Identification and removal of a promiscuous CD4+ T cell epitope from the C1 domain of factor VIII


Jones TD, Phillips WJ, Smith BJ, Bamford CA, Nayee PD, Baglin TP, Gaston JS, Baker MP.


The development of inhibitors in hemophiliacs is a severe complication of factor VIII (FVIII) replacement therapy and is a process driven by FVIII specific T helper cells.

To finely map T cell epitopes within the whole FVIII protein in order to investigate the possibility of engineering FVIII variants with reduced propensity for inhibitor development.

T cell lines were generated from five patients with severe hemophilia who had developed inhibitors, and were screened for T cell proliferation against pools of overlapping peptides spanning the entire B domain deleted (BDD) FVIII sequence. Positive peptide pools were decoded by screening individual peptides against the T cell lines. Positive peptides, and mutants thereof, were tested for their ability to bind major histocompatibility complex (MHC) Class II and stimulate T cell proliferation in a panel of healthy donors. The activities of the corresponding mutant proteins were assessed via chromogenic assay.

One peptide, spanning FVIII amino acids 2098-2112, elicited a vigorous response from one hemophiliac donor, induced strong T cell responses in the panel of healthy donors and bound to a number of HLA-DR alleles. Mutations were made in this peptide that removed its ability to stimulate T cells of healthy donors and to bind to MHC Class II while retaining full activity when incorporated into a mutant BDD-FVIII protein.

Fine T cell epitope mapping of the entire FVIII protein is feasible, although challenging, and this knowledge may be used to create FVIII variants which potentially have reduced immunogenicity.

The full paper is available here


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