|Budget Amount *help
¥4,400,000 (Direct Cost : ¥4,400,000)
Fiscal Year 1994 : ¥1,800,000 (Direct Cost : ¥1,800,000)
Fiscal Year 1993 : ¥2,600,000 (Direct Cost : ¥2,600,000)
To develop a new diagnostic method for ovarian cancers at an carly stage, we produced murine monoclonal antibodies (MoAbs) against ovarian cancer-derived molecules which are lmmunogenic in patients. Five different strategies of immunizing antigen/screening antigen combination were tried. In all strategies, immune complexes (ICs) purified from ascites of 30 patients with various ovarian cancers were used as immunizing antigens and/or screening antigens in ELISA.The purification of ICs includes ammonium sulphate precipitation. Sephacryl S300 gel filtration, and protein A affinity chromatography. As immunizing antigens, serum-free conditioned medium of a serous cystadenocarcinoma cell line (HOC/BR) or tumor tissue of ovarian cancers were also used. ICs purified from pooled sera of normal healthy controls were used as negative screening antigens. We screened more than 8,000 supernatants of hybridomas in to fusion experiments, and established 7 MoAbs which react with ascites LCs of ovarian cancers, but not with IgG or ICs of normal controls. Among them, 2 MoAbs (2Flt and 7E2 clones) can also detect circulating ICs in peripheral blood from patients with ovarian cancers by sandwich ELISA using protein A precoated micropiates. Conditioned medium of HOC/BR cell line inhibited the reactivity of 3 MoAbs (2Fll, 7E2, and H5 clones), indicating that these antibodies detect ovarian cancer-derived molecules involved in the patients' ICs. These molecules are, therefore, considered to express tumor-associated epitopes reacted by patients' IgG.
(competitivc inhibition ELISA)その結果,透析・濃縮されたこの培養上清中には2F11,7E2,H5の反応を阻害する分子が存在することが確認され,これらのモノクローナル抗体の対応抗原は実際の卵巣癌でも可溶性成分として産生されている可溶性が強く示唆された。