Studies on the mechanism of action of an antibody-targetted drug-carrier conjugate.
A conjugate of methotrexate-substituted human serum albumin (HSA) coupled to a monoclonal antibody (791T/36) recognizing osteogenic sarcoma cell lines has been reported previously to have good cytotoxicity and specificity in vitro (Garnett et al., 1983). Cytotoxicity was assessed by the median inhibition (IC50) of [75Se]selenomethionine uptake resulting from a 24-h incubation of conjugate with antigen-bearing 788T or 791T osteogenic sarcoma cell lines. In the present work, the properties of this conjugate have been investigated to determine whether cytotoxicity is optimal with respect to binding activity, and aspects of the mechanism of action investigated by the effects of specific inhibitors of biochemical processes on conjugate cytotoxicity. Conjugate cytotoxicity was reduced by ammonium chloride suggesting that endocytosis and an acidic internal compartment were involved in the mechanism of action. The specific inhibitors of proteinases leupeptin and E64 also reduced conjugate cytotoxicity, while the inhibitors pepstatin A and chymostatin had no effect, demonstrating that cysteine proteinases were involved. The inhibitor of methotrexate transport, folinic acid, reduced the cytotoxicity of conjugate more than that of free methotrexate whereas folic acid had no effect on either, indicating that the methotrexate transport system may still be involved but in a different manner to the free drug. The cytotoxicity of these conjugates is probably near optimal for maximum selectivity.
D000911:Antibodies, Monoclonal / Q000008:administration & dosage*; D001692:Biological Transport; D002470:Cell Survival / Q000187:drug effects; D000922:Immunotoxins; D002955:Leucovorin / Q000494:pharmacology; D008727:Methotrexate / Q000008:administration & dosage* / Q000493:pharmacokinetics / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D012709:Serum Albumin / Q000008:administration & dosage*; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D000922:Immunotoxins; D011480:Protease Inhibitors; D012709:Serum Albumin; D002955:Leucovorin; D008727:Methotrexate