Eicosanoid Gene Therapy

描述(申请人提供)：该项目将为原发性开角型青光眼(POAG)提供基于慢病毒载体的基因治疗。在之前的资金周期中，我们建立了一个强大的载体平台和实验能力，用于将基因输送到涉及房水动力学的前房结构。我们发现慢病毒载体可以有效、无毒地转导小梁网络，并获得与POAG慢性化程度相适应的持续表达。在家猫和食蟹猴身上建立了两个大型动物模型。在这里，我们探索了二十烷类化合物的基因治疗潜力，这是一种强大的脂类信号分子，包括前列腺素、血栓素类和白三烯。我们推测，由于PGF21激动剂已被证明具有降低眼压的活性，我们可以通过基因治疗来操纵眼睛中的前列腺素生理学，以实现持续的眼压降低。在初步研究中，我们在体外和体内设计了一个完整的前列腺素生物合成和反应途径，最初的重点是环氧合酶-2(COX-2)和PGF21途径，因为它在眼压生理学中的已知作用以及它作为药物靶点(拉坦前列素和相关药物)的成功开发。我们获得了合成的、序列优化的cDNA，这对于克服极端的mRNA易感性是必不可少的。这种基因疗法可以持续、无毒地降低猫的眼压。我们将开发和使用慢病毒载体进行组织培养试验、灌流人类供体眼和两种动物模型，以建立实用的、完全翻译的降低眼压基因疗法。先进的慢病毒载体表达变量和可选择的PG通路将被系统地研究，以了解二十烷基类化合物对前房生理的影响，并最大限度地发挥基因治疗的潜力。与公共卫生相关。该项目将开发基于慢病毒载体的基因治疗原发性开角型青光眼(POAG)。我们将确定前列腺素的最佳生物合成和反应途径，以持续降低眼压，并获得有关前列腺素在眼睛中的作用的新知识。

DESCRIPTION (provided by applicant): This project will provide lentiviral vector-based gene therapy for primary open angle glaucoma (POAG). In the previous funding cycle we established a robust vector platform and experimental capacity for gene delivery to anterior chamber structures involved in aqueous humor dynamics. We showed that lentiviral vectors could effectively and non-toxically transduce the trabecular meshwork and achieve sustained expression commensurate to the chronicity of POAG. Two large animal models were established in the domestic cat and Cynomolgus monkey. Here we have explored the gene therapy potential of eicosanoids, the potent lipid- derived signaling molecules that include the prostaglandins, thromboxanes and leukotrienes. We hypothesize that since PGF21 agonists have proven intraocular pressure (IOP)-lowering activity, we can manipulate prostaglandin physiology in the eye via gene therapy to achieve sustained IOP reduction. In preliminary studies, we engineered a complete prostaglandin biosynthesis and response pathway in vitro and in vivo, with initial focus on cyclooxygenase-2 (COX-2) and on the PGF21 pathway because of its known role in IOP physiology and its successful exploitation as a pharmacologic target (latanoprost and related drugs). We derived synthetic, sequence optimized cDNAs that were essential to overcoming extreme mRNA liability. This gene therapy results in sustained, non-toxic IOP reduction in the cat. We will develop and use lentiviral vectors in tissue culture assays, perfused human donor eyes, and the two animal models to establish a practical, fully translational IOP-reduction gene therapy. Advanced lentiviral vector expression variables and alternative PG pathways will be systematically studied to learn about the effects of eicosanoids on anterior chamber physiology and maximize gene therapy potential. PUBLIC HEALTH RELEVANCE. The project will develop lentiviral vector-based gene therapy for primary open angle glaucoma (POAG). We will determine optimal prostaglandin biosynthesis and response pathways for sustained IOP reduction and we will gain new knowledge about the roles of prostaglandins in the eye.