Costa Kárita Antunes, Salles Mariana Vallim, Whitebirch Chris, Chiang John, Sallum Juliana Maria Ferraz
Department of Ophthalmology and Visual Sciences, Federal University of São Paulo (UNIFESP), São Paulo, Brazil.
Casey Eye Institute Molecular Diagnostic Laboratory, Oregon Health and Science University (OHSU), Portland, OR USA.
Int J Retina Vitreous. 2017 Sep 11;3:33. doi: 10.1186/s40942-017-0087-6. eCollection 2017.
Retinal dystrophies constitute a group of diseases characterized by clinical variability and pronounced genetic heterogeneity. Retinitis pigmentosa is the most common subtype of hereditary retinal dystrophy and is characterized by a progressive loss of peripheral field vision (Tunnel Vision), eventual loss of central vision, and progressive night blindness. The characteristics of the fundus changes include bone-spicule formations, attenuated blood vessels, reduced and/or abnormal electroretinograms, changes in structure imaged by optical coherence tomography, and subjective changes in visual function. The different syndromic and nonsyndromic forms of retinal dystrophies can be attributed to mutations in more than 250 genes. Molecular diagnosis for patients with retinitis pigmentosa has been hampered by extreme genetic and clinical heterogeneity between retinitis pigmentosa and other forms of retinal dystrophies. Next generation sequencing (NGS) technologies are among the most promising techniques to identify pathogenic variations in retinal dystrophies.
The purpose of this study was to discover the molecular diagnosis for Brazilian patients clinically diagnosed with a retinitis pigmentosa pattern of inheritance by using NGS technologies.
Sixteen patients with the clinical diagnosis of retinitis pigmentosa were included in the study. Their DNA was sequenced in a panel with 132 genes related to retinal dystrophies using the Illumina platform. Sequence analysis and variation calling was performed using Soft Genetics, NextGene, and Geneticist Assistant software. The criteria for pathogenicity analysis were established according to the results of prediction programs (Polyphen 2, Mutation taster and MetaCore™) and comparison of pathogenic variations found with databases.
The identified potentially pathogenic variations were all confirmed by Sanger sequencing. There were 89 variations predicted as pathogenic, but only 10 of them supported the conclusion of the molecular diagnosis. Five of the nine patients were autosomal dominant RP (56%), two (22%) were autosomal recessive RP, and two (22%) were X-linked RP. Nine of the 16 patients (56%) had probably positive or positive results.
The Next Generation Sequencing used in this study allowed the molecular diagnosis to be confirmed in 56% of the patients and clarified the inheritance pattern of the patient's retinal dystrophies.
视网膜营养不良是一组具有临床变异性和显著遗传异质性的疾病。视网膜色素变性是遗传性视网膜营养不良最常见的亚型，其特征是周边视野逐渐丧失（管状视野）、最终中心视力丧失以及进行性夜盲。眼底改变的特征包括骨针状形成、血管变细、视网膜电图降低和/或异常、光学相干断层扫描成像的结构变化以及视觉功能的主观变化。视网膜营养不良的不同综合征型和非综合征型可归因于250多个基因的突变。视网膜色素变性患者的分子诊断一直受到视网膜色素变性与其他形式视网膜营养不良之间极端遗传和临床异质性的阻碍。下一代测序（NGS）技术是识别视网膜营养不良致病变异最有前景的技术之一。
本研究的目的是通过使用NGS技术为临床诊断为视网膜色素变性遗传模式的巴西患者发现分子诊断方法。
本研究纳入了16例临床诊断为视网膜色素变性的患者。使用Illumina平台在一个包含132个与视网膜营养不良相关基因的面板中对他们的DNA进行测序。使用Soft Genetics、NextGene和Geneticist Assistant软件进行序列分析和变异检测。根据预测程序（Polyphen 2、Mutation taster和MetaCore™）的结果以及与数据库中发现的致病变异进行比较，确定致病性分析标准。
鉴定出的潜在致病变异均通过桑格测序得到证实。有89个变异被预测为致病，但其中只有10个支持分子诊断结论。9例患者中有5例为常染色体显性视网膜色素变性（56%），2例（22%）为常染色体隐性视网膜色素变性，2例（22%）为X连锁视网膜色素变性。16例患者中有9例（56%）可能为阳性或阳性结果。
本研究中使用的下一代测序使56%的患者得以确诊分子诊断，并明确了患者视网膜营养不良的遗传模式。
