1997 MINUTES
LUPUS ANTICOAGULANT/PHOSPHOLIPID-DEPENDENT ANTIBODIES SUBCOMMITTEE
Friday, 6 June, 1997, 8:00-12:00
Giotto I, Fortezza da Basso
Florence, Italy
Chair: T. Barbui, Italy
Co-Chairs: J. Brandt, USA; S. Machin, UK; R. Roubey, USA;
I. Scharrer, Germany; D. Triplett, USA
Attendance: approximately 325
The Subcommittee Meeting consisted of three parts:
Part 1
Prof. D. A. TRIPLETT updated the methodology for the diagnosis of lupus anticoagulants. Major points of discussion were the heterogeneity of the lupus anticoagulants and the lack of nomenclature and classification (which should take into account their differences in antigenic targets, association with thrombosis, underlying disease and types of laboratory tests employed for their detection). The laboratory methodology for the detection of lupus anticoagulants is a major task of the Subcommittee, which has repeatedly proposed recommendations (last time in 1995). Triplett underlined the current methodological problems: the failure to comply with the recommendations; the simplification of diagnosis (for example, by adopting integrated systems); the presence of pseudo-factor deficiencies and that of true factor deficiencies; the absence of reference standards and materials; the need to quantify the titer of the lupus anticoagulants. He reviewed the ongoing clinical trials (Italian Registry and WAPS) and proposed, as future work of the Subcommittee, the identification of physiopathological mechanisms of thrombosis, the development of reference standards, the quantification of the inhibitors and the development of tests (or combination of tests) able to predict the risk of thrombosis.
Prof. R. A. S. ROUBEY updated the current methodology for the detection of anticardiolipin (aCL) and anti-b2-glycoprotein I (ab2-GPI) antibodies by immunoassays. First, he gave the general principles for the applicability of enzyme immunoassays to aCL and ab2-GPI antibodies: the technical evaluation should consider precision, accuracy, analytical sensitivity and specificity (cross-reactivity), robustness and non-specific binding. The clinical evaluation should take into account: normal range diagnostic sensitivity, specificity and predictive values and also ROC curves. Roubey emphasized the importance of the autoantibody characteristics, such as the antigenic specificity (b2-GPI, prothrombin, other proteins and the role of phospholipids), titer, isotype, subclass, affinity, and avidity. The assessment of the immunoassays could be phospholipid-based (conventional or modified cardiolipin ELISA) or protein-based (ab2-GPI ELISA or immunoblot). Other immunoassays could be developed for measuring anti-prothrombin antibodies. In conclusion, he gave the recommendations and suggested the future directions and the role of the Subcommittee.
Prof. P. de GROOT reported on a large clinical study performed in almost 200 SLE-positive patients that assessed the association between thromboembolic events and antiphospholipid antibodies measured either via immunoassays (aCL, ab2-GPI and anti-prothrombin antibodies) or via coagulation tests (lupus anticoagulants). Only the presence of lupus anticoagulants was statistically associated with both venous and arterial thrombosis. Adsorption experiments were performed to investigate the contribution of ab2-GPI and anti-prothrombin antibodies to the expression of the lupus anticoagulant activity. De Groot concluded that there is no direct clinical need to use ELISAs to detect ab2-GPI and anti-prothrombin antibodies and that, in a large majority of patients, lupus anticoagulant activity is caused by a combination of antibodies with different specificities.
Prof. J. ARNOUT discussed the pathophysiological mechanisms of antiphospholipid antibodies. Monoclonal antibodies (MoAbs) were raised against b2-GPI and their effect on the binding of b2-GPI to a phospholipid surface was studied by real-time biospecific interaction analysis. MoAbs (and their Fab2 fragments) provided with phospholipid-dependent anticoagulant activity were able to form stable bivalent complexes on the surface; conversely, MoAbs lacking this property failed to stabilize the binding. Arnout hypothesized that this might occur also on a partially activated physiological surface, in this way potentiating cell activation via the Fc receptor.
