High throughput screen for discovery of N-acetyltransferase 8 Like (NAT8L) inhibitors

项目总结 Canavan病(CD)是一种罕见的常染色体隐性遗传性脑白质营养不良症，由基因突变引起 天冬氨酸酯酶基因(ASPA)，导致酶活性丧失和浓度增加 底物N-乙酰天冬氨酸(NAA)在大脑中。大脑中NAA的积累导致海绵状 白质变性，异常髓鞘形成，脑水肿，巨头畸形，严重的认知和运动 赤字，并最终导致死亡。没有治愈的方法，也没有治疗CD的标准疗程。治疗 目前仅限于支持性护理和症状管理。小鼠ASPA基因的遗传缺失 已经被证明可以重现患者身上出现的许多CD病症状。对这项提议很重要， 天冬氨酸N-乙酰转移酶8(Nat8L)编码基因的缺失，该酶催化 从天冬氨酸和乙酰辅酶A生物合成NAA，预防CD小鼠模型中的脑白质营养不良。这些 小鼠的NAA水平显著降低，没有星形胶质细胞空泡化、星形胶质细胞增生症或 小脑或前脑出现脱髓鞘。在脑池内观察到了类似的治疗效果。 将反义寡核苷酸应用于NAT8L。这些结果强烈表明，对NAT8L的抑制 在CD的治疗中将是有用的。然而，目前还没有临床上使用的NAT8L抑制剂 已知的NAT8L抑制剂效力低(IC50值在μM-mm范围内)和/或拥有 防止大脑渗透的羧酸盐部分。这项建议旨在进行高通量筛选 (HTS)用于人类NAT8L小分子抑制剂的400,000个大而多样的化合物文库。 成功鉴定易处理的HIT化合物，然后进行初步结构优化 导致发现具有未来治疗学发展潜力的有前途的NAT8L抑制剂 对于CD。我们准备通过执行以下三个具体目标来抓住这个机会：(目标1) 进行Nat8L HTS和HIT确认分析；(目标2)进行HIT集群、初步SAR和ADME 分析；(目标3)进行初步的引线优化。

PROJECT SUMMARY Canavan disease (CD) is rare autosomal recessive leukodystrophy caused by mutations in the aspartoacylase gene (ASPA), leading to loss of enzyme activity and increased concentrations of the substrate N-acetylaspartate (NAA) in the brain. Accumulation of brain NAA results in spongiform degeneration of white matter, aberrant myelination, brain edema, macrocephaly, severe cognitive and motor deficits and ultimately death. There is no cure, nor is there a standard course of treatment for CD. Treatment is currently limited to supportive care and symptom management. Genetic deletion of the ASPA gene in mice has been shown to reproduce many of the CD disease symptoms seen in patients. Important to this proposal, deletion of the gene encoding for aspartate N-acetyltransferase 8 (NAT8L), the enzyme that catalyzes the biosynthesis of NAA from aspartate and acetyl CoA, prevented leukodystrophy in a CD mouse model. These mice showed substantial reduction in NAA levels and no evidence of astroglial vacuolation, astrogliosis, or demyelination in the cerebellum or forebrain. Similar therapeutic benefits were observed with intracisternal administration of antisense oligonucleotide to NAT8L. These results strongly suggest that inhibition of NAT8L would be useful in the treatment of CD. Currently, however, there are no NAT8L inhibitors that are clinically available; known NAT8L inhibitors have low potency (IC50 values in the μM - mM range) and/or possess carboxylate moieties that prevent brain penetration. This proposal aims to conduct high throughput screening (HTS) of a large and diverse 400,000 compound library for small molecule inhibitors of human NAT8L. Successful identification of tractable hit compounds followed by preliminary structural optimization should lead to the discovery of promising lead NAT8L inhibitors with potential for future development of therapeutics for CD. We are poised to seize this opportunity by executing the following three Specific Aims: (Aim 1) conduct NAT8L HTS and hit confirmation assays; (Aim 2) conduct hit clustering, preliminary SAR, and ADME profiling; (Aim 3) conduct preliminary lead optimization.