Background <p>Dexrazoxane, a putative iron chelator, is effective in preventing doxorubicin-induced cardiotoxicity. However, dexrazoxane is also a catalytic inhibitor of topoisomerase 2b (Top2b), a key mediator of doxorubicin toxicity. Preclinical studies have shown that dexrazoxane induces Top2b degradation, and early administration (8&#xa0;h before doxorubicin) can prevent doxorubicin-induced cardiotoxicity. In this study, we investigated the dose–response relationship and time course of dexrazoxane-induced Top2b degradation in human volunteers.</p> Methods <p>Twenty-five healthy female volunteers received an intravenous infusion of dexrazoxane at doses ranging from 100&#xa0;mg/m² to 500&#xa0;mg/m². Blood samples were collected hourly from time zero to 12&#xa0;h, as well as at 24- and 48-h post-infusion. Peripheral blood mononuclear cells (PBMCs) were isolated, nuclear fractions were extracted, and Top2b expression was analyzed by western blot using Lamin B1 as a control. A linear mixed-effects model was used to assess differences among the five dose groups.</p> Results <p>Dexrazoxane infusion led to a rapid and sustained reduction of Top2b in PBMCs, lasting up to 12&#xa0;h. Statistical analysis revealed a significant difference in Top2b levels among the five dose groups (<i>p</i> = 0.0002). Subgroup analysis identified a significant difference between the 100&#xa0;mg/m² and 500&#xa0;mg/m² groups (<i>p</i> = 0.005). However, topoisomerase 2a (Top2a), the molecular target of doxorubicin’s tumor-killing effect, remained unchanged following dexrazoxane infusion.</p> Conclusions <p>Findings from this dose–response and time-course study can inform the design of future clinical trials investigating the efficacy of early dexrazoxane administration in preventing doxorubicin-induced cardiotoxicity while minimizing the risk of tumor protection.</p> Trial registration <p>(Funded by the National Institute of Health, RO1HL151993; PHOENIX trials,&#xa0;ClinicalTrials.gov number,&#xa0;NCT03930680.)</p>