The ever-increasing drug-resistant tuberculosis (TB) has invigorated the focus on the discovery and development of novel therapeutic agents and treatment options. Thiazolidinone-based compounds have shown good antitubercular properties in vitro. Here, we report the design and synthesis of a number of new derivatives inspired by the structure of thiazolidine-2,4-dione (TZD). The TZD-based hybrids with the thiosemicarbazone or the pyridinecarbohydrazone moiety were synthesised and their antimycobacterial activity was investigated against the reference H37Rv and two wild Mycobacterium tuberculosis (Mtb) strains. In further studies, a two-drug interaction analysis was also performed for assessing their synergism with the current first-line drugs used for the treatment of TB. It was found that some of the compounds showed high antimycobacterial activity with MICs (0.078-0.283 µM) and a synergistic effect with isoniazid or rifampicin, thereby demonstrating their potential as a promising scaffold for the development of novel coadjuvants for the effective treatment of TB.

Streptomyces sp. MJM3502 is a promising producer of rufomycins, which are a class of potent anti-tuberculosis lead compounds. Although the structure, activity, and mechanism of the main rufomycin 4/6 and its analogs have been extensively studied, a significant gap remains in our understanding of the genome sequence and biosynthetic pathway of Streptomyces sp. MJM3502, and its metabolic engineering has not yet been reported. This study established the genetic manipulation platform for the strain. Using CRISPR/Cas9-based technology to in-frame insert the strong kasO∗p promoter upstream of the rufB and rufS genes of the rufomycin BGC, we increased rufomycin 4/6 production by 4.1-fold and 2.8-fold, respectively. Furthermore, designing recombinant strains by inserting the kasO∗p promoter upstream of the biosynthetic genes encoding cytochrome P450 enzymes led to new rufomycin derivatives. These findings provide the basis for enhancing the production of valuable natural compounds in Streptomyces and offer insights into the generation of novel active natural products via synthetic biology and metabolic engineering.

In this work, novel 2-substituted-3-((1-substituted-1H-1,2,3-triazol-4-yl) methoxy) quinoxaline analogues were designed, synthesized, and various analytical techniques, viz., 1H NMR, 13C NMR, and Mass spectrometry, were deployed in the structure confirmation of the final compounds. Synthesized derivatives were evaluated for their antimycobacterial activity against Mycobacterium tuberculosis (Mtb) H37Rv. Target molecules mainly consist of methyl substituent in the second position of quinoxaline moiety (QM series) or phenyl substituent in the second position (QP series). Among the forty-two compounds synthesized and evaluated for anti-mycobacterial activity, the MIC values ranged between 5.58 μg/mL to >100 μg/mL. Among QM series compounds, QM7, with MIC 5.58 μg /mL, was the most active compound. Among the QP series derivatives, the intermediate QP-Acy with MIC 23.39 μg /mL was the most promising. Most of the analogues tested in the QP series are less potent than the QM series. All the synthesized molecules showed good drug-likeness when evaluated using the SWISS ADME tool. QM7 was evaluated for docking studies using the crystal structure of enoyl-acyl carrier (INH-A) enzyme PDB: 4TZK, and it showed significant docking scores and interactions. MD simulations were carried out to assess the stability of the protein QM7 complex. Single crystals were grown for QM1, QM6, and QPb from these forty-two compounds, and their structures were solved using OLEX. The corresponding CCDC numbers for these compounds are 2,388,310, 2,388,309, and 2,388,291, respectively.

There is a growing interest in longitudinal omics data paired with some longitudinal clinical outcome. Given a large set of continuous omics variables and some continuous clinical outcome, each measured for a few subjects at only a few time points, we seek to identify those variables that co-vary over time with the outcome. To motivate this problem we study a dataset with hundreds of urinary metabolites along with Tuberculosis mycobacterial load as our clinical outcome, with the objective of identifying potential biomarkers for disease progression. For such data clinicians usually apply simple linear mixed effects models which often lack power given the low number of replicates and time points. We propose a penalized regression approach on the first differences of the data that extends the lasso + Laplacian method [Li and Li (Network-constrained regularization and variable selection for analysis of genomic data. Bioinformatics 2008;24:1175-82.)] to a longitudinal group lasso + Laplacian approach. Our method, PROLONG, leverages the first differences of the data to increase power by pairing the consecutive time points. The Laplacian penalty incorporates the dependence structure of the variables, and the group lasso penalty induces sparsity while grouping together all contemporaneous and lag terms for each omic variable in the model.

With an automated selection of model hyper-parameters, PROLONG correctly selects target metabolites with high specificity and sensitivity across a wide range of scenarios. PROLONG selects a set of metabolites from the real data that includes interesting targets identified during EDA.

An R package implementing described methods called "prolong" is available at https://github.com/stevebroll/prolong. Code snapshot available at 10.5281/zenodo.14804245.

The first total synthesis of the alkaloid brevianamide S has been achieved in eight steps. This natural product, isolated from Aspergillus versicolor, exhibits selective antibacterial activity against Bacille Calmette-Guérin (BCG), a commonly used surrogate for Mycobacterium tuberculosis. Brevianamide S is proposed to act through a novel, yet-to-be-elucidated mechanism, making it a promising lead in the development of next-generation antitubercular agents. Our approach employs a bidirectional synthetic strategy, involving a bespoke alkenyl-alkenyl Stille cross-coupling reaction and a double aldol condensation. This represents a flexible and efficient platform for the future synthesis of structurally diverse analogues.

