Additive Manufacturing of SmartEx QD 100 Designed Oral Three-Dimensional Printlets Containing Isoniazid for Immediate Gastric Release by Selective Laser Sintering.

The selection of appropriate materials and compatibility of selected materials with drugs and formulations are limiting steps in three-dimensional printing technology. In this study, SmartEx QD 100 (SM QD 100) was introduced as a novel, coprocessed, unexplored excipient that can be used in SLS-mediated 3D printing. The current study aimed to evaluate the feasibility of fabricating SM QD 100 containing INH-embedded SLS-mediated immediate gastric release tablets. The prepared physical mixtures were subjected to the fabrication of 3D printlets by using SLS-mediated 3D printing. The fabricated 3D printlets were subjected to physicochemical characterization by using various analytical techniques. After oral administration of sintered 3D printlets to rabbits, samples were collected and pharmacokinetic parameters were analyzed using the developed LC-APCI-MS/MS method. The optimized batch was able to release 100% INH within 15 min, which confirmed the immediate gastric release. Similarly, sintered 3D printlets were stable under accelerated stability conditions for three months. Finally, the pharmacokinetic parameters revealed the rate and extent of absorption of INH from sintered 3D printlets. As evidenced by in vitro and in vivo analyses, SM QD 100 was able to sinter SLS-mediated INH-embedded stable immediate gastric release tablets. SM QD 100 is a novel material for SLS-mediated 3D printing in pharmaceutical applications.

Natural deep eutectic solvent for the simultaneous derivatization and microextraction of isoniazid from human plasma.

BACKGROUND: Personalized medicine is a rapidly revolving field that offers new opportunities for tailoring disease treatment to individual patients. The main idea behind this approach is to carefully select safe and effective medications and treatment plant based on each patient's unique pharmacokinetic profile. Isoniazid is a first-line anti-tuberculosis drug that has interindividual variability in its metabolic processing, leading to significant differences in plasma concentrations among patients receiving equivalent doses. This variability necessitates the creation of individualized treatment regimens as part of personalized medicine to achieve more effective therapy. RESULTS: In this work, a deep eutectic solvent-based liquid-liquid microextraction approach for the separation and determination of isoniazid in human plasma by high-performance liquid chromatography with UV-Vis detection was developed for the first time. A new natural deep eutectic solvent based on thymol as a hydrogen bond donor and 4-methoxybenzaldehyde as a hydrogen bond acceptor was proposed as the extraction solvent. The developed microextraction procedure assumed two simultaneous processes during the mixing of the sample and extraction solvent: the derivatization of the polar analyte in the presence of 4-methoxybenzaldehyde (component of the natural deep eutectic solvent) with the formation of a hydrophobic Schiff base (1); mass transfer of the Schiff base from the sample phase to the extraction solvent phase (2). Under optimal conditions, the limits of detection and quantification were 20 and 60 µg L-1, respectively. The RSD value was <10 %, the extraction recovery was 95 %. SIGNIFICANCE: In this work, the possibility of isoniazid derivatization in the natural deep eutectic solvent phase with the formation of the Schiff base was presented for the first time. The approach provided the separation and preconcentration of polar isoniazid without the use of expensive derivatization agents and solid-phase extraction cartridges. The formation of the Schiff base was confirmed by mass spectrometry.

Permeability of TB drugs through the mycolic acid monolayer: a tale of two force fields.

Tuberculosis (TB) treatment becomes challenging due to the unique cell wall structure of Mycobacterium tuberculosis (M. tb). Among various components of the M.tb cell wall, mycolic acid (MA) is of particular interest because it is speculated to exhibit extremely low permeability for most of the drug molecules, thus helping M.tb to survive against medical treatment. However, no quantitative assessment of the thermodynamic barrier encountered by various well-known TB drugs in the mycolic acid monolayer has been performed so far using computational tools. On this premise, our present work aims to probe the permeability of some first and second line TB drugs, namely ethambutol, ethionamide, and isoniazid, through the modelled mycolic acid monolayer, using molecular dynamics (MD) simulation with two sets of force field (FF) parameters, namely GROMOS 54A7-ATB (GROMOS) and CHARMM36 (CHARMM) FFs. Our findings indicate that both FFs provide consistent results in terms of the mode of drug-monolayer interactions but significantly differ in the drug permeability through the monolayer. The mycolic acid monolayer generally exhibited a higher free energy barrier of crossing with CHARMM FF, while with GROMOS FF, better stability of drug molecules on the monolayer surface was observed, which can be attributed to the greater electrostatic potential at the monolayer-water interface, found for the later. Although both the FF parameters predicted the highest resistance against ethambutol (permeability values of 8.40 × 10-34 cm s-1 and 9.61 × 10-31 cm s-1 for the CHARMM FF and the GROMOS FF, respectively), results obtained using GROMOS were found to be consistent with the water solubility of drugs, suggesting it to be a slightly better FF for modelling drug-mycolic acid interactions. Therefore, this study enhances our understanding of TB drug permeability and highlights the potential of the GROMOS FF in simulating drug-mycolic acid interactions.

