Selective macrophage activation by muramyldipeptide bound to monoclonal antibodies specific for mouse tumor cells

Summary IgM monoclonal antibodies directed against tumor cells which do not mediate antibody-dependent macrophage cytotoxicity (ADMC) even when they are cytotoxic in the presence of complement, have been shown to render macrophages tumoricidal when they carry an immunomodulating agent, i.e., muramyldipeptide (MDP).This statement is based on experiments using two IgM monoclonal antibodies selected for their ability to bind L1210 leukemia cells (F₂-10-23-IgM) and 3LL Lewis lung carcinoma cells (6B6-IgM) specifically, as shown by flow cytofluorometry analysis.The MDP-IgM conjugates, containing 45 MDP molecules per IgM molecule, were prepared by allowing MDP-hydroxy-succinimide ester to react with IgM monoclonal antibodies.The MDP-IgM conjugates are shown to bind to relevant tumor cells and to induce the activation of thioglycolate-elicited peritoneal mouse macrophages leading to 80% growth inhibition of target cells at optimum concentrations of bound MDP. These concentrations of bound MDP were 10 times lower than the concentration of free MDP, giving a maximum activation that is limited to 20% growth inhibition.No macrophage activation was evidenced when tumor cells were incubated in the presence of irrelevant MDP-IgM conjugates and macrophages or when macrophages were preincubated in the presence of MDP-IgM conjugates and then incubated in the presence of relevant or irrelevant tumor cells but in the absence of the MDP-IgM conjugates.The reported results are discussed with reference to the mechanism of activation of macrophage by muramyldipeptide and to the usefulness of such MDP-IgM conjugates as potential antitumor agents in cancer therapy.Abbreviations ADMC antibody dependent macrophage mediated cytotoxicity - F-GAM fluoresceinylthiocarbamyl goat anti mouse antibody - -Man-BSA -mannopyranosyl-phenylthiocarbamyl bovine serum albumin containing some 25 mannose residues (neoglycoprotein) - MDP muramyldipeptide, 2-acetamido-3(2-0-d-lactyl-l-alanyl-d-glutamyl amine) glucopyranose - MDP-F₂-10-23-IgM Murine monoclonal antibody specific of L1210 leukemia cells and substituted with 45 MDP molecules - MDP—6B6-IgM Murine monoclonal antibody specific of 3LL Lewis lung carcinoma cells and substituted with 45 MDP molecules - MEM minimal essential medium - TDM Trehalose-dimycolate