Latent infection by Mycobacterium tuberculosis (Mtb) impedes effective tuberculosis therapy and eradication. The protein PerM is essential for chronic Mtb infections in mice and acts via the divisome protein FtsB to modulate cell division. Using transgenic co-expression in Escherichia coli, we studied the Mtb PerM-FtsB interaction in isolation from other Mtb proteins, engineering PerM to enhance expression in the E. coli membrane. Using fluorescence microscopy in E. coli, we observed that the previously reported PerM-dependent instability of Mtb FtsB required a segment of FtsB predicted to bind cell-division proteins FtsL and FtsQ. Furthermore, we found that the stability of membrane-localized PerM hinged on its interaction with a conserved, C-terminal helix in FtsB. We also observed that removing this helix disrupted PerM-FtsB interaction using single-molecule tracking. Molecular dynamics results supported the observation that FtsB stabilized PerM and suggested that interactions at the PerM-FtsB interface differ from our initial structure prediction in a way that is consistent with PerM sequence conservation. Although narrowly conserved, the PerM-FtsB interaction emerges as a potential therapeutic target for persistent infections by disrupting the regulation of cell division. Integrating protein structure prediction, molecular dynamics, and single-molecule microscopy, our approach is primed to screen potential inhibitors of the PerM-FtsB interaction and can be straightforwardly adapted to explore other putative interactions.IMPORTANCEOur research reveals significant insights into the dynamic interaction between the proteins PerM and FtsB within Mycobacterium tuberculosis, contributing to our understanding of bacterial cell division mechanisms crucial for infection persistence. By combining innovative fluorescence microscopy and molecular dynamics, we established that the stability of these proteins is interdependent; molecular dynamics placing PerM-FtsB in the context of the mycobacterial divisome shows how disrupting PerM-FtsB interactions can plausibly impact bacterial cell wall synthesis. These findings highlight the PerM-FtsB interface as a promising target for novel therapeutics aimed at combating persistent bacterial infections. Importantly, our approach can be adapted for similar studies in other bacterial systems, suggesting broad implications for microbial biology and antibiotic development.

There is a growing body of data describing the high burden of respiratory sequelae seen among tuberculosis survivors, including children, adolescents, and adults. This group of sequelae are known as post-tuberculosis lung disease and include parenchymal damage, airway disease, and pulmonary vascular disease. It is thought that approximately half of pulmonary tuberculosis survivors have ongoing structural pathology, lung function impairment, or respiratory symptoms after the resolution of active disease. Post-tuberculosis lung disease has been associated with adverse patient outcomes, including persistent symptoms and functional impairment, ongoing health seeking, and impacts on income and employment. There is still much to understand about the epidemiology and nature of post-tuberculosis lung disease, but in this Review we focus on strategies for prevention, diagnosis, and care to inform the ongoing work of tuberculosis-affected communities, health-care providers, researchers, and policy makers. We summarise recent data, highlight evidence gaps, and suggest key research priorities for those working in the field.

Multiple in vitro potency assays are used to evaluate compounds against Mycobacterium tuberculosis, but a consensus on clinically relevant assays is lacking. We aimed to identify an in vitro assay signature that predicts preclinical efficacy and early clinical outcome. Thirty-one unique in vitro assays were compiled for 10 TB drugs. In vitro EC50 values were compared to pharmacokinetic-pharmacodynamic (PK-PD)-model-derived EC50 values from mice evaluated via multinomial regression. External validation of best-performing in vitro assay combinations was performed using five new TB drugs. Best-performing assay signatures for acute and subacute infections were described by assays that reproduce conditions found in macrophages and foamy macrophages and chronic infection by the ex vivo caseum assay. Subsequent simulated mouse bacterial burden over time using predicted in vivo EC50 was within 2-fold of observations. This study helps us identify clinically relevant assays and prioritize successful drug candidates, saving resources and accelerating clinical success.

In this work, a simple and sensitive ratiometric fluorescence probe to detect isoniazid (INH) was developed on the basis of carbon dots and MnO2 nanosheets (MnO2 NS). Nitrogen-doped carbon dots (FMT-CDs) were synthesized by microwave method using formononetin (FMT) in ammonia water. The oxidase-like activity of MnO2 NS was utilized to oxidize non-fluorescent o-phenylenediamine (OPD) to 2, 3-diaminophenazine (DAP) with orange fluorescence. It was demonstrated that DAP could effectively quench the fluorescence of FMT-CDs through the inner filtering effect (IFE). Due to the reducing nature of INH, MnO2 NS was decomposed and thus lost its oxidase-like activity after the addition of INH. Therefore, the production of DAP was reduced, and the fluorescence intensity was decreased, while IFE on FMT-CDs was weakened and the fluorescence of FMT-CDs was recovered. Based on the above principle, a sensitive method for the detection of INH was established based on the fluorescence intensity ratio of the two fluorescent substances, FMT-CDs and DAP, and was successfully applied to the detection of INH in human urine samples. The limit of detection (LOD) could reach to 0.16 μM in human urine samples.

Tuberculosis (TB) is one of the leading causes of death worldwide and a major public health problem. Immune evasion mechanisms and antibiotic resistance highlight the need to better understand this disease and explore alternative treatment approaches. Mycobacterial infection modulates the macrophage response and metabolism to persist and proliferate inside the cell. Cannabinoid receptor type 2 (CB2) is expressed mainly in leukocytes and modulates the course of inflammatory diseases. Therefore, our study aimed to evaluate the effects of the CB2-selective agonist GP1a on irradiated Mycobacterium bovis-BCG (iBCG)-induced J774A.1 macrophage activation. We observed increased expression of CB2 in macrophages after iBCG stimulation. The pretreatment with CB2-agonists, GP1a, JWH-133, and GW-833972A (10 µM), reduced iBCG-induced TNF-α and IL-6 release by these cells. Moreover, the CB2-antagonist AM630 (200 nM) treatment confirmed the activity of GP1a on CB2 by scale down its effect on cytokine production. GP1a pretreatment (10 µM) also inhibited the iBCG-induced production of inflammatory mediators as prostaglandin (PG)E2 and nitric oxide by macrophages. Additionally, GP1a pretreatment also reduced the transcription of proinflammatory genes (inos, il1b, and cox2) and genes related to lipid metabolism (dgat1, acat1, plin2, atgl, and cd36). Indeed, lipid droplet accumulation was reduced by GP1a treatment, which was partially blockade by AM630 pretreatment. Finally, GP1a pretreatment reduced the activation of the NF-κB signaling pathway. In conclusion, the activation of CB2 by GP1a modulated the macrophage response to iBCG by reducing inflammatory mediator levels and metabolic reprogramming.

