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CRISPR-dead Cas9 interference (CRISPRi) has become a valuable tool for precise gene regulation. In this study, CRISPRi was designed to target the inhA gene of Mycobacterium smegmatis (Msm), a gene necessary for mycolic acid synthesis. Our findings revealed that sgRNA2 induced with 100 ng/ml aTc achieved over 90% downregulation of inhA gene expression and inhibited bacterial viability by approximately 1,000-fold. Furthermore, CRISPRi enhanced the susceptibility of M. smegmatis to isoniazid and rifampicin, which are both 50% and 90% lower than those of the wild-type strain or other strains, respectively. This study highlights the ability of CRISPRi to silence the inhA gene, which impacts bacterial viability and drug susceptibility. The findings provide valuable insights into the utility of CRISPRi as an alternative tool for gene regulation. CRISPRi might be further assessed for its synergistic effect with current anti-tuberculosis drugs and its possible implications for combating mycobacterial infections, especially drug-resistant tuberculosis. [ABSTRACT FROM AUTHOR]

The rapid emergence of various forms of drug‐resistant tuberculosis (TB) has massively impacted people's health globally. According to the WHO, TB care and facilities have been downgraded by 21% since the COVID‐19 pandemic, which has further slowed down the progress toward the "End TB Strategy." Multiple studies have shown drug‐resistant tuberculosis to be resilient to front‐line antituberculosis drugs, including isoniazid (INH), thus constantly prompting researchers worldwide to probe new potent chemical entities to replace/supplement the current artillery of anti‐TB therapeutics. The molecular hybridization (MH) technique has recently been adopted by many synthetic and medicinal chemists to combine different pharmacophoric units into a single molecular architecture which in most cases has been reported to exhibit a better pharmacological profile as compared to its precursors. The present review provides a concise account of the MH hybridization strategies directed at the structural manipulations of isoniazid to develop new anti‐TB agents and their structure‐activity relationships. Furthermore, their toxicity profiles and docking studies with relevant receptors are also briefly discussed. [ABSTRACT FROM AUTHOR]

The Mycobacterium cell wall is a capsule‐like structure comprising of various layers of biomolecules such as mycolic acid, peptidoglycans, and arabinogalactans, which provide the Mycobacteria a sort of cellular shield. Drugs like isoniazid, ethambutol, cycloserine, delamanid, and pretomanid inhibit cell wall synthesis by inhibiting one or the other enzymes involved in cell wall synthesis. Many enzymes present across these layers serve as potential targets for the design and development of newer anti‐TB drugs. Some of these targets are currently being exploited as the most druggable targets like DprE1, InhA, and MmpL3. Many of the anti‐TB agents present in clinical trials inhibit cell wall synthesis. The present article covers a systematic perspective of developing cell wall inhibitors targeting various enzymes involved in cell wall biosynthesis as potential drug candidates for treating Mtb infection. [ABSTRACT FROM AUTHOR]

The inhibin alpha (INHA) serves as a marker for the number of fully developed ovarian follicles and plays a crucial role in regulating the secretion of pituitary FSH (follicle-stimulating hormone) and the frequency of ovulation. This study aims to examine the effect of INHA gene polymorphisms on the litter size of thin-tailed sheep. Detection of single nucleotide polymorphisms (SNPs) in the INHA gene was performed using PCR and DNA sequencing techniques. A total of 45 ewes were included in the study. Three SNPs were identified: g.236311141G>C, g.236311367G>A, and g.236311368G>A. Further investigation of the g.236311367G > A variant revealed that individuals with the GA genotype had a significantly higher litter size than those with the AA or GG genotype (p<0.05). SNPs at positions g.236311141G/C and g.236311368G/A were non-synonymous mutations resulting in amino acid changes p.A225P and V301I, respectively. Our results suggest that g.236311367G>A loci may serve as a potential molecular marker for improving the litter size trait in thin-tailed sheep. [ABSTRACT FROM AUTHOR]

Abstract CRISPR-dead Cas9 interference (CRISPRi) has become a valuable tool for precise gene regulation. In this study, CRISPRi was designed to target the inhA gene of Mycobacterium smegmatis (Msm), a gene necessary for mycolic acid synthesis. Our findings revealed that sgRNA2 induced with 100 ng/ml aTc achieved over 90% downregulation of inhA gene expression and inhibited bacterial viability by approximately 1,000-fold. Furthermore, CRISPRi enhanced the susceptibility of M. smegmatis to isoniazid and rifampicin, which are both 50% and 90% lower than those of the wild-type strain or other strains, respectively. This study highlights the ability of CRISPRi to silence the inhA gene, which impacts bacterial viability and drug susceptibility. The findings provide valuable insights into the utility of CRISPRi as an alternative tool for gene regulation. CRISPRi might be further assessed for its synergistic effect with current anti-tuberculosis drugs and its possible implications for combating mycobacterial infections, especially drug-resistant tuberculosis.

In this study, we report the synthesis and characterization using various spectroscopic techniques of a series of six new furan-based Schiff base derivatives as potential antitubercular agents. The strategic incorporation of the furan scaffold and nitro group, both known for their diverse biological activities, provided a strong rationale for their evaluation against tuberculosis. Molecular docking simulations were employed to assess the binding affinities of these compounds towards enoyl-acyl carrier protein reductase (InhA), a validated and essential target in M. tuberculosis. Remarkably, all synthesized compounds exhibited superior binding scores (–6.074 to –9.939 kcal/mol), compared to the clinically used antitubercular drug isoniazid (–4.585 kcal/mol), indicating their potential as potent InhA inhibitors. Furthermore, in silico absorption, distribution, metabolism, and excretion (ADME) predictions were performed to evaluate the drug-likeness of these compounds. The findings highlight the balanced combination of potent target binding and drug-like characteristics achieved by these furan-based Schiff base hybrids. [ABSTRACT FROM AUTHOR]

Tuberculosis (TB) caused by the Mycobacterium tuberculosis (MTB) pathogen is a serious contagious illness that endangers the public's health. However, TB is treatable but due to the result of ineffective and protracted medication, MDR‐MTB and XDR‐MTB were able to develop, making their eradication highly challenging and necessitating potential treatments to be cured. This study aimed to identify the novel inhibitor for the InhA enzyme, a potential target of MTB. Initially, 2721 phytochemicals libraries were screened for ADME/Tox profiling and 49 phytochemicals were filtered out. Moreover, these phytochemicals with reference inhibitor triclosan were docked against InhA (PDB ID: 1BVR)., were showing binding energies ranging from −9.16–−6.3 kcal/mol, whereas triclosan showed −6.90 kcal/mol of binding energy. The docking results reveal that irilone (−9.16 kcal/mol) and aurantio‐obtusin (−8.84 kcal/mol) exhibit promising binding energy and perfectly occupy the binding site of InhA. Furthermore, molecular dynamics simulation (MDS) and MMGBSA studies showed better affinities of selected compounds in terms of stability, compactness, flexibility, and total BFE in complexes with InhA. This study concluded that irilone and aurantio‐obtusin could be promising InhA inhibitors based on their impressive pharmacokinetic profiling, toxicity, binding affinity, and stability. However experimental validation is a subject of further research. [ABSTRACT FROM AUTHOR]

We evaluated the ability of FluoroType MTBDR version 2 (FTv2; Hain Lifescience), a second-step real-time PCR assay, to simultaneously detect Mycobacterium tuberculosis complex (MTBC) DNA and mutations conferring resistance to rifampicin (RIF) and isoniazid (INH), in pulmonary and extrapulmonary samples from patients and compared them with corresponding cultures. FTv2 MTBC was evaluated on 1815 and 432 samples from Denmark (DK) and Germany (DE), respectively. RIF and INH resistance mutations were assessed in the German samples and 110 samples from Sierra Leone and subsequently compared to phenotypic antimicrobial susceptibility testing and a composite reference DNA (CRD) based on the GenoType MTBDR line-probe assay and Sanger sequencing or whole-genome sequencing. Of the 584 (557 smear-negative) Danish and 277 (85 smear-negative) German sputum samples, 42 (16) and 246 (54) were culture positive, and 44 (18) and 222 (35) were FTv2 positive, providing an FTv2 sensitivity and specificity of 0.86 (0.63) and 0.98 (DK), 0.90 (0.65) and 1.00 (DE), respectively. The count, sensitivities, and specificities for all pulmonary samples were 1434, 0.79, and 0.99 (DK) and 347, 0.86, and 1.00 (DE), respectively; for extrapulmonary samples, 381, 0.33, 0.99 (DK) and 83, 0.50, and 1.00 (DE). The valid count, sensitivity, and specificity compared with CRD for detecting resistance mutations were RIF 355, 0.99, 0.96, and INH 340, 1.00, and 0.98, respectively. FTv2 reliably detects MTBC DNA in pulmonary and extrapulmonary samples and detects resistance mutations for INH and RIF resistance in inhA promoter, katG, and rpoB genes. [ABSTRACT FROM AUTHOR]

A set of alkyl-/1H-1,2,3-triazole-based dimers was strategically designed and synthesized to evaluate their in vitro anti-mycobacterial activities against Mycobacterium tuberculosis and the non-tuberculous Mycobacterium abscessus strains. Systematic variations in the nature (alkyl/1H-1,2,3-triazole) and positioning of the linker were implemented based on the docking scores observed in the binding sites identified in the crystal structures of InhA from M. tuberculosis and M. abscessus. However, the in vitro evaluation results revealed that the synthesized compounds did not exhibit inhibitory effects on the growth of mycobacteria, even at the highest tested concentrations. The elevated lipophilicity values determined through ADMET studies for these synthesized dimers might be a contributing factor to their poor activity profiles.