Design and Synthesis of Isatin-Tagged Isoniazid Conjugates with Cogent Antituberculosis and Radical Quenching Competence: In-vitro and In-silico Evaluations.

In pursuit of potential chemotherapeutic alternates to combat severe tuberculosis infections, novel heterocyclic templates derived from clinically approved anti-TB drug isoniazid and isatin have been synthesized that demonstrate potent inhibitory action against Mycobacterium tuberculosis, and compound 4i with nitrophenyl motif exhibited the highest anti-TB efficacy with a MIC value of 2.54 µM/ml. Notably, the same nitro analog 4i shows the best antioxidant efficacy among all the synthesized compounds with an IC50 value of 37.37 µg/ml, suggesting a synergistic influence of antioxidant proficiency on the anti-TB action. The titled compounds exhibit explicit binding affinity with the InhA receptor. The befitting biochemical reactivity and near-appropriate pharmacokinetic proficiency of the isoniazid conjugates is reflected in the density functional theory (DFT) studies and ADMET screening. The remarkable anti-TB action of the isoniazid cognates with marked radical quenching ability may serve as a base for developing multi-target medications to confront drug-resistant TB pathogens.

Synthesis of indole-functionalized isoniazid conjugates with potent antimycobacterial and antioxidant efficacy.

Aim: Developing potent medicinal alternates for tuberculosis (TB) is highly desirable due to the advent of drug-resistant lethal TB strains.Methods & results: Novel indole-isoniazid integrates have been synthesized with promising antimycobacterial action against the H37Rv strain, and the nitro analogs 4e and 4j show the highest efficacy with a minimum inhibitory concentration of 1.25 µg/ml. The molecular docking studies against InhA support the experimental findings. Indole conjugates display remarkable radical quenching efficiency, and compounds 4e and 4j demonstrate maximum IC50 values of 50.19 and 52.45 µg/ml, respectively. Pharmacokinetic analysis anticipated appreciable druggability for the title compounds.Conclusion: The notable bioaction of the indole-isoniazid templates projects them as potential lead in developing anti-TB medications with synergetic antioxidant action.

[Box: see text].

Synthesis and characterisation of antimicrobial metal-organic frameworks as multi-drug carriers.

Antibiotic resistance is a significant global concern, necessitating the development of either new antibiotics or advanced delivery methods. With this in mind, we report on the synthesis and characterisation of a new family of Metal-Organic Frameworks (MOFs), OnG6 MOFs, designed to act as multi-drug carriers for bacterial infection treatment. OnG6 is based on the pro-drug 4,4'-azodisalicylic acid (AZDH4), which in vivo produces two equivalents of para-aminosalicylic acid (ASA), a crucial drug for M. tuberculosis treatment. X-ray and computational studies revealed that OnG6 MOFs are mesoporous MOFs with etb topology and an [M2(AZD)] formula (M = Zn, OnG6-Zn; Mg, OnG6-Mg; Cu, OnG6-Cu; and Co, OnG6-Co), featuring 1-dimensional channel type pores of 25 Å diameter. OnG6 MOFs are the first reported MOFs bearing the ligand AZDH4, joining the family of mesoporous MOFs arranged in a honeycomb pattern. They absorb isoniazid (INH) and ciprofloxacin (CIPRO) with the former being a specific antibiotic for M. tuberculosis, and the latter being a broader-spectrum antibiotic. The stability of the MOFs and their capacity for antibiotic uptake depend on the nature of the metal ion, with OnG6-Mg demonstrating the highest drug absorption. The antimicrobial activity of these species was assessed against S. aureus and E. coli, revealing that the carriers containing CIPRO displayed optimal efficacy.

