Isoniazid (isonicotinic acid hydrazide, INH) is a key drug used to treat tuberculosis (TB), which continues to be the world's most lethal infectious disease. Nevertheless, the efficacy of INH has diminished because of the emergence of Mycobacterium tuberculosis (Mtb) strains that are resistant to INH. Our goal in this study was to modify INH to reduce this significant resistance chemically. We synthesized INH-based hydrazones (IP1-IP13) through the reaction of INH with in-house obtained benzaldehydes carrying a piperidine or piperazine ring in refluxing ethanol. Upon confirmation of their proposed structures by various spectral techniques, IP1-IP13 were evaluated for their antimycobacterial capacity against Mtb H37Rv strain and INH-resistant clinical isolates with katG and inhA mutations using the Microplate Alamar Blue Assay (MABA). The compounds were additionally tested for their cytotoxicity. The obtained data indicated that the compounds with moderately increased lipophilicity compared to INH (IP7-IP13) were promising antitubercular drug candidates, exhibiting drug-like properties and negligible cytotoxicity. Out of these, IP11 (N'-(4-(4-cyclohexylpiperazin-1-yl)benzylidene)isonicotinohydrazide) emerged as the most promising derivative, demonstrating the lowest MIC values against all Mtb strains tested. Subsequently, the target molecules were evaluated for their capacity to inhibit enoyl acyl carrier protein reductase (InhA), the main target enzyme of INH. Except for IP11 demonstrating 81% InhA inhibition at a concentration of 50 μM, direct InhA inhibition was shown not to be the primary mechanism responsible for the antitubercular activity of the compounds. The binding mechanism of IP11 to InhA was analyzed through molecular docking and molecular dynamics simulations. Altogether, our research identified a novel approach to modify INH to address the challenges posed by the rising prevalence of drug-resistant Mtb strains.
Keywords: InhA; Mycobacterium tuberculosis; N-acylhydrazone; antimycobacterial; cytotoxicity; molecular modeling.