Dr. M. Aiach presented an update of the protein S database, 204 mutations (140 unique events). There are only 3 large deletions, 99 missense mutations, as well as frameshift mutations, etc. There are mainly type I/III deficiencies, very few type II, all of the latter mutations in N terminal region. It was decided to look into the possibility of publishing the update as a SSC communication in Thrombosis and Haemostasis and also to explore the possibility of preparing a website for the database.

Dr. I. Jennings discussed the practical issues of APCR/Factor V Leiden tests, under the UK laboratory quality control scheme, NEQAS. In this routine quality control exercise many laboratories could not reproducibly detect heterozygotes with functional assays or even with PCR. Forty-seven centers (mainly UK) were sent fresh whole blood. Different extraction techniques were used with different primer sets for Factor V Leiden and PT 20210A. Finally, different endpoints (SSCP, restriction) were used. In the first study of only the APCR clotting test, five out of 47 tests resulted in the incorrect diagnosis of Factor V Leiden. A later study included genetic testing for FV Leiden and PT 20210A. Once again, there were many incorrect diagnoses. A further survey underway includes polymorphisms of the MTHFR gene, and there are still reporting errors. The conclusion is that proficiency testing for these common polymorphisms is very important in the routine laboratory context. There was some discussion whether identifying these polymorphisms is clinically valuable.

Dr. J. Emmerich summarized results of a meta-analysis of eight case control studies of venous thrombosis concerned with interaction of Factor V Leiden and PT 20210A. There were 2310 cases in total and 3206 controls. An overall OR for thrombosis for Factor V Leiden was 5.01, while that for PT 20210A was 3.88. Both mutations combined gave an OR 24.1. Age of first onset was significantly younger, 36 compared to 40.5 years. The OR for PE was approximately 1.0 for both Factor V Leiden and PT 20210A. Synergism was demonstrated between oral contraceptives for both Factor V Leiden and PT 20210A. It is planned to submit this as a SSC communication for publication in Thrombosis and Haemostasis.

NEW RISK FACTORS

Dr. A.K. Ohlin presented her results on association of thrombomodulin gene mutations and venous thromboembolism. Seven hundred patients (Swedish/USA/France) have had their thrombomodulin gene screened. Thirteen cases were found to be heterozygous for a mutation. A start has been made characterizing gene mutations by in vitro transfection but further work is required. Dr. G. Kunz presented results of thrombomodulin mutations in arterial disease. In a case control study of 104 patients, six different mutations were identified. Three promoter polymorphisms have been evaluated by reporter-gene analysis and one, —33G to A, had reduced reporter activity. A coding sequence mutation Ala25Thr was found in two individuals and evaluated in a large case control study of myocardial infarction (MI). The results suggest Ala25Thr is a risk, ~2 fold, for MI. There is also evidence that an insertion/frameshift mutation in the coding sequence results in reduced expression of thrombomodulin in vitro and in vivo. It was concluded that thrombomodulin gene mutations may be important in MI but more work is required to clarify this.

Dr. R. Simmonds summarized the polymorphic nature of the EPCR gene. This gene is a candidate risk factor for venous and arterial thrombosis. The sequence of the EPCR gene has been completed. There are four exons coding for the receptor sparing 6kbp. Four common polymorphisms have been identified in healthy normals and their population frequencies determined, as a prelude to clinical studies. Dr. E. Faioni also addressed this issue with two clinical studies. Two hundred and two survivors of MI/190 controls were used as well as 209 patients with DVT/402 controls. A 23bp insertion was identified and its consequences evaluated in the clinical studies. Adjusted OR of 2.5 for MI and 2.2 for DVT were obtained, but there were wide CIs. This is the first indication that mutation of the EPCR may have a role in disease.

Dr. F.C. Church discussed nomenclature issues in SERPINS. First, he reviewed the recent advances in SERPINS, basic and clinical. An international committee has been set up chaired by Gary Silverman to develop a logical classification system, to link websites, and to form a SERPINS society. There are approximately 400 known SERPINs. These will be grouped as plants, insects, nematodes, and certain types of factors, e.g., PAI-1. Dr. Church asked for people with an interest in this area to contact him during the meeting.

Dr. F. Bernardi discussed the HR2 Factor V gene allele. Approximately 10% of subjects are carriers and are widely distributed. Polymorphisms between exon 8-25 are linked and form the HR2 allele. This seems to be associated with increased risk of thrombosis, OR around two, but this has been inconsistent. A large study of Italian patients suggests the HR2 might be a risk factor for coronary artery disease. The functionally important change is that HR2 alters APCR. The isoforms of factor differing in glycosylation, Factor Va1 and Factor Va₂, seem to be altered in their relative distribution in the HR2 haplotype and this could explain the effect on function.