OP0084 Oxidation of β<sub>2</sub>-Glycoprotein I is Strongly Associated with the Presence of Anti-Domain I Antibodies in Patients With Antiphospholipid Syndrome
Authors: M.G. Raimondo, C. Pericleous, A. Radziszewska, M.O. Borghi, P.L. Meroni, I. Giles, A. Rahman, Y. Ioannou
Journal: Annals of the Rheumatic Diseases (2015)

<h3>Background</h3> Beta-2 glycoprotein I (β<sub>2</sub>GPI) represents the major antigenic target for antiphospholipid antibodies (aPL), the hallmark of antiphospholipid syndrome (APS). β<sub>2</sub>GPI contains five homologous domains, with domain I (DI) being identified as the main antigenic epitope for pathogenic anti-β<sub>2</sub>GPI (aβ<sub>2</sub>GPI). It has been reported that APS patients show an elevated amount of total β<sub>2</sub>GPI as compared with healthy donors and other autoimmune disease control groups. Moreover, in healthy individuals, β<sub>2</sub>GPI mainly exists in its biochemically reduced form whilst the post-translational oxidized structure (fish-hook linear configuration) is elevated in patients with APS. Antibodies targeting β<sub>2</sub>GPI-DI (aDI) represent a key pathogenic sub-population of aPL, thus their detection may allow the identification of patients at highest clinical risk. <h3>Objectives</h3> Our hypothesis is that the different redox state of β<sub>2</sub>GPI, from reduced (circular) to oxidised (linear), might be responsible for the exposure of DI, thus providing the antigenic drive for the aDI antibodies. However, no study has investigated the association between the circulating oxidized form of β<sub>2</sub>GPI and the presence of aDI in APS patients. <h3>Methods</h3> Serum samples from 40 patients who fulfilled the classification criteria for APS were tested (34 primary and 6 secondary APS). A sandwich ELISA for quantifying total β<sub>2</sub>GPI levels within serum was performed for all samples. An in-house standard (consisting of pooled serum from 10 healthy controls) was used to obtain a standard curve in each ELISA. Biochemically reduced β<sub>2</sub>GPI was detected via labelling free thiols within serum proteins with a biotinylated free-thiol binding reagent and then, via coating on a streptavidin plate, detecting the presence of labelled β<sub>2</sub>GPI with an aβ<sub>2</sub>GPI antibody, based on published methods. Then samples were screened for IgG aDI, aβ<sub>2</sub>GPI and anti-cardiolipin (aCL). <h3>Results</h3> The positivity for aDI, aβ<sub>2</sub>GPI and aCL IgG was 60%, 75% and 80% respectively. The relative proportion of reduced β<sub>2</sub>GPI was expressed as a percentage of that observed in the in house-standard after correction for the total amount of β<sub>2</sub>GPI. A significant negative correlation was observed between proportion of reduced β<sub>2</sub>GPI and aDI levels (Spearman coefficient r = -0.409, p=0.008) and aCL titre (r = -0.317, p=0.045). There was no significant correlation between proportion of reduced β<sub>2</sub>GPI and aβ<sub>2</sub>GPI (r = -0.059, p=0.713). <h3>Conclusions</h3> The strong association between the oxidative state of β<sub>2</sub>GPI and aDI positivity supports the hypothesis that modulation of the redox state of β<sub>2</sub>GPI affects the production of antibodies against the DI epitope, which become exposed in the oxidised protein. However, it remains to be proven if the exposure of DI within the oxidised (hence linearized) form of β<sub>2</sub>GPI acts as the antigenic drive for development of aDI autoantibodies or whether the presence of these antibodies stabilizes the antigen-antibody complex maintaining the linear configuration of the molecule. <h3>Disclosure of Interest</h3> None declared