Introduction: B-domain modification can improve production of recombinant factor VIII (rFVIII) proteins. However, the engineered junction results in non-native peptide sequences with the potential to elicit immune responses via major histocompatibility complex class-II (MHC-II)-binding and CD4+ T cell activation.
Aim: Assess the immunogenic potential of B-domain junction peptides of turoctocog alfa and other B-domain-modified rFVIII proteins using in silico and in vitro immunogenicity assessment techniques.
Methods: Peptides with amino acid sequences identical to the B-domain junction of turoctocog alfa, simoctocog alfa and moroctocog alfa were evaluated by in silico peptide-MHC-II binding prediction, in vitro peptide-MHC-II-binding measurement and in vitro T cell-activation assays. Moreover, turoctocog alfa was assessed for peptide presentation on dendritic cells (DCs) using MHC-associated peptide proteomics.
Results: In silico analysis predicted virtually no neo-epitopes in the B-domain junction for turoctocog alfa, whereas some were predicted for simoctocog alfa and moroctocog alfa. Turoctocog alfa and moroctocog alfa peptides showed minimal capacity to bind high-frequency MHC-II molecules in vitro, whereas simoctocog alfa peptide demonstrated some degree of binding to approximately half of the MHC-II molecules tested. In line with this, no B-domain peptides from turoctocog alfa were found to be presented on MHC-II complexes on DCs. B-domain junction peptides from all 3 compounds induced T cell responses in only a few percentages of donors.
Conclusion: All 3 junction peptides were found to have a low immunogenicity potential, suggesting that modification of the B-domain does not constitute an increased immunogenicity risk for any of the products examined.
Keywords: blood coagulation factor inhibitors; haemophilia A; immunogenicity; in silico; in vitro techniques; recombinant factor VIII.
© 2018 John Wiley & Sons Ltd.