Inhibitors Of Mycobacterium Tuberculosis Research Articles

2-Aryl-Benzoimidazoles as Type II NADH Dehydrogenase Inhibitors of Mycobacterium tuberculosis.

The nonproton pumping type II NADH dehydrogenase in Mycobacterium tuberculosis is essential for meeting the energy needs in terms of ATP under normal aerobic and stressful hypoxic environmental states. Type II NADH dehydrogenase conduits electrons into the electron transport chain in Mycobacterium tuberculosis, which results in ATP synthesis. Therefore, the inhibition of NDH-2 ensures the abolishment of the entire ATP synthesis machinery. Also, type II NADH dehydrogenase is absent in the mammalian genome, thus making it a potential target for antituberculosis drug discovery. Herein, we have screened a commercially available library of drug-like molecules and have identified a hit having a benzimidazole core moiety (6, H37Rv mc26230; minimum inhibitory concentration (MIC) = 16 μg/mL and ATP IC50 = 0.23 μg/mL) interfering with the oxidative phosphorylation pathway. Extensive medicinal chemistry optimization resulted in analogue 8, with MIC = 4 μg/mL and ATP IC50 = 0.05 μg/mL against the H37Rv mc26230 strain of Mycobacterium tuberculosis. Compounds 6 and 8 were found to be active against mono- and multidrug-resistant mycobacterium strains and demonstrated a bactericidal response. The Peredox-mCherry experiment and identification of single-nucleotide polymorphisms in mutants of CBR-5992 (a known type II NADH dehydrogenase inhibitor) were used to confirm the molecules as inhibitors of the type II NADH dehydrogenase enzyme. The safety index >10 for the test active molecules revealed the safety of test molecules.

A modified BPaL regimen for tuberculosis treatment replaces linezolid with inhaled spectinamides.

The Nix-TB clinical trial evaluated a new 6 month regimen containing three oral drugs; bedaquiline (B), pretomanid (Pa), and linezolid (L) (BPaL regimen) for the treatment of tuberculosis (TB). This regimen achieved remarkable results as almost 90% of the multidrug-resistant or extensively drug-resistant TB participants were cured but many patients also developed severe adverse events (AEs). The AEs were associated with the long-term administration of the protein synthesis inhibitor linezolid. Spectinamide 1599 is also a protein synthesis inhibitor of Mycobacterium tuberculosis with an excellent safety profile, but it lacks oral bioavailability. Here, we propose to replace L in the BPaL regimen with spectinamide (S) administered via inhalation and we demonstrate that inhaled spectinamide 1599, combined with BPa --BPaS regimen--has similar efficacy to that of the BPaL regimen while simultaneously avoiding the L-associated AEs. The BPaL and BPaS regimens were compared in the BALB/c and C3HeB/FeJ murine chronic TB efficacy models. After 4-weeks of treatment, both regimens promoted equivalent bactericidal effects in both TB murine models. However, treatment with BPaL resulted in significant weight loss and the complete blood count suggested the development of anemia. These effects were not similarly observed in mice treated with BPaS. BPaL and BPa, but not the BPaS treatment, also decreased myeloid to erythroid ratio suggesting the S in the BPaS regimen was able to recover this effect. Moreover, the BPaL also increased concentration of proinflammatory cytokines in bone marrow compared to mice receiving BPaS regimen. These combined data suggest that inhaled spectinamide 1599 combined with BPa is an effective TB regimen without L-associated AEs.

Indazole to 2‐Cyanoindole Scaffold Progression for Mycobacterial Lipoamide Dehydrogenase Inhibitors Achieves Extended Target Residence Time and Improved Antibacterial Activity

AbstractTuberculosis remains a leading cause of death from a single infection worldwide. Drug resistance to existing and even new antimycobacterials calls for research into novel targets and unexplored mechanisms of action. Recently we reported on the development of tight‐binding inhibitors of Mycobacterium tuberculosis (Mtb) lipoamide dehydrogenase (Lpd), which selectively inhibit the bacterial but not the human enzyme based on a differential modality of inhibitor interaction with these targets. Here we report on the striking improvement in inhibitor residence time on the Mtb enzyme associated with scaffold progression from an indazole to 2‐cyanoindole. Cryo‐EM of Lpd with the bound 2‐cyanoindole inhibitor 19 confirmed displacement of the buried water molecule deep in the binding channel with a cyano group. The ensuing hours‐long improvement in on‐target residence time is associated with enhanced antibacterial activity in axenic culture and in primary mouse macrophages. Resistance to 2‐cyanoindole inhibitors involves mutations within the inhibitor binding site that have little effect on inhibitor affinity but change the modality of inhibitor‐target interaction, resulting in fast dissociation from Lpd. These findings underscore that on‐target residence time is a major determinant of antibacterial activity and in vivo efficacy.

