Department of Ophthalmology and Vision Science, University of California Davis, Davis, California, USA.
Department of Cell Biology and Human Anatomy, University of California Davis, Davis, California, USA.
Hum Gene Ther. 2021 Jul;32(13-14):682-693. doi: 10.1089/hum.2020.281. Epub 2021 Apr 8.
The suprachoroid is a potential space located between the sclera and choroid of the eye, which provides a novel route for ocular drug or viral vector delivery. Suprachoroidal injection of adeno-associated virus (AAV)8 using transscleral microneedles enables widespread transgene expression in eyes of nonhuman primates, but may cause intraocular inflammation. We characterized the host humoral and cellular immune responses after suprachoroidal delivery of AAV8 expressing green fluorescent protein (GFP) in rhesus macaques, and found that it can induce mild chorioretinitis that resolves after systemic corticosteroid administration, with recovery of photoreceptor morphology, but persistent immune cell infiltration after 3 months, corresponding to a loss of GFP expression from retinal pigment epithelial cells, but persistent expression in scleral fibroblasts. Suprachoroidal AAV8 triggered B cell and T cell responses against GFP, but only mild antibody responses to the viral capsid compared to intravitreal injections of the same vector and dose. Systemic biodistribution studies showed lower AAV8 levels in liver and spleen after suprachoroidal injection compared with intravitreal delivery. Our findings suggest that suprachoroidal AAV8 primarily triggers host immune responses to GFP, likely due to sustained transgene expression in scleral fibroblasts outside the blood-retinal barrier, but elicits less humoral immune reactivity to the viral capsid than intravitreal delivery due to lower egress into systemic circulation. As GFP is not native to primates and not a clinically relevant transgene, suprachoroidal AAV delivery of human transgenes may have significant translational potential for retinal gene therapy.
脉络膜上腔是位于眼球巩膜和脉络膜之间的潜在空间，为眼部药物或病毒载体的输送提供了新的途径。使用经巩膜微针进行脉络膜上腔注射腺相关病毒 (AAV)8 可以使非人类灵长类动物的眼睛广泛表达转基因，但可能导致眼内炎症。我们描述了在恒河猴中脉络膜上腔注射表达绿色荧光蛋白 (GFP) 的 AAV8 后宿主体液和细胞免疫反应的特征，发现它可以诱导轻度脉络膜视网膜炎，在全身皮质类固醇给药后消退，感光细胞形态恢复，但在 3 个月后仍有免疫细胞浸润，这与视网膜色素上皮细胞中 GFP 表达的丧失相对应，但巩膜成纤维细胞中仍有表达。脉络膜上腔 AAV8 引发 GFP 的 B 细胞和 T 细胞反应，但与相同载体和剂量的玻璃体内注射相比，针对病毒衣壳的抗体反应仅轻微。全身生物分布研究表明，与玻璃体内给药相比，脉络膜上腔注射后肝脏和脾脏中的 AAV8 水平较低。我们的研究结果表明，脉络膜上腔 AAV8 主要引发宿主对 GFP 的免疫反应，可能是由于巩膜成纤维细胞中外源性基因表达持续存在于血视网膜屏障之外，但由于进入全身循环的量较少，对病毒衣壳的体液免疫反应比玻璃体内给药少。由于 GFP 不是灵长类动物的天然基因，也不是临床相关的转基因，因此脉络膜上腔 AAV 传递人类转基因可能具有重要的视网膜基因治疗转化潜力。
