A series of 33 (E)-N'-benzylidenepyrazine-2-carbohydrazides and their derivatives were synthesized and tested for biological activity. Benzylidene derivatives with 2-OH substitution on the phenyl ring (18: R = 2-OH, 21: R = 2,3-diOH, and 22: R = 2,4-diOH) exhibit various biological activities. Compounds 18 and 21 demonstrate antimycobacterial activity against Mycobacterium tuberculosis H37Ra, M. tuberculosis H37Rv, and M. aurum, with minimum inhibitory concentration values ranging from 15.625 to 62.5 μg mL-1. Compounds 18, 21, and 22 show mild cytotoxicity on several human cell lines (IC50 ranging from 70.2 to 500 μM). Crystallographic studies confirm the (E)-configuration of compound 18 and a nearly planar molecular conformation. Due to their structural similarity with salicylaldehyde isonicotinoyl hydrazone (SIH), a known iron chelator, selected compounds were tested for iron-chelating properties, revealing comparable or superior activity. Mechanistic assays targeting enoyl-[acyl carrier protein] reductase (InhA), isocitrate lyase (ICL), and lipid/mycolic acid biosynthesis show no significant inhibition, suggesting a nonspecific mechanism potentially linked to iron chelation. A correlation is observed between chelating activity and cytotoxicity, while antimycobacterial activity appears to involve additional mechanisms. Pharmacokinetic studies with compound 18 reveal no specific plasma metabolites, and no significant metabolites are detected after incubation with human liver microsomes.

• Keywords
• antimycobacterials, antitumor agents, iron chelators, medicinal chemistry, pyrazine‐2‐carbohydrazides,
• MeSH
• Anti-Bacterial Agents * pharmacology   chemistry   chemical synthesis   MeSH
• Antitubercular Agents * pharmacology   chemistry   chemical synthesis   MeSH
• Hydrazines * chemistry   pharmacology   chemical synthesis   MeSH
• Humans MeSH
• Microbial Sensitivity Tests MeSH
• Molecular Structure MeSH
• Mycobacterium tuberculosis drug effects   MeSH
• Cell Line, Tumor MeSH
• Cell Proliferation drug effects   MeSH
• Antineoplastic Agents * pharmacology   chemistry   chemical synthesis   MeSH
• Pyrazines * chemistry   pharmacology   chemical synthesis   MeSH
• Drug Screening Assays, Antitumor MeSH
• Dose-Response Relationship, Drug MeSH
• Structure-Activity Relationship MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Anti-Bacterial Agents * MeSH
• Antitubercular Agents * MeSH
• carbohydrazide MeSH Browser
• Hydrazines * MeSH
• Antineoplastic Agents * MeSH
• Pyrazines * MeSH

Drug resistance is a growing problem for many pathogens, including mycobacteria. Small heterocyclic molecules are among the leading scaffolds for developing potential antimycobacterial agents. Therefore, based on the molecular hybridization approach, we have prepared an extensive series of N-substituted 5-(3,5-dinitrophenyl)-1,3,4-oxadiazol-2-amine derivatives. We also investigated their isosteres and acyclic synthetic precursors. The compounds were evaluated for their in vitro activity against Mycobacterium tuberculosis (Mtb) H37Rv, a panel of multidrug- and extensively drug-resistant Mtb isolates and two nontuberculous mycobacterial strains (NTM; M. avium and M. kansasii). The ability to inhibit mycobacterial growth was quantified using minimum inhibitory concentration (MIC) values. Many compounds achieved MIC values ≤ 0.03 µM for NTM and Mtb, regardless of their resistance profile. The highest activity was associated with oxadiazole and thiadiazole scaffolds with benzylamino or C5-C9 alkylamino substitution. The experimentally confirmed mechanism of action of these compounds consists of disruption of mycobacterial cell wall biosynthesis via inhibition of decaprenylphosphoryl-β-D-ribose 2'-epimerase (DprE1). In vitro toxicity evaluation was performed in a hepatocyte model (HepG2), while in vivo toxicity was evaluated using Danio rerio embryos. These findings identify a promising new chemotype with potent, broad-spectrum and selective antimycobacterial activity, including efficacy against resistant strains, and support its further development as a potential therapeutic candidate.

• MeSH
• Antitubercular Agents * pharmacology   chemical synthesis   chemistry   toxicity   MeSH
• Zebrafish MeSH
• Humans MeSH
• Microbial Sensitivity Tests MeSH
• Mycobacterium tuberculosis drug effects   MeSH
• Oxadiazoles * pharmacology   chemical synthesis   chemistry   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Antitubercular Agents * MeSH
• Oxadiazoles * MeSH

3,5-Dinitrobenzylsulfanyl tetrazoles and 1,3,4-oxadiazoles, previously identified as having high in vitro activities against both replicating and nonreplicating mycobacteria and favorable cytotoxicity and genotoxicity profiles were investigated. First we demonstrated that these compounds act in a deazaflavin-dependent nitroreduction pathway and thus require a nitro group for their activity. Second, we confirmed the necessity of both nitro groups for antimycobacterial activity through extensive structure-activity relationship studies using 32 structural types of analogues, each in a five-membered series. Only the analogues with shifted nitro groups, namely, 2,5-dinitrobenzylsulfanyl oxadiazoles and tetrazoles, maintained high antimycobacterial activity but in this case mainly as a result of DprE1 inhibition. However, these analogues also showed increased toxicity to the mammalian cell line. Thus, both nitro groups in 3,5-dinitrobenzylsulfanyl-containing antimycobacterial agents remain essential for their high efficacy, and further efforts should be directed at finding ways to address the possible toxicity and solubility issues, for example, by targeted delivery.

• MeSH
• Antitubercular Agents pharmacology   chemistry   MeSH
• Microbial Sensitivity Tests MeSH
• Mycobacterium tuberculosis * MeSH
• Nitroreductases MeSH
• Oxadiazoles pharmacology   chemistry   MeSH
• Mammals MeSH
• Tetrazoles pharmacology   chemistry   MeSH
• Structure-Activity Relationship MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antitubercular Agents MeSH
• Nitroreductases MeSH
• Oxadiazoles MeSH
• Tetrazoles MeSH