A novel anti-OX40 human monoclonal antibody that blocks OX40/OX40L signaling and depletes OX40(+) T cells.

Tumor necrosis factor receptor superfamily member 4 (TNFRSF4), also known as OX40, plays a crucial role in the regulation of T-cell immune responses under normal physiological conditions. Abnormal expression of OX40 and its cognate ligand OX40L (TNFSF4) have been associated with various autoimmune diseases, indicating that blocking the OX40/OX40L pathway could be a promising strategy for the treatment of a broad range of T cell-mediated autoimmune diseases. Here, we screened and characterized a fully human anti-OX40 antibody (JY007) from a naïve human scFv phage library. JY007 has an affinity constant of 7.71 nmol/L and effectively inhibited the OX40-OX40L interaction at both molecular and cellular levels, with IC(50) values of 1.088 and 10.12 nmol/L, respectively. Furthermore, JY007 demonstrated the ability to deplete activated T lymphocytes through antibody-dependent cellular cytotoxicity (ADCC) activity, with an EC(50) of 5.592 pmol/L. The combination of ADCC and its antagonist activity against OX40 suggests potential efficacy in suppressing inflammatory responses mediated by the OX40/OX40L pathway. Additionally, we employed molecular docking, site-directed mutagenesis, and competitive ELISA to pinpoint the epitopes on OX40. The results revealed that JY007 binds to Pro(37), Ser(38), and Asp(40) of OX40. Interestingly, we also found that the most potent anti-OX40 antibody drug in the clinical stage, KHK4083, binds to different OX40 amino-acid residues, including Asp(74), Lys(82), Asp(117), Ser(118), Tyr(119), and Lys(120). This divergence suggests that the novel monoclonal antibody JY007 holds promise as a potential therapeutic option for patients with atopic dermatitis and may find broad applications in the treatment of autoimmune diseases. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s42995-025-00284-y.