Exploring the Pathogenic Mechanisms of Batten's disease MFSD8 mutations using patient iPSC derived neurons.

MFSD8基因的突变导致CLN7Batten病(BD)，这是一种罕见的神经退行性溶酶体储存障碍，其特征是一种名为类脂的脂类物质在大脑中积聚。此外，最近发现MFSD8突变与额颞叶痴呆(FTD)有关，其他BD相关基因与帕金森病(ATP13A2)和FTD的其他亚型(GRN、CTSD)有关。CLN7 BD属于一种更广泛的称为神经元蜡样脂褐素沉着症(NCL)的儿科疾病，这些疾病具有共同的组织病理学，即神经元中蜡样物质积聚，以及包括痴呆、失明、癫痫和过早死亡在内的神经症状。MFSD8基因编码CLN7蛋白，这是一种功能未知的膜蛋白，定位于溶酶体。导致CLN7 BD的突变是功能丧失，许多已知的突变会导致过早停止或点突变，从而破坏蛋白质的稳定。我们之前在患者成纤维细胞中的研究表明，CLN7的缺失会减少溶酶体水解酶的活性、自噬通量和自体荧光物质的积累。虽然这些研究有助于我们理解CLN7是如何导致疾病的，但导致神经退化的机制尚不清楚。此外，还没有在相关的患者来源的神经元模型中进行研究。在这里，我们建议使用来自不同BD患者的几个IPSC系来探索CLN7突变引起的神经毒性的机制。我们的初步数据表明，CLN7 BD系在体外可以发育为皮质神经元，但表现为溶酶体表型和减少的轴突延伸。在目标1中，我们计划使用这个已建立的神经元模型来确定CLN7的丢失如何影响蛋白质组的稳定性和蛋白质的溶解性，以发现新的途径并产生关于CLN7 BD中细胞死亡如何发生的假设。在目标2中，我们将评估旨在恢复CLN7神经元蛋白质稳态的救援机制，以及将野生型星形胶质细胞与CLN7-BD神经元共培养，以评估胶质细胞在痴呆中的作用。由于几乎没有关于CLN7如何导致神经功能障碍的机制信息，我们的研究有可能发现CLN7 BD和其他具有相似病理机制的NCLS的新治疗途径。除了NCLS，MFSD8的突变与FTD有关，FTD是一种与年龄相关的神经退行性疾病，因此我们的研究可能与更常见的与年龄相关的神经退行性疾病有关。由于有越来越多的证据表明溶酶体功能障碍是常见的散发性疾病，包括帕金森氏病和其他联核病，我们的研究可能会对该领域产生广泛的影响。

Mutations in the MFSD8 gene cause CLN7 Batten’s disease (BD), a rare neurodegenerative lysosomal storage disorder characterized by the accumulation of a lipopigment called ceroid in the brain. Furthermore, MFSD8 mutations were recently associated with frontotemporal dementia (FTD), and other BD-associated genes are linked to Parkinson’s disease (ATP13A2) and other subtypes of FTD (GRN, CTSD). CLN7 BD falls within a broader category of pediatric diseases called neuronal ceroid lipofuscinoses (NCL) that share common histopathology of ceroid accumulation in neurons and neurological symptoms including dementia, blindness, seizures, and premature death. The MFSD8 gene encodes the CLN7 protein, a membrane protein of unknown function that is localized to lysosomes. Mutations that cause CLN7 BD are loss of function and many of the known mutations result in premature stops or point mutations that destabilize the protein. Our previous studies in patient fibroblasts demonstrated that loss of CLN7 reduces lysosomal hydrolase activity, autophagic flux, and the accumulation of autofluorescent material. While these studies contributed to our understanding of how CLN7 leads to disease, the mechanisms that lead to neurodegeneration are not known. Furthermore, there are no studies done in relevant patient-derived neuronal models. Here, we propose to explore the mechanisms of neurotoxicity caused by CLN7 mutations, using a several iPSC-lines derived from distinct BD patients. Our preliminary data indicates that CLN7 BD lines can develop into cortical neurons in vitro, but demonstrate lysosomal phenotypes and reduced neurite extensions. In aim 1, we plan to use this established neuronal model to determine how loss of CLN7 influences the stability of the proteome and protein solubility to discover novel pathways and generate hypotheses for how cell death occurs in CLN7 BD. In aim 2, we will assess rescue mechanisms that aim to restore protein homeostasis in CLN7 neurons, as well as co-culturing wild-type astrocytes with CLN7-BD neurons to assess the role of glia cells in dementia. Since almost no mechanistic information is known about how CLN7 causes neurological dysfunction, our studies have the potential to uncover novel therapeutic pathways for CLN7 BD and other NCLs that share similar pathologies. Beyond NCLs, mutations in MFSD8 are associated with FTD, an age-related neurodegenerative disorder, and therefore our studies may be relevant to more common age-related neurodegeneration. Since there is accumulating evidence for lysosomal dysfunction common sporadic diseases including Parkinson’s disease and other synucleinopathies, our studies may have a broad impact on the field.