A Novel BIV vector pseudotyped with thogoto virus gp75

一种新型 BIV 载体，用 thogoto 病毒 gp75 假型化

• 批准号: 7108387
• 负责人: TIANCI LUO
• 金额: $ 14.98万
• 依托单位: ADVANCED VISION THERAPIES, INC.
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2008-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7108387
• 关键词: Lentivirus; Orthomyxoviridae; biotechnology; complement inhibitors; complement pathway regulation; evaluation /testing; eye fundus photography; gene delivery system; laboratory mouse; microinjections; ophthalmoscopy; retina; technology /technique development; transfection /expression vector; virus envelope; virus infection mechanism; virus receptors

DESCRIPTION (provided by applicant): Lentivirus-based gene transfer systems represent a promising gene delivery technology, as they integrate into the genome of the target cell and mediate sustained expression of the transferred gene. Advanced Vision Therapies, Inc. (AVT) has developed a proprietary lentiviral vector system based on the bovine immunodeficiency virus (BIV), an animal lentivirus not associated with human disease. The BIV vectors combine the transduction efficiency of the HIV-based vectors with the safety advantages of animal-based lentiviral vector systems. Importantly, BlV-vector mediated delivery of an anti-angiogenic transgene efficiently blocked retinal neovascularization in a relevant rodent model, suggesting that the AVT vector is suitable for clinical applications. Lentiviruses are routinely pseudotyped with heterologous viral envelopes to broaden vector tropism. The most widely used envelope is derived from the vesciular stomatitis virus glycoprotein (VSV-G). However, VSV-G has several limitations including cytotoxicity and inactivation by human complement. Therefore, a variety of alternative envelopes have been explored for use in pseudotyping lentiviral vectors including the baculovirus gp64. AVT was recently successful in the generation of a gp64 envelope protein-pseudotyped BIV vector. High vectors titers were obtained, the vectors were stable, and importantly, gp64 can be constitutively expressed in cells without toxicity (an important consideration in the development of a BIV producer cell line). However, gp64 is inactivated by human complement. In this Phase I application, AVT will explore the use of a novel envelope derived from the Thogoto virus. Thogoto virus is transferred to human blood through ticks, and therefore, may display resistance to human complement. Interestingly, the Thogoto gp75 envelope glycoprotein displays significant homology to the baculovirus gp64 envelope glycoprotein, potentially due to the tropism of Thogoto for arthropods and human hosts. AVT has successfully generated a high titer BIV vector pseudotyped with the Thogoto virus gp74 envelope glycoprotein. This Phase I project will focus on the further evaluation of this novel vector. There are 3 specific aims for this Phase I project. Specific Aim 1: Evaluation of cellular tropism of Thogoto virus gp75- pseudotyped BIV vectors. A panel of cell lines and primary cells will be evaluated for transduction efficiency using Thogoto gp75-pseudotyped GFP. Specific Aim 2: Evaluation of Thogoto-pseudotyped BIV vectors in rodents. Both VSV-G and baculovirus gp64 envelope glycoprotein- pseduotyped vectors specifically transduce RPE cells following a subretinal injection in rodents. The tropism of the Thogoto-pseudotyped vector will be evaluated in rodents. Specific Aim 3: Determination of the human complement resistance of the Thogoto-pseudotyped BIV vector. Phase II studies will further evaluate BIV vector physical properties, including stability, Thogoto gp75 cytotoxicity, and vector purification strategies.

描述（由申请人提供）：基于慢病毒的基因转移系统代表了一种有前途的基因递送技术，因为它们整合到靶细胞的基因组中并介导转移基因的持续表达。 Advanced Vision Therapies, Inc. (AVT) 开发了一种基于牛免疫缺陷病毒 (BIV) 的专有慢病毒载体系统，BIV 是一种与人类疾病无关的动物慢病毒。 BIV 载体结合了基于 HIV 的载体的转导效率和基于动物的慢病毒载体系统的安全优势。重要的是，BlV载体介导的抗血管生成转基因的传递有效地阻断了相关啮齿动物模型中的视网膜新生血管形成，这表明AVT载体适合临床应用。慢病毒通常用异源病毒包膜进行假型化，以扩大载体趋向性。最广泛使用的包膜源自水泡性口炎病毒糖蛋白（VSV-G）。然而，VSV-G 有一些局限性，包括细胞毒性和人补体失活。因此，人们已经探索了多种替代包膜用于假型慢病毒载体，包括杆状病毒 gp64。 AVT 最近成功生成了 gp64 包膜蛋白假型 BIV 载体。获得了高载体滴度，载体稳定，重要的是，gp64可以在细胞中组成型表达而无毒性（开发BIV生产细胞系的重要考虑因素）。然而，gp64 会被人类补体灭活。在此一期应用中，AVT 将探索使用源自 Thogoto 病毒的新型包膜。托戈托病毒通过蜱虫转移到人类血液中，因此可能对人类补体表现出抗性。有趣的是，Thogoto gp75 包膜糖蛋白与杆状病毒 gp64 包膜糖蛋白表现出显着的同源性，这可能是由于 Thogoto 对节肢动物和人类宿主的向性。 AVT 已成功生成用 Thogoto 病毒 gp74 包膜糖蛋白假型化的高滴度 BIV 载体。该第一阶段项目将侧重于对这种新型载体的进一步评估。第一阶段项目有 3 个具体目标。具体目标 1：评估 Thogoto 病毒 gp75 假型 BIV 载体的细胞向性。将使用 Thogoto gp75 假型 GFP 评估一组细胞系和原代细胞的转导效率。具体目标 2：在啮齿动物中评估 Thogoto 假型 BIV 载体。 VSV-G 和杆状病毒 gp64 包膜糖蛋白假型载体在啮齿动物视网膜下注射后特异性转导 RPE 细胞。 Thogoto 假型载体的趋向性将在啮齿类动物中进行评估。具体目标 3：确定 Thogoto 假型 BIV 载体的人类补体抗性。 II 期研究将进一步评估 BIV 载体的物理特性，包括稳定性、Thogoto gp75 细胞毒性和载体纯化策略。