Evaluation of monoclonal antibody DF3 conjugated with ricin as a specific immunotoxin for in vitro purging of human bone marrow.
DF3 is an IgG1 monoclonal antibody (MAb) generated against a Mr 350,000-400,000 glycoprotein expressed by approximately 80% of human breast cancers. We have coupled MAb DF3 to ricin. Purification of the immunotoxin (DF3-IT) was obtained by affinity and size exclusion chromatography. DF3 antigen-positive breast cancer cell lines (ZR-75-1, BT-20, and MCF-7) and DF3 antigen-negative lung cancer cell lines (A549 and CALU6) were tested for cytotoxicity with metabolic labeling and clonogenic assays. The cells were exposed for 3 h to different concentrations of DF3-IT, MAb DF3, ricin, and a combination of unconjugated MAb DF3 and ricin. In the presence of 100 mM lactose, DF3-IT specifically inhibited protein synthesis of lines expressing DF3 antigen on their cell surface. Moreover, clonogenic survival experiments demonstrated that DF3-IT at a concentration of 1 x 10(-9) M specifically kills 2.6-2.8 log of ZR-75-1 and BT-20 cells and 1.6 log of MCF-7 cells. At the same concentration, nonspecific toxicity of DF3-IT resulted in a 30% reduction of bone marrow granulocyte and macrophage colony formation. In bone marrow-purging experiments, tumor cells were mixed with an excess of bone marrow cells and treated with DF3-IT or ricin. Tumor cell clonogenic survival assays demonstrated that the presence of bone marrow cells had no detectable effect on activity or specificity of the DF3-IT. These results thus indicate that MAb DF3 is an effective vehicle to specifically deliver toxins to cancer cells which express DF3 antigen on their surface and that DF3-IT may be useful for in vitro purging of bone marrow.
Tondini
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Department of Medicine, Dana-Farber Cancer Institute, Boston, Massachusetts 02115.
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Hayes
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Kufe
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D000888:Antibodies, Anti-Idiotypic / Q000276:immunology / Q000494:pharmacology*; D000951:Antigens, Neoplasm / Q000276:immunology*; D001853:Bone Marrow / Q000187:drug effects / Q000276:immunology*; D001943:Breast Neoplasms / Q000276:immunology / Q000378:metabolism / Q000628:therapy*; D003114:Colony-Forming Units Assay; D004338:Drug Combinations; D005455:Fluorescent Antibody Technique; D006098:Granulocytes / Q000166:cytology; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology* / Q000494:pharmacology; D008175:Lung Neoplasms / Q000378:metabolism; D009363:Neoplasm Proteins / Q000096:biosynthesis; D012276:Ricin / Q000276:immunology / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000276:immunology / Q000378:metabolism
D000888:Antibodies, Anti-Idiotypic; D000951:Antigens, Neoplasm; D004338:Drug Combinations; D007074:Immunoglobulin G; D009363:Neoplasm Proteins; C048844:anti-IgG; D012276:Ricin