Blackjack (Bidens pilosa L.) has a bioactive compound, one of which is polyacetylene, which can inhibit the growth of antimicrobials in general. This study aims to conduct an in silico study to determine the antibacterial activity against Mycobacterium Tuberculosis of 20 derivatives polyacetylene; TPC was determined by Folin–Ciocalteu colorimetric method using gallic acid as standard, and various concentrations of the extract solutions were measured at 741 nm. TFC was calculated using an aluminum chloride colorimetric assay. Quercetin was used as standard, and the absorbance was measured at 426 nm. Antioxidant activity was evaluated using DPPH scavenging 517 nm and Frap Assay 596 nm. The molecular docking results showed that the compounds (5-(2-Phenylethynyl)-2-b-glucosylmethyl-thiophene, Glucopyranosyloxy-3-hydroxy-6(E)-tetradecane-8,10,12-triyne and Phenylhexa-1,3,5-triyn-1-yl acetate were most potentially anti-tuberculosis on the target protein InhA with the binding energy produced

BACKGROUND: Isoniazid (INH) is one of the first-line drugs used for the treatment of Mycobacterium tuberculosis (Mtb). This study aimed to determine the mutations in katG and inhA promoter regions associated with INH resistance in clinical isolates of Mtb from Tehran, Iran. METHODS: This descriptive cross-sectional study was conducted in the tuberculosis (TB) center of Tehran in 2020. 50 samples obtained from these patients were cultured on Löwenstein-Jensen medium (LJ), then, INH-resistant strains and their minimum inhibitory concentration (MIC) were determined using the proportional method. To determine the INH-resistance mutations, sequencing was performed following the amplification of both katG and inhA genes using real-time polymerase chain reaction (PCR). The genetic relationships were determined based on each strain's genetic pattern using the primers related to enterobacterial repetitive intergenic consensus PCR (ERIC-PCR), which is a proper tool used for typing Mtb strains. In this study, the phylogenetic tree was plotted using NTSYS software. RESULTS: Five INH-resistant Mtb strains were isolated from 50 patients with TB in Tehran. All the studied resistant strains (100%) showed a mutation in codon 315 of the katG gene; none of them exhibited any mutation in the promoter. I335T mutation was observed in one INH-resistant strain (20%). The phylogenetic tree of the strains indicated seven clusters as well as 31 patterns in the strains. The strain with two mutations in 335 and 315 had MIC > 8. CONCLUSION: KatG mutation could result in a high level of INH resistance. Therefore, routine identification of this mutation can help in determining the INH resistance, thereby preventing further propagation of these strains.

Blackjack (Bidens pilosa L.) has a bioactive compound, one of which is polyacetylene, which can inhibit the growth of antimicrobials in general. This study aims to conduct an in silico study to determine the antibacterial activity against Mycobacterium Tuberculosis of 20 derivatives polyacetylene; TPC was determined by Folin--Ciocalteu colorimetric method using gallic acid as standard, and various concentrations of the extract solutions were measured at 741 nm. TFC was calculated using an aluminum chloride colorimetric assay. Quercetin was used as standard, and the absorbance was measured at 426 nm. Antioxidant activity was evaluated using DPPH scavenging 517 nm and Frap Assay 596 nm. The molecular docking results showed that the compounds (5-(2-Phenylethynyl)-2-b-glucosylmethyl-thiophene, Glucopyranosyloxy-3-hydroxy-6(E)-tetradecane-8,10,12-triyne and Phenylhexa-1,3,5-triyn-1-yl acetate were most potentially anti-tuberculosis on the target protein InhA with the binding energy produced <-6.55 Kcal/mol and the lowest inhibitory cost of 1.838x10-6. The compound has amino acids similar to natural ligands and standard compounds of isoniazid. The given ADME-T predictions showed good pharmacokinetic properties and safe levels of toxicity (5-(2-Phenylethynyl)-2-b-glucosylmethyl-thiophene, Glucopyranosyloxy-3-hydroxy-6(E)-tetradecane-8,10,12-triyne in hepatotoxic and LD50. The highest antioxidant properties in blackjack are produced by FRAP assay 722.905±7.985 mMTE/gr. This proves that blackjack has antioxidant activity and is required for bioassayguided isolation testing against Mycobacterium tuberculosis. [ABSTRACT FROM AUTHOR]

Antimicrobial resistance is a global health threat that requires innovative strategies against drug-resistant bacteria. Our study focuses on enoyl-acyl carrier protein reductases (ENRs), in particular FabI, FabK, FabV, and InhA, as potential antimicrobial agents. Despite their promising potential, the lack of clinical approvals for inhibitors such as triclosan and isoniazid underscores the challenges in achieving preclinical success. In our study, we curated and analyzed a dataset of 1412 small molecules recognized as ENR inhibitors, investigating different structural variants. Using advanced cheminformatic tools, we mapped the physicochemical landscape and identified specific structural features as key determinants of bioactivity. Furthermore, we investigated whether the compounds conform to Lipinski rules, PAINS, and Brenk filters, which are crucial for the advancement of compounds in development pipelines. Furthermore, we investigated structural diversity using four different representations: Chemotype diversity, molecular similarity, t-SNE visualization, molecular complexity, and cluster analysis. By using advanced bioinformatics tools such as matched molecular pairs (MMP) analysis, machine learning, and SHAP analysis, we were able to improve our understanding of the activity cliques and the precise effects of the functional groups. In summary, this chemoinformatic investigation has unraveled the FAB inhibitors and provided insights into rational antimicrobial design, seamlessly integrating computation into the discovery of new antimicrobial agents. [ABSTRACT FROM AUTHOR]

The inhibin alpha (INHA) serves as a marker for the number of fully developed ovarian follicles and plays a crucial role in regulating the secretion of pituitary FSH (follicle-stimulating hormone) and the frequency of ovulation. This study aims to examine the effect of INHA gene polymorphisms on the litter size of thin-tailed sheep. Detection of single nucleotide polymorphisms (SNPs) in the INHA gene was performed using PCR and DNA sequencing techniques. A total of 45 ewes were included in the study. Three SNPs were identified: g.236311141G>C, g.236311367G>A, and g.236311368G>A. Further investigation of the g.236311367G > A variant revealed that individuals with the GA genotype had a significantly higher litter size than those with the AA or GG genotype (pA loci may serve as a potential molecular marker for improving the litter size trait in thin-tailed sheep.

