PDF(Pubmed)
Abstract：
CRISPR-Cas9-based therapeutic genome editing approaches hold promise to cure a variety of human diseases. Recent findings demonstrate pre-existing immunity for the commonly used Cas orthologs from Streptococcus pyogenes (SpCas9) and Staphylococcus aureus (SaCas9) in humans, which threatens the success of this powerful tool in clinical use. Thus, a comprehensive investigation and potential risk assessment are required to exploit the full potential of the system. Here, we investigated existence of immunity to SpCas9 and SaCas9 in control rhesus macaques (Macaca mulatta) alongside monkeys transplanted with either lentiviral transduced or CRISPR-SpCas9 ribonucleoprotein (RNP)-edited cells. We observed significant levels of Cas9 antibodies in the peripheral blood of all transplanted and non-transplanted control animals. Transplantation of ex vivo transduced or SpCas9-mediated BCL11A enhancer-edited cells did not alter the levels of Cas9 antibodies in rhesus monkeys. Following stimulation of peripheral blood cells with SpCas9 or SaCas9, neither Cas9-specific T cells nor cytokine induction were detected. Robust and durable editing frequencies and expression of high levels of fetal hemoglobin in BCL11A enhancer-edited rhesus monkeys with no evidence of an immune response (>3 years) provide an optimistic outlook for the use of ex vivo CRISPR-SpCas9 (RNP)-edited cells.
摘要：
基于CRISPR-Cas9的治疗性基因组编辑方法有望治愈多种人类疾病。最近的研究结果表明，人类对来自化脓性链球菌（SpCas9）和金黄色葡萄球菌（SaCas9）的常用Cas直向同源物已经存在免疫力，这威胁到这个强大的工具在临床使用中的成功。因此，需要进行全面调查和潜在风险评估，以充分利用系统的潜力。这里,我们调查了对照恒河猴(Macacamulatta)以及移植了慢病毒转导或CRISPR-SpCas9核糖核蛋白(RNP)编辑细胞的猴子对SpCas9和SaCas9的免疫力的存在。我们在所有移植和非移植对照动物的外周血中观察到显著水平的Cas9抗体。离体转导或SpCas9介导的BCL11A增强子编辑的细胞的移植不会改变猕猴中Cas9抗体的水平。在用SpCas9或SaCas9刺激外周血细胞之后,既没有检测到Cas9特异性T细胞也没有检测到细胞因子诱导。在BCL11A增强子编辑的恒河猴中，稳健而持久的编辑频率和高水平的胎儿血红蛋白表达，没有免疫反应的证据（>3年），为使用离体CRISPR-SpCas9（RNP）编辑的细胞提供了乐观的前景。
