Bioengineered factor VIII gene therapy for hemophilia A

描述（由申请人提供）：本提案的总体目标是对血友病A患者进行基因治疗临床试验，包括遗传修饰的造血干细胞（HSC）。在最近的一系列研究中，我们已经表明，基因工程改造的HSC的移植可以恢复因子VIII（fVIII）的活性，在血友病A小鼠的治疗水平，人HSC很容易与编码基因工程改造的fVIII转基因的重组慢病毒载体转导。我们的基因治疗方法使用埃默里大学设计的生物工程fVIII转基因，并含有关键的高表达序列，我们已经证明这是必要的，足以达到治疗/治愈性fVIII表达水平。在临床前鼠研究中使用生物工程fVIII构建体（命名为ET-3），我们常规地在植入1 - 5%遗传修饰的造血细胞的小鼠中观察到正常的fVIII活性水平（1单位/ml）。这些植入水平与基于人类HSC的临床基因治疗试验中已经达到的水平相似。相比之下，我们和其他人已经表明，使用全人fVIII序列的类似策略在血友病A小鼠临床前模型中不能达到这些稳健的fVIII活性水平。我们已经使用ET-3转基因产生了广泛的临床前数据，这些数据证明了用编码ET-3的慢病毒载体进行基因工程改造的HSC，加上非清髓性移植方案，可用于治疗和可能治愈血友病A。此外，还举行了一次有利的IND前会议，为最终确定我们的临床前数据包和载体生产提供了必要的指导。我们现在建议1）使用临床级（GMP）lentivector产品生成后期临床前数据，2）完成批准进行试点基因治疗临床试验所需的监管申报，3）在拟议试验中招募和治疗7例血友病A患者。有四个组织为实现这些目标结成了伙伴关系，其中包括：i）Expression Therapeutics，LLC，一家基于高表达fVIII技术成立的生物技术公司，ii）Emory大学，其中已经进行了大部分临床前研究，iii）亚特兰大儿童保健，Emory大学基因治疗计划的财政支持者和Expression Therapeutics的投资者，以及iv）Lentigen公司，一家致力于慢病毒载体成功临床应用的公司。Lentigen将生产用于拟定研究（包括临床试验）的临床lentivecto产品，该试验将由在干细胞移植和血友病A治疗方面具有临床专业知识的专家主要研究者在埃默里大学进行。目前II期SBIR提案的主要里程碑是将ET-3基因治疗概念和项目从后期临床前研究阶段转化为首次人体试验。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to conduct a gene therapy clinical trial incorporating genetically modified hematopoietic stem cells (HSCs) for persons with hemophilia A. In a series of recent studies, we have shown that the transplantation of genetically-engineered HSCs can restore factor VIII (fVIII) activity to curative levels in hemophilia A mice and that human HSCs are readily transduced with recombinant lentivector encoding a genetically-engineered fVIII transgene. Our gene therapy approach uses a bioengineered fVIII transgene designed at Emory University and contains critical high-expression sequences, which we have shown are necessary and sufficient to achieve therapeutic/curative fVIII expression levels. Using the bioengineered fVIII construct, designated ET-3, in preclinical murine studies we routinely observe normal fVIII activity levels (1 unit/ml) i mice engrafted with 1 - 5% genetically-modified hematopoietic cells. These engraftment levels are similar to those that have already been achieved in human HSC-based clinical gene therapy trials. In contrast, we and others have shown that similar strategies using fully human fVIII sequences do not achieve these robust fVIII activity levels in the hemophilia A mouse preclinical model. We have generated extensive preclinical data using the ET-3 transgene, which demonstrate proof-of-concept that HSCs genetically engineered with ET-3-encoding lentivector, coupled with a non-myeloablative transplant regimen, can be used to treat and possibly cure hemophilia A. In addition, a favorable pre-IND meeting was held that provided the necessary guidance needed to finalize our preclinical data package and vector manufacturing. We now propose to 1) generate late-stage preclinical data using clinical-grade (GMP) lentivector product, 2) complete the regulatory submissions necessary for approval to conduct a pilot gene therapy clinical trial and 3) recruit and treat 7 patients with hemophilia A in the proposed trial. Four organizations have partnered to accomplish these aims, including: i) Expression Therapeutics, LLC, a biotechnology company founded on the high expression fVIII technology, ii) Emory University, where the majority of the preclinical studies have been performed, iii) Children's Healthcare of Atlanta, a financial supporter of the Gene Therapy Program at Emory University and investor in Expression Therapeutics, and iv) Lentigen Corporation, a company dedicated to the successful clinical application of lentivectors. Lentigen will generate the clinical lentivecto product that will be used in the proposed studies including the clinical trial, and the trial will e conducted at Emory University by expert principal investigators with clinical expertise in stem cell transplantation and the treatment of hemophilia A. The chief milestone of the current Phase II SBIR proposal is the translation of the ET-3 gene therapy concept and project from the late-pre-clinical study phase into a first-in-man trial.