Multifunctional Fe/Cu Dual-Single Atom Nanozymes with Enhanced Peroxidase Activity for Isoniazid Detection and Levofloxacin Degradation.

The design of single-atom nanozymes with dual active sites to increase their activity and for the detection and degradation of contaminants is rare and challenging. In this work, a single-atom nanozyme (FeCu-NC) based on a three-dimensional porous Fe/Cu dual active site was developed as a colorimetric sensor for both the quantitative analysis of isoniazid (INH) and the efficient degradation of levofloxacin (LEV). FeCu-NC was synthesized using a salt template and freeze-drying method with a three-dimensional hollow porous structure and dual active sites (Fe-Nx and Cu-Nx). In terms of morphology and structure, FeCu-NC exhibits excellent peroxidase-like activity and catalytic properties. Therefore, a colorimetric sensor was constructed around FeCu-NC for sensitive and rapid quantitative analysis of INH with a linear range of 0.9-10 µM and a detection limit as low as 0.3 µM, and the sensor was successfully applied to the analysis of INH in human urine. In addition, FeCu-NC promoted the efficient degradation of LEV by peroxymonosulfate activation, with a degradation rate of 90.4% for LEV at 30 min. This work sheds new light on the application of single-atom nanozymes to antibiotics for colorimetric sensing and degradation.

The use of green protic ionic liquids in the crystallization of isoniazid: Evaluation of physicochemical and biological properties of drug.

This study evaluated the synthesis of protic ionic liquids (PILs), 2-hydroxy ethylammonium formate (2-HEAF) and 2-hydroxy ethylammonium acetate (2-HEAA), and their applicability in the crystallization process of the active pharmaceutical ingredient isoniazid (INH) as anti-solvent. Isoniazid is an antibiotic used in the treatment of tuberculosis infections, being used as a first-line chemotherapeutic agent against Mycobacterium tuberculosis. Futhermore, this investigation was conducted in order to evaluate how these PILs can influence the habit, solubility, stability, and therapeutic efficiency of the obtained isoniazid crystals. The 2-HEAF and 2-HEAA PILs were easily formed in reactions between ethanolamine and carboxylic acids (formic or acetic acid), and they have no toxicity against Artemia salina. The PILs were able to crystallize isoniazid, influencing the crystal habit and size. The greatest variations in the hydrogen signals of the NH2 and NH groups of the amine and low variations in the chemical shifts of the hydrogens of the cation of the ethanolamine group from 2-HEAA and 2-HEAF indicate that PILs establish possibly weak interactions with INH. The obtained crystals were amorphous and showed higher solubility in water than standard INH. Moreover, these crystals showed therapeutic efficiency inantimycobacterial activity to inhibit the growth of Mycobacterium tuberculosis. The INH:2-HEAF only degraded 5.1 % (w/w), however, INH:2-HEAA degraded 32.8 % (w/w) after 60 days in an accelerated atmosphere. Then, the 2-HEAA and 2-HEAF were able to crystallize isoniazid, being a new application for these PILs. The used PILs also influenced the characteristics of isoniazid crystals.

Novel isoniazid-hydrazone derivatives induce cell growth inhibition, cell cycle arrest and apoptosis via mitochondria-dependent caspase activation and PI3K/AKT inhibition.

In this study, seven isoniazid-hydrazone derivatives (3a-g) were synthesized and their structures elucidated by chromatographic techniques, and then the antiproliferative effects of these compounds on various cancer cells were tested. The advanced anticancer mechanism of the most potent compound was then investigated. Antiproliferative activities of the synthesized compounds were evaluated on human breast cancer MCF-7, lung cancer A-549, colon cancer HT-29, and non-cancerous mouse fibroblast 3T3-L1 cell lines by XTT assay. Flow cytometry analysis were carried out to determine cell cycle distribution, apoptosis, mitochondrial membrane potential, multi-caspase activity, and expression of PI3K/AKT signaling pathway. The XTT results showed that all the title molecules displayed cytotoxic activity at varying strengths in different dose ranges, and among them, the strongest cytotoxic effect and high selectivity were exerted by 3d against MCF-7 cells with the IC50 value of 11.35 µM and selectivity index of 8.65. Flow cytometry results revealed that compound 3d induced apoptosis through mitochondrial membrane disruption and multi-caspase activation in MCF-7 cells. It also inhibited the cell proliferation via inhibition of expression of PI3K/AKT and arrested the cell cycle at G0/G1 phase. In conclusion, all these data disclosed that among the synthesized compounds, 3d is notable for in vivo anticancer studies.