Indazole to 2-Cyanoindole Scaffold Progression for Mycobacterial Lipoamide Dehydrogenase Inhibitors Achieves Extended Target Residence Time and Improved Antibacterial Activity.

Tuberculosis remains a leading cause of death from a single infection worldwide. Drug resistance to existing and even new antimycobacterials calls for research into novel targets and unexplored mechanisms of action. Recently we reported on the development of tight-binding inhibitors of Mycobacterium tuberculosis (Mtb) lipoamide dehydrogenase (Lpd), which selectively inhibit the bacterial but not the human enzyme based on a differential modality of inhibitor interaction with these targets. Here we report on the striking improvement in inhibitor residence time on the Mtb enzyme associated with scaffold progression from an indazole to 2-cyanoindole. Cryo-EM of Lpd with the bound 2-cyanoindole inhibitor 19 confirmed displacement of the buried water molecule deep in the binding channel with a cyano group. The ensuing hours-long improvement in on-target residence time is associated with enhanced antibacterial activity in axenic culture and in primary mouse macrophages. Resistance to 2-cyanoindole inhibitors involves mutations within the inhibitor binding site that have little effect on inhibitor affinity but change the modality of inhibitor-target interaction, resulting in fast dissociation from Lpd. These findings underscore that on-target residence time is a major determinant of antibacterial activity and in vivo efficacy.

Unleashing the potential of cheminformatic analysis for Mycobacterium tuberculosis inhibitors: Insights into chemical space and structural diversity

Mycobacterium tuberculosis (Mtb) remains a major global health concern, causing millions of infections and deaths annually. Drug-resistant tuberculosis poses significant challenges, necessitating the search for new Mtb inhibitors. In this context, a cheminformatic analysis of the currently available antimycobacterial chemical space can be performed to discover structural diversity and new scaffolds. This study conducts a comprehensive cheminformatic analysis of two datasets containing Mtb inhibitors (Mtbs and Phytochemicals), comparing them with Approved drugs and Nutraceuticals datasets. The analysis includes physicochemical properties, molecular scaffolds, structural fingerprints, and structural similarities. The analysis of physicochemical property distributions revealed that Mtb inhibitors are generally less polar or equally polar compared to approved drugs and have similar flexibility. The visual representation of the property space illustrates that the Mtbs dataset is confined to a narrower area, whereas certain compounds within the Phytochemical dataset broaden the scope of the traditional medicinal space. Scaffold analysis identifies several promising candidates in both the Mtbs and Phytochemical datasets. Additionally, employing SkelSpheres descriptors and rubber band scaling for similarity analysis reveals a limited depiction of the chemical space. The analysis of structural diversity indicates that the compounds in the Mtbs dataset exhibit less structural diversity. These findings underscore the necessity to broaden the chemical space of Mtb-targeting molecules by incorporating representative molecules from diverse scaffolds to effectively tackle the issue of drug resistance.

Bactericidal Permeability-Increasing Protein (BPI) Inhibits Mycobacterium tuberculosis Growth.

Bactericidal permeability-increasing protein (BPI) is a multifunctional cationic protein produced by neutrophils, eosinophils, fibroblasts, and macrophages with antibacterial anti-inflammatory properties. In the context of Gram-negative infection, BPI kills bacteria, neutralizes the endotoxic activity of lipopolysaccharides (LPSs), and, thus, avoids immune hyperactivation. Interestingly, BPI increases in patients with Gram-positive meningitis, interacts with lipopeptides and lipoteichoic acids of Gram-positive bacteria, and significantly enhances the immune response in peripheral blood mononuclear cells. We evaluated the antimycobacterial and immunoregulatory properties of BPI in human macrophages infected with Mycobacterium tuberculosis. Our results showed that recombinant BPI entered macrophages, significantly reduced the intracellular growth of M. tuberculosis, and inhibited the production of the proinflammatory cytokine tumor necrosis factor-alpha (TNF-α). Furthermore, BPI decreased bacterial growth directly in vitro. These data suggest that BPI has direct and indirect bactericidal effects inhibiting bacterial growth and potentiating the immune response in human macrophages and support that this new protein's broad-spectrum antibacterial activity has the potential for fighting tuberculosis.

