Engineering a Therapeutic Protein to Enhance the Study of Anti-Drug Immunity

Abstract

The development of anti-drug antibodies represents a significant barrier to the utilization of protein-based therapies for a wide variety of diseases. While the rate of antibody formation can vary depending on the therapeutic employed and the target patient population receiving the drug, the antigen-specific immune response underlying the development of anti-drug antibodies often remains difficult to define. This is especially true for patients with hemophilia A who, following exposure, develop antibodies against the coagulation factor, factor VIII (FVIII). Models capable of studying this response in an antigen-specific manner have been lacking. To overcome this challenge, we engineered FVIII to contain a peptide (323–339) from the model antigen ovalbumin (OVA), a very common tool used to study antigen-specific immunity. FVIII with an OVA peptide (FVIII-OVA) retained clotting activity and possessed the ability to activate CD4 T cells specific to OVA323–339 in vitro. When compared to FVIII alone, FVIII-OVA also exhibited a similar level of immunogenicity, suggesting that the presence of OVA323–339 does not substantially alter the anti-FVIII immune response. Intriguingly, while little CD4 T cell response could be observed following exposure to FVIII-OVA alone, inclusion of anti-FVIII antibodies, recently shown to favorably modulate anti-FVIII immune responses, significantly enhanced CD4 T cell activation following FVIII-OVA exposure. These results demonstrate that model antigens can be incorporated into a therapeutic protein to study antigen-specific responses and more specifically that the CD4 T cell response to FVIII-OVA can be augmented by pre-existing anti-FVIII antibodies.

Therapeutic proteins immunogenicity: a peptide point of view

“…FVIII is administrated to patients who suffer from hemophilia A, a congenital bleeding disorder caused by the lack of this factor in circulation, resulting in ineffective coagulation upon injury. FVIII administration, however, leads to ADA development in about 30% of the patients and thus various studies focused on this protein [34]. Various endogenous proteins bind to FVIII during its internalization and these different complexes impact the epitope presentation, thus the study investigated the recognition of FVIII peptides by FVIII-specific CD4 + T-cell hybridoma clones, modifying the microenvironment during the FVIII uptake, more in detail, FVIII was compared with FVIII complexed with von Willebrand factor (VWF, the plasma chaperon of FVIII), FVIII activated by thrombin (activation that occurs at the bleeding site), and FVIII accompanied by a concurrent blockade of APCs receptors.…”