Isolation by flow cytometry of a human ovarian tumor cell subpopulation exhibiting a high glutathione content phenotype and increased resistance to adriamycin

Abstract

We have used monochlorobimane as a quantitative marker by which cells of naturally high or low GSH contents were purified by fluorescence-activated cell sorting (FACS). The cell line chosen for this purpose, MLS, was a human ovarian tumor cell line established from a patient who had received extensive chemotherapy and showed evidence of ‘multidrug’ resistance. Cells of a specified volume were sorted into subpopulations containing the 1% most dim (low GSH) and 1% most bright (high GSH) cells. With an increasing number of sortings, cell subpopulations emerged with progressively lower (dim) and higher (bright) GSH content as compared to the parent population. After 4 sortings, GSH contents were 10.6 ± 0.8, 5.1 ± 0.4, and 7.2 ± 0.7 X 10- −18 moles/μm 3 for MLS/bright, MLS/dim and MLS/parent respectively. The high and low GSH phenotypes were of limited stability reverting to the parent phenotype by the sixth week following the last FACS. Cells with high GSH content were more resistant to adriamycin than cells with low GSH content, for example, at 1 log cell kill MLS/bright was 1.6 fold more resistant than MLS/dim. An ADR resistant variant of the MLS line, designated MLS/SDR R 2 , established by twice treating MLS cells with 1 μg/ml ADR for 2 hr, also showed increased GSH content (1.3-fold) and ADR resistance as compared with the parent line. These results illustrate the possible importance of tumor cell GSH status in determining the response to chemotherapy of a heterogenous population of tumor cells with diverse GSH contents.