Peroxynitrite imaging in ferroptosis-mediated drug-induced liver injury with a near-infrared fluorescence probe.

BACKGROUND: Over-consumption of drugs can result in drug-induced liver damage (DILI), which can worsen liver failure. Numerous studies have shown the significant role ferroptosis plays in the pathophysiology of DILI, which is typified by a marked imbalance between the generation and breakdown of lipid reactive oxygen species (ROS). The content of peroxynitrite (ONOO-) rapidly increased during this process and was thought to be a significant marker of early liver injury. Therefore, the construction of fluorescence probe for the detection and imaging of ONOO- holds immense importance in the early diagnosis and treatment of ferroptosis-mediated DILI. RESULTS: We designed a probe DILI-ONOO based on the ICT mechanism for the purpose of measuring and visualizing ONOO- in ferroptosis-mediated DILI processes and associated studies. This probe exhibited significant fluorescence changes with good sensitivity, selectivity, and can image exogenous and endogenous ONOO- in cells with low cytotoxicity. Using this probe, we were able to show changes in ONOO- content in ferroptosis-mediated DILI cells and mice models induced by the intervention of acetaminophen (APAP) and isoniazid (INH). By measuring the concentration of ferroptosis-related indicators in mice liver tissue, we were able to validate the role of ferroptosis in DILI. It is worth mentioning that compared to existing alanine transaminase (ALT) and aspartate aminotransferase (AST) detection methods, this probe can achieve early identification of DILI prior to serious liver injury. SIGNIFICANCE: This work has significant reference value in researching the relationship between ferroptosis and DILI and visualizing research. The results indicate a strong correlation between the progression of DILI and ferroptosis. Additionally, the use of DILI-ONOO shows promise in investigating the DILI process and assessing the effectiveness of medications in treating DILI.

Experimental and Computational Studies on the Interaction of DNA with Hesperetin Schiff Base CuII Complexes.

The interactions with calf thymus DNA (CT-DNA) of three Schiff bases formed by the condensation of hesperetin with benzohydrazide (HHSB or L1H3), isoniazid (HIN or L2H3), or thiosemicarbazide (HTSC or L3H3) and their CuII complexes (CuHHSB, CuHIN, and CuHTSC with the general formula [CuLnH2(AcO)]) were evaluated in aqueous solution both experimentally and theoretically. UV-Vis studies indicate that the ligands and complexes exhibit hypochromism, which suggests helical ordering in the DNA helix. The intrinsic binding constants (Kb) of the Cu compounds with CT-DNA, in the range (2.3-9.2) × 106, from CuHTSC to CuHHSB, were higher than other copper-based potential drugs, suggesting that π-π stacking interaction due to the presence of the aromatic rings favors the binding. Thiazole orange (TO) assays confirmed that ligands and Cu complexes displace TO from the DNA binding site, quenching the fluorescence emission. DFT calculations allow for an assessment of the equilibrium between [Cu(LnH2)(AcO)] and [Cu(LnH2)(H2O)]+, the tautomer that binds CuII, amido (am) and not imido (im), and the coordination mode of HTSC (O-, N, S), instead of (O-, N, NH2). The docking studies indicate that the intercalative is preferred over the minor groove binding to CT-DNA with the order [Cu(L1H2am)(AcO)] > [Cu(L2H2am)(AcO)] ≈ TO ≈ L1H3 > [Cu(L3H2am)(AcO)], in line with the experimental Kb constants, obtained from the UV-Vis spectroscopy. Moreover, dockings predict that the binding strength of [Cu(L1H2am)(AcO)] is larger than [Cu(L1H2am)(H2O)]+. Overall, the results suggest that when different enantiomers, tautomers, and donor sets are possible for a metal complex, a computational approach should be recommended to predict the type and strength of binding to DNA and, in general, to macromolecules.

Injectable isoniazid-loaded bone cement based on hydrazone bonds achieving long-term release and decent mechanical properties.

