Molecular engineering of factor VIII gene for rAAV delivery

描述（由申请人提供）：在人类人口中，大约每5000名男性中就有1人患有凝血障碍，血友病a。这种疾病主要是由位于x染色体上的因子VIII基因缺乏引起的，传统医学难以治疗。目前通过静脉输注因子VIII浓缩物治疗A型血友病是非常昂贵的，并且具有开发抑制剂的潜在副作用。另一方面，基因疗法可以潜在地防止当前治疗方法的这些局限性。虽然重组腺相关病毒（rAAV）载体很有希望传递因子VIII基因，但由于其大小限制（限制在~5kb）， AAV载体技术在A型血友病中的应用滞后于其他遗传性疾病。为了改善利用rAAV载体的因子VIII基因传递，我们开发了新的工程因子VIII分子，用于双载体和单载体策略。本提案的具体目标是：1)。开发一种高效分泌的因子VIII重链分子；2）. 开发一种新的rAAV传递小因子VIII基因；3)。目的分析对工程因子VIII转基因产物的免疫应答。这一建议的成功可能导致使用AAV载体的血友病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.