Normalizing E:I imbalance in Rett Syndrome by Modulation of Late Response Genes

项目总结和职业发展摘要 Rett综合征（RTT）是一种毁灭性的神经发育障碍，目前没有 可行的治疗策略。大多数RTT病例是由转录中的功能缺失突变引起的。 甲基CpG结合蛋白2（MECP 2），Mecp 2的敲除（Mecp 2-KO）导致RTT-1。 比如啮齿类动物的表型和病理。RTT模型小鼠的脑的特征是获得性的， 小头畸形，这不是由于神经萎缩，而是减少突触和随后的 神经元结构的简化。因此，临床前治疗策略，已获得最 注意力是那些旨在使突触密度正常化的注意力，通常通过替代迟发反应来实现。 (LR)或直接调节其受体。在这方面，RTT小鼠模型是指导 治疗设计;然而，考虑到将数据从小鼠转换到人类的挑战，我们 最近采取了不同的策略。使用8例RTT患者的皮质和小脑组织， 进行RNA测序（seq）分析，以筛选植根于人类的新干预点。 患者数据，然后对小鼠中的相关命中进行反向建模。令人兴奋的是，我们发现了毒蕈碱乙酰胆碱 受体4（M4）基因在RTT患者中显著降低。这一发现很突出，因为M4调节剂 已被证明可以挽救与RTT和M4重叠的精神分裂症等疾病的症状领域， KO小鼠具有与Mecp 2-KO小鼠相似的认知和社交缺陷。这就提出了一个令人兴奋的可能性， 减弱的M4表达和信号传导可能是RTT患者的潜在认知和社会表型。 在目标1中，我们建议进一步开发初步数据，表明认知和社会表型， Mecp 2-KO小鼠对M4阳性变构调节剂（PAM）有反应。在目标2中，我们建议 结论性地将M4信号传导与患者的RTT联系起来，通过对来自 已临床诊断患有RTT，但没有MECP 2突变（MECP 2- 突变阴性）。最后，在目标3中，我们提出将我们的M4-PAM与谷氨酸能调节剂共同施用。 和GABA能信号传导，以确定是否可以挽救互补的症状域。 我的职业目标是成为一个实验室的负责人，研究重点是儿科疾病， 神经系统，我已经确定了四个方面的培训不足，我建议补救，以协助 在这个角色上取得长期的成功。这四个方面是：1）电生理学培训2）下一步培训 代测序分析3）临床暴露于RTT和4）Mentor经验。删除这些 由于技术和概念上的障碍，我制定了一个培训计划，将正式的教学培训与 实践指导，目标是将这些弱点转化为优势。预计完工后， 建议的项目和培训计划将使我处于一个理想的位置，以获得终身教职 因此，我相信这份申请非常适合K 01职业发展奖。

PROJECT SUMMARY AND CAREER DEVELOPMENT ABSTRACT Rett syndrome (RTT) is a devastating neurodevelopmental disorder for which there are currently no viable treatment strategies. The majority of RTT cases result from loss of function mutations in a transcription factor known as methyl CpG binding protein 2 (MECP2), and knockout of Mecp2 (Mecp2-KO) results in RTT- like phenotypes and pathologies in rodents. The brains of RTT-model mice are characterized by acquired microcephaly, which results not from neural atrophy, but rather a reduction in synapses and the subsequent simplification of the neuronal arbor. As such, the preclinical treatment strategies that have garnered the most attention are those designed to normalize synaptic density, often through the replacement of late response (LR) or direct modulation of their receptors. In this capacity RTT-mouse models are valuable tools to guide therapeutic design; however, given the challenges associated with translating data from mice to humans, we have recently adopted a different strategy. Using cortical and cerebellar tissue from 8 RTT patients, we performed RNA-sequencing (seq) analysis to screen for novel points of intervention that are rooted in human patient data, and then back-modeled relevant hits in mice. Excitingly, we identified the muscarinic acetylcholine receptor 4 (M4) as a gene significantly decreased in RTT patients. This finding is salient, as M4-modulators have been shown to rescue symptom domains in diseases like schizophrenia that overlap with RTT and M4- KO mice have cognitive and social deficits similar to Mecp2-KO mice. This raises the exciting possibility that attenuated M4 expression and signaling could be underlying cognitive and social phenotypes in RTT patients. In Aim 1, we propose to further develop preliminary data indicating that cognitive and social phenotypes in Mecp2-KO mice are responsive to M4 positive allosteric modulators (PAMs). In Aim 2, we propose to conclusively link M4-signaling to RTT in patients by perform an RNA-seq analysis on autopsy samples from patients that have been clinically diagnosed with RTT, but who do not have mutations in MECP2 (MECP2- mutation negative). Finally, in Aim 3 we propose co-administer our M4-PAM with modulators of glutamatergic and GABAergic signaling, to establish whether a complimentary symptom domains can be rescued. My career goal is to become the head of a lab with a research focus on pediatric diseases of the nervous system, and I have identified four areas of insufficient training that I propose to remedy to assist in achieving long term success in that role. The four areas are: 1) Training in electrophysiology 2) Training next generation sequencing analysis 3) Clinical exposure to RTT and 4) Mentor experience. To remove these technical and conceptual barriers, I have developed a training plan that integrates formal didactic training with hands-on mentorship, with the goal of turning these weaknesses into strengths. It is anticipated that completion of the proposed project and training plan will place me in an ideal position to receive a tenure track faculty position, and as such, I believe that this application is well suited for a K01 Career Development Award.