Separate mechanisms underlying the multidrug resistant (MDR) phenotype were identified in 2 independent approaches to select tumour cells resistant to low concentrations of doxorubicin (Dox) from the sensitive T cell leukemia cell line CCRF‐CEM. The CEM/A7 cell line was selected at an initial concentration of 0.005 μg/ml of Dox and maintained at 0.07 μg/ml. In contrast, the CEM/A5 line was selected using an initial concentration of 0.01 μg/ml and maintained in Dox at a concentration of 0.05 μg/ml. P‐glycoprotein expression was demonstrated in the CEM/A7 line but not the CEM/A5 line. Amplification of the mdr1 gene was not observed in the CEM/A7 cell line. Both cell lines showed cross‐resistance to a number of structurally unrelated cytotoxic drugs including anthracyclines and etoposide (VP‐16), although only the CEM/A7 line was cross resistant to Vinca alkaloids. Immunoblots of total cell lysates of the CEM/A5 line have revealed almost undetectable levels of topoisomerase 11 α and β in this line. Cytogenetic analyses of both lines revealed numerous karyotypk abnormalities which were present in the parental cell line as well as both resistant cell lines. The CEM/A7 line also demonstrated a duplication of part of the long arm of chromosome 7 which included the region containing the mdr1 gene, a finding not seen in the parental or CEM/AS line. CEM/AS, however, demonstrated an abnormality of chromosome 7, outside the region of the mdr1 gene, and it also contained a deletion of the short arm of chromosome 2. Abnormalities in this latter region of genome have been associated with non‐P‐glycoprotein‐mediated MDR. © 1994 Wiley‐Liss, Inc.