Abstact

Dysregulation of the CD40-CD40L axis is implicated in autoimmune diseases. Early clinical trials targeting CD40L with antibodies failed due to Fc-mediated side effects. To address this, we developed an anti-CD40L Fab fragment, Fab20, designed to block B-cell activation. Fab20 was evaluated for its binding properties, CD40-CD40L inhibition, and effects on human B-cell activation and differentiation using immunoassays, cryo-electron microscopy, flow cytometry, and cell cultures. Fab20 binds CD40L with a dissociation constant of 70 nM. Structural analysis revealed a "propeller-like" structure consisting of three Fabs binding to the CD40L trimer, sterically blocking parts of the CD40 binding site. Fab20 effectively inhibited B-cell activation, maintaining naive B cells in their inactive state, and suppressed antibody (IgG) production over 14 days. Fab20 represents a promising novel therapeutic approach for treating autoimmune diseases driven by CD40-CD40L dysregulation. Its mechanism of action, coupled with the absence of Fc-mediated effects, suggests a favorable safety profile.