A dire need exists for novel drugs to treat Mycobacterium tuberculosis infection. In an effort to build on our early efforts targeting the MenG enzyme within the menaquinone biosynthetic pathway, we have pursued the optimization of diaryl amide JSF-2911 to address its poor metabolic stability and modest in vitro potency. A hit evolution campaign focused on modification of the amine substructure within this hit compound, resulting in a range of analogues that have been profiled extensively. Among these derivatives, JSF-4536 and JSF-4898 demonstrated significantly improved biological profiles, notably offering submicromolar MIC values versus M. tuberculosis and promising values characterizing the mouse liver microsome stability, aqueous solubility, and mouse pharmacokinetic profile. JSF-4898 enhanced the efficacy of rifampicin in a subacute model of M. tuberculosis infection in mice. The findings suggest a rationale for the further optimization of MenG inhibitors to provide a novel therapeutic strategy to address M. tuberculosis infection.

The urgent need for safer and innovative antitubercular agents remains a priority for the scientific community. In pursuit of this goal, we designed and evaluated novel 5-phenylfuran-2-carboxylic acid derivatives targeting Mycobacterium tuberculosis (Mtb) salicylate synthase (MbtI), a key enzyme, absent in humans, that plays a crucial role in Mtb virulence. Several potent MbtI inhibitors demonstrating significant antitubercular activity and a favorable safety profile were identified. Structure-guided optimization yielded 5-(3-cyano-5-isobutoxyphenyl)furan-2-carboxylic acid (1e), which exhibited strong MbtI inhibition (IC50 = 11.2 μM) and a promising in vitro antitubercular activity (MIC99 = 32 μM against M. bovis BCG). Esters of 1e were effectively loaded into poly(2-methacryloyloxyethyl phosphorylcholine)-poly(2-(diisopropylamino)ethyl methacrylate) (PMPC-PDPA) polymersomes (POs) and delivered to intracellular mycobacteria, resulting in reduced Mtb viability. This study provides a foundation for the use of POs in the development of future MbtI-targeted therapies for tuberculosis.

Tuberculosis (TB) is notoriously difficult to treat, likely due to the complex host-pathogen interactions driven by pathogen heterogeneity. An understudied area of TB pathogenesis is host responses to Mycobacterium tuberculosis bacteria (Mtb) that are limited in zinc ions. This distinct population resides in necrotic granulomas and sputum and could be the key player in tuberculosis pathogenicity. In this study, we tested the hypothesis that macrophages differentiate between Mtb grown under zinc limitation or in the standard zinc-replete medium. Using several macrophage infection models, such as murine RAW 264.7 and murine bone marrow-derived macrophages (BMDMs), as well as human THP-1-derived macrophages, we show that macrophages infected with zinc-limited Mtb have increased bacterial burden compared with macrophages infected with zinc-replete Mtb. We further demonstrate that macrophage infection with zinc-limited Mtb trigger higher production of reactive oxygen species (ROS) and cause more macrophage death. Furthermore, the increased ROS production is linked to the increased phagocytosis of zinc-limited Mtb, whereas cell death is not. Finally, transcriptional analysis of RAW 264.7 macrophages demonstrates that macrophages have more robust pro-inflammatory responses when infected with zinc-limited Mtb than zinc-replete Mtb. Together, our findings suggest that Mtb's access to zinc affects their interaction with macrophages and that zinc-limited Mtb may be influencing TB progression. Therefore, zinc availability in bacterial growth medium should be considered in TB drug and vaccine developments.

Novel 2-phenylquinolin-4(1H)-one threaded 1,2,3- triazoles were designed, synthesized and evaluated for in vitro activity against Mycobacterium tuberculosis which could be putatively through inhibition of carbonic anhydrase β. Molecules were synthesized in simple Schottan Baumann reaction for amide synthesis. Purified compounds were screened for antitubercular and antibacterial activities. Among them, 1-((1-(2-methoxyphenyl)-1H-1,2,3-triazol-4-yl)methyl)-2-phenylquinolin-4(1H)-one 9j with 2-methoxy at the ortho position of phenyl ring indicated significant antitubercular activity with MIC value of 6.25, 3.12 and 3.12 μg/ml antimicrobial activity against Mycobacterium tuberculosis, gram positive and gram negative strain. The molecular docking and dynamics studies demonstrated that the compound 9j occupied the Zn-binding site of the enzyme with docking energy of -6.2 kcal mol-1. In silico ADME studies indicated that the synthesized compounds have good drug likeliness. The findings explore and present a potential series of antimycobacterial agents in the hope of developing new and advanced therapeutics for tuberculosis.

