Molecular analysis of two human doxorubicin-resistant cell lines: evidence for differing multidrug resistance mechanisms.

The molecular characteristics of two human doxorubicin-resistant cell lines were examined specifically for MDR1 gene amplification by Southern analysis and for overexpression of its messenger RNA. The 285-fold doxorubicin-resistant colon adenocarcinoma subline, LoVo/DR5, was found to overexpress the mRNA for P-glycoprotein without the concomitant requirement of MDR1 gene amplification, suggesting that relatively high levels of P-glycoprotein mediated multiple drug resistance may occur by transcriptional activation of the gene. Despite a similar in vitro selection strategy and in contrast to LoVo/DR cells, the 220-fold doxorubicin-resistant fibrosarcoma subline, HT1080/DR4, did not overexpress P-glycoprotein mRNA nor was the MDR1 gene amplified. In-gel renaturation studies were performed to determine the nature of a putative HSR-bearing chromosome 7 found in HT1080/DR4 cells; however, at a level of sensitivity nearing 20 copies of an amplified DNA fragment per haploid genome, no amplified sequences could be detected. These results suggest that doxorubicin resistance is multifactorial and alternative mechanisms of multiple drug resistance remain to be determined. LoVo/DR5 cells should prove to be a useful model for investigating transcriptional activation of the MDR1 gene; HT1080/DR4 cells should be an excellent model for the study of non-P-glycoprotein mediated multiple drug resistance.