Two different mechanisms that contribute to multidrug resistance (MDR) were found in derivatives of the human squamous lung cancer cell line SW-1573. The parental cell line has a low amount of mdr1 P-glycoprotein mRNA. In three independent selections for doxorubicin resistance, MDR variants arose in which mdr1 P-glycoprotein mRNA and protein was not detectable. Selection on higher doxorubicin concentrations gave rise to variants containing high levels of mdr1 mRNA, due to transcriptional activation of the mdr1 gene. Upon continued selection for higher levels of doxorubicin resistance, the mdr1 gene became amplified, resulting in an additional increase in the level of mdr1 mRNA. The cross-resistance pattern of the sublines that lack mdr1 P-glycoprotein expression is different from that seen in the mdr1 overexpressing cells. Both types of MDR cell lines are resistant to doxorubicin, daunorubicin, etoposide, colchicine, gramicidin D, and vincristine. However, in the non-P-glycoprotein-mediated MDR cell lines, resistance levels are lower and a preferential resistance for etoposide is seen.