Disclosing the true nature of hesperetin’s antigenotoxicity in vivo within the Drosophila melanogaster somatic cells through the extensive genotoxical and structure-based studies

Abstract

Previously unreported genotoxic and antigenotoxic potentials of hesperetin (Hes) were revealed by treating the Drosophila melanogaster (dm) whose DNA has been altered by means of O6-ethylguanine (dmGO6-Et) and O4-ethylthymine (dmTO4-Et) lesions appearance, caused by ethyl methanesulfonate (EMS), a proven alkylating agent and mutagen. Therefore, Hes potencies were determined by means of the comet assay on somatic cells level, where compound exerted no genotoxic effects but acted genotoxically as a Topoisomerase IIα (dmTopIIα) catalytic inhibitor by invading the Binding and Cleavage Domain and stabilizing the noncovalent dmTopIIα-plasmid DNA (dmPDNA) complex, as verified by the kinetoplast DNA (dmK-DNA) decatenation assays. Hes’s structure-based alignment caused compound’s A and C rings to occupy the area normally invaded by EMS, thus making a spatial barrier for the dmGO6-Et or dmTO4-Et lesions formation: the A ring C7-OH group formed hydrogen bonds (HBs) with either dmGO6 (dHB = 2.576 Å) or guanine’s N7 nitrogen (dmGN7, dHB = 2.737 Å), whereas the A ring C5-OH group formed an HB with dmTO4 (dHB = 3.548 Å). Furthermore, Hes likewise acted as a mixed-type competitive inhibitor of dmATPase, as verified by the catalytic, FRET, and structure-based studies where it affected the dmATPase dimerization and the hydrolysis of ATP, denying the metabolic energy for the catenation of ethylated G-dmDNA segment, the formation of dmTO4-Et-G-dmDNA phosphotyrosine intermediate (dmTO4-Et-GdmDNA- PTyr785I), and the passage of ethylated T-dmDNA segment through the temporarily broken dmTO4-Et-G-dmDNA-PTyr785I, processes seen as comets. Conclusively, Hes may be used in anticancer therapy controlling the effects of alkylating agents.