Factor v inhibitors associated with hemorrhagic symptoms bind to a limited region of the factor va light chain
Factor V inhibitors most commonly occur as (1 ) spontaneously arising autoantibodies in previously normal patients, or (2) cross-reacting alloantibodies arising after exposure to topical bovine thrombin preparations containing bovine factor V. We previously demonstrated that a spontaneous factor V inhibitor associated with hemorrhagic symptoms bound to the aminoterminal region of the second C-type domain (C2) of the light chain of factor V. This resulted in loss of procoagulant activity as well as phosphatidylserine-specHic binding. We have now investigated a total of twelve patients with factor V inhibitors, including seven autoantibodies and five antibodies arising after exposure to bovine thrombin preparations. Eight patients presented with hemorrhagic manifestations (five autoantibodies and three alloantibodies). Four had no clinical manifestations, but were found to have a factor V inhibitor during evaluation of abnormal screening coagulation assays. Using recombinant human factor V (rHFV) deletion mutants, all twelve inhibitors were found to bind epitopes contained within the light chain of factor Va. All eight hemorrhagic inhibitors and one non-hemorrhagic inhibitor were further mapped to the C2 domain. One of the remaining non-hemorrhagic antibodies appeared to recognize an epitope in the C1 domain, but the other two antibodies did not bind to any construct smaller than the light chain. Using recombinant factor V/factor VIII chimeras, we found that seven of the eight hemorrhagic inhibitors bound to the amino terminal third of the 02 domain, while the eighth antibody bound to a larger epitope in the same region. The non-hemorrhagic inhibitor did not bind to any of the chimeras. IgQ preparations from the hemorrhagic Inhibitors neutralized factor Va procoagulant activity in a prothrombinase assay, but this inhibitory effect could be abrogated by pre-incubation of the inhibitor with purified rHFV C2 domain. In addition, none of the inhibitors neutralized the procoagulant activity of a recombinant chimera that replaced the epitope(s) recognized by the inhibitors with the corresponding region of the factor VIII molecule. IgG fractions from the non-hemorrhagic inhibitors had no effect on factor V activity in the same assay system. In summary, inhibitors associated with hemorrhagic symptoms bound to the amino terminal region of the 02 domain, regardless of whether they were autoantibodies or alloantibodies. Interestingly, this region of the bovine factor V 02 domain, spanning 51 amino acids, is identical in primary sequence to the human protein with the exception of 3 amino acids.
Ortel, TL; Moore, KD; Quinn-Allen, M; Kane, WH
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