Biochemical and pharmacological characterization of MCF-7 drug-sensitive and Adr R multidrug-resistant human breast tumor xenografts in athymic nude mice

Abstract

The phenotypic expression of multidrug resistance by the doxorubicin-selected Adr R human breast tumor cell line is associated with overexpression of plasma membrane P-170 glycoprotein and increased cytosolic selenium-dependent GSH-peroxidase activity relative to the parental MCF-7 wild-type line (WT). To determine whether doxorubicin resistance by Adr R cells persists in vivo, and to further investigate the possibility of biochemical differences between WT and Adr R solid tumors, both tumor cell lines were grown as subcutaneous xenografts in athymic nude mice. Tumorigenicity depended upon cell inoculation burden, and tumor incidence was similar for both cell lines (> 80% tumor takes at 10 7 cells/mouse) at 14 days, provided 17β-estradiol was supplied to the animals bearing the WT tumors. However, the growth rate for the Adr R xenografts was only about half that of WT xenografts. Doxorubicin (2–8 mg/kg, i.p., injected weekly) significantly diminished the growth of the WT′tumors, but Adr R solid tumors failed to respond to doxorubicin. The accumulation of 14C-labeled doxorubicin was 2-fold greater in WT xenografts that in Adr R, although there were no differences in host organ drug levels in mice bearing either type of tumors. Membrane P-170 glycoprotein mRNA was detected by slot-blot analysis in the Adr R tumors, but not in WT. Electron spin resonance 5,5-dimethylpyrroline- N-oxide-spin-trapping experiments with microsomes and mitochondria from WT and Adr R xenographs demonstrated a 2-fold greater oxygen radical (superoxide and hydroxyl) formation from activated doxorubicin with WT xenographs compared to Adr R. Selenium-dependent glutathione (GSH)-peroxidase, superoxide dismutase and GSH- Saryltranferase activities in Adr R xenographs were elevated relative to WT. Although the activities of the latter two enyzmes were similar to those measured in both tumor cell lines, GSH-peroxidase activities were elevated 70-fold (WT) and 10-fold (Adr R) in xenografts compared to tumor cells. In contrast, in both WT and Adr R solid tumors in vivo, catalase, NAD(P)H-oxidoreductases, and glutathione disulfide (GSSG)-reductase activities, and GSH and GSSG levels were not markedly different, and were essentially the same as in cells in vitro. Like the MDR cells in culture, Adr R tumor xenografts were extremely resistant to doxorubicin and retained most of the characteristics of the altered phenotype. These results suggest that WT and Adr R breast tumor xenografts provide a useful model for the study of biochemical and pharmacological mechanisms of drug resistance by solid tumors in vivo.