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Immune response to FVIII in hemophilia patients
tion have normal levels of FVIII:Ag, although the protein is non-functional,1 while patients with large deletion/intron inversions have no circulating protein.2 Due to such differences in protein expression, patients are also heterogeneous as far as the education of their immune system against endogenous FVIII is concerned.
To date, the prevention or treatment of bleeds in hemo- philia A patients relies on the intravenous administration of therapeutic FVIII. Therapeutic FVIII is purified from pools of plasma from healthy blood donors or originates from recombinant technology. While differences exist between plasma-derived and recombinant FVIII products, as well as among recombinant products, in terms of struc- ture, glycosylation pattern,3 ability to bind von Willebrand factor (VWF),4 the endogenous chaperone for FVIII, all the available products share the property of inducing neutral- izing immunoglobulin G (IgG), termed ‘FVIII inhibitors’, in a substantial number of patients. The occurrence of FVIII inhibitors following replacement therapy is a serious clinical problem that complicates patients’ management and reduces their quality of life, as well as being a major society issue owing to the high costs associated with the treatment of bleeding when FVIII cannot be used.5 Several factors have been identified as increasing the risk of a patient developing FVIII inhibitors, in particular genetic risk factors such as a family history of inhibitor develop- ment,6 the type of gene abnormality causing the hemo- philia A and the ensuing severity of the disease,7,8 HLA-DR haplotypes9,10 and polymorphisms in a restricted set of immune genes.11-14 Nevertheless, it is, to date, impossible to predict with certainty whether a given patient will develop FVIII inhibitors.
Over the last 20 years, a large body of the research ded- icated to deciphering the immunogenicity of FVIII has been based on the ‘danger theory’ proposed by Polly Matzinger almost 25 years ago.15 Researchers have attempted to elucidate the nature of the danger signals that are adjuvants of the immune response to exogenous FVIII in 5-30% of patients with hemophilia A (including all severities of the disease) following replacement thera- py. Here, we review the evidence on the presence of dan- ger signals associated with FVIII administration to ques- tion the notion that developing an immune response to FVIII requires danger signals. Furthermore, we challenge the idea that developing an immune response to FVIII is unequivocally pathogenic and propose that the develop- ment of FVIII inhibitors in a substantial number of patients results from the inability of the immune system to mount a counteractive tolerogenic response.
Danger signals as adjuvants of the anti-factor VIII immune response in patients with hemophilia A
The immune response to therapeutic FVIII is believed to be a classical immune response against an exogenous anti- gen, wherein some of the intravenously administered FVIII transiently accumulates in secondary lymphoid organs, as observed in spleens of FVIII-deficient mice,16,17 is internalized by antigen-presenting cells18,19 and presented to naïve FVIII-specific CD4+ T cells. Upon activation, FVIII-specific T cells proliferate and provide help to naïve FVIII-specific B cells that differentiate into memory B cells or plasmocytes secreting inhibitory anti-FVIII IgG. According to the “infectious non-self model” introduced by C Janeway20 and later the “danger theory” coined by P Matzinger,21 antigen-presenting cells need to sense danger
signals concomitant with antigen uptake to mature appro- priately, activate naïve T cells and induce the immune response. In the context of the anti-FVIII immune response, severe/recurrent bleeds, trauma, surgery, infec- tion or vaccination have been proposed as potential sources of danger signals that may trigger the immune system.22 In recent years, experimental investigations per- formed in FVIII-deficient mice, a model of severe hemo- philia A, and clinical observations in hemophilia A patients have challenged most of these hypotheses, as dis- cussed below.
Factor VIII dose matters
The dose of administered FVIII is one of the few param- eters that has shown consistent association with the development of an anti-FVIII immune response. Seminal work in FVIII-deficient mice clearly demonstrated that increasing the dose of intravenously injected FVIII results in a proportional increase in the intensity of the immune response.23 The experimental data were confirmed by the RODIN study in which the intensity of FVIII treatment and mean dose of administered FVIII correlated with the incidence of inhibitor development in patients with severe hemophilia.24 Conversely, the reduction in the amount of FVIII internalized by antigen-presenting cells and, conse- quently, presented to CD4+ T cells has been hypothesized as a mechanism by which von Willebrand factor may play an immuno-protective role towards therapeutic FVIII.25 On the other hand, the daily administration of high-dose FVIII to inhibitor-positive patients undergoing immune tolerance induction protocols results in the eradication of FVIII inhibitors in two-thirds of the cases. This suggests that the antigen dose may not be as important in deter- mining immunogenicity.
An enticing recent concept proposes that the immuno- genicity of a protein depends on its discontinuous pres- ence in the organism. This ‘discontinuity theory’ first pro- posed by Pradeu et al. states that “the key to the induction of an immune response is antigenic difference in a time- dependent context”. It is hypothesized that the intermit- tent appearance of a foreign antigen triggers strong and long-lasting immune responses, whereas the persistence of the same foreign antigen over time or its progressive introduction over a long period leads to tolerance.26 This theory is particularly suitable in the case of therapeutic FVIII, a molecule with a short half-life (<15 hours) that is administered either on-demand or every 2 to 3 days in patients under prophylactic treatment, and thus intermit- tently appears and disappears from the circulation along cycles of intravenous injections.
Factor VIII is not an alarmin
Danger signals are provided by endogenous (damage- associated molecular patterns, DAMP) or exogenous (pathogen-associated molecular patterns, PAMP) triggers, collectively referred to as ‘alarmins’, that, upon binding to pattern-recognition receptors, lead to the maturation of antigen-presenting cells which is crucial for the activation of naïve T cells. Early studies on FVIII immunogenicity investigated the possibility that FVIII directly induces the maturation of antigen-presenting cells. While the first domain of hemagglutinin (HA1) was found to induce sig- nificantly higher levels of anti-HA1 IgG in intravenously injected mice when fused to the FVIII light chain,27 co- incubation of full-length FVIII with Toll-like receptor 2-
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