Vaccine Discovery.

New strategies are required to reduce the worldwide burden of tuberculosis. Intracellular survival and replication of Mycobacterium tuberculosis after macrophage phagocytosis is a fundamental step in the complex host–pathogen interactions that lead to granuloma formation and disease. Greater understanding of how the bacterium survives and thrives in these environments will inform novel drug and vaccine discovery programs. Here, we use in-depth RNA sequencing of Mycobacterium bovis BCG from human THP-1 macrophages to describe the mycobacterial adaptations to the intracellular environment. We identify 329 significantly differentially regulated genes, highlighting cholesterol catabolism, the methylcitrate cycle and iron homeostasis as important for mycobacteria inside macrophages. Examination of multi-functional gene families revealed that 35 PE/PPE genes and five cytochrome P450 genes were upregulated 24 h after infection, highlighting pathways of potential significance. Comparison of the intracellular transcriptome to gene essentiality and immunogenicity studies identified 15 potential targets that are both required for intracellular survival and induced on infection, and eight upregulated genes that have been demonstrated to be immunogenic in TB patients. Further insight into these new and established targets will support drug and vaccine development efforts.

Alkyltriphenylphosphonium turns naphthoquinoneimidazoles into potent membrane depolarizers against mycobacteria.

Due to its central role in energy generation and bacterial viability, mycobacterial bioenergetics is an attractive therapeutic target for anti-tuberculosis drug discovery. Building upon our work on antimycobacterial dioxonaphthoimidazoliums that were activated by a proximal positive charge and generated reactive oxygen species upon reduction by Type II NADH dehydrogenase, we herein studied the effect of a distal positive charge on the antimycobacterial activity of naphthoquinoneimidazoles by incorporating a trialkylphosphonium cation. The potency-enhancing properties of the linker length were affirmed by structure-activity relationship studies. The most active compound against M. tb H37Rv displayed good selectivity index (SI = 34) and strong bactericidal activity in the low micromolar range, which occurred through rapid bacterial membrane depolarization that resulted in depletion of intracellular ATP. Through this work, we demonstrated a switch of the scaffold's mode-of-action via relocation of positive charge while retaining its excellent antibacterial activity and selectivity.

Antituberculosis Targeted Drug Delivery as a Potential Future Treatment Approach.

Mycobacterium tuberculosis (Mtb) is the microorganism that causes tuberculosis. This infectious disease has been around for centuries, with the earliest record of Mtb around three million years ago. The discovery of the antituberculosis agents in the 20th century has managed to improve the recovery rate and reduce the death rate tremendously. However, the conventional antituberculosis therapy is complicated by the development of resistant strains and adverse drug reactions experienced by the patients. Research has been conducted continuously to discover new, safe, and effective antituberculosis drugs. In the last 50 years, only two molecules were approved despite laborious work and costly research. The repurposing of drugs is also being done with few drugs; antibiotics, particularly, were found to have antituberculosis activity. Besides the discovery work, enhancing the delivery of currently available antituberculosis drugs is also being researched. Targeted drug delivery may be a potentially useful approach to be developed into clinically accepted treatment modalities. Active targeting utilizes a specifically designed targeting agent to deliver a chemically conjugated drug(s) towards Mtb. Passive targeting is very widely explored, with the development of multiple types of nanoparticles from organic and inorganic materials. The nanoparticles will be engulfed by macrophages and this will eliminate the Mtb that is present in the macrophages, or the encapsulated drug may be released at the sites of infections that may be in the form of intra- and extrapulmonary tuberculosis. This article provided an overview on the history of tuberculosis and the currently available treatment options, followed by discussions on the discovery of new antituberculosis drugs and active and passive targeting approaches against Mycobacterium tuberculosis.

Architecture of the mycobacterial succinate dehydrogenase with a membrane-embedded Rieske FeS cluster

Complex II, also known as succinate dehydrogenase (SQR) or fumarate reductase (QFR), is an enzyme involved in both the Krebs cycle and oxidative phosphorylation. Mycobacterial Sdh1 has recently been identified as a new class of respiratory complex II (type F) but with an unknown electron transfer mechanism. Here, using cryoelectron microscopy, we have determined the structure of Mycobacterium smegmatis Sdh1 in the presence and absence of the substrate, ubiquinone-1, at 2.53-Å and 2.88-Å resolution, respectively. Sdh1 comprises three subunits, two that are water soluble, SdhA and SdhB, and one that is membrane spanning, SdhC. Within these subunits we identified a quinone-binding site and a rarely observed Rieske-type [2Fe-2S] cluster, the latter being embedded in the transmembrane region. A mutant, where two His ligands of the Rieske-type [2Fe-2S] were changed to alanine, abolished the quinone reduction activity of the Sdh1. Our structures allow the proposal of an electron transfer pathway that connects the substrate-binding and quinone-binding sites. Given the unique features of Sdh1 and its essential role in Mycobacteria, these structures will facilitate antituberculosis drug discovery efforts that specifically target this complex.

