Disseminated gene delivery to the CNS by human iPSC-derived neural stem cells

 描述(由申请人提供)：神经干细胞(NSCs)在某些情况下在大脑内迁移的自然能力，长期以来一直使它们成为治疗中枢神经系统疾病的候选细胞。将人类体细胞重新编程为组织特异性干细胞的方法的发展，为将NSC疗法用于许多脑部疾病提供了巨大的希望。在人类疾病的小鼠模型上进行的NSC移植实验已经证明，在几种类型的疾病中至少可以纠正一些病理，为基于NSC的方法提供了原理证明。然而，有效翻译的一个重要障碍是嫁接水平较低。由于缺陷的性质，神经遗传性疾病在中枢神经系统具有全球分布的损害，因此治疗需要供体细胞的弥漫性分布。然而，要实现这一点，需要更好地了解神经干细胞移植后的特性。我们已经将人类患者成纤维细胞重新编程为多能干细胞(IPSCs)，并从中衍生出NSCs，并对HU-iPS-NSCs进行了基因矫正。我们建议进行异种移植研究，以评估工程HU-iPS-NSC对移植后分布和存活的影响。我们将在一种具有良好特征的人类溶酶体储存疾病(LSD)的小鼠模型中，测试在大脑中传递可扩散蛋白的治疗效果。有50个个体LSD，它们是影响中枢神经系统的所有遗传性儿童遗传病的主要原因。原则上，可以对大多数LSD使用通用的治疗策略，其基础是观察到溶酶体酶从基因校正的细胞中输出，并被突变细胞吸收，以恢复丢失的酶活性。初步研究表明，HU-iPS-NSCs移植到NOD-SCID小鼠脑内的水平较低。拟议的实验将调查移植后供体细胞分散和存活的基本特征，以解决神经干细胞植入不足的问题。我们已经获得了令人信服的初步数据，证明了拟议实验的可行性，我们将使用定量分析来测量植入量、分化为成熟神经细胞类型和病理变化。这一系列研究的长期战略是将NSC移植到小鼠大脑中，达到可以在未来的翻译研究中在脑部疾病的大型动物模型中进行测试的水平，并最终进入比小鼠大脑大约3000倍的人脑。显然，在这一问题上的进展需要在理解神经干细胞植入的生物学方面取得重大进展，并制定加强植入的战略。

 DESCRIPTION (provided by applicant): The natural ability of neural stem cells (NSCs) to migrate within the brain, under certain circumstances, has long made them a candidate for treatment of CNS diseases. The development of methods to reprogram human somatic cells into tissue specific stem cells has provided great hope for using NSC therapies for many brain disorders. NSC transplantation experiments in mouse models of human diseases have demonstrated correction of at least some pathology in several types of diseases, providing proof-of-principle for the NSC- based approach. However, a significant barrier to effective translation is the low level of engraftment that occurs. Neurogenetic diseases have globally distributed lesions in the CNS due to the nature of the defect, thus treatment requires disseminated distribution of the donor cells. Achieving that will, however, require much better understanding of the post-transplantation properties of NSCs. We have reprogrammed human patient fibroblasts into pluripotent stem cells (iPSCs), derived NSCs from them, and genetically corrected the hu-iPS- NSCs. We propose xenograft studies to evaluate the effects of engineering hu-iPS-NSCs on post-transplant distribution and survival. We will test the therapeutic effectiveness of delivering a diffusible protein within the brain, in a well-characterized mouse model of a human lysosomal storage disease (LSD). There are >50 individual LSDs and they are responsible for a large portion of all inherited childhood genetic diseases that affect the CNS. A common treatment strategy can be used, in principle, for most of the LSD's, based on the observation that lysosomal enzymes are exported from genetically corrected cells and taken up by mutant cells to restore the missing enzymatic activity. Preliminary studies indicate the hu-iPS-NSCs engraft in the NOD- SCID mouse brain at low levels. The proposed experiments will investigate basic features of post-transplant dispersion and survival of the donor cells to address the problem of inadequate NSC engraftment. We have obtained compelling preliminary data demonstrating the feasibility of the proposed experiments and we will use quantitative analyses to measure amounts of engraftment, differentiation into mature neural cell types, and changes in pathology. The long-term strategy of this line of research is to develop NSC transplants in the mouse brain to a level where they can be tested in large animal models of brain diseases in future translational studies and eventually into the human brain which is ~3,000 times larger than the mouse brain. It is clear that progress on this problem needs significant advances in understanding the biology of NSC engraftment and development of strategies to enhance engraftment.