N-Acetylaspartate (NAA) is the second most abundant organic metabolite in the brain, but its physiological significance remains enigmatic. Toxic NAA accumulation appears to be the key factor for neurological decline in Canavan disease—a fatal neurometabolic disorder caused by deficiency in the NAA-degrading enzyme aspartoacylase. To date clinical outcome of gene replacement therapy for this spongiform leukodystrophy has not met expectations. To identify the target tissue and cells for maximum anticipated treatment benefit, we employed comprehensive phenotyping of novel mouse models to assess cell type-specific consequences of NAA depletion or elevation. We show that NAA-deficiency causes neurological deficits affecting unconscious defensive reactions aimed at protecting the body from external threat. This finding suggests, while NAA reduction is pivotal to treat Canavan disease, abrogating NAA synthesis should be avoided. At the other end of the spectrum, while predicting pathological severity in Canavan disease mice, increased brain NAA levels are not neurotoxic per se. In fact, in transgenic mice overexpressing the NAA synthesising enzyme Nat8l in neurons, supra-physiological NAA levels were uncoupled from neurological deficits. In contrast, elimination of aspartoacylase expression exclusively in oligodendrocytes elicited Canavan disease like pathology. Although conditional aspartoacylase deletion in oligodendrocytes abolished expression in the entire CNS, the remaining aspartoacylase in peripheral organs was sufficient to lower NAA levels, delay disease onset and ameliorate histopathology. However, comparable endpoints of the conditional and complete aspartoacylase knockout indicate that optimal Canavan disease gene replacement therapies should restore aspartoacylase expression in oligodendrocytes. On the basis of these findings we executed an ASPA gene replacement therapy targeting oligodendrocytes in Canavan disease mice resulting in reversal of pre-existing CNS pathology and lasting neurological benefits. This finding signifies the first successful post-symptomatic treatment of a white matter disorder using an adeno-associated virus vector tailored towards oligodendroglial-restricted transgene expression.

ñ-乙酰天冬氨酸（NAA）是大脑中第二大最丰富的有机代谢物，但其生理学意义仍然是个谜。有毒的NAA积累似乎是Canavan病神经衰弱的关键因素，Canavan病是由NAA降解酶天冬氨酸酰化酶缺乏引起的致命性神经代谢疾病。迄今为止，针对这种海绵状白细胞营养不良的基因替代疗法的临床结果尚未达到预期。为了鉴定目标组织和细胞以获得最大的预期治疗益处，我们采用了新型小鼠模型的综合表型分析来评估NAA耗竭或升高的细胞类型特异性后果。我们表明，NAA缺乏会导致神经功能受损，从而影响旨在保护人体免受外部威胁的无意识防御反应。这一发现表明，虽然降低NAA对治疗Canavan病至关重要，但应避免废止NAA的合成。在频谱的另一端，虽然预测了Canavan病小鼠的病理严重程度，但脑中NAA水平的升高本身并不具有神经毒性。实际上，在过表达NAA合成酶的转基因小鼠中神经元中的Nat8l，超生理性NAA水平与神经功能缺损无关。相反，仅在少突胶质细胞中消除天冬氨酸酰化酶表达会引起像病理一样的Canavan病。尽管少突胶质细胞中条件性天冬氨酸酰化酶的缺失消除了整个中枢神经系统的表达，但周围器官中剩余的天冬氨酸酰化酶足以降低NAA水平，延缓疾病发作并改善组织病理学。但是，条件性和完全天冬氨酸酰化酶敲除的可比终点表明，最佳的Canavan疾病基因替代治疗应能恢复少突胶质细胞中天冬氨酸酰化酶的表达。根据这些发现，我们执行了ASPA基因替代疗法针对Canavan病小鼠中的少突胶质细胞，可逆转先前存在的中枢神经系统病理，并具有持久的神经功能。该发现标志着使用针对少突胶质细胞限制性转基因表达而设计的腺相关病毒载体，首次成功地对白质病进行了症状后治疗。