HUMANIZATION AND PHARMACOKINETICS OF A MONOCLONAL-ANTIBODY WITH SPECIFICITY FOR BOTH E-SELECTIN AND P-SELECTIN

E- and P-selectin (CD62E and CD62P) are cell adhesion molecules that m ediate leukocyte-endothelial cell and leukocyte-platelet interactions and are involved in leukocyte recruitment during inflammation. We prev iously developed a murine mAb, EP-5C7 (Or mEP-5C7), that binds and blo cks both E- and P-selectin. When used in humans, murine mAbs have shor t circulating half-lives and generally induce potent human anti-mouse Ab responses. We therefore engineered a humanized, complementarity det ermining region-grafted version of mEP-5C7 incorporating human gamma 4 heavy and kappa light chain constant regions (HuEP5C7.g4). HuEP5C7.g4 retains the specificity and avidity of mEP-5C7, binding to human E-an d P-selectin but not to human L-selectin, and blocking E-and P-selecti n-mediated adhesion. Surprisingly, when administered to rhesus monkeys , HuEP5C7.g4 was eliminated from the circulation very rapidly, even fa ster than the original murine Ah. To isolate the cause of the short se rum half-life of HuEP5C7.g4, several Ab variants were constructed. A c himeric IgG4 Ab was made by replacing the humanized V regions with mur ine V regions. A humanized IgG2 Ab, HuEP5C7.g2, was also made by repla cing the human gamma 4 with a gamma 2 constant region. Results from ph armacokinetic studies in rhesus monkeys demonstrated that the chimeric IgG4 is also rapidly eliminated rapidly from serum, similar to the hu manized IgG4 Ab, while the humanized IgG2 Ab displays a long circulati on half-life, typical of human Abs.