Canine G-CSF gene transfer

描述（由申请人提供）：循环中性粒细胞减少症是一种罕见的疾病，发生在人和灰牧羊犬。复发性严重中性粒细胞减少症导致细菌感染，缩短预期寿命。长期每日给药重组G-CSF可消除严重复发性中性粒细胞减少症，为犬和患者提供治疗。本研究将研究使用两种不同的方法通过输送G-CSF对灰牧羊犬进行循环造血的治疗。在一个目的，我们将研究慢病毒介导的犬G-CSF的体内递送。在另一个目标中，G-CSF将通过在生物隔离装置中使用转导异体细胞的细胞植入物来递送，从而消除对免疫抑制的需要。我们之前的研究表明，长期给药重组犬G-CSF可以消除患病犬的复发性造血循环，我们的数据显示，给药编码犬G-CSF的慢病毒后，患病犬的中性粒细胞产量增加。我们将编码慢病毒的G-CSF注射到灰牧羊犬和正常狗、大鼠的肌肉中。我们将研究在生物隔离装置中植入转导的血管平滑肌细胞，以在大鼠、正常狗和灰牧羊犬中长期安全表达G-CSF。选择这些靶细胞是为了便于培养、可移植性和允许转导基因长期稳定表达的潜力。

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.