Aim: The configuration found programmed recreation ligand-receptor relations were worn during rework also rationale of investigate in order to establish budding bio-energetic candidate seeing that (Enoyl-ACP)-Enoyl-acyl carrier protein reductase (InhA) antagonist with in silico analog intend advance. Materials and Methods: InhA go towards NADH-dependent Enoyl ACP (Co-A) reductase enzyme folks also extend acyl fatty acid, which be progenitor of mycolic acid also myco-bacterial cells component. Quinazolin-4-one have been used to therapy a set of misery a set in human renowned mechanism exemplifies to facilitate various biological activities. Assortment of literature quinazolin-4(3H)-one derivatives exhibited anti-tubercular action have been raising targets of some quinazolin-4-one as well as exhibits all most various biological activity with increased anti-tubercular activity at 3rd position. Results: This study suggested that the designed quinazolin-4-one derivatives systematically investigate requisite affinity and drug likeness property beside the Enoyl ACP (InhA). The interface of lately deliberate candidates (QT1 to QDT8) among admirable binding interaction of alongside preferred (PDB ID: 4TZK) Macromolecule with good ADMET Properties. Conclusion: Further may launch a consistent medicine or sustain potential escort acknowledged and could exist additional taken for experimental studies. [ABSTRACT FROM AUTHOR]

Background: High-dose isoniazid is recommended in the 9–12 months short-course regimen for multidrug-resistant tuberculosis with inhA mutation. However, there is insufficient evidence to support the assumption of genotypic-phenotypic concordance. This study aimed to identify the genetic mutations associated with high-level phenotypic isoniazid resistance. Methods: Clinical isolates from patients with drug-resistant tuberculosis were profiled by whole-genome sequencing and subjected to minimum inhibitory concentration (MIC) testing using MGIT based-method. MICs were performed in concentration ranges based on the mutation present: isolates with no isoniazid resistance-conferring mutations and H37Rv, 0.016–0.256 µg/ml; inhA, 0.256–4.0 µg/ml, katG 1.0–16.0 µg/ml; and inhA + katG, 4.0–64.0 µg/ml. Isolates demonstrating resistance at the upper limit of the concentration range were tested up to the maximum of 64.0 µg/ml. Bootstrap of the mean MICs was performed to increase the robustness of the estimates and an overlap index was used to compare the distributions of the MICs for each mutation profile. Results: A total of 52 clinical isolates were included in this analysis. Bootstrap MIC means for inhA, katG and inhA + katG were 33.64 (95% CI, 9.47, 56.90), 6.79 (4.45, 9.70) and 52.34 (42.750, 61.66) µg/ml, respectively. There was high overlap between inhA and inhA + katG mutations (eta = 0.45) but not with inhA and katG (eta = 0.19). Furthermore, katG showed poor overlap with inhA + katG mutations (eta = 0.09). Unexpectedly, 4/8 (50.0%) of all InhA mutants demonstrated high-level resistance, while 20/24 (83.3%) of katG mutants demonstrated moderate-level resistance. Conclusions: InhA mutations demonstrated unexpectedly high MICs and showed high overlap with inhA + katG. Contrary to the common belief that katG mutants are associated with high-level resistance, this mutation primarily showed moderate-level resistance.

Ghyzlane EL Haddoumi,1,2 Mariam Mansouri,1,2 Houda Bendani,1,2 El Mehdi Bouricha,1,2 Ilham Kandoussi,1,2 Lahcen Belyamani,2– 4 Azeddine Ibrahimi1– 3 1Biotechnology Lab (MedBiotech), Rabat Medical and Pharmacy School, University Mohammed V, Rabat, Morocco; 2Centre Mohammed VI for Research & Innovation (CM6), Rabat, Morocco; 3Mohammed VI University of Health Sciences (UM6SS), Casablanca, Morocco; 4Emergency Department, Military Hospital Mohammed V, Rabat, MoroccoCorrespondence: Azeddine Ibrahimi, Mohammed VI University of Health Sciences (UM6SS), Casablanca, Morocco, Tel +212660240131, Email aibrahimi@um6ss.maPurpose: The enoyl-acyl carrier protein reductase (InhA) is one of the important key enzymes employed in mycolic acids biosynthesis pathway and an important component of mycobacterial cell walls. This enzyme has also been identified as major target of isoniazid drug, except that isoniazid needs to be activated first by the catalase peroxidase (KatG) protein to form the isonicotinoyl-NAD (INH-NAD) adduct that inhibits the action of InhA enzyme. However, this activation becomes more difficult and unreachable with the problem of mutation-related resistance caused mainly by acquired mutations in KatG and InhA protein. Our main interest in this study is to identify direct InhA inhibitors using computer-aided drug design.Methods: Computer-aided drug design was used to solve this problem by applying three different approaches including mutation impact modelling, virtual screening and 3D-pharmacophore search.Results: A total of 15 mutations were collected from the literature, then a 3D model was generated for each of them and their impact was predicted. Of the 15 mutations, 10 were found to be deleterious and have a direct effect on flexibility, stability and SASA of the protein. In virtual screening, from 1,000 similar INH-NAD analogues obtained by the similarity search method, 823 compounds passed toxicity filter and drug likeness rules, which were then docked to the wild-type of InhA protein. Subsequently, 34 compounds with binding energy score better than that of INH-NAD were selected and docked against the 10 generated mutated models of InhA. Only three leads showed a lower binding affinity better than the reference. The 3D-pharmacophore model approach was used to identify the common features between those three compounds by generating a pharmacophoric map.Conclusion: The result of this study may pave the way to develop more potent mutant-specific inhibitors to overcome this resistance.Keywords: molecular docking, CADD, Mycobacterium tuberculosis, InhA, mutational profile

The enoyl reductase from Mycobacterium tuberculosis (MtInhA) was shown to be a major target for isoniazid, the most prescribed first-line anti-tuberculosis agent. The MtInhA (EC 1.3.1.9) protein catalyzes the hydride transfer from the 4S hydrogen of β-NADH to carbon-3 of long-chain 2-trans-enoyl thioester substrates (enoyl-ACP or enoyl-CoA) to yield NAD+ and acyl-ACP or acyl-CoA products. The latter are the long carbon chains of the meromycolate branch of mycolic acids, which are high-molecular-weight α-alkyl, β-hydroxy fatty acids of the mycobacterial cell wall. Here, stopped-flow measurements under single-turnover experimental conditions are presented for the study of the transient of reactants. Single-turnover experiments at various enzyme active sites were carried out. These studies suggested isomerization of the MtInhA:NADH binary complex in pre-incubation and positive cooperativity that depends on the number of enzyme active sites occupied by the 2-trans-dodecenoyl-CoA (DD-CoA) substrate. Stopped-flow results for burst analysis indicate that product release does not contribute to the rate-limiting step of the MtInhA-catalyzed chemical reaction. The bearings that the results presented herein have on function-based anti-tuberculosis drug design are discussed.

Abstract Background Different anesthetics may have opposite effects on the immune system, thus affecting the prognosis of tumor patients. Cell-mediated immunity forms the primary defense against the invasion of tumor cells, so manipulation of the immune system to produce an enhanced anti-tumor response could be utilized as an adjuvant oncological therapy. Sevoflurane has proinflammatory effects, while propofol, has anti-inflammatory and antioxidant effects. Therefore, we compared the overall survival (OS) and disease-free survival (DFS) of patients with esophageal cancer under total intravenous anesthesia and inhalation anesthesia. Methods This study collected the electronic medical records of patients undergoing esophagectomy from January 1, 2014 to December 31, 2016. According to the intraoperative anesthetics, the patients were divided into total intravenous anesthesia (TIVA) group or inhalational anesthesia (INHA) group. Stabilized inverse probability of treatment weighting (SIPTW) was used to minimize differences. Kaplan–Meier survival curve was established to evaluate the correlation between different anesthesia methods in overall survival and disease-free survival of patients undergoing esophageal cancer surgery. Results A total of 420 patients with elective esophageal cancer were collected, including 363 patients eligible for study (TIVA, n = 147, INHA, n = 216). After SIPTW there were no significant differences between two groups in overall survival and disease-free survival. However, the adjuvant therapy was statistically significant in improving OS, and the degree of differentiation was correlated with OS and DFS. Conclusions In conclusion, there were no significant difference in overall survival and disease-free survival between total intravenous anesthesia and inhalational anesthesia in patients undergoing esophageal cancer surgery.

Antimicrobial resistance is a global health threat that requires innovative strategies against drug-resistant bacteria. Our study focuses on enoyl-acyl carrier protein reductases (ENRs), in particular FabI, FabK, FabV, and InhA, as potential antimicrobial agents. Despite their promising potential, the lack of clinical approvals for inhibitors such as triclosan and isoniazid underscores the challenges in achieving preclinical success. In our study, we curated and analyzed a dataset of 1412 small molecules recognized as ENR inhibitors, investigating different structural variants. Using advanced cheminformatic tools, we mapped the physicochemical landscape and identified specific structural features as key determinants of bioactivity. Furthermore, we investigated whether the compounds conform to Lipinski rules, PAINS, and Brenk filters, which are crucial for the advancement of compounds in development pipelines. Furthermore, we investigated structural diversity using four different representations: Chemotype diversity, molecular similarity, t-SNE visualization, molecular complexity, and cluster analysis. By using advanced bioinformatics tools such as matched molecular pairs (MMP) analysis, machine learning, and SHAP analysis, we were able to improve our understanding of the activity cliques and the precise effects of the functional groups. In summary, this chemoinformatic investigation has unraveled the FAB inhibitors and provided insights into rational antimicrobial design, seamlessly integrating computation into the discovery of new antimicrobial agents.