A robust and easily manufactured high-strength and long-term release hydrazone-based isoniazid acrylic (HIA) bone cement is reported. The mechanical strength of HIA bone cement is similar to that of normal polymethyl methacrylate (PMMA) bone cement, far surpassing that of traditional isoniazid-containing antibiotic-loaded bone cement (INH bone cement). Isoniazid is connected to the bone cement through bioorthogonal hydrazone chemistry, and it possesses release properties superior to those of INH bone cement, allowing for the sustained release of isoniazid for up to 12 weeks. In vivo and in vitro studies also indicate that HIA cement exhibits better biocompatibility than INH bone cement. The results of this study not only signify progress in the realm of antimicrobial bone cement for addressing bone tuberculosis but also enhance our capacity to create and comprehend high-performing antimicrobial bone cement.

AI-driven design of customized 3D-printed multi-layer capsules with controlled drug release profiles for personalized medicine.

Personalized medicine aims to effectively and efficiently provide customized drugs that cater to diverse populations, which is a significant yet challenging task. Recently, the integration of artificial intelligence (AI) and three-dimensional (3D) printing technology has transformed the medical field, and was expected to facilitate the efficient design and development of customized drugs through the synergy of their respective advantages. In this study, we present an innovative method that combines AI and 3D printing technology to design and fabricate customized capsules. Initially, we discretized and encoded the geometry of the capsule, simulated the dissolution process of the capsule with classical drug dissolution model, and verified it by experiments. Subsequently, we employed a genetic algorithm to explore the capsule geometric structure space and generate a complex multi-layer structure that satisfies the target drug release profiles, including stepwise release and zero-order release. Finally, Two model drugs, isoniazid and acetaminophen, were selected and fused deposition modeling (FDM) 3D printing technology was utilized to precisely print the AI-designed capsule. The reliability of the method was verified by comparing the in vitro release curve of the printed capsules with the target curve, and the f2 value was more than 50. Notably, accurate and autonomous design of the drug release curve was achieved mainly by changing the geometry of the capsule. This approach is expected to be applied to different drug needs and facilitate the development of customized oral dosage forms.

Unravelling performance of honeycomb structures as drug delivery systems for the isoniazid drug using DFT-D3 correction dispersion and molecular dynamic simulations.

In this study, the potential of aluminum nitride (h-AlN), boron nitride (h-BN) and silicon carbide (h-SiC) nanosheets as the drug delivery systems (DDS) of isoniazid (INH) was scrutinized through density functional theory (DFT) and molecular dynamic (MD) simulations. We performed DFT periodic calculations on the geometry and electronic features of nanosheets adsorbed with INH by the DFT functional (DZP/GGA-PBE) employed in the SIESTA code. In the energetically favorable model, an oxygen atom of the C-O group of the INH molecule interacts with a Si atom of the h-SiC at 2.077 Å with an interaction energy of -1.361 eV. Charge transfer (CT) calculation by employing the Mulliken, Hirshfeld and Voronoi approaches reveals that the monolayers and drug molecules act as donors and acceptors, respectively. The density of states (DOS) calculations indicate that the HOMO-LUMO energy gap (HLG) of the h-SiC nanosheet declines significantly from 2.543 to 1.492 eV upon the adsorption of the INH molecule, which causes an electrical conductivity increase and then produces an electrical signal. The signal is linked to the existence of INH, demonstrating that h-SiC may be an appropriate sensor for INH sensing. The decrease in HLG for the interaction of INH and h-SiC is the uppermost (up to 41%) representing the uppermost sensitivity, whereas the sensitivity trend is σ(h-SiC) > σ(h-AlN) > σ(h-BN). Quantum theory of atoms in molecules (QTAIM) investigations is employed to scrutinize the nature of the INH/nanosheet interactions. The QTAIM analysis reveals that the interaction of the INH molecule and h-SiC has a partially covalent nature, while INH/h-AlN model electrostatic interaction occurs in the system and noncovalent and electrostatic interaction for the INH/h-BN model. Finally, the state-of-the-art DFT-MD simulations utilized in this study can mimic ambient conditions. The results obtained from the MD simulation show that it takes more time to bond the INH drug and h-SiC, and the INH/h-SiC system becomes stable. The results of the current research demonstrate the potential of h-SiC as a suitable sensor and drug delivery platform for INH drugs to remedy tuberculosis.

