Anti-factor Xa antibodies in patients with antiphospholipid syndrome and their effects upon coagulation assays

Abstract

IntroductionThe aim of this study was to examine the prevalence and functional effects of antibodies directed against Factor (F)Xa and other serine proteases (SP) in patients with antiphospholipid syndrome (APS).MethodsSerum from patients with APS (n = 59), systemic lupus erythematosus (SLE; n = 106), other autoimmune rheumatic disease (ARD; n = 63) and 40 healthy controls (HC) were tested for IgG activity against thrombin (Thr), FXa, FVIIa, phosphatidylserine (PS)/FXa and antithrombin (AT)-III by enzyme-linked immunosorbent assay (ELISA). Anti-FXa positive IgG were purified to measure their avidity by chaotropic ELISA and functional effects upon clotting time (FXa-ACT) and FXa enzymatic activity (± AT-III).ResultsAnti-FXa IgG were found in patients with SLE (49.1%) and APS (33.9%) (P <0.05) but not in ARD controls and HC. In contrast, anti-Thr and anti-PS/FXa IgG were identified in other ARD and anti-FVIIa IgG were low in all groups. The avidity of APS-IgG to FXa was significantly higher than SLE-IgG (P <0.05). Greatest prolongation of FXa-ACT was observed with APS-IgG and greatest inhibitory effect upon FXa enzymatic activity was found with APS-IgG followed by SLE-IgG compared to HC-IgG. ATIII inhibition of FXa was significantly reduced by APS-IgG compared with HC and SLE (P <0.05) and did not correlate with binding to AT-III.ConclusionAPS anti-FXa IgG have higher avidity to FXa and greater effects upon the enzymatic and coagulant activity of FXa compared with SLE anti-FXa IgG. Further studies of anti-FXa antibodies in APS, SLE and other non-autoimmune thrombotic disease cohorts are now required to evaluate whether targeting FXa with selective inhibitors in patients bearing anti-FXa antibodies may be an effective treatment strategy.

Factor Xa Mediates Calcium Flux in Endothelial Cells and is Potentiated by Igg From Patients With Lupus and/or Antiphospholipid Syndrome

“…Our previous findings of the effects of APS-IgG on FXa in vitro 17 are summarised in Table 2 . APS IgG significantly reduced FXa activity (P < 0.0001 for APS versus HC, and P = 0.0008 for APS versus SLE IgG) and antithrombin-III mediated inhibition of FXa activity (P < 0.0001 for APS versus SLE and APS versus HC IgG) and significantly prolonged the clotting time compared to HC IgG (p < 0.0001) and SLE/APS- IgG (p = 0.04).…”

“…Further purification of aFXa IgG was performed by passage of IgG though a heparin column to remove IgG that binds non-specifically to the negatively charged resin, followed by purification of the remaining IgG fraction though an immobilised FXa column and subsequent dialysis to PBS. The presence of IgG directed against cardiolipin 14 , β2-glycoprotein I (β2-GPI) 14 thrombin 16 and FXa was measured by ELISA as previously described 17 . The cut-off of positivity for all ELISAs was determined from a cohort of forty HC.…”

“…Effects of FXa-reactive IgG on FXa-activated clotting time (ACT) was measured as described previously 17 . The effects of anti-FXa reactive IgG on FXa activity were studied by digestion of a chromogenic substrate S-2765 (Chromogenix; DiaPharma) both in the absence and presence of ATIII and the degree of colour change used to quantify the activity of FXa 17 .…”

“…More recently, we have shown that serum from patients with APS and patients with SLE (without APS) had significantly increased IgG reactivity for FXa compared with controls 17 . Polyclonal IgG purified from serum of patients with APS showed higher avidity binding to FXa and greater effects upon the enzymatic and coagulant activity of FXa compared with polyclonal IgG isolated from patients with SLE who lacked APS.…”

Thrombotic events in patients with antiphospholipid syndrome treated with rivaroxaban: a series of eight cases

“…The presence and functional effects of anti-factor Xa are promising because some NOACs directly inhibit factor Xa. It could be possible that this group of patients would be eligible for their use [18].…”