Treatment options against Mycobacterium abscessus infections are very limited. New compounds are needed to cure M. abscessus pulmonary diseases. While the mycolic acid biosynthetic pathway has been largely exploited for the treatment of tuberculosis, this metabolic process has been overlooked in M. abscessus, although it offers many potential drug targets for the treatment of this opportunistic pathogen. Herein, the authors review the role of the MmpL3 membrane protein and the enoyl-ACP reductase InhA involved in the transport and synthesis of mycolic acids, respectively. They discuss their importance as two major vulnerable drug targets in M. abscessus and report the activity of MmpL3 and InhA inhibitors. In particular, they focus on NITD−916, a direct InhA inhibitor against M. abscessus, particularly warranted in the context of multidrug resistance. There is an increasing body of evidence validating the mycolic acid pathway as an attractive drug target to be further exploited for M. abscessus lung disease treatments. The NITD−916 studies provide a proof-of-concept that direct inhibitors of InhA are efficient in vitro, in macrophages and in zebrafish. Future work is now required to improve the activity and pharmacological properties of these inhibitors and their evaluation in pre-clinical models. [ABSTRACT FROM AUTHOR]

Introduction: Mycobacterium tuberculosis (MTB), the causative agent of tuberculosis, has been a global threat to human beings for several decades. Treating tuberculosis has become more difficult as the prevalence of drug-resistant tuberculosis has increased globally. Evidence suggests that the comprehensive landscape of resistance mechanisms in MTB is ambiguous. More importantly, little is known regarding the series of events connected to resistance mechanisms in MTB before exposure to anti-TB drugs, during exposure to the drugs, and finally, when the MTB becomes resistant after exposure, upon analyses of its genome. Methods: We used the wild-type strain of MTB (H37Rv) in an in vitro model for generating induced resistance using a sub-inhibitory concentration of isoniazid, and the generated resistance-associated variants (RAVs) were identified using the whole genome sequencing method. Results: The detection of an inhA promoter mutation (fabG1-15C>T), which results in increased production of InhA protein, was found to be a major mechanism for developing resistance to isoniazid in the first place. We observed adaptation of MTB resistance mechanisms in high isoniazid stress by alteration and abolishment of KatG due to the detection of katG S315N, the common region of mutation that confers isoniazid resistance, along with katG K414N, katG N138S, and katG A162E. Furthermore, we detected the ahpC-72C>T and ahpC 21C>A mutations, but further investigation is needed to determine their role in compensating for the loss of KatG activity. Discussion: This suggests that increased InhA production is themainmechanism where there are low levels of isoniazid, whereas the alteration of KatG was found to be utilized in mycobacterium with a high concentration of isoniazid. Our work demonstrates that this in vitro approach of generating induced resistance could provide clinically relevant information after the fabG1-15C>T mutation, which is the common mutation found in clinical isolates. Moreover, other mutations detected in this work can also be found in clinical isolates. These findingsmay shed light on the impact of isoniazid in generating RAV and the resistance mechanism scenario that mycobacterium used under various isoniazid-pressuring conditions. More research is needed to understand better the role of RAV and mechanical resistance events within the mycobacterium genome in promoting a promising drug prediction platform that could lead to the right treatment for patients with MDR-TB and XDR-TB. [ABSTRACT FROM AUTHOR]

Background: Mycobacterium tuberculosis is a bacterium that has historically had a substantial impact on human health. Despite advances in understanding and management of tuberculosis (TB), the disease remains a crucial problem that necessitates ongoing work to discover effective drugs, minimize transmission, and improve global health outcomes. Methods: The purpose of this study is to use molecular docking and absorption, distribution, metabolism, excretion, and toxicity (ADMET) analyses to explore the molecular interactions of different proteins that are involved in mycolic acid biosynthesis (HadAB, InhA, KasA, FabD, and beta-ketoacyl-acyl carrier protein synthase III) of M. tuberculosis with Demospongiae metabolites. The docking findings were evaluated using the glide gscore, and the top 10 compounds docked against each protein receptor were chosen. Furthermore, the selected compounds underwent ADMET analysis, indicating that they have the potential for therapeutic development. Results: Among the selected compounds, makaluvamine G showed the highest binding affinity against HadAB, psammaplysin E showed highest binding affinity against InhA, pseudotheonamide D showed the highest binding affinity against KasA protein, dinordehydrobatzelladine B showed the highest binding affinity against FabD, and nagelamide X showed the highest binding affinity against beta-ketoacyl-acyl carrier protein synthase III. Additionally, molecular mechanics generalized born surface area (MM-GBSA) binding free energy and molecular dynamics simulations were used to support the docking investigations. Conclusion: The results of the study suggest that these compounds may eventually be used to treat TB. However, computer validations were included in this study, and more in vitro research is required to turn these prospective inhibitors into clinical drugs. [ABSTRACT FROM AUTHOR]

The enoyl reductase from Mycobacterium tuberculosis (MtInhA) was shown to be a major target for isoniazid, the most prescribed first-line anti-tuberculosis agent. The MtInhA (EC 1.3.1.9) protein catalyzes the hydride transfer from the 4S hydrogen of β-NADH to carbon-3 of long-chain 2-trans-enoyl thioester substrates (enoyl-ACP or enoyl-CoA) to yield NAD+ and acyl-ACP or acyl-CoA products. The latter are the long carbon chains of the meromycolate branch of mycolic acids, which are high-molecular-weight α-alkyl, β-hydroxy fatty acids of the mycobacterial cell wall. Here, stopped-flow measurements under single-turnover experimental conditions are presented for the study of the transient of reactants. Single-turnover experiments at various enzyme active sites were carried out. These studies suggested isomerization of the MtInhA:NADH binary complex in pre-incubation and positive cooperativity that depends on the number of enzyme active sites occupied by the 2-trans-dodecenoyl-CoA (DD-CoA) substrate. Stopped-flow results for burst analysis indicate that product release does not contribute to the rate-limiting step of the MtInhA-catalyzed chemical reaction. The bearings that the results presented herein have on function-based anti-tuberculosis drug design are discussed. [ABSTRACT FROM AUTHOR]

Enoyl acyl carrier protein reductase (InhA) is a crucial enzyme for the biosynthesis of mycolic acids which are major compartments of the Mycobacterium tuberculosis (Mtb) cell wall. Direct inhibition of InhA without activation by drug-NADH adduct has clinical utility to overcome drug resistance. We aimed at the in silico identification of triclosan derivatives with the potential inhibitory effect of S94A-InhA as a clinically important mutant form. Caver Web 1.0 server was used to analyze the ligand transport through access tunnels. Two macrocyclic triclosan derivatives (4 and 6) could be identified with more energy-favorable transfer routes toward the enzyme active site. Molecular dynamics (MD) simulations (50 ns) of the best-scored compounds revealed the stability of enzyme structure upon binding to 4 and 6. Compound 4 could better retain enzyme stability upon target binding. Results of intermolecular H-bond analysis indicated that both complexes were mediated through hydrophobic contacts. Declined solvent accessible surface area (SASA) for the apo and bound enzyme states indicated non-destabilizing behavior and no structural relaxation. Electrostatic and van der Waals interactions between triclosan derivatives and their surroundings were used to acquire binding free energies through the linear interaction energy (LIE) method based on MD simulations (Average Δ G b , − 7. 8 3 ± 0. 7 2 kcal/mol and − 8. 8 9 ± 0. 6 4 kcal/mol). Both of the triclosan derivatives showed relatively stable energy variations and their steady accommodation inside enzyme active site could be confirmed during 50 ns. These results may be implicated in further structure-guided approaches against drug-resistant Mtb. Two macrocyclic triclosan derivatives (4 and 6) were identified as top-scored binders with superior energy-favorable transfer routes toward active site of Mtb S94A-InhA (Clinically frequent mutant enzyme). MD trajectories of compounds 4 & 6 confirmed stable binding conformations through cooperative hydrophobic interactions with LIE-based average bindig free energies of –7.83±0.72 and –8.89±0.64 kcal/mol, respectively. Dynamic behaviour of the mutant enzyme showed non-destabilizing function with no structural relaxation upon binding to triclosan derivatives. [ABSTRACT FROM AUTHOR]

Background: Mycobacterium tuberculosis is a bacterium that has historically had a substantial impact on human health. Despite advances in understanding and management of tuberculosis (TB), the disease remains a crucial problem that necessitates ongoing work to discover effective drugs, minimize transmission, and improve global health outcomes. Methods: The purpose of this study is to use molecular docking and absorption, distribution, metabolism, excretion, and toxicity (ADMET) analyses to explore the molecular interactions of different proteins that are involved in mycolic acid biosynthesis (HadAB, InhA, KasA, FabD, and beta-ketoacyl-acyl carrier protein synthase III) of M. tuberculosis with Demospongiae metabolites. The docking findings were evaluated using the glide gscore, and the top 10 compounds docked against each protein receptor were chosen. Furthermore, the selected compounds underwent ADMET analysis, indicating that they have the potential for therapeutic development. Results: Among the selected compounds, makaluvamine G showed the highest binding affinity against HadAB, psammaplysin E showed highest binding affinity against InhA, pseudotheonamide D showed the highest binding affinity against KasA protein, dinordehydrobatzelladine B showed the highest binding affinity against FabD, and nagelamide X showed the highest binding affinity against beta-ketoacyl-acyl carrier protein synthase III. Additionally, molecular mechanics generalized born surface area (MM-GBSA) binding free energy and molecular dynamics simulations were used to support the docking investigations. Conclusion: The results of the study suggest that these compounds may eventually be used to treat TB. However, computer validations were included in this study, and more in vitro research is required to turn these prospective inhibitors into clinical drugs.