Crystal structure and Hirshfeld surface analysis of 5-hy-droxy-penta-nehydrazide.

Carb-oxy-hydrazides are widely used in medicinal chemistry because of their medicinal properties and many drugs have been developed containing this functional group. A suitable inter-mediate to obtain potential hydrazide drug candidates is the title compound 5-hy-droxy-penta-nehydrazide, C5H12N2O2 (1). The aliphatic compound can react both via the hydroxyl and hydrazide moieties forming derivatives, which can inhibit Mycobacterium tuberculosis catalase-peroxidase (KatG) and consequently causes death of the pathogen. In this work, the hydrazide was obtained via a reaction of a lactone with hydrazine hydrate. The colourless prismatic single crystals belong to the ortho-rhom-bic space group Pca21. Regarding supra-molecular inter-actions, the compound shows classic medium to strong inter-molecular hydrogen bonds involving the hydroxyl and hydrazide groups. Besides, the three-dimensional packing also shows weak H⋯H and C⋯H contacts, as investigated by Hirshfeld surface analysis (HS) and fingerprint plots (FP).

Exploring Scaffold Hopping for Novel 2-(Quinolin-4-yloxy)acetamides with Enhanced Antimycobacterial Activity.

Utilizing a scaffold-hopping strategy from the drug candidate telacebec, a novel series of 2-(quinolin-4-yloxy)acetamides was synthesized and evaluated as inhibitors of Mycobacterium tuberculosis (Mtb) growth. These compounds demonstrated potent activity against drug-sensitive and multidrug-resistant strains (MIC ≤ 0.02 μM). Leading compounds were evaluated against a known qcrB resistant strain (T313A), and their loss in activity suggested that the cytochrome bc1 complex is the likely target. Additionally, these structures showed high selectivity regarding mammalian cells (selectivity index > 500) and stability across different aqueous media. Furthermore, some of the synthesized quinolines demonstrated aqueous solubility values that exceeded those of telacebec, while maintaining low rates of metabolism. Finally, a selected compound prevented Mtb growth by more than 1.7 log10 colony forming units in a macrophage model of tuberculosis (TB) infection. These findings validate the proposed design and introduce new 2-(quinolin-4-yloxy)acetamides with potential for development in TB drug discovery campaigns.

Computational drug repositioning identifies niclosamide and tribromsalan as inhibitors of Mycobacterium tuberculosis and Mycobacterium abscessus

Tuberculosis (TB) is still a major global health challenge, killing over 1.5 million people each year, and hence, there is a need to identify and develop novel treatments for Mycobacterium tuberculosis (M. tuberculosis). The prevalence of infections caused by nontuberculous mycobacteria (NTM) is also increasing and has overtaken TB cases in the United States and much of the developed world. Mycobacterium abscessus (M. abscessus) is one of the most frequently encountered NTM and is difficult to treat. We describe the use of drug-disease association using a semantic knowledge graph approach combined with machine learning models that has enabled the identification of several molecules for testing anti-mycobacterial activity. We established that niclosamide (M. tuberculosis IC90 2.95 μM; M. abscessus IC90 59.1 μM) and tribromsalan (M. tuberculosis IC90 76.92 μM; M. abscessus IC90 147.4 μM) inhibit M. tuberculosis and M. abscessus in vitro. To investigate the mode of action, we determined the transcriptional response of M. tuberculosis and M. abscessus to both compounds in axenic log phase, demonstrating a broad effect on gene expression that differed from known M. tuberculosis inhibitors. Both compounds elicited transcriptional responses indicative of respiratory pathway stress and the dysregulation of fatty acid metabolism.

Synthesis of steroidal inhibitors for Mycobacterium tuberculosis

Oxidised derivatives of cholesterol have been shown to inhibit the growth of Mycobacterium tuberculosis (Mtb). The bacteriostatic activity of these compounds has been attributed to their inhibition of CYP125A1 and CYP142A1, two metabolically critical cytochromes P450 that initiate degradation of the sterol side chain. Here, we synthesise and characterise an extensive library of 28 cholesterol derivatives to develop a structure-activity relationship for this class of inhibitors. The candidate compounds were evaluated for MIC with virulent Mtb and in binding studies with CYP125A1 and CYP142A1 from Mtb.

A high-throughput target-based screening approach for the identification and assessment of Mycobacterium tuberculosis mycothione reductase inhibitors.

