The TNFa-binding domain of the therapeutic antibody adalimumab elicits CD4 T-cell responses in Rheumatoid Arthritis patients

Mateusz Makuch¹Josine Van Beek^1,2Carla A Wijbrandts³Marja Aalbers¹Philippe Stas⁴Alexander Meijer⁵Anja ten Brinke¹T Rispens¹Paul Peter Tak^6,7Gertjan Wolbink^1,3Janine Schuurman⁸Paul W.H.I. Parren^10,9S. Marieke Van Ham^1,11*

• ¹Department of Immunopathology, Sanquin Research, Amsterdam, Netherlands
• ²Centre for Immunology of Infectious Diseases and Vaccines, National Institute for Public Health and the Environment (RIVM), Bilthoven, Netherlands
• ³Department of Rheumatology, Reade, Amsterdam, Netherlands
• ⁴ImmunXperts SA, Gosselies, Belgium
• ⁵Department of Plasma Proteins, Van Creveld Laboratory of UMC Utrecht and Sanquin Research, Amsterdam, Netherlands
• ⁶Department of Rheumatology, Amsterdam Rheumatology and Immunology Center (ARC), Amsterdam, Netherlands
• ⁷Candel Therapeutics, Needham, Massachusetts, United States
• ⁸Genmab (Netherlands), Utrecht, Netherlands, Netherlands
• ⁹Department of Immunology, Leiden University Medical Center (LUMC), Leiden, Netherlands
• ¹⁰Lava Therapeutics, Utrecht, Netherlands
• ¹¹Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, Gelderland, Netherlands

Treatment efficacy of patients receiving anti-TNF antibodies is limited by the formation of anti-drug antibodies. These are observed in most adalimumab-treated rheumatoid arthritis patients, despite the adjuvant-free and human sequence-derived nature of the antibody. The class switched phenotype and high affinity of these antibodies suggest CD4 T-cell involvement in their formation. In this study, we investigated the potential epitopes in the functional domain of adalimumab and assessed their actual HLA II presentation and elicitation induction of CD4 T-cell responses in exposed patients. The binding strength of overlapping adalimumab-derived peptides to 27 DR and 14 DQ HLA alleles was predicted in silico. 10 strong and 44 medium-binding 10-mer peptides were identified within the variable regions of the heavy and light chain of adalimumab. HLA-DR-mediated antigen presentation of selected peptides by monocyte-derived dendritic cells was determined by mass spectrometry of the peptide pool eluted from isolated HLA-DR complexes. Binding of the variable region peptides of heavy (H41-62) and light chains (L18-39) was demonstrated. The presence of actual adalimumab-specific CD4 T-cells in adalimumab-experienced patients was investigated via peptide stimulation of peripheral blood mononuclear cells and assessment of T-cell proliferation assays. Anti-adalimumab CD4 T-cell responses were observed against four variable region peptides in a group of adalimumab-experienced RA patients. Some of these responses were also present in healthy control donors. This is the first study that identifies immunologically relevant CD4 T-cell epitopes in the variable region of the human therapeutic antibody adalimumab based on RA patients' reactivity. Modification of these epitopes or concomitant therapy that targets or prevents adalimumab-specific T cell responses could be beneficial for patients with significant anti-drug responses.