The chemotherapeutic, doxorubicin, is currently used empirically in the treatment of AIDS-related Kaposi's sarcoma (AIDS-KS). Although often employed in a chemotherapeutic cocktail (doxorubicin, bleomycin, vincristine) single-agent therapy has recently been attempted with liposome encapsulated doxorubicin. Although doxorubicin's mechanism of action against AIDS-KS is unknown, we hypothesized that doxorubicin's ability to undergo redox cycling is associated with its clinical efficacy. The current study was conducted to investigate the effects of doxorubicin on selected xenobiotic-associated biochemical responses of three cellular populations: KS lesional cells, nonlesional cells from the KS donors, and fibroblasts obtained from HIV Ϫ aged matched men. Our results show that during doxorubicin challenge, there are strong positive correlations between cellular glutathione (GSH) levels and viability (r ϭ 0.94), NADPH levels and viability (r ϭ 0.93), and GSH and NADPH levels (r ϭ 0.93), and demonstrate that as a consequence of their abilities to maintain cellular thiol redox pools HIV Ϫ donor cells are significantly less susceptible to doxorubicin's cytotoxic effects relative to AIDS-KS cells. Additional studies further supported the contribution of reduced thiols in mediating doxorubicin tolerance. While pretreatment with the GSH precursor, N-acetylcysteine was cytoprotective for all cell groups during doxorubicin challenge, GSH depletion markedly enhanced doxorubicin's cytotoxic effects. Studies to investigate the effects of a hydroxyl scavenger and iron chelator during doxorubicin challenge showed moderate cytoprotection in the AIDS-KS cells but deleterious effects in the HIV Ϫ control cells. Inactivation of the longer lived membrane generated ROI in the cytoprotective deficient AIDS-KS cells, as well as an impairment of endogenous defenses in the HIV Ϫ donor control cells, may account for these scavenger and chelator associated findings. In summary, our findings show that doxorubicin mediates, at least in part, its AIDS-KS cellular cytotoxic effects by a redox related mechanism, and provides a biochemical rationale for doxorubicin's clinical efficacy in AIDS-KS treatment.