Successful tuberculosis therapy requires treatment with an unwieldy multidrug combination for several months. Thus, there is a growing need to identify novel genetic vulnerabilities that can be leveraged to develop new, more effective antitubercular drugs. Consequently, recent efforts to optimize tuberculosis (TB) therapy have exploited Mycobacterium tuberculosis (Mtb) chemical genetics to identify pathways influencing antibiotic efficacy, novel mechanisms of antibiotic action, and new targets for TB drug discovery. However, the influence of the complex host environment on these interactions remains largely unknown, leaving the therapeutic potential of the identified targets unclear. In this study, we leveraged a library of conditional mutants targeting 467 essential Mtb genes to characterize the chemical-genetic interactions (CGIs) with TB drugs directly in the mouse infection model. We found that these in vivo CGIs differ significantly from those identified in vitro. Both drug-specific and drug-agnostic effects were identified, and many were preserved during treatment with a multidrug combination, suggesting numerous strategies for enhancing therapy. This work also elucidated the complex effects of pyrazinamide (PZA), a drug that relies on aspects of the infection environment for efficacy. Specifically, our work supports the importance of coenzyme A synthesis- inhibition during infection, as well as the antagonistic effect of iron limitation on PZA activity. In addition, we found that inhibition of thiamine and purine synthesis increases PZA efficacy, suggesting additional therapeutically exploitable metabolic dependencies. Our findings present a map of the unique in vivo CGIs, characterizing the mechanism of PZA activity in vivo and identifying potential targets for TB drug development.

Simpler, safer, and faster chemotherapeutic regimens for tuberculosis and other respiratory mycobacterial infections are an urgent need. Many current therapies suffer from suboptimal drug exposure and dose-limiting systemic adverse effects, challenges that could be addressed via controlled delivery of drugs to the primary site of infection. We sought to address this need by designing a flexible formulation platform that targets drugs to the lung macrophages that concentrate at infectious foci. Our approach is based on an encapsulation strategy in which drugs or specifically designed prodrugs are captured in the hydrophobic core of a glucan-lipid particle (GLP). We show chemically diverse antimycobacterial drugs can be efficiently and stably encapsulated within GLP and that these microparticles can be engineered to release drugs upon low pH or reducing conditions that occur upon phagocytosis by macrophages. Encapsulated formulations of clofazimine, isoniazid, and linezolid retain activity against intracellular Mycobacterium tuberculosis (Mtb) in an ex vivo model, demonstrating efficient drug delivery and release. Intranasal administration of GLP-clofazimine to Mtb-infected mice effectively concentrates the drug in the lung and reduces bacterial burden, whereas GLP-delivered linezolid was systemically distributed and failed to inhibit bacterial growth in the lung. This work establishes GLPs as a promising platform for targeted antibiotic delivery to the lung and also illustrates pharmacokinetic parameters that must be considered in future development.

Tuberculosis (TB) causes an estimated 10.8 million cases each year and remains one of the leading causes of infectious death. Effective treatment is complicated due to the lengthy drug regimen required to prevent relapse and treatment failure. A primary challenge is delivering drugs effectively to lung granulomas, where TB bacteria can persist. Here, we developed yeast-derived glucan lipid microparticles (GLPs) as a novel delivery system to efficiently encapsulate and deliver TB drugs directly to lung tissue via intranasal administration. Of the formulations evaluated, GLP-encapsulated clofazimine achieved increased lung drug levels and reduced bacterial burden in TB-infected mice. The use of GLPs offers a promising approach to improve TB treatment by enabling targeted drug delivery to infection sites within the lungs.

In the search for new antituberculosis drugs with novel mechanisms of action, we evaluated the antimycobacterial activity of a panel of eight phenolic acids against four pathogenic mycobacterial model species, including Mycobacterium tuberculosis. We demonstrated that salicylic acid (SA), as well as the iodinated derivatives 5-iodo-salicylic acid (5ISA) and 3,5-diiodo-salicylic acid (3,5diISA), displayed promising antitubercular activities. Remarkably, using a genetically encoded mycobacterial intrabacterial pH reporter, we describe for the first time that SA, 5ISA, 3,5diISA, and the anti-inflammatory drug aspirin (ASP) act by disrupting the intrabacterial pH homeostasis of M. tuberculosis in a dose-dependent manner under in vitro conditions mimicking the endolysosomal pH of macrophages. In contrast, the structurally related second-line anti-TB drug 4-aminosalicylic acid (PAS) had no pH-dependent activity and was strongly antagonized by l-methionine supplementation, thereby suggesting distinct modes of action. Finally, we propose that SA, ASP, and its two iodinated derivatives could restrict M. tuberculosis growth in a pH-dependent manner by acidifying the cytosol of the bacilli, therefore making such compounds very attractive for further development of antibacterial agents.

This study investigates how Strongyloides stercoralis (Ss) infection impacts pulmonary tuberculosis (PTB) treatment outcomes, disease severity, and bacterial burdens in PTB patients with Ss coinfection.

We used chest x-rays and sputum smear grades to assess lung conditions and bacterial loads in 483 PTB patients. Ss infection was confirmed by seropositivity, and cytokine and profibrotic factor levels were analyzed using multiplex enzyme-linked immunosorbent assay. Treatment outcomes were categorized as favorable (cure without recurrence) or unfavorable (treatment failure or TB recurrence) during treatment or within 12 months postcure.

PTB patients coinfected with Ss had significantly higher bacterial loads, increased risk of bilateral lung lesions, and greater likelihood of cavitary disease compared with those without Ss infection. The coinfected individuals exhibit significantly increased levels of cytokines (interleukin [IL]-4, IL-5, IL-13, interferon [IFN]-α, and IFN-β) and profibrotic factors (vascular endothelial growth factor, epidermal growth factor [EGF], fibroblast growth factor 2 [FGF-2], and PDGF-AB/BB [platelet-derived growth factor]) and significantly diminished levels of cytokines (IFN-γ and IL-2).

This study underscores the exacerbating impact of Ss coinfection on PTB severity and treatment outcomes, emphasizing the need for integrated management strategies for affected patients.