Prof. C. N. CHESTERMAN discussed the effect of isolated IgG with lupus anticoagulant activity on thrombin generation under flow conditions. Patients' IgG was able to increase the generation of thrombin 1.2 to 5.6-fold over controls. Opposite results were obtained under static conditions. IgG with lupus anticoagulant activity was also able to protect FVa from inactivation by aPC. Patients' IgG facilitated the interaction of prothrombin to the phosholipid surface in flow but not in static conditions. This may have important implications in shifting the balance toward thrombosis in vivo.
Part 2
Dr. A. TRIPODI discussed the problems regarding the monitoring of oral anticoagulation by means of the INR system in patients with aPL antibodies. He proposed a multicenter collaboration to investigate the issue of laboratory control of oral anticoagulation therapy in patients with aPL antibodies. Plasmas from patients with and without oral anticoagulation will be collected in different centers and the INR will be evaluated centrally with commercial reagents previously calibrated against the International Reference Preparation for thromboplastin to determine the ISI. Results will be analyzed to assess the extent of interference of aPL antibodies on PT-INR and to identify the reagents more likely to be affected.
Prof. S. MACHIN discussed current knowledge on the treatment of thrombosis and reported on the ongoing clinical trials. He defined four different types of aPL-positive patients who could be enrolled in appropriate clinical studies: 1. patients without thrombosis, to establish the possible role of primary prophylaxis and the best treatment of high-risk situations; 2. patients with a single thrombotic event, for whom the duration and intensity of oral anticoagulation has to be defined, also in consideration of possible concomitant risk factors (i.e., FV Leiden mutation); 3. patients with recurrent thrombosis despite long-term, high-dose warfarin treatment (although their number is small, the clinical impact is very high); 4. patients with cerebrovascular thrombosis (a prospective trial is ongoing). He concluded by suggesting that an International Registry with data of ongoing clinical trials be reviewed at the annual SSC meeting.
Prof. M. GREAVES provided an overview of the current knowledge on the pathophysiology of recurrent miscarriages, updated the therapeutic approaches, and discussed the risk-benefits of steroids, aspirin, heparin alone or in combination. Based on the results of a recent clinical trial, the combination aspirin/heparin (5,000 x twice daily) appears associated with a better pregnancy outcome. He reported about a randomized trial (aspirin vs. low molecular weight heparin) which is currently in progress at the Liverpool Women's Hospital. Finally, among the questions to be solved in the future, he underlined: the need to have predictive laboratory tests; the necessity to confirm the advantage of heparin/aspirin over the other treatments; the role for low-molecular weight heparins; the impact for short- and long-term side effects of heparin therapy in the mother; the optimal dose and duration of treatment; and the possible role of intravenous immunoglobulins.
Part 3
Prof. F. WISLØFF discussed an integrated coagulation system that can be used in patients with lupus anticoagulants, which incorporates in a single assay the screening, mixing, and confirmatory procedures. This system has been proposed for the aPTT and the dRVVT. Wisløff proposed to investigate the aPTT-based system in the setting of the forthcoming ISLA-5, with the aim to define the cut-off between normal and pathological (97.5 vs. 99th percentile) and to serve as a reference for semi-quantification of lupus anticoagulants.
Prof. M. C. BOFFA summarized the first meeting of the European Forum on aPL antibodies. Three projects were presented, standardization of aPL ELISA, standardization of ab2-GPI antibodies, and registry of cases of familial antiphospholipid syndrome. She summarized the results of the questionnaire sent by A. Tincani to 29 centers to investigate the current methodology employed for the detection of aPL antibodies.
Dr. G. FINAZZI provided an update of the WAPS study, the aim of which is to investigate long-term, high-dose warfarin treatment in the secondary prevention of arterial and venous thromboembolism. The trial is due to start with patient enrollment in July 1997.
A booklet with the reports of each presentation was distributed during the Subcommittee meeting.