In vitro characterization of five humanized OKT3 effector function variantantibodies

Orthoclone OKT 3 (mOKT3) is a highly effective agent for the reversal of st eroid-resistant renal allograft rejection. However, its wider use has been limited by the development of a human anti-mouse antibody response (HAMA) a nd by the "cytokine release syndrome" (CRS). CRS has been associated with T cell/monocyte activation and, secondarily, with activation of the compleme nt cascade. These processes are mediated through Abs' Fc regions by their a bilities to cross-link T cells and mononuclear cells and to activate comple ments. To alleviate these problems, a group of five huIgG1- and huIgG4-base d OKT3 wildtype antibodies and their corresponding Fc mutants with altered residues at amino acids 234, 235, and 318, reported to be required for Fc g amma RI and Fc gamma RII binding and complement fixation, were constructed. Characterization of these humanized OKT3 Abs, denoted huOKT3 gamma 1, huOK T3 gamma 4, huOKT3 gamma 1(A(234), A(235)), huOKT3 gamma 4(A(234), A(235)) and huOKT3 gamma 1(A(318)), has demonstrated that huOKT3 gamma 1(A(234), A( 235)) and huOKT3y4(A(234), A(235)) and have at least a 100-fold reduced bin ding to Fc gamma RI and Fc gamma RII. As expected, they are much less poten t in the induction of T cell activation and cytokine release, yet retain in vitro immunosuppressive effects as potent as those of mOKT3. Unexpectedly, while huORT3 gamma 1(A(318)) did not show any reduction in its ability to bind C1q and to fix a complement huOKT3 gamma 1(A(234), A(235)) was complet ely inactive. The in vitro characteristics of huOKT3 gamma 1(A(234), A(235) ) are consistent with recent in vivo studies, in which this Ab showed great ly reduced HABU and CRS with the retention of its ability to reverse ongoin g graft rejection in man. (C) 2000 Academic Press.