Mechanisms of circuit failure and treatments in patient-derived neurons in autism

描述(由申请人提供)：缺乏有效的临床前模型来探索细胞机制，限制了严重自闭症的治疗发展。为了开发新的临床前模型和治疗方法，对患者来源的神经元中的细胞表型和对潜在药物的反应进行适当的研究将是有利的。这项研究的长期目标是阐明严重自闭症病理基础的分子机制，这将成为为这些目前几乎没有治疗选择的患者开发创新治疗方法的基础。我们最近发现了一种新的自闭症机制，涉及到内体Na+/H+交换器(NHEs)。我们现在已经从具有一系列NHE6突变的患者中产生了诱导多能干细胞系(IPSCs)。这项应用的目的是阐明NHE6在人类轴突发育中的作用，并测试患者来源的神经元对可用的、基于机制的药物的反应，这些药物可能随后用于患者的治疗。NHE6调节内质体质子的外流。在我们对人类神经元的初步研究中，NHE6缺乏会导致内体管腔的过度酸化以及轴突生长和分支的缺陷。这种机制是偶然的，因为有许多广为人知的FDA批准的药物针对内膜内的pH。我们的中心假设是，NHE6的缺乏抑制了轴突的生长和分支，这是由于极化膜的增加减少所致。这项研究的基本原理是，它构成了开发新的治疗方法的第一个关键步骤 由NHE6表达减少和/或轴突生长或树枝缺陷引起的严重自闭症和相关疾病的个人。我们已经发表了关于NHE6在小鼠模型中的功能的研究，但目前过渡到患者来源的组织是有必要的。筛选潜在的人类治疗药物最好是在人类(和患者来源的)组织上进行。我们提出了三个具体的目标：(1)确定NHE6在调节内体管腔酸性和内体循环中的作用。(2)确定患者来源的神经细胞中的NHE6突变对RAB10相关SVS在轴突生长中的作用的干扰程度。(3)确定外源性生长因子对患者来源神经元轴突生长和分支缺陷的可逆性。这项拟议的研究具有创新性，因为我们正在研究自闭症的一种新的细胞机制，即调节AX-ON生长中的内体内pH。这项研究具有重要意义，因为它将导致在患者来源的神经元中开发急需的临床前模型，并可能治疗严重的自闭症和相关疾病，这是一个紧迫的公共卫生问题。

DESCRIPTION (provided by applicant): A lack of validated preclinical models for exploring cellular mechanisms has limited treatment development in severe autism. In order to develop new preclinical models and therapeutics, rig- orous studies of cell phenotypes and cellular responses to potential agents in patient-derived neurons will be advantageous. The long-term goal of this research is to elucidate the molecular mechanisms underlying the pathology of severe autism that will serve as the basis to develop innovative treatments for these patients with few current therapeutic options. We recently identi- fied a novel mechanism in autism involving endosomal Na+/H+ exchangers (NHEs). We have now generated induced pluripotent stem cell lines (iPSCs) from patients with a range of NHE6 mutations. The objective for this application is to elucidate the role of NHE6 in human axon de- velopment and to test the response of patient-derived neurons to available, mechanism-based agents that may subsequently serve as treatments in patients. NHE6 regulates the efflux of pro- tons from endosomes. In our preliminary studies in human neurons, NHE6 deficiency leads to over-acidification of the endosomal lumen and defects in axon growth and branching. This mechanism is fortuitous because there are a number of well-known FDA-approved agents that target intra-endosomal pH. Our central hypothesis is that deficiency of NHE6 inhibits axon growth and branching due to diminished polarized membrane addition. The rationale for this re- search is that it constitutes the first critical steps toward the development of new treatments for individuals with severe autism and related disorders caused by reductions in NHE6 expression and/or by defects in axonal growth or arborization. We have published studies on NHE6 function in mouse models, yet the transition to patient-derived tissues is warranted at this time. Screening of potential human therapeutic agents is best performed on human (and patient-derived) tis- sues. Three specific aims are proposed: (1) Determine the role of NHE6 in regulating endosome lumen acidity and endosome recycling. (2) Determine the extent to which the function of RAB10-associated SVs in axon growth is perturbed by NHE6 mutation in patient-derived neu- rons. (3) Determine the reversibility of axon growth and branching defects in patient-derived neurons by exogenous growth factors. The proposed research is innovative because we are studying a novel cellular mechanism in autism, namely, regulation of intra-endosomal pH in ax- on growth. This research is significant because it will lead to the development of critically need- ed preclinical models in patient-derived neurons and potential treatments for severe autism and related disorders, an urgent public health problem.