Antibodies to Specific Citrullinated Proteins in Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory disease of synovial joints with a prevalence between 0.5% and 1% and leading to progressive disability if untreated. Studies of rheumatoid factor (RF) in the 1940s and 1950s, leading to the discovery of its antigenic target being the Fc region of immunoglobulin (Ig)G, led to RA being considered an autoimmune disease. However, the presence of RF is not unique to RA. Two immunofluorescent serologic assays, the antiperinuclear factor and antikeratin antibody tests, described in 1964 and 1979, respectively, did have high diagnostic specificity for RA but variable sensitivity. A key finding in the early 1990s was that the antigenic target of these two assays was filaggrin. The fact that RA sera were not reactive with recombinant filaggrin, or with the (pro)filaggrin, present in cultured buccal mucosal cells implied the need for a post-translational modification; this was identified in the late 1990s as the conversion of arginine residues into citrulline. Citrullination is mediated by peptidylarginine deiminase (PAD) enzymes, which are active in a number of physiologic situations, such as terminal epithelial cell differentiation, but also at sites of inflammation. A cyclic citrullinated peptide (CCP) of filaggrin was found to have high diagnostic sensitivity (68%) and specificity (98%); however, filaggrin does not occur in the joint, and it is widely assumed that the reactivity of RA sera with citrullinated filaggrin is due to cross-reactivity. An assay with improved diagnostic performance was derived by screening a library of citrulline-containing peptides for reactivity with RA sera to identify those yielding the highest sensitivity and specificity. Known as the anti-CCP2 enzyme-linked immunosorbent assay (ELISA), this is used in clinical immunology laboratories around the world and remains a “gold-standard” assay. Although the identity of the peptides it contains remains a commercial secret, it is thought they are nonphysiologic and do not occur in the human joint where production of anticitrullinated protein/peptide antibodies (ACPA) has been demonstrated to occur. Therefore, despite the usefulness of the anti-CCP2 ELISA, both in the diagnosis and prognosis of RA, it provides no insight into the real in vivo antigenic targets of ACPA.