Biotransformation of isoniazid produced isonicotinic acid (1), isonicotinic acid N-oxide (2), and isonicotinamide (3) which were isolated by column chromatography using silica gel and Sephadex LH 20 and elucidated using various spectroscopies. This is the first report for isolation of 2. Antituberculosis activity was evaluated against Mycobacterium tuberculosis strains: drug sensitive (DS), multiple drug resistant (MDR) and extensively drug resistant (XDR). 1-3 and isoniazid showed MICs of 63.49, 0.22, 15.98 and 0.88 µM, respectively, against the DS strain. For the MDR strain, 2 and 3 exhibited MICs of 28.06 and > 1000 µM, respectively, while 1 was inactive. Moreover, 2 had an MIC of 56.19 µM against XDR strain, while 1 and 3 were inactive. Docking simulation using enoyl ACP reductase (InhA) revealed favorable protein-ligand interactions. In silico study of pharmacokinetics and hepatotoxicity predicted 1-3 to have good oral bioavailability and 2 to have a lower hepatoxicity probability than isoniazid. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

A desirable and established target for creating antituberculosis drug lead molecules is the Mycobacterium tuberculosis enoyl-acyl carrier protein reductase InhA. Additionally, direct inhibitors of InhA continue to be effective against InhA variants with mutations linked to isoniazid resistance, providing the possibility of activity against MDR isolates. Initially, we isolated monoterpenoid 1,8-cineole from leaf parts of the Callistemon citrinus by aqueous ethanol extract, followed functionalization with several functional groups, and we created a novel 1,8-cineole analogue as a novel compound for treating tuberculosis infection. This study used a structure-based virtual screening of 20 structures to identify novel lead InhA inhibitors as potential antituberculosis drugs. Compound (SG01) was identified as a lead candidate for the synthesis and has been characterized by IR, 1H, 13C NMR, and mass spectroscopy. Using a microplate Alamar Blue assay method, the compound was also tested for their in vitro antimycobacterial activity against the H37Rv strain. This compound offers a desirable starting point for ligand optimization attempts to find new and potent anti-tuberculosis drugs that target InhA. [ABSTRACT FROM AUTHOR]

Drug-resistant (DR) tuberculosis (TB) is a global threat to health security and TB control programs. Since conventional drug susceptibility testing (DST) takes several weeks, we have developed a molecular method for the rapid identification of DR strains of Mycobacterium Tuberculosis (M.tb) utilizing DNA bio-chips. DNA bio-chips were prepared by immobilizing oligonucleotides (probes) on highly microporous polycarbonate track-etched membranes (PC-TEM) as novel support. Bio-chip was designed to contain 15 specific probes to detect mutations in three genes (rpoB, embB, and inhA). A sensitive and specific chemiluminescence based bio-chip assay was developed based on multiplex PCR followed by hybridization on bio-chip. Fifty culture isolates were used to evaluate the ability of in-house developed bio-chip to detect the mutations. Bio-chip analysis shows that 37.7% of samples show wild type sequences, 53.3% of samples were monoresistance showing resistance to either rifampicin (RMP), isoniazid (INH), or ethambutol (EMB). 4.4% of samples were polydrug resistant showing mutations in both the rpoB gene and embB gene while 4.4% of samples were multidrug-resistant (MDR), harboring mutations in the rpoB and inhA genes. The results were compared with DST and sequencing. Compared to sequencing, bio-chip assay shows a sensitivity of 96.5% and specificity of 100% for RMP resistance. For EMB and INH, the results were in complete agreement with sequencing. This study demonstrates the first-time use of PC-TEMs for developing DNA bio-chip for the detection of mutations associated with drug resistance in M.tb. Developed DNA bio-chip accurately detected different mutations present in culture isolates and thus provides detailed and reliable data for clinical diagnosis. [ABSTRACT FROM AUTHOR]

Background: Different anesthetics may have opposite effects on the immune system, thus affecting the prognosis of tumor patients. Cell-mediated immunity forms the primary defense against the invasion of tumor cells, so manipulation of the immune system to produce an enhanced anti-tumor response could be utilized as an adjuvant oncological therapy. Sevoflurane has proinflammatory effects, while propofol, has anti-inflammatory and antioxidant effects. Therefore, we compared the overall survival (OS) and disease-free survival (DFS) of patients with esophageal cancer under total intravenous anesthesia and inhalation anesthesia. Methods: This study collected the electronic medical records of patients undergoing esophagectomy from January 1, 2014 to December 31, 2016. According to the intraoperative anesthetics, the patients were divided into total intravenous anesthesia (TIVA) group or inhalational anesthesia (INHA) group. Stabilized inverse probability of treatment weighting (SIPTW) was used to minimize differences. Kaplan–Meier survival curve was established to evaluate the correlation between different anesthesia methods in overall survival and disease-free survival of patients undergoing esophageal cancer surgery. Results: A total of 420 patients with elective esophageal cancer were collected, including 363 patients eligible for study (TIVA, n = 147, INHA, n = 216). After SIPTW there were no significant differences between two groups in overall survival and disease-free survival. However, the adjuvant therapy was statistically significant in improving OS, and the degree of differentiation was correlated with OS and DFS. Conclusions: In conclusion, there were no significant difference in overall survival and disease-free survival between total intravenous anesthesia and inhalational anesthesia in patients undergoing esophageal cancer surgery. [ABSTRACT FROM AUTHOR]

IntroductionMycobacterium tuberculosis (MTB), the causative agent of tuberculosis, has been a global threat to human beings for several decades. Treating tuberculosis has become more difficult as the prevalence of drug-resistant tuberculosis has increased globally. Evidence suggests that the comprehensive landscape of resistance mechanisms in MTB is ambiguous. More importantly, little is known regarding the series of events connected to resistance mechanisms in MTB before exposure to anti-TB drugs, during exposure to the drugs, and finally, when the MTB becomes resistant after exposure, upon analyses of its genome.MethodsWe used the wild-type strain of MTB (H37Rv) in an in vitro model for generating induced resistance using a sub-inhibitory concentration of isoniazid, and the generated resistance-associated variants (RAVs) were identified using the whole genome sequencing method.ResultsThe detection of an inhA promoter mutation (fabG1−15C>T), which results in increased production of InhA protein, was found to be a major mechanism for developing resistance to isoniazid in the first place. We observed adaptation of MTB resistance mechanisms in high isoniazid stress by alteration and abolishment of KatG due to the detection of katG S315N, the common region of mutation that confers isoniazid resistance, along with katG K414N, katG N138S, and katG A162E. Furthermore, we detected the ahpC−72C>T and ahpC 21C>A mutations, but further investigation is needed to determine their role in compensating for the loss of KatG activity.DiscussionThis suggests that increased InhA production is the main mechanism where there are low levels of isoniazid, whereas the alteration of KatG was found to be utilized in mycobacterium with a high concentration of isoniazid. Our work demonstrates that this in vitro approach of generating induced resistance could provide clinically relevant information after the fabG1−15C>T mutation, which is the common mutation found in clinical isolates. Moreover, other mutations detected in this work can also be found in clinical isolates. These findings may shed light on the impact of isoniazid in generating RAV and the resistance mechanism scenario that mycobacterium used under various isoniazid-pressuring conditions. More research is needed to understand better the role of RAV and mechanical resistance events within the mycobacterium genome in promoting a promising drug prediction platform that could lead to the right treatment for patients with MDR-TB and XDR-TB.