Bioenergetic Inhibitors: Antibiotic Efficacy and Mechanisms of Action in Mycobacterium tuberculosis

Development of novel anti-tuberculosis combination regimens that increase efficacy and reduce treatment timelines will improve patient compliance, limit side-effects, reduce costs, and enhance cure rates. Such advancements would significantly improve the global TB burden and reduce drug resistance acquisition. Bioenergetics has received considerable attention in recent years as a fertile area for anti-tuberculosis drug discovery. Targeting the electron transport chain (ETC) and oxidative phosphorylation machinery promises not only to kill growing cells but also metabolically dormant bacilli that are inherently more drug tolerant. Over the last two decades, a broad array of drugs targeting various ETC components have been developed. Here, we provide a focused review of the current state of art of bioenergetic inhibitors of Mtb with an in-depth analysis of the metabolic and bioenergetic disruptions caused by specific target inhibition as well as their synergistic and antagonistic interactions with other drugs. This foundation is then used to explore the reigning theories on the mechanisms of antibiotic-induced cell death and we discuss how bioenergetic inhibitors in particular fail to be adequately described by these models. These discussions lead us to develop a clear roadmap for new lines of investigation to better understand the mechanisms of action of these drugs with complex mechanisms as well as how to leverage that knowledge for the development of novel, rationally-designed combination therapies to cure TB.

Molecular Cloning, Purification and Characterization of Mce1R of Mycobacterium tuberculosis.

The mce1 operon of Mycobacterium tuberculosis, important for lipid metabolism/transport, host cell invasion, modulation of host immune response and pathogenicity, is under the transcriptional control of Mce1R. Hence characterizing Mce1R is an important step for novel anti-tuberculosis drug discovery. The present study reports functional and in silico characterization of Mce1R. In this work, we have computationally modeled the structure of Mce1R and have validated the structure by computational and experimental methods. Mce1R has been shown to harbor the canonical VanR-like structure with a flexible N-terminal 'arm', carrying conserved positively charged residues, most likely involved in the operator DNA binding. The mce1R gene has been cloned, expressed, purified and its DNA-binding activity has been measured in vitro. The Kd value for Mce1R-operator DNA interaction has been determined to be 0.35 ± 0.02 µM which implies that Mce1R binds to DNA with moderate affinity compared to the other FCD family of regulators. So far, this is the first report for measuring the DNA-binding affinity of any VanR-type protein. Despite significant sequence similarity at the N-terminal domain, the wHTH motif of Mce1R exhibits poor conservancy of amino acid residues, critical for DNA-binding, thus results in moderate DNA-binding affinity. The N-terminal DNA-binding domain is structurally dynamic while the C-terminal domain showed significant stability and such profile of structural dynamics is most likely to be preserved in the structural orthologs of Mce1R. In addition to this, a cavity has been detected in the C-terminal domain of Mce1R which contains a few conserved residues. Comparison with other FCD family of regulators suggests that most of the conserved residues might be critical for binding to specific ligand. The max pKd value and drug score for the cavity are estimated to be 9.04 and 109 respectively suggesting that the cavity represents a suitable target site for novel anti-tuberculosis drug discovery approaches.

A fragment-based approach to assess the ligandability of ArgB, ArgC, ArgD and ArgF in the L-arginine biosynthetic pathway of Mycobacterium tuberculosis

The L-arginine biosynthesis pathway consists of eight enzymes that catalyse the conversion of L-glutamate to L-arginine. Arginine auxotrophs (argB/argF deletion mutants) of Mycobacterium tuberculosis are rapidly sterilised in mice, while inhibition of ArgJ with Pranlukast was found to clear chronic M. tuberculosis infection in a mouse model. Enzymes in the arginine biosynthetic pathway have therefore emerged as promising targets for anti-tuberculosis drug discovery. In this work, the ligandability of four enzymes of the pathway ArgB, ArgC, ArgD and ArgF is assessed using a fragment-based approach. We identify several hits against these enzymes validated with biochemical and biophysical assays, as well as X-ray crystallographic data, which in the case of ArgB were further confirmed to have on-target activity against M. tuberculosis. These results demonstrate the potential for more enzymes in this pathway to be targeted with dedicated drug discovery programmes.

Multifunctional Enzymes as Targets for the Treatment of Tuberculosis: Paving the Way for New Anti-TB Drugs.

In spite of the medical and technological developments of the last centuries, Tuberculosis (TB) has remained a challenging disease, with a limited number of therapeutic options, particularly in light of the increase in drug-resistant cases. The search for new molecules continues, with several candidates currently in clinical testing and ongoing efforts to identify novel targets. This work summarizes recent developments on anti-TB therapy, starting by discussing the current epidemiologic status and presenting an overview of the history of anti-tuberculosis drug discovery. Special attention is dedicated to five multifunctional enzymes that are regarded as promising targets for new anti-TB drugs: 5-aminoimidazole-4-carboxamide ribonucleotide transformylase/IMP cyclohydrolase (ATIC); 3,4-dihydroxy-2-butanone 4-phosphate synthase (DHBPS)/GTP cyclohydrolase II (GCHII); glutamine dependent NAD+ Synthetase (NadE); chorismate synthase (CS); and Tryptophan synthase (TS). These enzymes are involved in metabolic pathways critical for the M. tuberculosis survival, growth or replication, but that are not expressed in humans or have significant differences in terms of functionality, which makes them appealing targets. Their function, structure, possible catalytic mechanisms, and current inhibition strategies and inhibitors are reviewed and discussed.