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.

Biomedical Research Centre University Hospital Hradec Králové Sokolská 581 500 05 Hradec Králové Czech Republic

Department of Biochemistry Faculty of Natural Sciences Comenius University in Bratislava Mlynská dolina Ilkovičova 6 842 15 Bratislava Slovakia

Department of Chemistry University of Milan Via Golgi 19 20133 Milan Italy

Department of Clinical Microbiology University Hospital Hradec Králové Sokolská 581 500 05 Hradec Králové Czech Republic

Department of Pharmaceutical Sciences University of Milan Via Mangiagalli 25 20133 Milan Italy

Faculty of Pharmacy in Hradec Králové Charles University Ak Heyrovského 1203 500 03 Hradec Králové Czech Republic

Faculty of Pharmacy University of Ljubljana Aškerčeva cesta 7 SI 1000 Ljubljana Slovenia

Zobrazit více v PubMed

WHO , Tuberculosis—Overview, Symptoms and Treatment, https://www.who.int/health‐topics/tuberculosis#tab=tab_1, (accessed: June, 2025).

Alsayed S. S. R., Gunosewoyo H., Int. J. Mol. Sci. 2023, 24, 5202. PubMed PMC

Brehm T. T., Kohler N., Schmiedel S., Terhalle E., Martensen J., Kalsdorf B., Kandulla J., Heyckendorf J., Kuhns M., Friesen I., Lange C., Inn. Med. 2023, 64, 701. PubMed

Shi W., Chen J., Feng J., Cui P., Zhang S., Weng X., Zhang W., Zhang Y., Emerg. Microbes Infect. 2014, 3, e58. PubMed PMC

Sun Q., Li X., Perez L. M., Shi W., Zhang Y., Sacchettini J. C., Nat. Commun. 2020, 11, 339. PubMed PMC

Bouz G., Slechta P., Jand’ourek O., Konecna K., Paterova P., Barta P., Novak M., Kucera R., Dal N. K., Fenaroli F., Zemanova J., Forbak M., Kordulakova J., Pavlis O., Kubickova P., Dolezal M., Zitko J., ACS Infect. Dis. 2023, 9, 79. PubMed

Zitko J., Servusova B., Paterova P., Mandikova J., Kubicek V., Kucera R., Hrabcova V., Kunes J., Soukup O., Dolezal M., Molecules 2013, 18, 14807. PubMed PMC

Nawrot D. E., Bouz G., Jandourek O., Konecna K., Paterova P., Barta P., Novak M., Kucera R., Zemanova J., Forbak M., Kordulakova J., Pavlis O., Kubickova P., Dolezal M., Zitko J., Eur. J. Med. Chem. 2023, 258, 115617. PubMed

Vergara F. M., Lima C. H., Henriques M., Candea A. L., Lourenco M. C., Mde L. F., Kaiser C. R., de Souza M. V., Eur. J. Med. Chem. 2009, 44, 4954. PubMed

d. S. B. Felipe A. R. Rodrigues I., Cavalcanti B. C., Pessoa C. Ó., Pinheiro A. C., Lima C. H. S., de Souza M. V. N., Euro. Chem. Bullet. 2014, 3, 358.

Abdel‐Aziz M., Abdel‐Rahman H. M., Eur. J. Med. Chem. 2010, 45, 3384. PubMed

Ferreira M. D., Candea A. L. P., Henriques M. D. M. D., Kaiser C. R., Lima C. H. D., de Souza M. V. N., Lett. Drug Des. Discovery 2010, 7, 275.

da Silva Y. K., Augusto C. V., de Castro Barbosa M. L., de Albuquerque Melo G. M., de Queiroz A. C., de Lima Matos Freire Dias T., Junior W. B., Barreiro E. J., Lima L. M., Alexandre‐Moreira M. S., Bioorg. Med. Chem. 2010, 18, 5007. PubMed

Jansova H., Kubes J., Reimerova P., Sterbova‐Kovarikova P., Roh J., Simunek T., Chem. Res. Toxicol. 2018, 31, 1151. PubMed

Alanine A., Anselm L., Steward L., Thomi S., Vifian W., Groaning M. D., Bioorg. Med. Chem. Lett. 2004, 14, 817. PubMed

Baddeley T. C., Howie R. A., Lima C. H. D., Kaiser C. R., de Souza M. V. N., Wardell J. L., Wardell S. M. S. V., Zeitschrift Fur Kristallographie 2009, 224, 506.