Background: INHA expression has been correlated with the development, growth, and progression of multiple cancer types. However, the biological role of INHA has not been investigated in patients with lung adenocarcinoma (LUAD). Here, we performed a comprehensive bioinformatics analysis of the LUAD dataset to determine the mechanisms underlying the regulation of tumorigenesis by INHA. Materials and methods: INHA expression and clinical information of patients with LUAD were obtained from The Cancer Genome Atlas (TCGA) database. Protein levels in LUAD cell lines and human lung epithelial cells were examined by western blotting. Next, the prognostic value of INHA in LUAD was assessed using Cox regression analysis, while the potential biological functions and the impact on the immune microenvironment of INHA were investigated using gene set enrichment analysis (GSEA) and single sample GSEA (ssGSEA). Finally, the effect of INHA on LUAD cell proliferation and invasion was determined in vitro and in vivo. Results: We found significantly high mRNA and protein expression levels of INHA in LUAD tissues and cell lines. Additionally, a higher expression of INHA was linked to a shorter overall survival (OS) and a worse pathological stage, while INHA expression was associated with immune cell infiltration and immune-related markers in the LUAD tumor microenvironment. LUAD with high INHA expression tends to be a cold tumor. Furthermore, GO and KEGG enrichment analysis indicated that INHA-related genes were enriched in the cell adhesion and immune signaling pathways of LUAD. INHA promoted LUAD cell proliferation and invasion, in vitro and in vivo, by inducing the EGFR pathway. Conclusion: Our findings revealed that INHA is overexpressed in LUAD and is linked to a poor prognosis. Our study demonstrates the potential of INHA as an immunotherapeutic and predictive biomarker in LUAD.

Metabolic syndrome is considered the precursor of type 2 diabetes mellitus. Tuberculosis is a leading infection that constitutes a global threat remaining a major cause of morbi-mortality in developing countries. People with type 2 diabetes mellitus are more likely to suffer from infection with Mycobacterium tuberculosis. For both type 2 diabetes mellitus and tuberculosis, there is pulmonary production of anti-inflammatory glucocorticoids mediated by the enzyme 11b-hydroxysteroid dehydrogenase type 1 (11b-HSD1). The adrenal hormone dehydroepiandrosterone (DHEA) counteracts the glucocorticoid effects of cytokine production due to the inhibition of 11b-HSD1. Late advanced tuberculosis has been associated with the suppression of the Th1 response, evidenced by a high ratio of cortisol/DHEA. In a murine model of metabolic syndrome, we determined whether DHEA treatment modifies the pro-inflammatory cytokines due to the inhibition of the 11b-HSD1 expression. Since macrophages express 11b-HSD1, our second goal was incubating them with DHEA and Mycobacterium tuberculosis to show that the microbicide effect was increased by DHEA. Enoyl-acyl carrier protein reductase (InhA) is an essential enzyme of Mycobacterium tuberculosis involved in the mycolic acid synthesis. Because 11b-HSD1 and InhA are members of a short-chain dehydrogenase/reductase family of enzymes, we hypothesize that DHEA could be an antagonist of InhA. Our results demonstrate that DHEA has a direct microbicide effect against Mycobacterium tuberculosis; this effect was supported by in silico docking analysis and the molecular dynamic simulation studies between DHEA and InhA. Thus, DHEA increases the production of proinflammatory cytokines in the lung, inactivates GC by 11b-HSD1, and inhibits mycobacterial InhA. The multiple functions of DHEA suggest that this hormone or its synthetic analogs could be an efficient co-adjuvant for tuberculosis treatment. [ABSTRACT FROM AUTHOR]

To extend our screening for novel antimycobacterial molecules, we have designed, synthesized, and biologically evaluated a library of 14 new hydrazide derivatives containing 1,3,4-oxadiazole core. A variety of mycobacterial strains, including some drug-resistant strains, were tested for antimycobacterial activity. Among the compounds tested, five showed high antimycobacterial activity (MIC values of 8 μg/mL) against M. tuberculosis H37Ra attenuated strain, and two derivatives were effective (MIC of 4 µg/mL) against pyrazinamide-resistant strains. Furthermore, the novel compounds were tested against the fungal C. albicans strain, showing no antimycotic activity, and thus demonstrating a good selectivity profile. Notably, they also exhibited low cytotoxicity against human SH-SY5Y cells. The molecular modeling carried out suggested a plausible mechanism of action towards the active site of the InhA enzyme, which confirmed our hypothesis. In conclusion, the active compounds were predicted in silico for ADME properties, and all proved to be potentially orally absorbed in humans. [ABSTRACT FROM AUTHOR]

Abstract Background Different types of anesthesia may affect cancer patient’s outcomes, we compared the overall survival (OS) and disease-free survival (DFS) of patients with pancreatic cancer under total intravenous and inhalation anesthesia. Methods The authors collected the electronic medical records of patients who had accepted at a pancreatectomy from January 1, 2010 to December 31, 2016. Patients respectively received total intravenous anesthesia (TIVA) or inhalational anesthesia (INHA). Stabilized inverse probability of treatment weighting (SIPTW)was used to minimize differences. Kaplan–Meier survival was established to analyze the influence of sort of anesthesia on disease-free and overall survival. We compare the effects of each sort of anesthesia on cancer recurrence or metastasis and all-cause mortality. Results A total of 463 patients who had undergone pancreatic cancer resection were collected in this study, of which 421 patients were available (TIVA group, n = 114 INHA group, n = 307). After SIPTW there were no significant differences between the two groups in disease-free survival (hazard ratio, 1.01, 95%CI, 0.78 to 1.29, P = 0.959) or overall survival (hazard ratio, 1.11, 95%CI, 0.87 to 1.42, P = 0.405). Conclusions In conclusion, the present study showed no significant difference in overall survival and disease-free survival between total intravenous anesthesia and volatile anesthesia.

A library of 25-series compounds was designed against Mycobacterium Tuberculosis (M.tb) to identify novel antitubercular drugs. In silico inhibition of InhA , an essential component of FAS-II, was successfully achieved. The drug ability, lead-likeness, and toxicity of the compounds were assessed using Swiss ADME, pkCSM, and Osiris Property Explorer, which revealed the potential for drug development of chalcone compounds. Through in silico research, it was confirmed that toxic-free compounds could bind to InhA. It was found that all of the compounds bind to InhA with binding affinities ranging from −7.78 to −10.29 kcal/mol−1 which is higher than the reference standard Isoniazid and Pyrazinamide. The top five compounds were synthesized from 15 toxic-free compounds. The structural characteristics of the compounds were determined using IR, NMR, and mass spectrometry techniques. These findings indicate that these substances are competitive, reversible, and specific InhA inhibitors of InhA. using the Alamar Blue assay method (H37RV, ATCC No. 27294), the in vitro anti-mycobacterial activity of each of the synthesized compounds against M.tb was evaluated. The two most powerful compounds were (2E)-3-[4-(benzyloxy)-3,5-dimethylphenyl] and (2E)-1-(3,5-dibromophenyl)-3-(3-phenoxyphenyl) prop-2-en-1-one. In the MABA Assay, the MIC for 1-(3,5-dibromophenyl) prop-2-en-1-one was 6.25 μg/ml. • A library of 25 chalcone compounds was designed for Mycobacterium tuberculosis (M.tb) inhibition, targeting the InhA enzyme. • In silico analysis revealed high binding affinities (−7.78 to −10.29 kcal/mol) for all compounds, outperforming Isoniazid and Pyrazinamide. • ADMET analysis indicated promising drug-likeness and safety profiles for 15 toxic-free compounds. • The top five compounds were synthesized and characterized using IR, NMR, and mass spectrometry. • In vitro testing showed significant anti-mycobacterial activity, with two leading compounds having MIC values of 6.25 μg/ml. [ABSTRACT FROM AUTHOR]

