Studies on the Mechanism of Antibody-Dependent Polymorphonuclear Leukocyte-Mediated Cytotoxicity

Abstract

Human polymorphonuclear leukocytes (PMN) were cytotoxic to mammalian tumor cells (LSTRA) in the presence of anti-LSTRA serum as determined by a ⁵¹Cr release assay. Cytotoxicity was observed with effector to target cell ratios as low as 0.25:1 and with incubation periods as short as 30 min. Under the conditions employed, human blood mononuclear cells had only minimal cytotoxic activity. Immunologic specificity was demonstrated by 1) the loss of activity when immune serum was preabsorbed with target cells or replaced with normal (preimmune) serum and 2) the lack of ⁵¹Cr release from an unrelated target cell. Viable PMN were required as demonstrated by the loss of cytotoxicity on heating or homogenization. Further, an intact PMN metabolic burst was essential for maximal activity since PMN from patients with chronic granulomatous disease were only minimally active. The myeloperoxidase-H₂O₂-halide system of the PMN was not responsible for cytotoxicity since activity was normal with myeloperoxidase deficient PMN, was preserved in the presence of azide, cyanide or catalase, and was only slightly dependent on halides. Lack of inhibition by heparin militated against a role for PMN granular cationic proteins. Cytotoxicity was blocked by inhibitors of glycolysis, but not by an inhibitor of oxidative phosphorylation. The use of chelating agents indicated a requirement for divalent actions, primarily Mg⁺⁺. Inhibition of cytotoxicity by the anti-tubulins, colchicine, and vincristine, suggested a role for microtubules. These studies suggest that antibody-dependent PMN-mediated tumor cell cytotoxicity requires specific antibody, viable PMN, a burst of PMN oxidative metabolism, glycolysis, divalent cations, and microtubular function. The PMN lysosomal constituents, myeloperoxidase and the cationic proteins, did not appear to be involved under the conditions employed.