DESCRIPTION (provided by applicant): The proposed study will elucidate mechanisms of immune responses to the infused factor VIII (FVIII) used to arrest bleeding in hemophilia A (HA) patients, in which ~25% of patients develop "inhibitors" (neutralizing antibodies) that can greatly complicate their treatment by causing life- and limb-threatening bleeds. The projects seek explanations for the observation that inhibitor incidence in black HA patients is twice that of white patients. Clinical collaborators include 15 U.S. hemophilia treatment centers (HTCs) with patient populations that include large numbers of African Americans, and the National HTC network in South Africa. DNA and plasma collected as part of the Hemophilia Growth and Development Study (HGDS) funded by the NIH/NICHD, and also from a Canadian Repository, will also be studied. Laboratory collaborators -- based in Los Angeles (CA), Seattle (WA), San Antonio (TX), and Burlington (VT) -- will investigate the genetic basis for inter-individual variability in inhibitor formation, and humoral and cellular aspects of immune responses to FVIII, using established assays that they developed. High-throughput DNA sequencing and other PCR-based assays will identify the F8 mutations causing hemophilia as well as the alleles of all F8 nonsynonymous-singlenucleotide polymorphisms (ns-SNPs), which are alterations in a single coding DNA base that result in amino acid changes in the FVIII protein. A previous analysis of F8 haplotypes (Viel et al, NEJM 2009) will be expanded and refined. In that study, black HA patients whose endogenous (self) FVIII proteins were allelically mismatched, at sites corresponding to ns-SNPs, with commonly infused FVIII proteins had a significantly higher frequency of inhibitors than black patients whose endogenous FVIII matched the therapeutic FVIII. The proposed study greatly increases the number of subjects, which will provide improved control for covariates and adequate statistical power to perform subgroup analyses. The study utilizes sensitive, haplotype-specific tests capable of identifying even low-titer (concentration) and non-neutralizing anti-FVIII antibodies, which are not detectable by conventional Bethesda assays. Microarrays will be used to genotype almost 2,000,000 SNPs and copy number variants for genome-wide analyses, concentrating initially on the subset located in known immune response genes. Finally, HLA-class II tetramer-based assays will test directly the hypothesis that amino acid sequences encoded by F8 loci containing ns-SNPs comprise immunodominant epitopes that, when allogeneically mismatched, stimulate T cells of HA patients. Antigenicity will also be tested directly by surface plasmon resonance assays to (1) evaluate binding of patient-derived antibodies to recombinant FVIII proteins corresponding to different F8 haplotypes, and (2) determine the epitopes recognized by these antibodies. If the hypotheses are confirmed, this will motivate preclinical testing of novel, less immunogenic FVIII replacement proteins that could ultimately eradicate this long known race-based disparity in hemophilia A, and potentially extend in concept to other areas of transfusion medicine and protein therapeutics. PUBLIC HEALTH RELEVANCE: In a collaboration of hemophilia treatment centers and expert laboratories, the mechanisms of immune response to factor VIII in hemophilia A and the risk factors for inhibitor development will be studied. The previously-suggested inhibitor risk of a possible mis-match between a patient's racially-determined background factor VIII structure and the factor VIII structure of exogenous therapeutic factor VIII products will be further studied. The possibility of a therapeutic factor VIII, better matched to minority recipients, will be explored.
|Effective start/end date||30/09/09 → 31/08/10|
- National Heart, Lung, and Blood Institute: $2,958,699.00
- National Heart, Lung, and Blood Institute: $3,548,764.00
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