Tuberculosis (TB) remains a significant global health threat with high mortality and efforts to meet WHO End TB Strategy milestones are off-track. It has become clear that TB is not a dichotomous infection with latent and active forms but presents along a disease spectrum. Subclinical TB plays a larger role in transmission than previously thought. Aerosol studies have shown that undiagnosed TB patients, even with paucibacillary disease, can be highly infectious and significantly contribute to TB spread. Encouraging clinical results have been seen with the M72/AS01E vaccine. If preliminary results can be confirmed in ongoing larger trials, modelling shows the vaccine can positively impact the epidemic. TB preventive therapy (TPT), especially for high-risk groups like people living with HIV and household contacts of drug-resistant TB patients, has shown efficacy but implementation is resource intensive. Treatment options for infectious patients have grown rapidly. New shorter, all-oral treatment regimens represent a breakthrough, but progress is threatened by rising resistance to bedaquiline. Many new chemical entities are entering clinical trials and raise hopes for all-new regimens that could overcome rising resistance rates to conventional agents. More research is needed on the management of complex cases, such as central nervous system TB and severe HIV-associated TB. Post-TB lung disease (PTLD) is an under-recognised but growing concern, affecting millions of survivors with lasting respiratory impairment and increased mortality. Continued investment in development of TB vaccines and therapeutics, treatment shortening, and management of TB sequelae is critical to combat this ongoing public health challenge.

To explore the potent anti-inflammatory and anti-tuberculosis potential of novel trimetallic Au-Pt-Cu nanofluids.

We employed BSA assay for anti-inflammatory assessment and tested anti-tuberculosis activity against Mycobacterium tuberculosis H37Rv. Advanced molecular docking, dynamics, and DFT analyses were conducted to reveal interaction mechanisms and electronic properties.

The nanofluids demonstrated impressive IC50 values (15.88 ± 0.09 μM and 0.54 ± 0.02 μg/ml), with strong binding affinities and stable interactions confirmed via molecular simulations. Favorable ADMET profiling highlights their promise as innovative therapeutic agents.

The emergence of new Mycobacterium tuberculosis (Mtb) strains resistant to the key drugs currently used in the clinic for tuberculosis treatment can substantially reduce the probability of therapy success, causing the relevance and importance of studies on the development of novel potent antibacterial agents targeting different vulnerable spots of Mtb. In this study, 28,860 compounds from the library of bioactive molecules were screened to identify novel potential inhibitors of β-ketoacyl-acyl carrier protein synthase I (KasA), one of the key enzymes involved in the biosynthesis of mycolic acids of the Mtb cell wall. In doing so, we used a structure-based virtual screening approach to drug repurposing that included high-throughput docking of the C171Q KasA enzyme with compounds from the library of bioactive molecules including the FDA-approved drugs and investigational drug candidates, assessment of the binding affinity for the docked ligand/C171Q KasA complexes, and molecular dynamics simulations followed by binding free energy calculations. As a result, post-modeling analysis revealed 6 top-ranking compounds exhibiting a strong attachment to the malonyl binding site of the enzyme, as evidenced by the values of binding free energy which are significantly lower than those predicted for the KasA inhibitor TLM5 used in the calculations as a positive control. In light of the data obtained, the identified compounds are suggested to form a good basis for the development of new antitubercular molecules of clinical significance with activity against the KasA enzyme of Mtb.Communicated by Ramaswamy H. Sarma.

Drug-resistant tuberculosis (DR-TB) represents a pressing global health issue, leading to heightened morbidity and mortality. Despite extensive research efforts, the escalation of DR-TB cases underscores the urgent need for enhanced prevention, diagnosis, and treatment strategies. This review delves deep into the molecular and genetic origins of different types of DR-TB, highlighting recent breakthroughs in detection and diagnosis, including Rapid Diagnostic Tests like Xpert Ultra, Whole Genome Sequencing, and AI-based tools along with latest viewpoints on diagnosis and treatment of DR-TB utilizing newer and repurposed drug molecules. Special emphasis is given to the pivotal role of novel drugs and discusses updated treatment regimens endorsed by governing bodies, alongside innovative personalized drug-delivery systems such as nano-carriers, along with an analysis of relevant patents in this area. All the compiled information highlights the inherent challenges of current DR-TB treatments, discussing their complexity, potential side effects, and the socioeconomic strain they impose, particularly in under-resourced regions, emphasizing the cost-effective and accessible solutions. By offering insights, this review aims to serve as a compass for researchers, healthcare practitioners, and policymakers, emphasizing the critical need for ongoing R&D to improve treatments and broaden access to crucial TB interventions.

Tuberculosis (TB), caused by the bacterium Mycobacterium tuberculosis (Mtb), is still a leading cause of mortality worldwide. Fifty-fungi from a marine-derived fungal library were screened for anti-Mtb activity, and an Aspergillus candidus strain with strong anti-Mtb activity was found. Three known flavones, chlorflavonin (1), dechlorflavonin (2), and bromoflavone (3), were isolated from this fungus. Chlorflavonin and bromoflavone showed inhibitory activity with MIC90 values of 2.6 and 1.2 μM, respectively. In combination with molecular docking, a series of new chlorflavonin derivatives (4-41) were rationally designed and semisynthesized. Three new derivatives substituted with (2)-chlorocinnamate (14), (3)-chlorocinnamate (15), and benzoate (18) at position 2' showed MIC90 values ranging from 0.7 to 1.0 μM, having the potential to be further explored as antitubercular agents.

