A cell line, NIH:OVCAR-3, has been established from the malignant ascites of a patient with progressive adenocarcinoma of the ovary after combination chemotherapy with cyclophosphamide, Adriamycin, and cisplatin. OVCAR-3 grows as a cobblestone-like monolayer with foci of multilayering, is tumorigenic in athymic mice, clones in agarose, and has an abnormal karyotype which includes a homogeneous staining region and a double minute chromosome. The cultured cells and xenografts contain cytoplasmic androgen- and estrogen-binding macromolecules with the specificity of the respective steroid hormone receptors. These components have sedimentation coefficients of 7 to 9S in low-salt sucrose-density gradients, have dissociation constants of 250 and 9.6 pM, and are present at concentrations of 30 and 28 fmol/mg cytosol protein characteristic of androgen and estrogen receptors, respectively. OVCAR-3 is resistant in vitro to clinically relevant concentrations of Adriamycin (5 X 10(-8) M), melphalan (5 X 10(-6) M), and cisplatin (5 X 10(-7) M) with survival compared to untreated controls of 43, 45, and 77%, respectively. Furthermore, there are multiple histological similarities between the patient's original tumor, the cell line, and the transplantable tumor. These data indicate that OVCAR-3 may be of use for investigations as to the significance of androgens and estrogens and the mechanisms of cytotoxic drug resistance in ovarian cancer.