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Affinity Cytotoxicity of Tumor Cells with Antibody-Glucose Oxidase Conjugates, Peroxidase, and Arsphenamine¹

Departments of Surgery [G. W. P., R. J. B.] and Medicine [K. L. P., W. T. S., C. W. P.], Washington University School of Medicine, St. Louis, Missouri 63110

Selective cytotoxicity was accomplished in two antibody-tumor cell models with antibody-glucose oxidase conjugates (Ab-GO) followed by treatment with horseradish peroxidase (HRP) and arsphenamine. The two cell models used were hapten [2,4,6-trinitrophenyl (TNP)]-substituted HeLa and HEp-2 cells with specifically purified antihapten (TNP) antibody and human colonic carcinoma cells (HT-29) with immunoglobulin G anti-carcinoembryonic antigen antibody. Brief treatment of TNP cells with anti-TNP antibody conjugated to glucose oxidase (0.07 to 70 µg/ml) followed by culture in medium with HRP (50 µg/ml) and arsphenamine (1 to 10 µg/ml) resulted in zero to 100% cell killing when compared with controls in a microcytotoxicity assay. Cytotoxicity was reduced or absent when (a) any of the three components (Ab-GO, HRP, or arsphenamine) were omitted; (b) cells not substituted with TNP were used; or (c) free hapten (dinitrophenyl-lysine) inhibited Ab-GO binding to TNP cells. Affinity cytotoxicity (73 to 90%) was also seen in HT-29 cells treated with anti-CEA antibody conjugated to glucose oxidase followed by treatment in HRP and arsphenamine (1 to 10 µg/ml). CEA, extracted from malignant ascitic fluid with perchloric acid, partially inhibited the cytotoxic action of the Ab-GO system, and normal goat immunoglobulin G-glucose oxidase caused significantly less killing, showing the selectivity of the reaction.

¹ Supported by Grant 5R01-CA12626 from the National Cancer Institute.

Received 3/ 7/74. Accepted 5/ 6/74.