CD6 monoclonal antibodies differ in epitope, kinetics and mechanism of action

摘要

Summary CD 6 is a type I T‐cell surface receptor that modulates antigen receptor signalling. Its activity is regulated by binding of its membrane proximal domain (domain 3) to a cell surface ligand, CD 166. CD 6 monoclonal antibodies ( mA bs) specific for the membrane distal domain (domain 1) perturb CD 6 function including itolizumab (Alzumab?), which has reached the clinic for treatment of autoimmune disease. We characterized molecular and functional properties of several CD 6 mA bs including itolizumab to define potential mechanisms of action. Epitope mapping using the crystal structure of CD 6 to design mutants identified two distinct binding sites on different faces of domain 1, one containing residue R77, crucial for MT 605 and T12.1 binding and the other, E63, which is crucial for itolizumab and MEM 98. Analysis of binding kinetics revealed that itolizumab has a lower affinity compared with other CD 6 domain 1 mA bs. We compared potential agonistic (triggering) and antagonistic (blocking) properties of CD 6 mA bs in assays where the mechanism of action was well defined. CD 6 domain 1 and 3 mA bs were equally effective in triggering interleukin‐2 production by a cell line expressing a chimeric antigen receptor containing the extracellular region of CD 6. CD 6 domain 1 mA bs hindered binding of multivalent immobilized CD 166 but were inferior compared with blocking by soluble CD 166 or a CD 6 domain 3 mA b. Characterization of CD 6 mA bs provides an insight into how their functional effects in vivo may be interpreted and their therapeutic use optimized.