CRISPR-Cas9-Mediated Correction of the 1.02 kb Common Deletion in CLN3 in Induced Pluripotent Stem Cells from Patients with Batten Disease
CRISPR-Cas9-Mediated Correction of the 1.02 kb Common Deletion in CLN3 in Induced Pluripotent Stem Cells from Patients with Batten Disease
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DOI：
10.1089/crispr.2017.0015
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发表时间：
2018-02-01
期刊：
影响因子：
3.7
通讯作者：
Wiley, Luke A.
中科院分区：
文献类型：
--
作者：
Burnight, Erin R.;Bohrer, Laura R.;Wiley, Luke A.
幼年神经元蜡样质脂褐质沉积症（Batten病）是一种罕见的进行性神经退行性疾病，由CLN 3突变引起。患者表现为早发性视网膜变性，随后出现癫痫、进行性运动缺陷、认知能力下降和过早死亡。大约85%的巴滕病患者至少有一个等位基因，该等位基因包含跨越外显子7和8的1.02 kb基因组缺失。这项研究证明了在两名独立患者产生的诱导多能干细胞中这种突变的基于CRISPR-Cas9的同源依赖性修复：一名纯合子和一名1.02 kb缺失的复合杂合子。我们的策略包括递送携带>3 kb DNA的构建体：野生型CLN 3序列和LoxP侧翼的嘌呤霉素抗性盒用于阳性选择。该策略导致在两个独立的患者系中在基因组DNA和mRNA水平上的校正。这些CRISPR校正的等基因细胞系将成为疾病建模和自体视网膜细胞替代的宝贵工具。
Juvenile neuronal ceroid lipofuscinosis (Batten disease) is a rare progressive neurodegenerative disorder caused by mutations in CLN3. Patients present with early-onset retinal degeneration, followed by epilepsy, progressive motor deficits, cognitive decline, and premature death. Approximately 85% of individuals with Batten disease harbor at least one allele containing a 1.02 kb genomic deletion spanning exons 7 and 8. This study demonstrates CRISPR-Cas9-based homology-dependent repair of this mutation in induced pluripotent stem cells generated from two independent patients: one homozygous and one compound heterozygous for the 1.02 kb deletion. Our strategy included delivery of a construct that carried >3 kb of DNA: wild-type CLN3 sequence and a LoxP-flanked, puromycin resistance cassette for positive selection. This strategy resulted in correction at the genomic DNA and mRNA levels in the two independent patient lines. These CRISPR-corrected isogenic cell lines will be a valuable tool for disease modeling and autologous retinal cell replacement.