Tuberculosis (TB) remains a persistent global health threat, with Mycobacterium tuberculosis (Mtb) continuing to be a leading cause of mortality worldwide. Despite efforts to control the disease, the emergence of multi-drug-resistant (MDR) and extensively drug-resistant (XDR) TB strains presents a significant challenge to conventional treatment approaches. Addressing this challenge requires the development of novel anti-TB drug molecules. This study employed de novo drug design approaches to explore new EthR ligands and ethionamide boosters targeting the crucial enzyme InhA involved in mycolic acid synthesis in Mtb. Leveraging REINVENT4, a modern open-source generative AI framework, the study utilized various optimization algorithms such as transfer learning, reinforcement learning, and curriculum learning to design small molecules with desired properties. Specifically, focus was placed on molecule optimization using the Mol2Mol option, which offers multinomial sampling with beam search. The study's findings highlight the identification of six promising compounds exhibiting enhanced activity and improved physicochemical properties through structure-based drug design and optimization efforts. These compounds offer potential candidates for further preclinical and clinical development as novel therapeutics for TB treatment, providing new avenues for combating drug-resistant TB strains and improving patient outcomes.

Tuberculosis (TB), caused by the Mycobacterium tuberculosis (M.tb), remains a serious medical concern globally. Resistant M.tb strains are emerging, partly because M.tb can survive within alveolar macrophages, resulting in persistent infection. Protein kinase G (PknG) is a mycobacterial virulence factor that promotes the survival of M.tb in macrophages. Targeting PknG could offer an opportunity to suppress the resistant M.tb strains.

In the present study, multiple computational tools were adopted to screen a library of 460,000 molecules for potential inhibitors of PknG of M.tb.

Seven Hits (1-7) were identified with binding affinities exceeding that of the reference compound (AX20017) towards the PknG catalytic domain. Next, the ADMETox studies were performed to identify the best hit with appropriate drug-like properties. The chromene glycoside (Hit 1) was identified as a potential PknG inhibitor with better pharmacokinetic and toxicity profiles rendering it a potential drug candidate. Furthermore, quantum computational analysis was conducted to assess the mechanical and electronic properties of Hit 1, providing guidance for further studies. Molecular dynamics simulations were also performed for Hit 1 against PknG, confirming the stability of its complex. In sum, the findings in the current study highlight Hit 1 as a lead with potential for development of drugs capable of treating resistant TB.

Tuberculosis (TB) remains a major cause of ill health and one of the leading causes of death worldwide, with about 1.25 million deaths estimated in 2023. Control measures have focused principally on early diagnosis, the treatment of active TB, and vaccination. However, the widespread emergence of anti-tuberculosis drug resistance remains the major public health threat to progress made in global TB care and control. Moreover, the Bacillus Calmette-Guérin (BCG) vaccine, the only licensed vaccine against TB in children, has been in use for over a century, and there have been considerable debates concerning its effectiveness in TB control. A multi-epitope vaccine against TB would be an invaluable tool to attain the Global Plan to End TB 2023-2030 target. A rational approach that combines several B-cell and T-cell epitopes from key lipoproteins was adopted to design a novel multi-epitope vaccine candidate. In addition, interactions with TLR4 were implemented to assess its ability to elicit an innate immune response. The conservation of the selected proteins suggests the possibility of cross-protection in line with the One Health approach to disease control. The vaccine candidate was predicted to be both antigenic and immunogenic, and immune simulation analyses demonstrated its ability to elicit both humoral and cellular immune responses. Protein-protein docking and normal-mode analyses of the vaccine candidate with TLR4 predicted efficient binding and stable interaction. This study provides a promising One Health approach for the design of multi-epitope vaccines against human and livestock tuberculosis. Overall, the designed vaccine candidate demonstrated immunogenicity and safety features that warrant further experimental validation in vitro and in vivo.

A major challenge in tuberculosis (TB) therapeutics is that antibiotic exposure leads to changes in the physiology of M. tuberculosis (Mtb), which may enable the pathogen to withstand treatment. While antibiotic-treated Mtb has been evaluated in in vitro experiments, it is unclear if and how long-term in vivo treatment with diverse antibiotics with varying treatment-shortening activity (sterilizing activity) affects Mtb physiologic processes differently. Here, we used SEARCH-TB, a pathogen-targeted RNA-sequencing platform, to characterize the Mtb transcriptome in the BALB/c high-dose aerosol infection mouse model following 4 weeks of treatment with three sterilizing and three non-sterilizing antibiotics. Certain transcriptional changes were shared among most antibiotics, including decreased expression of genes associated with protein synthesis and metabolism and the induction of certain genes associated with stress responses. However, the magnitude of this shared response differed between antibiotics. Sterilizing antibiotics rifampin, pyrazinamide, and bedaquiline generated a more quiescent Mtb state than did non-sterilizing antibiotics isoniazid, ethambutol, and streptomycin, as indicated by the decreased expression of genes associated with translation, transcription, secretion of immunogenic proteins, metabolism, and cell wall synthesis. Additionally, we identified distinguishing transcriptional effects specific to each antibiotic, indicating that different mechanisms of action induce distinct patterns in response to cellular injury. In addition to elucidating the Mtb physiologic changes associated with antibiotic stress, this study demonstrates the value of SEARCH-TB as a highly granular pharmacodynamic assay that reveals antibiotic effects that are not apparent based on culture alone.

