CRISPR/Cas9-mediated metabolic pathway reprogramming in a novel humanized rat model ameliorates primary hyperoxaluria type 1
CRISPR/Cas9-mediated metabolic pathway reprogramming in a novel humanized rat model ameliorates primary hyperoxaluria type 1
复制标题
新型人源化大鼠模型中 CRISPR/Cas9 介导的代谢途径重编程可改善 1 型原发性高草酸尿症
DOI：
10.1016/j.kint.2020.04.049
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发表时间：
2020
影响因子：
19.6
通讯作者：
Geng Hongquan
中科院分区：
文献类型：
--
作者：
Zheng Rui;Li Yueyan;Wang Liren;Fang Xiaoliang;Zhang Junqi;He Lei;Yang Lei;Li Dali;Geng Hongquan
原发性高尿酸I型是由丙氨酸乙醛酸氨基转移酶基因（AGXT）突变引起的，导致乙醛酸积累，随后产生草酸盐和尿石症。在这里，我们通过CRISPR/Cas9系统产生了一种新的原发性高尿酸I型大鼠模型，该模型在部分人源化的Agxt基因中携带D205 N突变。AgxtD 205 N突变大鼠表现出不可检测的丙氨酸乙醛酸氨基转移酶蛋白表达，在1月龄时出现高尿酸血症，并在乙二醇刺激后表现出严重的肾草酸钙沉积。这表明我们的新模型比现有的动物模型与人类疾病更相关。为了测试该模型是否可用于开发创新疗法，靶向羟酸氧化酶1（负责将乙醇酸代谢为乙醛酸）的SaCas 9通过腺相关病毒载体递送到患有原发性高尿酸血症1型的新生大鼠中。这种方法在肝脏中的Hao 1基因中产生了近30%的插入缺失，导致治疗组的尿草酸盐水平比对照组低42%，并防止患有原发性高尿症1型的大鼠发生严重的肾钙质沉着症至少12个月。因此，我们的研究结果表明，这种部分人源化的AgxtD 205 N大鼠品系是1型原发性高尿酸血症的高性能模型，用于理解病理学和开发新的治疗方法，例如通过基因组编辑重新编程代谢途径。
Primary hyperoxaluria type I is caused by mutations in the alanine glyoxylate aminotransferase gene (AGXT), leading to accumulation of glyoxylate and subsequent production of oxalate and urolithiasis. Here, we generated a novel rat model of primary hyperoxaluria type I that carries a D205N mutation in the partially humanizedAgxtgene through the CRISPR/Cas9 system. TheAgxtD205Nmutant rats showed undetectable alanine glyoxylate aminotransferase protein expression, developed hyperoxaluria at 1 month of age and exhibited severe renal calcium oxalate deposition after ethylene glycol challenge. This suggests our novel model is more relevant to the human disease than existing animal models. To test whether this model could be used for the development of innovative therapeutics, SaCas9 targeting hydroxyacid oxidase 1, responsible for metabolizing glycolate into glyoxylate, was delivered via adeno-associated viral vectors into newborn rats with primary hyperoxaluria type 1. This approach generated nearly 30% indels in theHao1gene in the liver, leading to 42% lower urine oxalate levels in the treated group than in the control group and preventing the rats with primary hyperoxaluria type 1 from undergoing severe nephrocalcinosis for at least 12 months. Thus, our results demonstrate that this partially humanizedAgxtD205Nrat strain is a high-performing model of primary hyperoxaluria type 1 for understanding pathology, and the development of novel therapeutics, such as reprogramming of the metabolic pathway through genome editing.