[Display omitted] • New coumarin-thiazole hybrids were synthesized as anti-TB agents. • The target compounds showed potent activity against Mtb H37Rv and Δ katG strains. • Compounds 2b and 3j inhibited mycobacterial growth inside human macrophages. • Compounds 2b and 3j showed potent inhibitory effect against InhA. • Compounds 2b and 3j showed good docking scores against InhA. Tuberculosis is a global serious problem that imposes major health, economic and social challenges worldwide. The search for new antitubercular drugs is extremely important which could be achieved via inhibition of different druggable targets. Mycobacterium tuberculosis enoyl acyl carrier protein reductase (InhA) enzyme is essential for the survival of M. tuberculosis. In this investigation, a series of coumarin based thiazole derivatives was synthesized relying on a molecular hybridization approach and was assessed against the wild type Mtb H37Rv and its mutant strain (Δ katG) via inhibiting InhA enzyme. Among the synthesized derivatives, compounds 2b , 3i and 3j were the most potent against wild type M. tuberculosis with MIC values ranging from 6 to 8 μg/ mL and displayed low cytotoxicity towards mouse fibroblasts at concentrations 8–13 times higher than the MIC values. The three hybrids could also inhibit the growth of Δ katG mutant strain which is resistant to isoniazid (INH). Compounds 2b and 3j were able to inhibit the growth of mycobacteria inside human macrophages, indicating their ability to penetrate human professional phagocytes. The two derivatives significantly suppress mycobacterial biofilm formation by 10–15 %. The promising target compounds were also assessed for their inhibitory effect against InhA and showed potent effectiveness with IC 50 values of 0.737 and 1.494 µM, respectively. Molecular docking studies revealed that the tested compounds occupied the active site of InhA in contact with the NAD+ molecule. The 4-phenylcoumarin aromatic system showed binding interactions within the hydrophobic pocket of the active site. Furthermore, H-bond formation and π −π stacking interactions were also recorded for the promising derivatives. [ABSTRACT FROM AUTHOR]

Metabolic syndrome is considered the precursor of type 2 diabetes mellitus. Tuberculosis is a leading infection that constitutes a global threat remaining a major cause of morbi-mortality in developing countries. People with type 2 diabetes mellitus are more likely to suffer from infection with Mycobacterium tuberculosis. For both type 2 diabetes mellitus and tuberculosis, there is pulmonary production of anti-inflammatory glucocorticoids mediated by the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). The adrenal hormone dehydroepiandrosterone (DHEA) counteracts the glucocorticoid effects of cytokine production due to the inhibition of 11β-HSD1. Late advanced tuberculosis has been associated with the suppression of the Th1 response, evidenced by a high ratio of cortisol/DHEA. In a murine model of metabolic syndrome, we determined whether DHEA treatment modifies the pro-inflammatory cytokines due to the inhibition of the 11β-HSD1 expression. Since macrophages express 11β-HSD1, our second goal was incubating them with DHEA and Mycobacterium tuberculosis to show that the microbicide effect was increased by DHEA. Enoyl-acyl carrier protein reductase (InhA) is an essential enzyme of Mycobacterium tuberculosis involved in the mycolic acid synthesis. Because 11β-HSD1 and InhA are members of a short-chain dehydrogenase/reductase family of enzymes, we hypothesize that DHEA could be an antagonist of InhA. Our results demonstrate that DHEA has a direct microbicide effect against Mycobacterium tuberculosis; this effect was supported by in silico docking analysis and the molecular dynamic simulation studies between DHEA and InhA. Thus, DHEA increases the production of pro-inflammatory cytokines in the lung, inactivates GC by 11β-HSD1, and inhibits mycobacterial InhA. The multiple functions of DHEA suggest that this hormone or its synthetic analogs could be an efficient co-adjuvant for tuberculosis treatment.

This paper discusses the scientific lives of Julien, Louis and Claude Poinssot (grandfather, father and son) and their important contribution to the study of ancient religions in North Africa, especially in Tunisia, including their intense engagement with the ancient Thugga. Julien, who had been trained as a notary, soon became a pioneering epigraphist of Proconsularis. As a result of the colonial context in which he worked and the bureaucratic problems he encountered, his archaeological career was brief, albeit intensive. Nevertheless, his vocation was inherited by his descendants, Louis and Claude, who eventually became Directors of Antiquities and of Museums. Their work left a remarkable legacy, including the Mahdia underwater excavations and the exploration of the Dougga Capitol.

Abstract Background With the advancement of next generation sequencing technologies (NGS), whole-genome sequencing (WGS) has been deployed to a wide range of clinical scenarios. Rapid and accurate classification of drug-resistant Mycobacterium tuberculosis (MTB) would be advantageous in reducing the amplification of additional drug resistance and disease transmission. Methods In this study, a long-read sequencing approach was subjected to the whole-genome sequencing of clinical MTB clones with susceptibility test profiles, including isoniazid (INH) susceptible clones (n = 10) and INH resistant clones (n = 42) isolated from clinical specimens. Non-synonymous variants within the katG or inhA gene associated with INH resistance was identified using Nanopore sequencing coupled with a corresponding analytical workflow. Results In total, 54 nucleotide variants within the katG gene and 39 variants within the inhA gene associated with INH resistance were identified. Consistency among the results of genotypic profiles, susceptibility test, and minimal inhibitory concentration, the high-INH resistance signature was estimated using the area under the receiver operating characteristic curve with the existence of Ser315Thr (AUC = 0.822) or Thr579Asn (AUC = 0.875). Conclusions Taken together, we curated lists of coding variants associated with differential INH resistance using Nanopore sequencing, which may constitute an emerging platform for rapid and accurate identification of drug-resistant MTB clones.

Isoniazid (INH) is one of the key molecules employed in the treatment of tuberculosis (TB), the most deadly infectious disease worldwide. However, the efficacy of this cornerstone drug has seriously decreased due to emerging INH-resistant strains of Mycobacterium tuberculosis (Mtb). In the present study, we aimed to chemically tailor INH to overcome this resistance. We obtained thirteen novel compounds by linking INH to in-house synthesized sulfonate esters via a hydrazone bridge (SIH1–SIH13). Following structural characterization by FTIR, 1H NMR, 13C NMR, and HRMS, all compounds were screened for their antitubercular activity against Mtb H37Rv strain and INH-resistant clinical isolates carrying katG and inhA mutations. Additionally, the cytotoxic effects of SIH1–SIH13 were assessed on three different healthy host cell lines; HEK293, IMR-90, and BEAS-2B. Based on the obtained data, the synthesized compounds appeared as attractive antimycobacterial drug candidates with low cytotoxicity. Moreover, the stability of the hydrazone moiety in the chemical structure of the final compounds was confirmed by using UV/Vis spectroscopy in both aqueous medium and DMSO. Subsequently, the compounds were tested for their inhibitory activities against enoyl acyl carrier protein reductase (InhA), the primary target enzyme of INH. Although most of the synthesized compounds are hosted by the InhA binding pocket, SIH1–SIH13 do not primarily show their antitubercular activities by direct InhA inhibition. Finally, in silico determination of important physicochemical parameters of the molecules showed that SIH1–SIH13 adhered to Lipinski's rule of five. Overall, our study revealed a new strategy for modifying INH to cope with the emerging drug-resistant strains of Mtb. [ABSTRACT FROM AUTHOR]

Inhibin is a molecule that belongs to peptide hormones and is excreted through pituitary gonadotropins stimulation action on the granulosa cells of the ovaries. However, the differential regulation of inhibin and follicle-stimulating hormone (FSH) on granulosa cell tumor growth in mice inhibin-deficient females is not yet well understood. The objective of this study was to evaluate the role of inhibin and FSH on the granulosa cells of ovarian follicles at the premature antral stage. This study stimulated immature wild-type (WT) and Inhibin-α knockout (Inha−/−) female mice with human chorionic gonadotropin (hCG) and examined hCG-induced gene expression changes in granulosa cells. Also, screening of differentially expressed genes (DEGs) was performed in the two groups under study. In addition, related modules to external traits and key gene drivers were determined through Weighted Gene Co-Expression Network Analysis (WGCNA) algorithm. The results identified a number of 1074 and 931 DEGs and 343 overlapping DEGs (ODEGs) were shared in the two groups. Some 341 ODEGs had high relevance and consistent expression direction, with a significant correlation coefficient (r2 = 0.9145). Additionally, the gene co-expression network of selected 153 genes showed 122 nodes enriched to 21 GO biological processes (BP) and reproduction and 3 genes related to genomic pathways. By using principal component analysis (PCA), the 14 genes in the regulatory network were fixed and the cumulative proportion of fitted top three principal components was 94.64%. In conclusion, this study revealed the novelty of using ODEGs for investigating the inhibin and FSH hormone pathways that might open the way toward gene therapy for granulosa cell tumors. Also, these genes could be used as biomarkers for tracking the changes in inhibin and FSH hormone from the changes in the nutrition pattern. [ABSTRACT FROM AUTHOR]