Tuberculosis (TB) has become the biggest threat to human society because of the rapid rise in resistance to the causative bacteria Mycobacterium tuberculosis (MTB) against the available anti-tubercular drugs. There is an urgent need to design new multi-targeted anti-tubercular agents to overcome the resistance species of MTB through computational design tools. With this aim in mind, we performed a combination of atom-based three-dimensional quantitative structure-activity relationship (3D-QSAR), six-point pharmacophore (AHHRRR), and molecular docking analysis on a series of fifty-eight anti-tubercular agents. The created QSAR model had a R2 value of 0.9521, a Q2 value of 0.8589, and a Pearson r-factor of 0.8988, all of which are statistically significant. This means that the model was effective at making predictions. We performed the molecular docking study for the data set of compounds with the two important anti-tubercular target proteins, Enoyl acyl carrier protein reductase (InhA) (PDBID: 2NSD) and Decaprenyl phosphoryl-β-D-Ribose 20-epimerase (DprE1) (PDBID: 4FDO). We used the similarity search principle to do virtual screening on 237 compounds from the PubChem database in order to find strong anti-tubercular agents that act against multiple targets. The screened compound, MK3, showed the highest docking score of -9.2 and -8.3 kJ/mol towards both the target proteins InhA and DprE1, which were picked for a 100 ns molecular-dynamic simulation study using GROMACS. The data showed that the compound MK3 was thermodynamically stable and effectively bound to both target proteins in their active binding pockets without much movement. The analysis of the highest occupied molecular orbital (HOMO), lowest unoccupied molecular orbital (LUMO), and energy gap predicts the molecular reactivity and stability of the identified molecule. Based on the result of the above studies, the proposed compound MK3 can be successfully used for the development of a novel multi-targeted anti-tubercular agent with high binding affinity and favourable ADME-T properties.

Background/Objectives: This study aims to develop a sustainable and environmentally friendly drug delivery system by synthesizing a novel drug-drug eutectic mixture (DDEM) of acetylsalicylic acid (ASA) and pyrazinamide (PZA) using a green and efficient mechanochemical approach. Methods: The DDEM was characterized using various techniques, including differential scanning calorimetry (DSC), thermogravimetry and differential thermal analysis (TG-DTA), powder X-ray diffraction (PXRD), Fourier transform infrared spectroscopy (FT-IR), and Raman spectroscopy. Binary phase diagrams and Tammann's triangle analysis determined the eutectic point. Density functional theory (DFT) calculations were performed on the starting compounds. The new system was evaluated for aqueous solubility, dissolution, and hygroscopicity. Results: A V-shaped binary phase diagram indicated the formation of a DDEM with a 2:1 molar ratio of ASA to PZA. A positive mixing enthalpy suggested a quasi-eutectic structure. The solubility of ASA and PZA increased by 61.5% and 85.8%, respectively, in the DDEM compared to the pure drugs. Conclusions: These findings highlight the potential of DDEMs to enhance drug properties and delivery. The synergistic interaction between ASA and PZA in the eutectic mixture may further improve therapeutic efficacy, warranting further investigation.

Drug discovery is inherently challenged by a multiple criteria decision making problem. The arduous path from hit discovery through lead optimization and preclinical candidate selection necessitates the evolution of a plethora of molecular properties. In this study, we focus on the hit discovery phase while beginning to address multiple criteria critical to the development of novel therapeutics to treat Mycobacterium tuberculosis infection. We develop a hybrid structure- and ligand-based pipeline for nominating diverse inhibitors targeting the β-ketoacyl synthase KasA by employing a Bayesian optimization-guided docking method and an ensemble model for compound nominations based on machine learning models for in vitro antibacterial efficacy, as characterized by minimum inhibitory concentration (MIC), and mouse pharmacokinetic (PK) plasma exposure. The application of our pipeline to the Enamine HTS library of 2.1M molecules resulted in the selection of 93 compounds, the experimental validation of which revealed exceptional PK (41%) and MIC (19%) success rates. Twelve compounds meet hit-like criteria in terms of MIC and PK profile and represent promising seeds for future drug discovery programs.

Mycobacterium tuberculosis (MTB) causing tuberculosis (TB) infection is a leading source of illness and death in developing nations, and the emergence of drug-resistant TB remains a significant global threat and a challenge in treating the disease. Mutations in the inhA and katG genes are connected to the principal molecular mechanism of isoniazid (INH) resistance, and continuous treatment of INH for more than a decade led to the evolution of INH resistant-TB (inhR-TB). Structure-based drug discovery approaches on traditional natural compounds are the contemporary source to identify significant lead molecules. This work focuses on discovering effective small compounds from natural compound libraries and applying pharmacophore-based virtual screening to filter out the molecules. The best-identified hit complexes were used for molecular dynamics simulations (MDS) to observe their stability and compactness. A three-dimensional e-pharmacophore hypothesis and screening generated 62 hits based on phase fitness scores from the pharmacophore-based virtual screening. Molecular docking experiments in Maestro's GLIDE module indicated that ZINC000002383126 and ASN22022 may be potential inhibitors of inhA and katG (native, inhA mutants S94A, Y158A, Y158F and Y158S and D137S, Y229F, S315T, W321F, and R418L mutants of katG). In addition, MDS analysis indicated that the native and mutant docked complexes of inhA and katG had good stability and remained compact in the binding pocket of the targets. In vitro studies can further validate the compounds that can act as INH competitive inhibitors.Communicated by Ramaswamy H. Sarma.

Spinal tuberculosis (TB), or Pott's disease, remains a significant global health issue, particularly in regions with high TB prevalence. Despite antitubercular drug therapy being the primary treatment, surgical intervention is often required in cases of spinal instability or neurological complications. This study aims to conduct a comprehensive bibliometric analysis of worldwide publications related to the surgical management of spinal TB and to compare contributions from orthopaedic surgery and neurosurgery in this field.

A bibliometric analysis was performed using data from the Scopus database, covering publications from 1896 to 2024. The search strategy focused on terms related to spinal TB and surgical interventions. The analysis included 1,857 publications, which were examined for trends, key contributors, and the evolution of surgical techniques. Metrics such as the number of publications, leading authors, affiliations, countries, and funding sponsors were compared between orthopaedic surgery and neurosurgery.

