Sublines of sarcoma 180 (S180) of varying sensitivity to adriamycin (ADR) have been selected in culture. The degree of resistance of these sublines ranged from 6- to 125-fold above that of parent S180 cells. ADR-resistant sublines demonstrated comparable degrees of cross-resistance to daunomycin (DNR), marcellomycin and AD 32, but each subline showed a uniform degree of tolerance toward actinomycin D and vincristine. Compared to the anthracycline-sensitive parent tumor, a 40% decrease in the intracellular steady-state level of [3H]-daunomycin was observed in all sublines regardless of the degree of resistance. The level of cell-associated DNR and ADR observed after administration of equipotent concentrations of drug was different for each cell line and increased in proportion to the drug concentration. Thus, altered drug permeability appeared to be of minimal importance in the expression of high levels of resistance. In addition, the extent of DNR metabolism by the anthracycline-resistant sublines was not sufficiently different from that seen in parent S180 cells to account for the observed tolerance to these agents.