Complement depletion in vitro limits monoclonal antibody 6-19-dependent complement-mediated killing of tumor cells in bone marrow.
Selective killing of nonhematopoietic tumor cells in bone marrow harvested for autologous bone marrow transplantation was studied using cultured neuroblastoma cells, monoclonal antibody 6-19, and baby rabbit serum as a source of complement. Monoclonal antibody 6-19, a murine IgG2a antibody, was selected for binding to a cell surface antigen on cultured human neuroblastoma cells that is not expressed by hematopoietic cells. The antigen is an 80-kd sialoglycoprotein present on a wide variety of nonhematopoietic tumors, and it is expressed by normal fibroblast and endothelial cells. The effect of the presence or absence of bone marrow mononuclear cells on target cell survival was evaluated using Hoechst 33342-stained neuroblastoma cells, trypan blue exclusion, and fluorescence microscopy. More than 4 logs of neuroblastoma cells were destroyed in the presence of a more than tenfold excess of bone marrow cells following two incubations with monoclonal antibody 6-19 and complement for 30 min at 37 degrees C. Monoclonal antibody concentrations greater than 5-10 micrograms/ml did not increase cell lysis. The destruction of tumor cells was limited by depletion of complement activity. Tumor cell killing increased with complement concentration and incubation duration but there was no additional cell lysis with incubations greater than 30 min. The percentage of target cells killed was significantly decreased when the target cells were treated in the presence of increasing concentrations of bone marrow mononuclear cells. This decrease in target cell destruction is a result of additional depletion of complement activity by bone marrow mononuclear cells. These results suggest that optimal purging of tumor cells by antibody and complement will be achieved following multiple brief incubations with fresh antibody and complement.
Duerst
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RE
Department of Pediatrics, University of Rochester School of Medicine and Dentistry, New York.
Rose
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Frantz
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CN
D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D001853:Bone Marrow / Q000473:pathology*; D002470:Cell Survival; D003165:Complement System Proteins / Q000494:pharmacology*; D003601:Cytotoxicity Tests, Immunologic / Q000379:methods*; D003602:Cytotoxicity, Immunologic; D006801:Humans; D009447:Neuroblastoma / Q000473:pathology*; D014407:Tumor Cells, Cultured / Q000473:pathology
D000911:Antibodies, Monoclonal; D003165:Complement System Proteins