This study identified a steady increase in the number of publications over time. Key topics evolved from basic surgical methods, including early spinal procedures, to integrating pharmacological approaches alongside surgical techniques, such as antitubercular drugs, advancing into imaging research and procedure research involving refined surgical methods like spinal fusion. The recent phase reflects a shift towards technology-driven approaches, including minimally invasive techniques, artificial intelligence, and machine learning. China emerged as the leading country with the most contributions based on author, affiliations, funding sponsors, and countries. Last, orthopaedic surgery had more publications (274) than neurosurgery (96).

In conclusion, spinal TB surgery has evolved significantly, with a notable shift towards advanced, technology-driven approaches. Orthopaedic surgery leads in research output compared to neurosurgery. This bibliometric analysis provides valuable insights into the global research landscape, guiding future studies in the management of spinal TB.

The development of granulomas with central necrosis harboring Mycobacterium tuberculosis (Mtb) is the hallmark of human tuberculosis (TB). New anti-TB therapies need to effectively penetrate the cellular and necrotic compartments of these lesions and reach sufficient concentrations to eliminate Mtb. BTZ-043 is a novel antibiotic showing good bactericidal activity in humans in a phase IIa trial. Here, we report on lesional BTZ-043 concentrations severalfold above the minimal-inhibitory-concentration and the substantial local efficacy of BTZ-043 in interleukin-13-overexpressing mice, which mimic human TB pathology of granuloma necrosis. High-resolution MALDI imaging further reveals that BTZ-043 diffuses and accumulates in the cellular compartment, and fully penetrates the necrotic center. This is the first study that visualizes an efficient penetration and accumulation of a clinical-stage TB drug in human-like centrally necrotizing granulomas and that also determines its lesional activity. Our results most likely predict a substantial bactericidal effect of BTZ-043 at these hard-to-reach sites in TB patients.

Tuberculosis (TB) remains a major global threat, with 10 million new cases and 1.5 million deaths each year. In multidrug-resistant tuberculosis (MDR-TB), resistance is most commonly observed against isoniazid (INH) and rifampicin (RIF), the two frontline drugs. Isoniazid resistance is predominantly linked to mutations in the InhA gene, which encodes an enzyme involved in mycolic acid synthesis, a vital component of the mycobacterial cell wall. Mutations in InhA reduce drug binding, rendering INH ineffective. These morbidity and mortality figures, along with the fact that the rise and global spread of drug-resistant TB, underscores the need for the discovery of novel therapeutics. In this direction, we have previously synthesized, characterized, and screened a library of diaryl ether dehydrozingerone derivatives against mycobacteria and identified two best hits, 7 and 14, based on bacteriostatic activities. The present study aimed to thoroughly investigate the antituberculosis potential of these compounds, particularly regarding drug-resistant TB. Our findings revealed that both compounds exhibited tuberculocidal activity against the standard laboratory strain Mycobacterium tuberculosis (M. tb) H37Rv, with minimal bactericidal concentrations (MBC) of 4μg/ml for compound 7 and 8 μg/ml for compound 14. Next, concentration vs time-kill kinetics of both these compounds showed concentration-dependent bactericidal activities against M. tb and complete pathogen eradication from culture at just 16× MIC. Both compounds were found to be suitable for combination regimens as their interactions with isoniazid and rifampicin against M. tb were observed to be synergistic. Additionally, 7 and 14 exhibited minimal hemolysis against human RBCs and less cytotoxicity was observed against three human cell lines up to 1000 μM. Molecular docking revealed that these compounds bind more effectively to M. tb InhA, including its mutant forms where isoniazid binding is impaired, outperforming both isoniazid and triclosan in binding affinity. Importantly 7 and 14 showed potent activity against drug-susceptible clinical isolates and two isoniazid-resistant M. tb clinical isolates equivalent to that against M. tb H37Rv. The most interesting observation was that both compounds were found to be effective against three multi-drug resistant (MDR) strains of M. tb, thereby depicting their potential against drug-resistant TB. An ex vivo assay on RAW 264 cells infected with M. tb demonstrated a significant reduction in bacterial load at 8× MIC, revealing the fact that these compounds are highly effective against intracellular M. tb H37Rv. To the best of our knowledge, this is the first study that reports promising antimycobacterial potential of 7 and 14 against drug-susceptible, isoniazid-resistant, and MDR tuberculosis which warrants further exploration considering the need for new anti-TB medicine.

Tuberculosis (TB) poses a significant threat to global health, with millions of new infections and approximately one million deaths annually. Various modeling efforts have emerged, offering tailored data-driven and physiologically-based solutions for novel and historical compounds. However, this diverse modeling panorama may lack consistency, limiting result comparability. Drug-specific models are often tied to commercial software and developed on various platforms and languages, potentially hindering access and complicating the comparison of different compounds.

This work introduces stormTB: SimulaTOr of a muRine Minimal-pbpk model for anti-TB drugs. It is a web-based interface for our minimal physiologically based pharmacokinetic (mPBPK) platform, designed to simulate custom treatment scenarios for tuberculosis in murine models. The app facilitates visual comparisons of pharmacokinetic profiles, aiding in assessing drug-dose combinations.

The mPBPK model, supporting 11 anti-TB drugs, offers a unified perspective, overcoming the potential inconsistencies arising from diverse modeling efforts. The app, publicly accessible, provides a user-friendly environment for researchers to conduct what-if analyses and contribute to collective TB eradication efforts. The tool generates comprehensive visualizations of drug concentration profiles and pharmacokinetic/pharmacodynamic indices for TB-relevant tissues, empowering researchers in the quest for more effective TB treatments. stormTB is freely available at the link: https://apps.cosbi.eu/stormTB.