Synthetic Anti-CCP Antibody Aggravated Severity of Animal Arthritis and Captured Citrullinated Antigen in the Serum of Patients with Rheumatoid Arthritis

Youngkyun Kim¹, Su-Jin Moon², Hyoju Yi² and Ji Hyeon Ju³, ¹1Convergent Research Consortium for Immunologic Disease, The Catholic University of Korea, Seoul, South Korea, ²Rheumatology, College of Medicine, The Catholic University of Korea, Seoul, South Korea, ³Rheumatology, College of Medicine, The Catholic University of Korea & Stanford University, Seoul & Palo Alto, CA

Meeting: 2012 ACR/ARHP Annual Meeting

Keywords: anti-CCP antibodies, citrulline and rheumatoid arthritis (RA)

Session Information

Title: Rheumatoid Arthritis - Human Etiology and Pathogenisis

Session Type: Abstract Submissions (ACR)

Background/Purpose: Auto-antibodies against citrullinated protein antigens (ACPA) have shown their relevance for the prognosis and diagnosis of rheumatoid arthritis (RA), and have been implicated in disease pathogenesis. However, the exact pathologic relevance of these antibodies remains unclear. To obtain insight into this question, we tried to synthesize ACPA and then apply it to animal model and human samples.

Methods: Using hybridoma technology, an IgG monoclonal antibody (mAb), 12G1 was developed against a cyclic-citrullinated peptide (CCP). Clones were selected on the basis of not reacting with control peptide (CRP) which was included one arginine instead of citrulline on the same peptide sequence and structure, but reacting well with synthetic CCP. After successful establishment of hybridoma for anti-CCP Ab, we analyzed the characteristics of this synthetic anti-CCP Ab. 12G1 was challenged to collagen induced arthritis (CIA) model. Western blot with 12G1 was done on the serum samples of RA patients (n=25) and non-RA control (n=23). Synovium of CIA and RA patients were immunostained with 12G1.

Results: Immunohistochmical analysis with 12G1 revealed positive staining result in CIA and RA patients in comparison with control samples of wild type mice and osteoarthritis. Western blot analysis of sera from RA patients had stronger 12G1-positive bands of citrullinated protein compared with non-RA controls. Average band density was four times higher in RA than non-RA (p=0.001). These western results showed the meaningful correlation with 2^nd generation commercial Anti-CCP Ab diagnostic kit (n=48, p=0.02). Moreover, to evaluate the direct role of generated anti-CCP Ab, mice were immunized with CII and then injected with 12G1 instead of second boosting. Immunized mice with 12G1 demonstrated significantly increased disease severity and incidence compared with mice challenged with negative control.

Conclusion: In conclusion, we generated a synthetic monoclonal antibody against CCP. This antibody exacerbated arthritis in CIA model and had the potential to detect citrullinated Ag in human RA serum. Application of this anti-CCP Ab may contribute to understanding the pathophysiology of RA and developing synthetic Ab-driven diagnostic strategy.

Disclosure:

Y. Kim,
None;

S. J. Moon,
None;

H. Yi,
None;

J. H. Ju,
None.

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