IMMUNOTOXIN SENSITIVITY OF CHINESE-HAMSTER OVARY CELLS EXPRESSING HUMAN TRANSFERRIN RECEPTORS WITH DIFFERING INTERNALIZATION RATES

Previous studies have shown that immunotoxin action is dependent upon selective binding to the target cell, internalization and then passage into the cytosol. It is important to define precisely how these criti cal steps are controlled so that the underlying relationship of each t o high cytotoxic effectiveness is understood. In order to evaluate the contribution of internalization rate and receptor number on immunotox in potency, the effects of an anti (transferrin receptor, TfR)/ricin A chain immunotoxin, 7D3-A, were assessed on a parent Chinese hamster o vary cell line developed in our laboratory with no TfR (TfR(neg)) and two lines transfected with either wild-type TfR (Tfr(wt)) or an intern alization-deficient (TfR(Delta 7-58del)) mutated human TfR. Potent, re ceptor-mediated cytotoxicity resulted from the action of 7D3-A on TfR( wt) cells (ID50 < 1 nM) while both TfR(neg) cells and TfR(Delta 7-58de l) were only minimally affected (ID50 > 100 nM). Butyrate up-regulatio n substantially increased receptor expression on the TfR(wt) and TfR(D elta 7-58del) cells, but no corresponding rise in sensitivity to 7D3-A was observed. In contrast, immunotoxin potency was increased by co-tr eatment of TfR(wt) cells with the carboxylic ionophore monensin and th e effect was even more pronounced for TfR(Delta 7-58del) cells. We con clude that internalization rate or intracellular destination is a much more important determinant of immunotoxin efficacy than receptor numb er.