Bruno I. J., Cole J. C., Kessler M., Luo J., Motherwell W. D., Purkis L. H., Smith B. R., Taylor R., Cooper R. I., Harris S. E., Orpen A. G., J. Chem. Inf. Comput. Sci. 2004, 44, 2133. PubMed

Ratledge C., Tuberculosis 2004, 84, 110. PubMed

Chao A., Sieminski P. J., Owens C. P., Goulding C. W., Chem. Rev. 2019, 119, 1193. PubMed PMC

Agoro R., Mura C., Pharmaceuticals 2019, 12, 75. PubMed PMC

Rodriguez G. M., Trends Microbiol. 2006, 14, 320. PubMed

Corce V., Gouin S. G., Renaud S., Gaboriau F., Deniaud D., Bioorg. Med. Chem. Lett. 2016, 26, 251. PubMed

Ibrahim O., O’Sullivan J., Biometals 2020, 33, 201. PubMed

Esposito B. P., Epsztejn S., Breuer W., Cabantchik Z. I., Anal. Biochem. 2002, 304, 1. PubMed

Zitko J., Dolezal M., Expert Opin. Ther. Pat. 2016, 26, 1079. PubMed

Sink R., Sosic I., Zivec M., Fernandez‐Menendez R., Turk S., Pajk S., Alvarez‐Gomez D., Lopez‐Roman E. M., Gonzales‐Cortez C., Rullas‐Triconado J., Angulo‐Barturen I., Barros D., Ballell‐Pages L., Young R. J., Encinas L., Gobec S., J. Med. Chem. 2015, 58, 613. PubMed

Kratky M., Novotna E., Stolarikova J., Svarcova M., Vinsova J., Eur. J. Pharm. Sci. 2022, 176, 106252. PubMed

Antil M., Gupta V., Protein Pept. Lett. 2022, 29, 1031. PubMed

Novotna E., Waisser K., Kunes J., Palat K., Skalova L., Szotakova B., Buchta V., Stolarikova J., Ulmann V., Pavova M., Weber J., Komrskova J., Haskova P., Vokral I., Wsol V., Arch. Pharm. 2017, 350, 1700020. PubMed

Vilcheze C., Morbidoni H. R., Weisbrod T. R., Iwamoto H., Kuo M., Sacchettini J. C., W. R. Jacobs Jr, J. Bacteriol. 2000, 182, 4059. PubMed PMC

Grzegorzewicz A. E., Pham H., Gundi V. A., Scherman M. S., North E. J., Hess T., Jones V., Gruppo V., Born S. E., Kordulakova J., Chavadi S. S., Morisseau C., Lenaerts A. J., Lee R. E., McNeil M. R., Jackson M., Nat. Chem. Biol. 2012, 8, 334. PubMed PMC

Karabanovich G., Dusek J., Savkova K., Pavlis O., Pavkova I., Korabecny J., Kucera T., Vlckova H. K., Huszar S., Konyarikova Z., Konecna K., Jand’ourek O., Stolarikova J., Kordulakova J., Vavrova K., Pavek P., Klimesova V., Hrabalek A., Mikusova K., Roh J., J. Med. Chem. 2019, 62, 8115. PubMed

Matviiuk T., Madacki J., Mori G., Orena B. S., Menendez C., Kysil A., Andre‐Barres C., Rodriguez F., Kordulakova J., Mallet‐Ladeira S., Voitenko Z., Pasca M. R., Lherbet C., Baltas M., Eur. J. Med. Chem. 2016, 123, 462. PubMed

Singh V., Dhar N., Pato J., Kolly G. S., Kordulakova J., Forbak M., Evans J. C., Szekely R., Rybniker J., Palcekova Z., Zemanova J., Santi I., Signorino‐Gelo F., Rodrigues L., Vocat A., Covarrubias A. S., Rengifo M. G., Johnsson K., Mowbray S., Buechler J., Delorme V., Brodin P., Knott G. W., Ainsa J. A., Warner D. F., Keri G., Mikusova K., McKinney J. D., Cole S. T., Mizrahi V., et al., Mol. Microbiol. 2017, 103, 13. PubMed

Chiarelli L. R., Salina E. G., Mori G., Azhikina T., Riabova O., Lepioshkin A., Grigorov A., Forbak M., Madacki J., Orena B. S., Manfredi M., Gosetti F., Buzzi A., Degiacomi G., Sammartino J. C., Marengo E., Kordulakova J., Riccardi G., Mikusova K., Makarov V., Pasca M. R., ACS Infect. Dis. 2020, 6, 313. PubMed

Fanali G., di Masi A., Trezza V., Marino M., Fasano M., Ascenzi P., Mol. Aspects Med. 2012, 33, 209. PubMed

Lopez‐Yerena A., Perez M., Vallverdu‐Queralt A., Escribano‐Ferrer E., Pharmaceutics 2020, 12, 1123.