HUMANIZED IGG1 AND IGG4 ANTI-CD4 MONOCLONAL-ANTIBODIES - EFFECTS ON LYMPHOCYTES IN THE BLOOD, LYMPH-NODES, AND RENAL-ALLOGRAFTS IN CYNOMOLGUS MONKEYS

Background. Optimizing therapeutic monoclonal antibody (mAb) depends o n the incorporation of the necessary effector functions and the develo pment of hypoantigenic ''humanized'' antibodies by genetic engineering , which then need to be tested in appropriate preclinical trials, Meth ods. Constructs of humanized OKT4A containing the complementarity-dete rmining region (CDR) of murine OKT4A and the framework and constant re gions of human light (kappa) and heavy chains (IgG1 and IgG4) were pre pared and tested in cynomolgus monkeys who received a renal allograft, A prophylactic course of CDR-OKT4A/human (h) IgG1 or CDR-OKT4A/hIgG4, either as high-dose single bolus (10 mg/kg) or as low-dose multiple i nfusion (1 mg/kg for 12 days) was given, and the effects on graft surv ival, immunohistology, circulating cells, and lymph node cells were as sessed, Results, The IgG1 isotype induced coating of T cells, modulati on of surface CD4 molecules, and profound depletion of CD4(+) lymphocy tes in peripheral blood, which persisted as long as the animals were f ollowed (up to 7 weeks). The IgG4 isotype induced only cell coating wi thout cell clearance or modulation. Iq lymph nodes, coating of lymphoc ytes (approximately 60%) was seen with both isotypes in the earliest s ample (6 hr), After 2 days, significant depletion of lymph node CD4 ce lls was evident, with a decrease in the CD4 to CD8 ratio in the IgG1-t reated group; no depletion occurred in the IgG4 group, The emigration of CD4(+) cells into the allograft was significantly delayed in the CD R-OKT4A/hIgG1-treated animals when compared with the CDR-OKT4A/hIgG4 g roup as judged by immunocytochemistry (23.8+/-13.2 days vs, 7.4+/-1.5 days, P<0.001) or interleukin-2-promoted T-cell outgrowth from allogra ft biopsies (22.2+/-11.0 days vs. 6.3+/-0.5 days, P<0.01). Conclusions , This study demonstrates that the in vivo effects of CDR-grafted OKT4 A are dependent on its isotype, The depleting mAb CDR-OKT4A/hIgG1 sign ificantly delays the entry of CD4(+) cells into the graft, inhibiting the early phase of rejection, However, graft rejection occurs when CD4 (+) cells eventually infiltrate the graft, even in the presence of dep ressed levels of circulating CD4(+) cells. Both isotypes demonstrated therapeutic efficacy: graft survival was prolonged over controls. In t he case of CDR-OKT4A/hIgG4, neither lymphocyte depletion, antigenic mo dulation, nor prevention of infiltration is necessary for a beneficial effect, which indicates that this mAb blocks CD4 function or renders the CD4(+) cell less responsive, The lack of depletion is a feature of potential clinical advantage in minimizing the risk of excessive immu nosuppression.