The growing anti-microbial resistance poses a global public health threat, impeding progress toward eradicating tuberculosis. Despite decades of active research, there is still a dire need for the discovery of drugs with novel modes of action and exploration of combination drug regimens. Within the European RespiriTB consortium, we explore Mycobacterium tuberculosis energy metabolism to identify new drug candidates that synergize with bedaquiline, with the aim of discovering more efficient combination drug regimens. In this study, we present the development of a high-throughput screening pipeline that led to the identification of M. tuberculosis mycothione reductase inhibitors.

Synthesis, biological activity and in silico study of alkyl eugenol derivatives as Mycobacterium tuberculosis inhibitors

Since ancient times, tuberculosis (TB) has been a fatal infectious disease, made even more difficult to treat with the emergence of drug-resistant strains. The present work aimed to evaluate the antituberculosis activity of newly synthesized eugenol derivatives, and their iron chelation ability as well. The mycobacterium H37Rv strain was sensitive to all tested compounds. The compounds 2-methoxy-4-propenyl-1-propynyloxybenzene (1), 3,5-bis(trifluoromethyl) benzyloxy-2-methoxy-4-propenylbenzene (3) and ethyl 2-methoxy-4-propenylphenylcarbonate (5) showed the best growth inhibitory activity at a concentration of 10 µg/mL. Also, a considerable iron chelation activity for 1 and 5 translated by EC50 values of 0.94 and 0.33 µg/mL, respectively. Generally, the synthesized derivatives illustrated an encouraging in silico pharmacokinetic profile. The molecular docking study revealed that compound 3 exhibited the excellent binding affinity within the active sites of MabA and PanK targets, involving hydrogen bond interactions, fluorine interactions and hydrophobic interactions. Our results suggest that compound 3 could potentially act via the inhibition of MabA and PanK proteins. While, derivatives 1 and 5 could have a potential iron chelation mechanism against mycobacterial growth.

NK-derived exosome miR-1249-3p inhibits Mycobacterium tuberculosis survival in macrophages by targeting SKOR1

Murine Natural Killer cells were cultivated in vitro to isolate NK-derived exosomes. Subsequent quantification via qPCR confirmed enrichment of miR-1249-3p. Ana-1 murine macrophages were cultured in vitro and subsequently inoculated with Mycobacterium tuberculosis (MTB) strain H37Rv. NK-exo and NK-exo miR-1249-3p were separately applied to the infection model, followed by immunological assays conducted post-48-hour co-culture. Western blot analyses corroborated that NK-exo exhibited exosomal marker proteins Granzyme A (GzmA), Granzyme B (GzmB), and Perforin (PFN), alongside a notable enrichment of miR-1249-3p. Functionally, NK-exo augmented the expression levels of Caspase-9,-8, and -3, as well as PARP, while attenuating the expression of NLRP3, ASC, and Cleaved-Caspase-1. Furthermore, qPCR demonstrated an up-regulation of Caspase-9, -8, and -3, along with pro-apoptotic factors Bax and Bid, and a concomitant down-regulation of the anti-apoptotic factor Bcl-2. The expression levels of inflammatory markers ASC, NLRP3, Cleaved-Caspase-1, and IL-1β were concomitantly decreased. ELISA findings indicated diminished levels of TNF-α and ROS secretion. NK-exo miR-1249-3p specifically targeted and attenuated the expression of SKOR-1, engendering up-regulation of apoptosis-associated proteins and down-regulation of inflammation-related proteins, consequently affecting cellular fate.Our empirical evidence substantiates that NK-exo induces macrophage apoptosis, thereby mitigating MTB survival. Furthermore, NK-exo miR-1249-3p directly targets and inhibits SKOR-1 expression, leading to macrophage apoptosis and consequently hampering the proliferation of MTB. The data implicate the potential therapeutic relevance of NK-exo and miR-1249-3p in managing drug-resistant tuberculosis.

A dual-targeting succinate dehydrogenase and F1Fo-ATP synthase inhibitor rapidly sterilizes replicating and non-replicating Mycobacterium tuberculosis

Mycobacterial bioenergetics is a validated target space for antitubercular drug development. Here, we identify BB2-50F, a 6-substituted 5-(N,N-hexamethylene)amiloride derivative as a potent, multi-targeting bioenergetic inhibitor of Mycobacterium tuberculosis. We show that BB2-50F rapidly sterilizes both replicating and non-replicating cultures of M.tuberculosis and synergizes with several tuberculosis drugs. Target identification experiments, supported by docking studies, showed that BB2-50F targets the membrane-embedded c-ring of the F1Fo-ATP synthase and the catalytic subunit (substrate-binding site) of succinate dehydrogenase. Biochemical assays and metabolomic profiling showed that BB2-50F inhibits succinate oxidation, decreases the activity of the tricarboxylic acid (TCA) cycle, and results in succinate secretion from M.tuberculosis. Moreover, we show that the lethality of BB2-50F under aerobic conditions involves the accumulation of reactive oxygen species. Overall, this study identifies BB2-50F as an effective inhibitor of M.tuberculosis and highlights that targeting multiple components of the mycobacterial respiratory chain can produce fast-acting antimicrobials.

