Fig. 1: Generation of ART5803, a humanized monovalent antibody, and its binding characterization to the GluN1-NTD.

a The proposed mechanism of action for the pathogenic autoantibody (bivalent, two-armed, red) and therapeutic antibody (monovalent, one-armed, blue). Bivalent pathogenic anti-NMDAR autoantibodies bind to GluN1-NTD and crosslink adjacent NMDARs, which triggers receptor internalization. Monovalent therapeutic antibodies bind to GluN1-NTD, block binding of pathogenic autoantibodies, and prevent crosslinking and internalization of NMDARs. b ART5803 schematic design. Variable regions (antigen binding sites) are shown in blue. “Hole” mutations (T366S / L368A / Y407V) are introduced in the CH3 domain of the parental antibody heavy chain. A “Knob” mutation (T366W) was introduced in the CH3 domain of the second Fc fragment. LALA mutations (L234A / L235A) are introduced in the CH2 domains to avoid antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell-mediated phagocytosis (ADCP) or complement-dependent cytotoxicity (CDC). c Competitive ELISA between biotinylated #003-102 Ab and ART5803, #003-102 Ab and control Ab on GluN1-NTD. Data are mean ± SD of n = 2 internal replicates. Source data are provided as a Source Data file. d Epitope mapping by X-ray co-crystallography. Ribbon model of #003-102 Ab (Fab’) (red) on NMDAR (green). NTD terminal domain, LBD Ligand Binding Domain. e The zoomed-in view of the Fab binding site of #003-102 Ab (red) at the R2 lobe of GluN1-NTD (green) as determined by X-ray crystallography. Interacting residues for #003-102 Ab are from the Vh domain except where indicated with VK. #003-102 Ab residues are labeled with the Kabat numbering system. f Space filling model of GluN1-NTD of NMDAR (green) with #003-102 Ab (red) epitope by X-ray crystallography and #003-102 Ab (pink) and ART5803 (blue) epitopes as determined by HDX-MS.