Anti-tumour activity of aminopterin-monoclonal antibody conjugates; in vitro and in vivo comparison with methotrexate-monoclonal antibody conjugates.

Two related folate antagonists aminopterin (AMN) and methotrexate (MTX) were used to produce drug-antibody conjugates and their tumouricidal effects compared in vitro and in vivo. Active ester derivatives were produced with the use of N-hydroxysuccinimide (NHS) and covalently coupled to monoclonal antibodies (MoAb) reactive with murine cell surface antigens; approximately 11 molecules of AMN or 13 molecules of MTX were specifically bound per molecule of anti-Ly-2.1, with good retention of antibody activity and protein recovery. AMN was a more effective inhibitor of tumour cell growth in vitro than MTX, and AMN-anti-Ly-2.1 conjugates were also more potent in vitro than MTX-anti-Ly-2.1 conjugates. Although there was some loss of drug activity on binding to antibody, AMN-MoAb conjugates were as toxic as free MTX. However, in contrast to free drugs (which can act on any target), the toxicity of drug-MoAb conjugates was entirely specific for the target cells. In addition, AMN-MoAb conjugates were effective anti-tumour agents in vivo, and in mice bearing established thymoma grafts AMN-MoAb conjugates inhibited tumour proliferation better than MTX-MoAb, free AMN or MTX or antibody alone. AMN coupled to specific MoAb is a potentially useful agent for immunotherapy and is of particular relevance in man as free AMN has been discarded because of its systemic toxicity. Now, coupled with antibody, there will be specific tumouricidal effects in the absence of toxicity.