Bioencapsulated Factor IX for Oral Tolerance in Hemophilia B

描述（由申请方提供）：血友病是由编码凝血因子VIII（血友病A）或因子IX（血友病B）的基因突变引起的X连锁出血性疾病。目前的治疗是基于血浆衍生或重组凝血因子浓缩物的静脉输注。最近，基因治疗取得了重大进展。治疗性凝血因子的抑制性抗体（“抑制剂”）的形成代表了治疗的严重并发症。通过目前的免疫耐受诱导方案消除抑制剂是昂贵的，并不总是有效的，并且对于针对因子IX（F.IX）的抗体不太成功。没有预防性方案来防止抑制剂形成。由于抑制剂的形成依赖于T辅助，因此CD4+ T细胞代表了耐受诱导的替代靶点。已知抗原施用的特定途径如粘膜（口服或鼻）诱导调节性T细胞和T细胞耐受性。该建议基于粘膜抗原给药或粘膜和全身抗原递送的组合可以防止抑制剂形成的假设。为了开发有效的、成本有效的和临床上可应用的口服耐受性方案，我们将产生在叶绿体中表达高水平的人F.IX（hF.IX）的转基因植物.将生物包封的hF.IX喂给小鼠以测试在重复静脉内注射hF.IX蛋白后对hF.IX抗体形成的预防。因为该方法基于对hF.IX衍生的肽表位特异性的CD4+ T细胞的抗原呈递，所以不需要口服递送的hF.IX的生物活性。将在具有大F.IX基因缺失的血友病B小鼠中评价优化方案的耐受性诱导。选择已知经常形成hF.IX抑制剂的小鼠品系用于这些研究。我们假设，最佳的活化调节性T细胞后，介绍了植物来源的hF.IX抗原将导致能够提供蛋白质或基因为基础的治疗血友病B没有抑制剂的形成。该方案还将应用于因既往蛋白质治疗而预先存在抑制剂的动物，以研究消除抑制剂的能力。在未来的研究中，口服耐受方案将进一步优化，并适用于预防血友病A治疗中的因子VIII抑制剂。

DESCRIPTION (provided by applicant): Hemophilia is the X-linked bleeding disorder caused by mutations in the gene encoding blood coagulation factor VIII (hemophilia A) or factor IX (hemophilia B). Current treatment is based on intravenous infusion of plasma-derived or recombinant clotting factor concentrates. More recently, substantial progress toward gene therapy has been made. Formation of inhibitory antibodies ("inhibitors") to the therapeutic clotting factor represents a serious complication of treatment. Elimination of inhibitors by current immune tolerance induction protocols is expensive, not always effective, and less successful for antibodies against factor IX (F.IX). No prophylactic protocols to prevent inhibitor formation are available. Since inhibitor formation is dependent on T help, CD4+ T cells represent an alternative target for tolerance induction. Specific routes of antigen administration such as mucosal (oral or nasal) are known to induce regulatory T cells and T cell tolerance. This proposal is based on the hypothesis that mucosal antigen administration or a combination of mucosal and systemic antigen delivery can prevent inhibitor formation. In order to develop an effective, cost- efficient, and clinically applicable oral tolerance protocol, we will generate transgenic plants expressing high levels of human F.IX (hF.IX) in the chloroplast. Bio-encapsulated hF.IX will be fed to mice to test for prevention of antibody formation to hF.IX upon repeated intravenous injected of hF.IX protein. Because this approach is based on antigen presentation to CD4+ T cells specific for hF.IX-derived peptide epitopes, biological activity of orally delivered hF.IX is not required. An optimized protocol will be evaluated for tolerance induction in hemophilia B mice with a large F.IX gene deletion. A strain of mice known to frequently form inhibitors to hF.IX is chosen for these studies. We hypothesize that optimal activation of regulatory T cells following presentation of plant-derived hF.IX antigen will result in the ability to provide protein- or gene-based therapy for hemophilia B without inhibitor formation. The protocol will also be applied to animals with pre-existing inhibitors due to prior protein therapy in order to investigate the ability to eliminate inhibitors. In future studies, the oral tolerance protocol will be further optimized and adapted to prevention of inhibitors to factor VIII in treatment of hemophilia A.