Selection and characterization of a novel factor XI inhibiting peptide by using phage display technology

[摘要] English: The role of factor XI in hemostasis can be seen as a combination of a procoagulantaction (the formation of fibrin) and an antifibrinolytic action (the protection of fibrin).High levels of factor XI lead to a prolonged down regulation of fibrinolysis andtherefore a risk of thrombosis (Meijers et ai, 2000). Under disease conditionsassociated with Disseminated Intravascular Coagulation (DIC), the continuousexposure to excess TF typically exhaust the available tissue factor pathwayinhibitor (TFPI), leading to rampant thrombin generation by factor XI feedback andtherefore also a risk of thrombosis (0sterud and Bjerlid, 2001).I selected possible .inhibitors of factor XI using phage display technology. I startedthe phage display selection by biopanning in immuno-tubes and eluted the factorXI binding phages non-specifically from the immuno-tube. I did four selectionrounds, to enrich the factor XI binding phages. I found only two strong factor XIbinding phage clones from a linear 12-mer phage library. Both phage clonesbound dose dependently and with a high affinity to factor XI. Both clones alsolengthened the partial thromboplastin time (aPTT) dose dependently.The amino acid sequences of the peptides displayed on these two clones indicatethat both peptides contain three amino acid sequences of HMWK and thrombin.One clone also contains a three amino acid sequence of factor XII. None of themcontains a three amino acid sequence of factor IX. I synthesize a linear peptidewith the corresponding sequence as the peptide displayed on the clone that wasprevented from binding to factor XI by both factor IX and thrombin.I characterized the peptide by studying its effect on the aPTT. This peptidelengthens the aPTT dose dependently. The lengthening in aPTT of our peptidehowever indicates that I have selected an inhibitor of the contact system factors ofcoagulation.In summary, this study shows that the phage display can be used to select novelfactor XI inhibitors from random peptide libraries. With further studies, this peptidemay be developed as an antithrombotic.

[发布机构] University of the Free State