Takahashi Kazuhisa, Igarashi Tsutomu, Miyake Koichi, Kobayashi Maika, Yaguchi Chiemi, Iijima Osamu, Yamazaki Yoshiyuki, Katakai Yuko, Miyake Noriko, Kameya Shuhei, Shimada Takashi, Takahashi Hiroshi, Okada Takashi
Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo 113-8602 Japan; Department of Ophthalmology, Nippon Medical School, Tokyo 113-8602, Japan.
Department of Biochemistry and Molecular Biology, Nippon Medical School, Tokyo 113-8602 Japan.
Mol Ther. 2017 Jan 4;25(1):296-302. doi: 10.1016/j.ymthe.2016.10.008.
The retina is an ideal target for gene therapy because of its easy accessibility and limited immunological response. We previously reported that intravitreally injected adeno-associated virus (AAV) vector transduced the inner retina with high efficiency in a rodent model. In large animals, however, the efficiency of retinal transduction was low, because the vitreous and internal limiting membrane (ILM) acted as barriers to transduction. To overcome these barriers in cynomolgus monkeys, we performed vitrectomy (VIT) and ILM peeling before AAV vector injection. Following intravitreal injection of 50 μL triple-mutated self-complementary AAV serotype 2 vector encoding EGFP, transduction efficiency was analyzed. Little expression of GFP was detected in the control and VIT groups, but in the VIT+ILM group, strong GFP expression was detected within the peeled ILM area. To detect potential adverse effects, we monitored the retinas using color fundus photography, optical coherence tomography, and electroretinography. No serious side effects associated with the pretreatment were observed. These results indicate that surgical ILM peeling before AAV vector administration would be safe and useful for efficient transduction of the nonhuman primate retina and provide therapeutic benefits for the treatment of retinal diseases.
视网膜因其易于接近和有限的免疫反应，是基因治疗的理想靶点。我们之前报道，在啮齿动物模型中，玻璃体内注射腺相关病毒（AAV）载体可高效转导视网膜内层。然而，在大型动物中，视网膜转导效率较低，因为玻璃体和内界膜（ILM）对转导起到了屏障作用。为了在食蟹猴中克服这些屏障，我们在注射AAV载体之前进行了玻璃体切除术（VIT）和ILM剥离。玻璃体内注射50 μL编码增强绿色荧光蛋白（EGFP）的三突变自互补AAV 2型载体后，分析转导效率。在对照组和VIT组中几乎未检测到GFP表达，但在VIT + ILM组中，在剥离的ILM区域内检测到强GFP表达。为了检测潜在的不良反应，我们使用彩色眼底照相术、光学相干断层扫描和视网膜电图对视网膜进行监测。未观察到与预处理相关的严重副作用。这些结果表明，在给予AAV载体之前进行手术性ILM剥离对于非人类灵长类动物视网膜的高效转导是安全且有用的，并为视网膜疾病的治疗提供治疗益处。
