Molecular engineering of factor VIII gene for rAAV delivery

描述（由申请方提供）：人类人群中约有1/5000的男性患有凝血障碍，血友病A。这种疾病主要是由位于X染色体上的因子VIII基因缺陷引起的，并且难以通过常规药物治疗。目前通过静脉输注因子VIII浓缩物治疗血友病A非常昂贵，并且具有产生抑制剂的潜在副作用。另一方面，基因治疗可以潜在地防止当前治疗的这些局限性。虽然重组腺相关病毒（rAAV）载体有希望用于递送因子VIII基因，但由于这种大小限制（限于~ 5 kb），将AAV载体技术应用于血友病A落后于其他遗传性疾病。为了改善利用rAAV载体的因子VIII基因递送，我们已经开发了用于双载体和单载体策略的新型工程化因子VIII分子。这项建议的具体目标是：1）。开发对分泌有效的因子VIII重链分子; 2）.开发和表征用于rAAV递送的新型小因子VIII基因; 3）。分析针对工程化因子VIII转基因产物的免疫应答。这一提议的成功可能会导致使用AAV载体进行血友病A的临床试验。

DESCRIPTION (provided by applicant): Approximately one in 5000 males in human population suffers from coagulation disorder, hemophilia A. This disease is primarily caused by deficiency in the factor VIII gene located in the X-chromosome and is difficult to treat by conventional medicine. Current treatment of hemophilia A by intravenous infusion of factor VIII concentrates is very costly and has a potential side effect of developing inhibitors. Gene therapy, on the other hand, can potentially prevent these limitations of current treatments. Although recombinant adeno-associated virus (rAAV) vectors are promising for deliver factor VIII gene, applying AAV vector technology to Hemophilia A lagged behind other genetic diseases because of this size constraint (limited to ~5kb). To improve factor VIII gene delivery utilizing rAAV vectors, we have developed novel engineered factor VIII molecules for use in both dual vectors and single vector strategy. The specific aims for this proposal are: 1). To develop a Factor VIII heavy chain molecule efficient for secretion; 2). To develop and characterize a novel mini factor VIII gene for rAAV delivery; 3). To analyze immune responses against the engineered factor VIII transgene products. The success of this proposal may lead to a clinical trial of hemophilia A using AAV vectors.