Phenanthroline-functionalized donor-acceptor covalent organic frameworks as photo-responsive nanozymes for visual colorimetric detection of isoniazid.

Development of metal-free nanozymes has raised concern for their extensive applications in photocatalysis and sensing fields. As novel metal-free nanomaterials, covalent organic frameworks (COFs) have engendered intense interest in the construction of nanozymes due to their structural controllability and molecular functionality. The formation of the molecular arrangement by embedding orderly donor-acceptors (D-A) linked in the framework topology to modulate material properties for highly efficient enzyme mimicking activity is of importance but challenging. Here, a strong D-A type of COF was designed and synthesized by integrating electron donor units (pyrene) and electron acceptor units (phenanthroline), named Py-PD COF. Using experiments and theoretical calculations, the introduction of a phenanthroline ring endowed the Py-PD COF with a narrowed band gap, and efficient charge transfer and separation. Further, the Py-PD COF exhibited a superior light-responsive oxidase-mimicking characteristic under visible light irradiation, which could catalyze the oxidation of 3,3',5,5-tetramethylbenzidine (TMB) and give the corresponding evolution of color. The nanoenzymatic activity of the Py-PD COF was light-regulated, which offers a fascinating advantage because of its high efficiency and spatial controllability. Based on previously mentioned characteristics, an "on-off" sensing platform for the colorimetric analysis of isoniazid (INH) could be constructed with a good linear relationship (2-100 µM) and a low limit of detection (1.26 µM). This research shows that not only is Py-PD COF an environmentally friendly compound for the colorimetric detection of INH, but it is also capable of providing the interesting D-A type COF-based material for designing an excellent nanozyme.

Assessing Polymorphic Purity of Rifampicin in Double and Triple-Drug Fixed-Dose Combination Products.

First-line tuberculostatic agents, Rifampicin (RIF), Isoniazid (ISH), Ethambutol (ETB), and Pyrazinamide (PZA) are generally administered as a fixed-dose combination (FDC) for improving patient adherence. The major quality challenge of these FDC products is their variable bioavailability, where RIF and its solid state are key factors. In this work, the analysis of the impact of the polymorphism in the performance of RIF in RIF-ISH and PZA-RIF-ISH combined products was carried out by an overall approach that included the development and validation of two methodologies combining near-infrared (NIR) spectroscopy and partial least squares (PLS) to the further evaluation of commercial products. For NIR-PLS methods, training and validation sets were prepared with mixtures of Form I/Form II of RIF, and the appropriate amount of ISH (for double associations) or ISH-PZA (for triple associations). The corresponding matrix of the excipients was added to the mixture of APIs to simulate the environment of each FDC product. Four PLS factors, reduced spectral range, and the combination of standard normal variate and Savitzky-Golay 1st derivative (SNV-D') were selected as optimum data pre-treatment for both methods, yielding satisfactory recoveries during the analysis of validation sets (98.5±2.0%, and 98.7±1.8% for double- and triple-FDC products, respectively). The NIR-PLS model for RIF-ISH successfully estimated the polymorphic purity of Form II in double-FDC capsules (1.02 ± 0.02w/w). On the other hand, the NIR-PLS model for RIF-ISH-PZA detected a low purity of Form II in triple FDC tablets (0.800 ± 0.021w/w), these results were confirmed by X-ray powder diffraction. Nevertheless, the triple-FDC tablets showed good performance in the dissolution test (Q=99-102%), implying a Form II purity about of 80% is not low enough to affect the safety and efficacy of the product.

Exploring the antimicrobial potential of isoniazid loaded Cu-based metal-organic frameworks as a novel strategy for effective killing of Mycobacterium tuberculosis.

