Preparation and properties of a drug‐carrier‐antibody conjugate showing selective antibody‐directed cytotoxicity in vitro

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Abstract
The preparation and properties of a drug‐carrier‐antibody preparation are reported. The antifolate chemotherapeutic agent methotrexate was covalently coupled to human serum albumin as a carrier. The carrier‐drug preparation was then chemically linked to a monoclonal antibody, raised originally against a human osteogenic sarcoma cell line, 79IT, in a manner permitting retention of antibody‐binding activity. The cytotoxic properties of the conjugate were tested in vitro in comparison with carrier‐methotrexate and free methotrexate against a panel of tumour cell lines containing both antigenically cross‐reactive cell lines and cell lines having low antigenic cross‐reactivity with the monoclonal antibody. The cytotoxicity tests demonstrated that coupling of methotrexate to carrier caused a loss of some drug activity but that coupling of the antibody to the carrier‐drug preparation permitted full expression of drug cytotoxicity against antibody‐reactive cell lines. It was further demonstrated that the conjugate was selective in its action and was preferentially cytotoxic towards antibody‐reactive cell types. The cytotoxicity against antibody‐reactive cell lines was shown by competitive inhibition by free antibody to be entirely dependent on antibody binding. A clonogenic assay showed that the conjugate was capable of killing >99% of 79IT target cells. These results indicate that a drug‐carrier antibody conjugate can be synthesized which has all the in vitro properties theoretically necessary for a successful antibody‐targeted cytotoxic agent.