Bioencapsulated Factor IX for Oral Tolerance in Hemophilia B

描述(申请人提供)：血友病是由编码凝血因子(血友病A)或凝血因子IX(血友病B)基因突变引起的X连锁出血性疾病。目前的治疗方法是静脉输注血浆衍生或重组凝血因子浓缩物。最近，基因治疗取得了实质性进展。形成针对治疗性凝血因子的抑制性抗体(“抑制物”)是治疗的严重并发症。通过目前的免疫耐受诱导方案消除抑制物是昂贵的，并不总是有效的，并且对抗因子IX(F.IX)抗体的成功率较低。目前还没有预防抑制剂形成的预防方案。由于抑制物的形成依赖于T细胞的帮助，因此，CD4+T细胞是诱导耐受的另一种靶点。已知特定的抗原给药途径，如粘膜(口腔或鼻腔)，可诱导调节性T细胞和T细胞耐受。这一建议是基于这样的假设，即粘膜抗原注射或粘膜和全身抗原输送的组合可以防止抑制物的形成。为了开发一种有效的、经济的和临床适用的口服耐受方案，我们将获得在叶绿体中高水平表达人F.IX(hF.IX)的转基因植株。将生物包裹的hF.IX喂给小鼠，以测试重复静脉注射hF.IX蛋白是否能防止hF.IX抗体的形成。因为这种方法是基于hF.IX衍生的多肽表位的抗原呈递给CD4+T细胞，所以口服hF.IX的生物活性是不必要的。将评估一种优化的方案，以诱导具有较大F.IX基因缺失的血友病B小鼠的耐受性。一种已知经常形成hF.IX抑制剂的小鼠品系被选为这些研究的对象。我们假设，在呈现植物来源的hF.IX抗原后，调节性T细胞的最佳激活将导致在没有抑制物形成的情况下提供基于蛋白质或基因的治疗血友病B的能力。该方案还将应用于由于先前的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，口服耐受性方案将进一步优化和调整，以预防血友病A治疗中凝血因子VIII的抑制剂。

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.