The spread of drug-resistant strains of Mycobacterium tuberculosis is a major threat to the global control of tuberculosis (TB), urging the need to constantly develop new anti-TB drugs. Tryptanthrins are convenient compounds for the development of candidate anti-TB drugs due to the easy synthesis, low toxicity, and antimycobacterial activity on both drug-susceptible and drug-resistant M. tuberculosis strains. Enoyl-acyl carrier protein reductase InhA was previously predicted in silico as a possible target for tryptanthrins, while spontaneous tryptanthrin-resistant M. smegmatis mutants were found to have mutations in the MSMEG_1963, MSMEG_4427, and MSMEG_5597 genes. Using the approaches of reverse genetics, we demonstrate that mutations in the MSMEG_1963 and MSMEG_5597 genes lead to a loss of function of their encoded transcriptional repressors and lead to resistance to tryptanthrins. We show that mutations in MSMEG_1963 and MSMEG_5597 lead to overexpression of MSMEG_1964 and MSMEG_5596, respectively, which encode enzymes potentially involved in redox inactivation of tryptanthrins. We also show that InhA is not a biotarget of tryptanthrins, as its overexpression does not affect susceptibility of mycobacteria to tryptanthrins. [ABSTRACT FROM AUTHOR]

A series of Triclosan-based hybrids and their Schiff base derivatives with isoniazid were designed through in silico modelling and synthesized using copper-catalysed azide-alkyne cycloaddition (CuAAC) reaction. These compounds were then evaluated against both Mycobacterium tuberculosis (Mtb) and Mycobacterium abscessus (Mab). However, none of the synthesized hybrids exhibited significant growth inhibition, with minimum inhibitory concentration (MIC) values consistently exceeding 100 µg/mL. To further investigate these findings, we conducted mechanistic studies including thermogravimetric analysis (TGA) and UV-Vis stability tests. TGA demonstrated thermal stability of the Schiff bases up to 270 °C, while UV-Vis analysis confirmed their chemical resilience across a wide pH spectrum, showing resistance to hydrolysis under acidic and basic conditions. The lack of observed antimicrobial activity may be attributed to the high lipophilicity of these molecules, as indicated by LogP values exceeding 5, which could limit their bioavailability. These findings suggest that although combining triclosan and isoniazid holds potential, further optimization of this hybrid strategy is necessary to improve efficacy., (© 2024 Wiley‐VCH GmbH.)

A set of alkyl-/1H-1,2,3-triazole-based dimers was strategically designed and synthesized to evaluate their in vitro anti-mycobacterial activities against Mycobacterium tuberculosis and the non-tuberculous Mycobacterium abscessus strains. Systematic variations in the nature (alkyl/1H-1,2,3-triazole) and positioning of the linker were implemented based on the docking scores observed in the binding sites identified in the crystal structures of InhA from M. tuberculosis and M. abscessus . However, the in vitro evaluation results revealed that the synthesized compounds did not exhibit inhibitory effects on the growth of mycobacteria, even at the highest tested concentrations. The elevated lipophilicity values determined through ADMET studies for these synthesized dimers might be a contributing factor to their poor activity profiles., Competing Interests: We declare we have no competing interests., (© 2024 The Author(s).)

Objective To analyze the characteristics of katG and inhA genetic mutations associated with INH-resistant Mycobacterium tuberculosis in Yunnan Province. Methods The INH-resistant Mycobacterium tuberculosis isolates were collected from drug-resistant monitoring sites in Yunnan Province, and the katG and inhA genes were amplified by PCR and then sequenced. Results Among the 88 MTB strains, the coincidence rate of INH-resistant genotype and phenotype was 82.95% （73/88）. 75.00% （66/88） had katG mutations and 9.09% （8/88） had inhA mutations. katG315 （67.05%） and inhA-15 （7.95%） were the top two mutation sites, and a high prevalence of katG Ser315Thr mutations （59.09%, 52/88） was observed. The frequency of the katG mutation was significantly higher in MDR-TB compared to INH mono-resistant isolates （χ² = 4.190, P = 0.041）, while katG315, inhA and inhA-15T mutations did not differ significantly between these two groups （P > 0.05）. Conclusion The most common mutation of INH-resistant Mycobacterium tuberculosis in Yunnan Province were katG315 and inhA-15, and katG mutation was higher among MDR isolates compared with INH mono-resistant strains. Molecular diagnostics based on gene mutations was a useful method for rapid detection of INH-resistant Mycobacterium tuberculosis in Yunnan Province. [ABSTRACT FROM AUTHOR]

Mycobacterium tuberculosis (M.tb.) enoyl-acyl carrier protein (ACP) reductase (InhA) is validated as a useful target for tuberculosis therapy and is considered an attractive enzyme to drug discovery. This study aimed to identify the novel inhibitor of the InhA enzyme, a potential target of M.tb. involved in the type II fatty acid biosynthesis pathway that controls mycobacterial cell envelope synthesis. We compiled 80 active compounds from Ruta graveolens and citrus plants belonging to the Rutaceae family for pharmacokinetics and molecular docking analyses. The chemical structures of the 80 phytochemicals and the 3D structure of the target protein were retrieved from the PubChem database and RCSB Protein Data Bank, respectively. The evaluation of druglikeness was performed based on Lipinski's Rule of Five, while the computed phytochemical properties and molecular descriptors were used to predict the ADMET of the compounds. Amongst these, 11 pharmacokinetically-screened compounds were further examined by performing molecular docking analysis with an InhA target using AutoDock 4.2. The docking results showed that gravacridonediol, a major glycosylated natural alkaloid from Ruta graveolens, might possess a promising inhibitory potential against InhA, with a binding energy (B.E.) of −10.80 kcal/mole and inhibition constant (Ki) of 600.24 nM. These contrast those of the known inhibitor triclosan, which has a B.E. of −6.69 kcal/mole and Ki of 12.43 µM. The binding efficiency of gravacridonediol was higher than that of the well-known inhibitor triclosan against the InhA target. The present study shows that the identified natural compound gravacridonediol possesses drug-like properties and also holds promise in inhibiting InhA, a key target enzyme of M.tb. [ABSTRACT FROM AUTHOR]

New medications are desperately needed to combat rising drug resistance among tuberculosis (TB) patients. New agents should ideally work through unique targets to avoid being hampered by preexisting clinical resistance to existing treatments. The enoyl-acyl carrier protein reductase InhA of M. tuberculosis is one of the most crucial targets since it is a promising target that has undergone extensive research for anti-tuberculosis drug development. A well-known scaffold for a variety of biological activities, including antitubercular activity, is the molecular linkage of a1,2,3-triazole with an acetamide group. As a result, in the current study, which was aided by ligand-based molecular modeling investigations, 1,2,3-triazolesweredesigned and synthesized adopting the CuAAC aided cycloaddition of 1-(4-(prop-2-yn-1-yloxy)phenyl)ethanone with appropriate acetamide azides. Standard spectroscopic methods were used to characterize the newly synthesized compounds. In vitro testing of the proposed compounds against the InhA enzyme was performed. All the synthesized inhibitors completely inhibited the InhA enzyme at a concentration of 10 µM that exceeded Rifampicin in terms of activity. Compounds 9, 10, and 14 were the most promising InhA inhibitors, with IC50 values of 0.005, 0.008, and 0.002 µM, respectively. To promote antitubercular action and investigate the binding manner of the screened compounds with the target InhA enzyme's binding site, a molecular docking study was conducted. [ABSTRACT FROM AUTHOR]

The inhibin a (INHA), inhibin ßA (INHBA) and bone morphogenetic protein 15 (BMP-15) were investigated as candidate genes for reproductive traits in four Iranian sheep breeds (Bahmaei, Lak Ghashghaei, Lori-Bakhtiari and Karakul). Based on the ovine sequences of BMP-15, INHA and INHBA genes, three pairs of primers were designed to identify single nucleotide polymorphisms (SNPs) in exon 2 of BMP-15, INHA and INHBA in multiparous ewes by DNA sequencing. Two SNPs were detected in exon 2 of the ovine BMP15 gene at positions 367 and 430, which lead to amino acid substitutions at position 231 and 252 in the BMP15 protein sequence, respectively. Substitution of Leucine to Proline at position 252 is predicted to affect the protein function. A synonymous mutation was found in the amplified fragment of exon 2 at position 752 in ovine INHBA gene. In addition, the c752C>T mutation was only found in heterozygous condition in only one Lori-Bakhtiari ewe, while other breeds were in wild type genotype for c752C>T mutation. The INHA gene was shown to be highly polymorphic. A total of 7 SNPs including 6 nucleotide substitutions and one insertion were found in the amplified fragment of the INHA locus. The insertion mutation was found in two animals of Bahmaei and Karakul breeds. Interestingly, homozygous condition for the mutant alleles in all identified SNPs in BMP15, INHA and INHBA loci was absent in these breeds. Generally, these breeds showed different genetic structures with regard to the identified SNPs in BMP15, INHBA and INHA genes. However, further research with larger sample size and phenotype data on reproductive performance is required to investigate the definitive effect of the identified mutations in this study. [ABSTRACT FROM AUTHOR]