Novel Gene Therapy Strategies for Canavan Disease

摘要 Canavan病(CD)是一种罕见的儿童脑白质营养不良症，由常染色体隐性突变引起 天冬氨酸酯酶(Aspa)基因。Canavan患者ASPA缺乏导致N- 乙酰天冬氨酸(NAA)，导致脑白质肿胀和海绵状变性。这个 这种致命疾病的临床表现包括精神运动迟缓、肌肉低张症、 巨头畸形、头部迟缓、癫痫发作和过早死亡。NAA的合成是在线粒体内进行的。 N-乙酰转移酶-8样酶(NAT8L)和ASPA对少突胶质细胞中的神经元进行水解。基因 ASPA缺乏症的替代疗法是目前治疗CD最有前景的策略。值得注意的是，我们 最近在Aspa基因敲除(CD-KO)中实现了对Canavan表型的完全治疗 老鼠模型。人重组腺相关病毒单次静脉注射 早期的aspa转基因(raav-haspa)完全解决了神经病理，导致动物得到治疗。 在运动功能测试中表现优于野人。然而，根据强有力的初步证据，我们现在 假设CD表型表现为与代谢功能障碍相关的继发性病因。在……里面 此外，我们最近发现，在体外野生型细胞中过表达aspa会导致异常 线粒体的形态和功能。这些发现需要进一步的临床前研究，重点是：1) 表征ASPA在中枢神经系统和外周器官细胞类型中过表达的可能毒性，2) 模拟ASPA内源生理水平的ASPA调节盒(S)的研制 避免治疗可能存在的不良影响，以及3)确定生理和 使用临床相关的非人类灵长类动物模型研究aspa过度表达的行为效应。我们的新产品 研究战略现在建立在我们目前有希望的进展的基础上，并推进我们的安全和 CD的有效基因治疗。

ABSTRACT Canavan disease (CD) is a rare childhood leukodystrophy caused by autosomal recessive mutations in the aspartoacylase (ASPA) gene. Deficiency of ASPA in Canavan patients leads to the accumulation of N- Acetyl-Aspartic Acid (NAA), resulting in swelling and spongy degeneration of white matter in the brain. The clinical manifestations of this fatal disease include psychomotor retardation, muscular hypotonia, macrocephaly, head lag, seizures, and early death. Synthesis of NAA is carried out in the mitochondria of neurons by N-acetyltransferase-8-like (NAT8L) and hydrolyzed in oligodendrocytes by ASPA. Gene replacement therapy for ASPA deficiency is currently the most promising strategy for treating CD. Notably, we have recently achieved full therapeutic correction of the Canavan phenotype in the Aspa knockout (CD-KO) mouse model. A single intravenous injection of recombinant adeno-associated virus packaged with the human ASPA transgene (rAAV-hASPA) at early ages completely resolves neuropathology, resulting in treated animals that outperform wild-types in motor function tests. However, based on strong preliminary evidence, we now hypothesize that the CD phenotype presents a secondary etiology related to metabolism dysfunction. In addition, we recently revealed that overexpression of ASPA in wild type cells in vitro resulted in abnormal mitochondrial shape and function. These findings necessitate further preclinical investigations that focus on: 1) characterizing the possible toxicity of ASPA overexpression in cell types of the CNS and peripheral organs, 2) developing ASPA regulatory cassette(s) that can mimic endogenous physiological levels of ASPA to circumvent adverse effects that may exist due to treatment, and 3) determining the physiological and behavioral effects of ASPA overexpression using a clinically relevant non-human primate model. Our new research strategy now builds on our current promising progress and advances our goals for a safe and effective gene therapy for CD.