A Tolerogenic Plasmid Inhibits Autoantibody Production Triggered by Immunization With Human Peptidyl Arginine Deiminase 4.

Abstract

Rheumatoid arthritis (RA) is usually preceded and likely mediated by autoantibodies to citrullinated proteins (ACPAs) that recognize citrulline residues on multiple proteins. Conversion of arginine into citrulline is performed by enzymes called peptidyl arginine deiminases (PADs). We have previously demonstrated that the human PAD4 (hPAD4) is a T cell target whose recognition provides help for the production of ACPAs by a hapten/carrier mechanism. Human PAD4 is thus an attractive therapeutic target to stop ACPA production. Here, we tested whether tolerization to hPAD4 may block the production of anti-citrullinated peptide antibodies. We used a tolerogenic plasmid encoding hPAD4 and transforming growth factor (TGF) β1, interleukin (IL) 10, and IL-2 in a model where C3H mice immunized with hPAD4 develop anti-citrullinated peptide antibodies. As controls, we used a plasmid encoding TGF-β1, IL-10, IL-2, or a plasmid without hPAD4 or cytokines. T cell activation in hPAD4-immunized mice injected with plasmids was analyzed by flow cytometry. Autoantibody profile was analyzed in hPAD4-immunized mice before and after plasmid injection by peptide array using 33 arginine-containing peptides and their 136 citrulline-substituted variants. Injection of the plasmid encoding hPAD4 and TGF-β1, IL-10, and IL-2 in hPAD4-immunized C3H mice significantly inhibited the T cell activation to hPAD4 and decreased antibody production to arginine and citrullinated peptides. Tolerizing to hPAD4 decreases the production of anti-citrullinated peptide antibodies in mice immunized with hPAD4. This suggests that tolerization to hPAD4 may allow one to switch off ACPAs in RA.