Protein a vectorized toxins—II. Preparation and “ in vitro” cytotoxic effect of protein A-ricin a chain conjugate on antibody coated human tumour cells

Protein A of Staphylococcus aureus was covalently bound to reduced ricin A chain toxin by N-succinimidyl 3-(2-pyridyldithio)propionate. The conjugate consisting mainly of one molecule of protein A bound to two molecules of A chains ( M r 107,000) was purified by tandem affinity chromatography on ConA-Sepharose 4B and IgG-Sepharose 4B. The purified protein A-A chain conjugate was able to bind and kill human lymphoma cells coated either with monoclonal mouse IgG2a anti-kappa antibody or with polyclonal rabbit anti-kappa antibody. The cytotoxic activity of protein A-A chain conjugate in conjunction with either mouse or rabbit anti-kappa antibodies was 10 times higher than that of rabbit IgG anti-mouse IgG coupled with A chain on Daudi cells coated with mouse anti-kappa antibody and 100 times higher than that of rabbit anti-kappa antibody coupled with A chain on non-coated Daudi cells. The cytotoxic effect of protein A-A chain conjugate on antibody-coated Daudi cells (9 × 10 −12 M) was comparable with that of ricin toxin on non-coated Daudi cells (2 × 10 −12 M). The results recommend the use of protein A-ricin A chain toxin conjugate as a unique specific toxin for the “ in vitro” killing of antibody-coated target cells.