Anti-GM2 monoclonal antibodies induce necrosis in GM2-rich cultures of a human glioma cell line

Abstract

The effects of four anti-GM2 monoclonal antibodies (DMAb-1, DMAb-2, DMAb-3, and DMAb-5) were studied on spheroid cultures from a human glioma cell line (D-54 MG) that is known to express high levels of GM2. The spheroids developed central necrosis 48 h after antibody exposures at concentrations greater than 6 micrograms/ml. No necrosis was found with antibodies that had been absorbed with GM2 prior to exposure or with unrelated cytotoxic antibodies. Immunohistochemistry showed that the necrosis started shortly after the antibodies were evenly distributed throughout the spheroids. Light and transmission electron microscopy revealed that a small portion of the cells, mainly in the periphery of the spheroids, was unaffected by antibody exposure. New monolayer cultures established from antibody-treated cells expressed a 50% lower GM2 content as shown by flow cytometry and determination of ganglioside content throughout at least 12 passages. Thus, the GM2-rich D-54 MG cell line has subpopulations of cells with lower GM2 content. Spheroids obtained from this subpopulation developed only minor necrosis after antibody treatment. These results show that GM2 antibodies cause severe necrosis of GM2-containing glioma cells in vitro, but the effect depends on the concentration of antigen, and a threshold number of GM2 molecules is required.