Development of highly efficient factor VIII mini-gene therapy

 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）载体有望用于递送因子VIII基因，但由于其大小限制（仅限于约5 kb）和因子VIII蛋白的低效分泌，将AAV载体技术应用于血友病A基因治疗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，我们将开发一种新的载体。 具有增强的表达和分泌的新的人因子VIII分子。这项建议的具体目标是：1）。开发具有改善的分泌和表达的人因子VIII分子; 2）.开发具有增强的比活性和最小的氨基酸改变的人因子VIII分子; 3）。目的优化重组腺相关病毒（AAV）因子VIII包装表达盒，并在血友病动物模型中进行临床前研究。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 gene therapy lagged behind other genetic diseases because of this size constraint (limited to ~5kb) and inefficient secretion of factor VIII protein. To improve factor VIII gene delivery utilizing rAAV vectors, we will develop a novel human factor VIII molecules with enhanced expression and secretion. The specific aims for this proposal are: 1). To develop a human factor VIII molecule with improved secretion and expression; 2). To develop a human factor VIII molecule with enhanced specific activity with minimal amino acid alteration; 3). To optimize the AAV factor VIII packaging and expression cassette and carry out preclinical studies in Hemophilia Animal Model. The success of this proposal may lead to a clinical trial of hemophilia A using AAV vectors.