New insights in Type I and II CD20 antibody mechanisms‐of‐action with a panel of novel CD20 antibodies

Summary Based on their mechanisms‐of‐action, CD 20 monoclonal antibodies ( mA bs) are grouped into Type I [complement‐dependent cytotoxicity ( CDC ) and antibody‐dependent cell‐mediated cytotoxicity ( ADCC )] and Type II [programmed cell death ( PCD ) and ADCC ] mA bs. We generated 17 new hybridomas producing CD 20 mA bs of different isotypes and determined unique heavy and light chain sequence pairs for 13 of them. We studied their epitope binding, binding kinetics and structural properties and investigated their predictive value for effector functions, i.e. PCD , CDC and ADCC . Peptide mapping and CD 20 mutant screens revealed that 10 out of these 11 new mA bs have an overlapping epitope with the prototypic Type I mA b rituximab, albeit that distinct amino acids of the CD 20 molecule contributed differently. Binding kinetics did not correlate with the striking differences in CDC activity among the mIgG 2c mA bs. Interestingly, chimerization of mA b m1 resulted in a mA b displaying both Type I and II characteristics. PCD induction was lost upon introduction of a mutation in the framework of the heavy chain affecting the elbow angle, supporting that structural changes within this region can affect functional activities of CD 20 mA bs. Together, these new CD 20 mA bs provide further insights in the properties dictating the functional efficacy of CD20 mA bs.