Synthesis and structure–activity relationships of aryl fluorosulfate-based inhibitors as novel antitubercular agents

To identify new compounds that can effectively inhibit Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), we screened, synthesized, and evaluated a series of novel aryl fluorosulfate derivatives for their in vitro inhibitory activity against Mtb. Compound 21b exhibited an in vitro minimum inhibitory concentration (MIC) of 0.06 µM against Mtb, no cytotoxicity against both HEK293T and HepG2 mammalian cell lines, and had good in vivo mouse plasma exposure and lung concentration with a 20 mg/kg oral dose, which supports advanced development as a new chemical entity for TB treatment.

The Mycobacterium tuberculosis MmpL3 inhibitor MSU-43085 is active in a mouse model of infection.

MmpL3 is a protein that is required for the survival of bacteria that cause tuberculosis (TB) and nontuberculous mycobacterial (NTM) infections. This report describes the discovery and characterization of a new small molecule, MSU-43085, that targets MmpL3 and is a potent inhibitor of Mycobacterium tuberculosis (Mtb) and M. abscessus survival. MSU-43085 is shown to be orally bioavailable and efficacious in an acute model of Mtb infection. However, the analog is inactive against Mtb in chronically infected mice. Pharmacokinetic and metabolite identification studies identified in vivo metabolism of MSU-43085, leading to a short half-life in treated mice. These proof-of-concept studies will guide further development of the MSU-43085 series for the treatment of TB or NTM infections.

Research Progress in HIV and Mycobacterium tuberculosis Inhibitors Containing Sulfonamide Moiety

Numerous studies have reported significant results regarding the efficacy of sulfonamides against Mycobacterium tuberculosis and various HIV strains. This scientific paper provides an overview of the progress made over a decade of sulfonamides against HIV-1 and mycobacteria and its development against extremely drug-resistant (XDR) and multidrug-resistant strains. The reviewed study offers novel insights into the structural design and structure-activity relationship (SAR) of sulfonamides that are effective and less toxic in combating HIV, Mycobacterium tuberculosis, and multidrug resistance.

Drug design and in-silico study of 2-alkoxylatedquinoline-3-carbaldehyde compounds: Inhibitors of Mycobacterium tuberculosis

Tuberculosis (TB) caused by Mycobacterium tuberculosis (MTB) is a deadly communicable disease that frequently affects the lungs. Current treatment protocols are bedeviled by extensive drug-resistant (XDR) and the evolution of multidrug-resistant (MDR-TB) strains. Virtual in-silico drug discovery tools were used to investigate thirty-two hypothetical 2-alkoxylatedquinoline-3-carbaldehyde compounds for screening against ten different diseases proteins based on drug-likeness, oral bioavailability, pharmacokinetics, global chemical reactivity and their theoretical binding affinities. Their chemical structures were optimized at the density functional theory (DFT) using Becke's three-parameter exchange functional with Lee–Yang–Parr correlation function (B3LYP) and the triple zeta basis set 6–311 in a vacuum using Gaussian 09 W software. Docking study using Pyrx and Discovery studio. Fourteen compounds; 4 - 6, 12 – 14, 19 – 22 and 27–30 complied with the established drug-likeness rules, however, five compounds 12, 13, 27, 28 and 29 exhibited no significant toxicity. Structural activity relationship revealed that shorter (n < 3) or longer (n > 5) alkyloxyl substituents at position-2 of the quinoline moiety reduces drug-likeness and increases toxicity. Individually, the binding energies obtained were (-8.9 kcal/ mole) against malaria for compound 12 and (-8.2 kcal/mole) against the diabetes for compound 29, both highest for the ten diseases investigated. Mycobacterium Tuberculosis proteins investigated. Molecular dynamics also confirms that 12 and 27 binds very well in the active pocket of Mycobacterium tuberculosis and calculated total free binding energy from MMPBSA is -97.53 ± 2.47 and -58.62 ± 2.94 kJ/mol respectively. The five lead compounds all had binding energies higher than the reference tuberculosis drugs; Isoniazid and Ethambutanol.

