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Adriamycin accumulation and metabolism were studied in three distinct groups of human ovarian cancer cell lines: those derived from previously untreated patients, those from clinically refractory (relapsed) patients, and those with induced resistance to adriamycin in vitro. The 2-hr [14C] adriamycin accumulation in cell lines from previously untreated patients (A2780 and A1847 [Eva et al., Nature, Lond. 295, 116 (1982)] and OVCAR-5 [National Institutes of Health human OVarian CAR-cinoma cell line no. 5]) was 11-14 ng/10(6) cells. 2780AD and 1847AD (variants with in vitro induced resistance to adriamycin) accumulated one-third as much adriamycin after 2 hr (4 ng/10(6) cells). However, three cell lines derived from clinically refractory patients accumulated the same amount of adriamycin as cell lines from untreated patients (8-13 ng/10(6) cells). A high-performance liquid chromatography (HPLC) assay for adriamycin and its analogs confirmed these results and demonstrated only parent drug (no metabolites) in any of the cell lines tested. These results demonstrate that the primary mechanism of adriamycin resistance in some ovarian cancer cells from clinically refractory patients is not enhanced metabolism of drug or a transport defect leading to a decreased net accumulation such as has been described for cells with in vitro induced resistance to adriamycin.

The optimum integration of chemotherapy and irradiation is of potential clinical significance in the treatment of ovarian cancer. A series of human ovarian cancer cell lines have been developed in which dose-response relationships to standard anticancer drugs have been determined, and the patterns of cross-resistance between these drugs and irradiation have been established. By stepwise incubation with drugs, sublines of A2780, a drug-sensitive cell line, have been made 100-fold, 10-fold, and 10-fold more resistant to Adriamycin (2780AD), melphalan (2780ME), and cisplatin (2780CP). Two additional cell lines, NIH:OVCAR-3nu(Ag+) and NIH:OVCAR-4(Ag+), were established from drug-refractory patients. 2780ME, 2780CP, OVCAR-3nu(Ag+), and OVCAR-4(Ag+) are all cross-resistant to irradiation, with DOS of 146, 187, 143, and 203, respectively. However, 2780AD remains sensitive to radiation, with a DO of 111, which is similar to that of A2780 (101). Glutathione (GSH) levels are elevated in 2780ME, 2780CP, OVCAR-3nu(Ag+), and OVCAR-4(Ag+) to 4.58, 6.13, 12.10, and 15.14 nmol/10(6) cells as compared to A2780, with 1.89 nmol/10(6) cells. However, the GSH level in 2780AD is only minimally higher than that in A2780 (2.94 nmol/10(6) cells). Buthionine sulfoximine, a specific inhibitor of GSH synthesis, significantly increases the radiation sensitivity of 2780ME (changing the DO from 143 to 95) and 2780CP to a lesser extent, suggesting that intracellular GSH levels may play an important role in the radiation response of certain neoplastic cells. These results suggest that the sequential use of irradiation following chemotherapy with melphalan and cisplatin may be less effective than a combined modality approach, which integrates radiation and chemotherapy prior to the development of drug resistance and cross-resistance to irradiation.