Tuberculosis (TB) remains one of the most infectious pathogens with the highest human mortality and morbidity. Biofilm formation during Mycobacterium tuberculosis (Mtb) infection is responsible for bacterial growth, communication, and, most essentially, increased resistance/tolerance to antibiotics leading to higher bacterial persistence. Thus, biofilm growth is presently considered a key virulence factor in the case of chronic disease. Metal-Organic Frameworks (MOFs) have recently emerged as a highly efficient system to improve existing antibiotics' therapeutic efficacy and reduce adverse effects. In this regard, we have synthesized Cu-MOF (IITI-3) using a solvothermal approach. IITI-3 was well characterized by various spectroscopic techniques. Herein, IITI-3 was first encapsulated with isoniazid (INH) to form INH@IITI-3 with 10 wt% loading within 1 hour. INH@IITI-3 was well characterized by PXRD, TGA, FTIR, and BET surface area analysis. Furthermore, the drug release kinetics studies of INH@IITI-3 have been performed at pH 5.8 and 7.4 to mimic the small intestine and blood pH, respectively. The results show that drug release follows first-order kinetics. Furthermore, the antimycobacterial activity of INH@IITI-3 demonstrated significant bacterial killing and altered the structural morphology of the bacteria. Moreover, INH@IITI-3 was able to inhibit the mycobacterial biofilm formation upon treatment and showed less cytotoxicity toward the murine RAW264.7 macrophages. Thus, this work significantly opens up new possibilities for the applications of INH@IITI-3 in biofilm infections in Mtb and further contributes to TB therapeutics.

Evaluating the Impact of the Tyr158 pKa on the Mechanism and Inhibition of InhA, the Enoyl-ACP Reductase from Mycobacterium tuberculosis.

InhA, the Mycobacterium tuberculosis enoyl-ACP reductase, is a target for the tuberculosis (TB) drug isoniazid (INH). InhA inhibitors that do not require KatG activation avoid the most common mechanism of INH resistance, and there are continuing efforts to fully elucidate the enzyme mechanism to drive inhibitor discovery. InhA is a member of the short-chain dehydrogenase/reductase superfamily characterized by a conserved active site Tyr, Y158 in InhA. To explore the role of Y158 in the InhA mechanism, this residue has been replaced by fluoroTyr residues that increase the acidity of Y158 up to ∼3200-fold. Replacement of Y158 with 3-fluoroTyr (3-FY) and 3,5-difluoroTyr (3,5-F2Y) has no effect on kcatapp/KMapp nor on the binding of inhibitors to the open form of the enzyme (Kiapp), whereas both kcatapp/KMapp and Kiapp are altered by seven-fold for the 2,3,5-trifluoroTyr variant (2,3,5-F3Y158 InhA). 19F NMR spectroscopy suggests that 2,3,5-F3Y158 is ionized at neutral pH indicating that neither the acidity nor ionization state of residue 158 has a major impact on catalysis or on the binding of substrate-like inhibitors. In contrast, Ki*app is decreased 6- and 35-fold for the binding of the slow-onset inhibitor PT504 to 3,5-F2Y158 and 2,3,5-F3Y158 InhA, respectively, indicating that Y158 stabilizes the closed form of the enzyme adopted by EI*. The residence time of PT504 is reduced ∼four-fold for 2,3,5-F3Y158 InhA compared to wild-type, and thus, the hydrogen bonding interaction of the inhibitor with Y158 is an important factor in the design of InhA inhibitors with increased residence times on the enzyme.

Doxorubicin-isoniazid conjugate regulates immune response and tumor microenvironment to enhance cancer therapy.

Immune checkpoint inhibitors (ICIs) represent a new class of immunotherapy drugs, and are used to relieve immune suppression or enhance the immune response through the blockade of checkpoint ligands or receptors. ICIs have achieved great success in clinical cancer treatment. Monoamine oxidase A (MAOA) is a potent immune checkpoint of immunotherapy. Recently, it has been reported that MAOA inhibitors could enhance CD8+ T cell activity by upregulating 5-HT autocrine pathway in T cells. In this study, we synthesized doxorubicin (DOX) and isoniazid (INH, a MAOA inhibitor) conjugates through a pH sensitive hydrazone bond. Results of the in vivo studies showed that DOX-INH could effectively enhance the activity of CD8+ T cells and perform a synergistic anti-tumor effect with PD-L1 small molecular inhibitor (BMS202). In addition, in an orthotopic 4T1 breast cancer model, it was demonstrated that DOX-INH could inhibit the epithelial-mesenchymal transition process by blocking Shh, IL-6, and TGF-ß signaling pathways, thereby inhibiting the growth and metastasis of breast cancer. Thus, a simple and effective small molecule conjugate produced by the combination of a chemotherapy drug and a MAOA inhibitor shows broad prospect in cancer therapy.

Isonicotinic acid N-oxide, from isoniazid biotransformation by Aspergillus niger, as an InhA inhibitor antituberculous agent against multiple and extensively resistant strains supported by in silico docking and ADME prediction.

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.