IgG antibodies to cyclic citrullinated peptides exhibit profiles specific in terms of IgG subclasses, Fc-glycans and a fab-Peptide sequence.

ABSTRACT: The Fc-glycan profile of IgG1 anti-citrullinated peptide antibodies (ACPA) in rheumatoid arthritis (RA) patients has recently been reported to be different from non-ACPA IgG1, a phenomenon which likely plays a role in RA pathogenesis. Herein we investigate the Fc-glycosylation pattern of all ACPA-IgG isotypes and simultaneously investigate in detail the IgG protein-chain sequence repertoire. IgG from serum or plasma (S/P, n?=?14) and synovial fluid (SF, n?=?4) from 18 ACPA-positive RA-patients was enriched using Protein G columns followed by ACPA-purification on cyclic citrullinated peptide-2 (CCP2)-coupled columns. Paired ACPA (anti-CCP2 eluted IgG) and IgG flow through (FT) fractions were analyzed by LC-MS/MS-proteomics. IgG peptides, isotypes and corresponding Fc-glycopeptides were quantified and interrogated using uni- and multivariate statistics. The Fc-glycans from the IgG4 peptide EEQFNSTYR was validated using protein A column purification. Relative to FT-IgG4, the ACPA-IgG4 Fc-glycan-profile contained lower amounts (p?=?0.002) of the agalacto and asialylated core-fucosylated biantennary form (FA2) and higher content (p?=?0.001) of sialylated glycans. Novel differences in the Fc-glycan-profile of ACPA-IgG1 compared to FT-IgG1 were observed in the distribution of bisected forms (n?=?5, p?=?0.0001, decrease) and mono-antennnary forms (n?=?3, p?=?0.02, increase). Our study also confirmed higher abundance of FA2 (p?=?0.002) and lower abundance of afucosylated forms (n?=?4, p?=?0.001) in ACPA-IgG1 relative to FT-IgG1 as well as lower content of IgG2 (p?=?0.0000001) and elevated content of IgG4 (p?=?0.004) in ACPA compared to FT. One ?-variable peptide sequence was significantly increased in ACPA (p?=?0.0001). In conclusion, the Fc-glycan profile of both ACPA-IgG1 and ACPA-IgG4 are distinct. Given that IgG1 and IgG4 have different Fc-receptor and complement binding affinities, this phenomenon likely affects ACPA effector- and immune-regulatory functions in an IgG isotype-specific manner. These findings further highlight the importance of antibody characterization in relation to functional in vivo and in vitro studies.

SUBMITTER: Lundstrom SL 

PROVIDER: S-EPMC4245247 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature