Exploração de aceptores de Michael contra proteases de interesse clínico: planejamento, triagem virtual e aplicação na resistência bacteriana e tripanossomíase africana

Abstract

The increasing rise in antibiotic resistance urgently demands the discovery of new drugs capable of combating pathogenic microorganisms or modulating resistance mechanisms. In parallel, neglected diseases, such as African trypanosomiasis, represent a critical challenge for developing countries. Both conditions share a common therapeutic target: serine and cysteine proteases, considered targets for the development of new pharmaceutical prototypes. Among the chemical classes with potential inhibitory effects on these enzymes, Michael acceptors have been highlighted. In this context, this study developed a series of compounds from virtual screening, containing α,β-unsaturated carbonyl structures with electrophilic centers ("warheads") to act against cysteine and serine proteases of clinical interest. Approximately 50 potential hits were identified, derived from acrylamides, α-substituted by cyan, fluorine and thiol. The synthesis of the selected compounds involved Knoevenagel, Horner– Wadsworth–Emmons and amide coupling reactions, resulting in 34 acrylic acid intermediates, 14 acrylamide esters and 13 acrylamides with yields ranging from 20-98%. The compounds were evaluated in enzymatic assays against the cysteine proteases cathepsins B, L and rhodesain, in addition to the serine proteases β-lactamase and trypsin, demonstrating selectivity for serine proteases. Additionally, synergy tests were performed against multidrug-resistant bacteria producing β-lactamases of classes A, B and C. The combination of the synthesized compounds with polymyxin B exhibited promising synergistic activity for compounds 6e, 2c and 11b, resulting in the reduction of bacterial growth of Klebsiella pneumoniae KPC P15-ST11 strains and also with other combinations and strains. The heterocyclic nucleus 5-bromoindole emerged as the recognition unit, since the active compounds present this substituent. Additionally, the in silico molecular dynamics assay revealed water-mediated hydrogen interactions at the catalytic site, in addition to direct hydrogen interactions with the NH of 5-bromoindole and Ser70. The compounds discovered here are expected to be applied in the design of future drugs against the β-lactamase-mediated bacterial resistance mechanism.