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Radioimmune Assay and Characteristics of Antibodies to Macromomycin (NSC 170105)¹

Department of Microbiology, The Medical College of Wisconsin, Milwaukee, Wisconsin 53233

A radioimmune assay for the antitumor agent, macromomycin, using purified, radioiodine-labeled macromomycin and antisera raised in rabbits against a carbodiimide-catalyzed macromycin-Limulus polyphemus hemocyanin complex has been developed. Radiolabeled macromomycin was prepared by direct iodination of the polypeptide antibiotic with the use of iodine monochloride or solid-state lactoper-oxidase. Antibody-bound drug was isolated from free macromomycin with dextran-coated, activated charcoal. The standard curve of the sequential saturation assay was linear on a logit-log plot and indicated a lower limit of sensitivity of approximately 100 pg macromomycin. The radioimmune assay was suitable for measuring macromomycin in the presence of other antitumor drugs, and detection of macromomycin was quantitative when it was added to normal human serum or urine. Drug binding to melanoma and mammary carcinoma cell surfaces could be inhibited by preincubating macromomycin with affinity-purified antimacromomycin antibodies. However, once the drug was bound to cell surfaces, addition of antimacromomycin antibodies did not result in removal of the drug from cell surfaces or in reversal of macromomycin-induced inhibition of thymidine incorporation into cellular DNA. Antimacromomycin antibodies and the radioimmune assay should provide useful tools for developing pharmacokinetic and toxicity studies of macromomycin, as well as for analyzing the mechanism(s) of action of the drug.

¹ This work was supported by grants from The Jane Coffin Childs Memorial Fund for Medical Research and The American Cancer Society, Milwaukee Division, Inc.

Received 11/15/76. Accepted 12/30/76.