Molecular Modeling of Enoyl Acyl Carrier Protein Reductase Inhibitors for Mycobacterium tuberculosis and their Pharmacokinetic Predictions

Tuberculosis (TB) is a deep public health concern worldwide worsened by reported multi drugresistant (MDR) and extensively drug- resistant (XDR) stralins of Mycobacterium tuberculosis, the causative agent of the disease. A new class of thiadiazole inhibitors were reported to inhibit the enoyl-acyl transporter protein reductase (InhA) of Mycobacterium tuberculosis (MTb). We performed here the computer-aided molecular design of novel thiadiazole (TDZ) inhibitors of InhA by in situ modifying the reference crystal structure of (S)-1-(5-((1-(2,6-difluorobenzyl)-1 H-pyrazol-3yl)amino)-1,3,4-thiadiazol-2-yl)-1-(4-methylthiazol-2-yl)ethanol-InhA (PDB code: 4BQP). Thus a training set of 15 hybrids with known inhibition potency \(\left(\mathrm{IC}_{50}^{\exp }\right)\) was selected to establish a onedescriptor quantitative structure-activity relationship (QSAR) model resulting in a linear correlation between the Gibbs free energy (GFE) during the formation of the InhA-TDZ complex and \(\mathrm{IC}_{50}^{\mathrm{exp}}\left(\mathrm{plC} \mathrm{C}_{50} \exp ==-0.29 \mathrm{x} \Delta \Delta \mathrm{G}_{\mathrm{com}}+8.13 ; \mathrm{n}=15 ; \mathrm{R}^2=0.92, \mathrm{R}^2{ }_{\mathrm{xv}}=0.91 ;\right.\) F-test of \(142.6 ; \sigma=0.21 ; \alpha&gt;\) \(\left.95 \% ; R^2-R_{x v}^2=0.01\right)\). The 3D pharmacophore model \((\mathrm{PH} 4)\) generated from the active conformations of TDZs ( \(\mathrm{pIC}_{50}^{\mathrm{exp}}=0.93 \times \mathrm{pIC}_{50}^{\text {pred }}+0.47 ; \mathrm{n}=15 ; \mathrm{R}^2=0.97 ; \mathrm{R}_{\mathrm{xv}}=0.94 ;\) F-test of \(215.45 ; \sigma=0.17 ; \alpha&gt;98 \% ; R^2-R_{x v}^2=0.03\) ) served as a virtual screening tool for new analogs from a virtual library (VL). The combination of molecular modeling and \(\mathrm{PH} 4\) in silico screening of (\(\mathrm{VL}\)) resulted in the identification of novel potent antitubercular agent candidates with favorable pharmacokinetic profiles of which the six best hits predicted inhibitory potencies \(\mathrm{IC}_{50}^{\text {pre }}\) in the sub nanomolar range \((0.1-0.2 \mathrm{nM})\).

Antimycobacterial Activities of Hydroxamic Acids and Their Iron(II/III), Nickel(II), Copper(II) and Zinc(II) Complexes.

Hydroxamic acid (HA) derivatives display antibacterial and antifungal activities. HA with various numbers of carbon atoms (C2, C6, C8, C10, C12 and C17), complexed with different metal ions, including Fe(II/III), Ni(II), Cu(II) and Zn(II), were evaluated for their antimycobacterial activities and their anti-biofilm activities. Some derivatives showed antimycobacterial activities, especially in biofilm growth conditions. For example, 20-100 µM of HA10Fe2, HA10FeCl, HA10Fe3, HA10Ni2 or HA10Cu2 inhibited Mycobacterium tuberculosis, Mycobacterium bovis BCG and Mycobacterium marinum biofilm development. HA10Fe2, HA12Fe2 and HA12FeCl could even attack pre-formed Pseudomonas aeruginosa biofilms at higher concentrations (around 300 µM). The phthiocerol dimycocerosate (PDIM)-deficient Mycobacterium tuberculosis H37Ra was more sensitive to the ion complexes of HA compared to other mycobacterial strains. Furthermore, HA10FeCl could increase the susceptibility of Mycobacterium bovis BCG to vancomycin. Proteomic profiles showed that the potential targets of HA10FeCl were mainly related to mycobacterial stress adaptation, involving cell wall lipid biosynthesis, drug resistance and tolerance and siderophore metabolism. This study provides new insights regarding the antimycobacterial activities of HA and their complexes, especially about their potential anti-biofilm activities.