Epitope Mapping Of Human Coagulation Factor VIII

Hemophilia A is the most severe hereditary blood coagulationdisorder to affect humans and is among the most common disorders ofthis type. One of the most severe complications of replacement therapy isthe development of inhibitory alloantibodies to the therapeutic F8protein,which occurs in approximately30%of severly affected patients withhemophilia A. These antibodies incease the morbidity and mortality andthe cost of the treatment of hemophilia A patients. Thus, developing amethod to reduce the incidence of inhibitory antibody developmentwould improve the management of patients with hemophilia A. However,for the medical community, it still remains a major challenge in thereduction of the immunogeneity of F8protein. This project focuses on thecomprehensive screening and identificatuion of potential epitopes thatlocated in the heavy and light chain of human coagulation factor VIII, andfinally specifies which epitopes are responsible for the immunogeneity ofthe heavy and light chain of factor VIII.Deletion and point mutagenesis and immunoblotting analysis wereintegrated in this project to screen and identify potential epitopes located in the heavy and light chain of F8protein. After a comprehensive analysis,the epitope of the heavy chain was localized in the A2domain and threeantigenic amino acid residues were specified: R484, R489and R593，theseresults accord with the related literature,which also confirm our newmethod is feasible; and the epitope of the light chain was localized in theA3domain and three previously unreported small segments of amino acidresidues were specified as antigenic sites: V79L80, Q91F92K93K94V95andT106Q107. These findings are fundamental to the development of lessimmumogenic bioengineering recombinant F8protein that retains bloodcoagulation activity.In conclusion, six antigenic sites of F8protein were identified in thisstudy with three of them that located in the light chain previouslyundocumented. This innovative finding is of great importance to reducethe incidence of inhibitor development in patients with hemophilia A.