Canine G-CSF gene transfer

描述(申请人提供)：循环性中性粒细胞减少症是一种罕见的疾病，在人类和灰色牧羊犬中都会发生。反复出现的严重中性粒细胞减少症会导致细菌感染和寿命缩短。长期每日给予重组G-CSF可消除严重的复发中性粒细胞减少症，并为狗和患者提供治疗。这项研究将研究用两种不同的方法注射G-CSF来治疗周期性造血的灰柯利犬。在一个目标中，我们将在体内研究慢病毒介导的犬G-CSF的传递。在另一个目标中，G-CSF将通过在生物隔离设备中使用转导的同种异体细胞的细胞植入来传递，从而消除了对免疫抑制的需要。我们之前已经证明，长期给予重组犬G-CSF可以消除受影响狗的反复造血循环，我们有数据显示，在感染编码犬G-CSF的慢病毒的狗中，中性粒细胞的产生增加。我们将编码慢病毒的G-CSF注射到灰色牧羊犬和正常狗和大鼠的肌肉中。我们将研究将转导的血管平滑肌细胞植入生物隔离装置中，以提供G-CSF在大鼠、正常狗和灰色牧羊犬中的安全长期表达。选择这些靶细胞是为了便于培养，可移植，以及允许转导基因长期稳定表达的潜力。 具体目的是：L。用G-CSF慢病毒感染灰牧羊犬、正常犬和大鼠，以确定载体介导的细胞因子表达的持续时间和病毒再给药的可行性。我们将确定病毒的组织分布。2.研究利用生物隔离装置向大鼠和犬体内输送持续治疗水平的G-CSF。我们相信，这项研究将证明基因转移可以用于循环性中性粒细胞减少症的长期治疗，并将应用于其他中性粒细胞减少症和对激素和凝血因子分泌反应的遗传性疾病的治疗。这项提议的总体目标是在一个可应用于患者治疗的大型动物模型中产生临床数据。

DESCRIPTION (provided by applicant): Cyclic neutropenia is a rare disease that occurs both in man and grey collie dogs. The recurrent severe neutropenia leads to bacterial infections and shortened life expectancy. Long-term daily administration of recombinant G-CSF eliminates the severe recurrent neutropenia and provides therapy for dogs and patients. This research will study the treatment of grey collie dogs with cyclic hematopoiesis by delivery of G-CSF using two different approaches. In one aim we will study in vivo lentivirus-mediated delivery of canine G-CSF. In another aim G-CSF will be delivered by cell implants employing transduced allogeneic cells in bioisolator devices that eliminate the need for immunosuppression. We have previously shown that long-term administration of recombinant canine G-CSF abolishes the recurrent hematopoietic cycling in affected dogs and we have data showing increased neutrophil production in an affected dog administered lentivirus encoding canine G-CSF. We will administer G-CSF encoding lentivirus to muscle of grey collie and normal dogs and rats. We will investigate implantation of transduced vascular smooth muscle cells in bio-isolator devices to provide safe long-term expression of G-CSF in rats, normal dogs and grey collies. These target cells were chosen for ease of culture, transplantability and potential to allow stable long-term expression of transduced genes. The Specific Aims are: l. To administer G-CSF lentivirus to grey collie, normal dogs and rats to determine the duration of vector-mediated cytokine expression and the feasibility of virus re-administration. We will determine the tissue distribution of virus. 2. To investigate the use of bioisolator devices to deliver sustained therapeutic levels of G-CSF to rats and dogs. We believe this research will demonstrate that gene transfer can be used for long-term treatment of cyclic neutropenia, and will have an application in the treatment of other neutropenias and genetic disorders responding to secreted hormones and clotting factors. The overall goal of this proposal is to generate clinical data in a large animal model that can be applied to the treatment of patients.