Cross‐reactivity of a human IgG₁ anticitrullinated fibrinogen monoclonal antibody to a citrullinated profilaggrin peptide

Abstract

Rheumatoid arthritis (RA) is the most common autoimmune rheumatic disease. It is characterized by persistent joint inflammation, resulting in loss of joint function, morbidity and premature mortality. The presence of antibodies against citrullinated proteins is a characteristic feature of RA and up to 70% of RA patients are anticitrullinated protein antibody (ACPA) positive. ACPA responses have been widely studied and are suggested to be heterogeneous, favoring antibody cross-reactivity to citrullinated proteins. In this study, we examined factors that may influence cross-reactivity between a commercial human anticitrullinated fibrinogen monoclonal antibody and a citrullinated peptide. Using a citrullinated profilaggrin sequence (HQCHQESTCit-GRSRGRCGRSGS) as template, cyclic and linear truncated peptide versions were tested for reactivity to the monoclonal antibody. Factors such as structure, peptide length and flanking amino acids were found to have a notable impact on antibody cross-reactivity. The results achieved contribute to the understanding of the interactions between citrullinated peptides and ACPA, which may aid in the development of improved diagnostics of ACPA.

Physical Characteristics of a Citrullinated Pro-Filaggrin Epitope Recognized by Anti-Citrullinated Protein Antibodies in Rheumatoid Arthritis Sera

“…Identifying a reactive region in amino acid residues 228–245, a cyclic and a linear peptide version were generated, however the two peptides showed similar characteristics [39]. These findings are in accordance to studies by Trier et al , describing the cross-reactivity of a human IgG 1 anti-citrullinated fibrinogen monoclonal antibody to cyclic and linear CCP1-derived truncated peptides [41]. …”

“…Thorough analysis of citrullinated epitopes reveal that peptide backbone primarily in combination with a Cit-Gly motif is essential for reactivity, although occasionally other amino acids are tolerated C-terminal to Cit [41]. Screenings of several epitopes prior to this study indicated that especially charged amino acids C-terminal to Cit are essential for antibody reactivity (data not shown) [41].…”

“…Previous studies describing reactivity of a human monoclonal antibody to citrullinated pro-filaggrin peptides indicated that antibody reactivity to these peptides is dependent on peptide conformation and peptide length [41]. To determine whether these results relate to RA patient sera as well, the reactivity of RA sera to cyclic and linear truncated peptides were analysed by streptavidin capture ELISA.…”

“…Cyclization of peptides was performed by traditional air oxidation. Following oxidation, all peptides were purified by reverse-phase high-performance liquid chromatography and peptide identity was confirmed with liquid chromatography-mass spectrometry as previously described and illustrated [41]. The peptides applied are listed in Table 1.…”

Anti-citrullinated protein antibodies cause arthritis by cross-reactivity to joint cartilage

“…The reactivity of ACPA is found to be governed by the conformational flexibility of the antigenic region and the amino acid residues surrounding the citrulline. The presence of a neutral amino acid with no or a small side chain (glycine and serine) on the N-terminal site of the citrulline often favors recognition as compared with rigid residues (proline) at this position (33). In agreement with these observations, there is a glycine residue next to the citrulline in the "RG-TG(CIT)" motif preferred by ACC1.…”