Mechanisms of Race-Based Differences in Factor VIII Immunogenicity in Hemophilia

血友病因子 VIII 免疫原性的种族差异机制

• 批准号: 7939678
• 负责人: Tom Eugene Howard
• 金额: $ 295.87万
• 依托单位: SEPULVEDA RESEARCH CORPORATION
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7939678
• 关键词: Academia; Address; African; African American; Alleles; Amino Acid Sequence; Amino Acids; Antibodies; Area; Arts; B-Lymphocyte Epitopes; B-Lymphocytes; Binding; Biological Assay; Blood coagulation; Blood specimen; C2 Domain; Canada; Cell Lineage; Characteristics; Clinical; Code; Collaborations; Communities; Core Facility; DNA; DNA Sequence; Data; Databases; Development; Disease; Endogenous Factors; Enrollment; Enzyme-Linked Immunosorbent Assay; Epitopes; Exons; Factor VIII; Frequencies; Funding; Future; Genetic; Genetic Polymorphism; Genotype; Growth and Development function; Haplotypes; Hemophilia A; Hemorrhage; Human; Immune; Immune Response Genes; Immune Tolerance; Immune response; Immunodominant Epitopes; Immunology; Incidence; Individual; Industry; Infusion procedures; Introns; Laboratories; Laboratory Research; Life; Limb structure; Los Angeles; Measures; Medicine; Methods; Microarray Analysis; Minority; Mutation; Patients; Peptides; Plasma; Population; Preclinical Testing; Prevalence; Proteins; Protocols documentation; Race; Regulatory T-Lymphocyte; Replacement Therapy; Research; Research Ethics Committees; Research Infrastructure; Research Personnel; Resources; Risk; Risk Factors; Sampling; Scientist; Signal Pathway; Site; South Africa; Structure; Subgroup; Surface Plasmon Resonance; T-Lymphocyte; Testing; Therapeutic; Time; Tissues; Transfusion; Variant; base; caucasian American; design; genetic risk factor; genetic variant; genome wide association study; genome-wide; immunogenic; immunogenicity; improved; inhibitor/antagonist; neutralizing antibody; novel; patient population; polarized cell; public health relevance; recombinant antihemophilic factor VIII; repository; response; therapeutic protein; treatment center

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.

描述（由申请人提供）： 拟议的研究将阐明对用于阻止血友病 A (HA) 患者出血的输注因子 VIII (FVIII) 的免疫反应机制，其中约 25% 的患者会产生“抑制剂”（中和抗体），从而使治疗变得更加复杂造成危及生命和肢体的出血。这些项目寻求对黑人 HA 患者的抑制剂发生率是白人患者的两倍这一观察结果的解释。临床合作者包括 15 个美国血友病治疗中心 (HTC)，其患者群体中包括大量非裔美国人，以及南非的国家 HTC 网络。作为由 NIH/NICHD 资助的血友病生长和发育研究 (HGDS) 的一部分以及从加拿大储存库收集的 DNA 和血浆也将进行研究。位于洛杉矶（加利福尼亚州）、西雅图（华盛顿州）、圣安东尼奥（德克萨斯州）和伯灵顿（佛蒙特州）的实验室合作者将研究抑制剂形成的个体间差异以及抑制剂形成的体液和细胞方面的遗传基础。使用他们开发的既定检测方法对 FVIII 进行免疫反应。高通量 DNA 测序和其他基于 PCR 的检测将鉴定导致血友病的 F8 突变以及所有 F8 非同义单核苷酸多态性 (ns-SNP) 的等位基因，这些基因是单个编码 DNA 碱基的改变，导致氨基酸FVIII 蛋白的变化。先前对 F8 单倍型的分析（Viel 等人，NEJM 2009）将得到扩展和完善。在该研究中，内源性（自身）FVIII 蛋白在 ns-SNP 对应的位点与通常输注的 FVIII 蛋白等位不匹配的黑人 HA 患者，其抑制剂的频率明显高于内源性 FVIII 与治疗性 FVIII 匹配的黑人患者。拟议的研究大大增加了受试者数量，这将为协变量提供更好的控制，并提供足够的统计能力来进行亚组分析。该研究利用敏感的单倍型特异性测试，甚至能够识别低滴度（浓度）和非中和性抗 FVIII 抗体，而传统的 Bethesda 检测无法检测到这些抗体。微阵列将用于对近 2,000,000 个 SNP 和拷贝数变异进行基因分型，以进行全基因组分析，最初集中于位于已知免疫反应基因的子集。最后，基于 HLA-II 类四聚体的测定将直接检验以下假设：由含有 ns-SNP 的 F8 位点编码的氨基酸序列包含免疫显性表位，当同种异体错配时，该表位会刺激 HA 患者的 T 细胞。还将通过表面等离子共振测定直接测试抗原性，以（1）评估患者来源的抗体与对应于不同F8单倍型的重组FVIII蛋白的结合，以及（2）确定这些抗体识别的表位。如果这些假设得到证实，这将推动新型、免疫原性较低的 FVIII 替代蛋白的临床前测试，最终消除 A 型血友病中这种长期已知的基于种族的差异，并有可能在概念上扩展到输血医学和蛋白质治疗的其他领域。 公共卫生相关性： 在血友病治疗中心和专家实验室的合作下，将研究血友病 A 中因子 VIII 的免疫反应机制以及抑制剂发展的危险因素。先前建议的抑制剂风险可能与患者的种族决定不匹配 背景VIII因子结构和外源治疗性VIII因子产品的VIII因子结构将进一步研究。将探讨更适合少数群体接受者的治疗因子 VIII 的可能性。