HIV Cure with CCr5 (-) Human IPS Hematopoietic Stem Cells

摘要 这项提议的目的是开发一种有效的方法来提供艾滋病毒的“功能疗法”-- 被感染的人。该方法是基于观察到缺乏CCR5表达的受试者可以 对艾滋病毒感染具有高度抵抗力。我们的假设是，造血CD34+干细胞和祖细胞 (HSPC)可通过CCR5基因突变而对HIV感染产生抵抗力。这些细胞被移植回 对于自体捐献者，将防止艾滋病毒复制，并起到“治愈”的作用。首先，纯化的人类CD34+细胞或 来自未感染个体的外周血单核细胞将被转化为诱导的多能干细胞 (IPS)细胞通过强调非整合载体的策略。这些诱导性多能性细胞将从基因上 经过修饰使CCR5的32bp自然突变与该受体表达缺失相关 在细胞表面。值得注意的是，天然缺乏CCR5表达的细胞被移植给了 几年后没有感染艾滋病毒的证据。 IPS来源的CCR5突变细胞将转化为CD34+细胞(即iPS来源的CD34+HSPC)和 然后分化为造血子代细胞。这些细胞将被评估对艾滋病毒的抵抗力 在细胞培养和移植到人源化小鼠后的感染和正常的细胞功能。相同 这些程序将使用来自艾滋病毒感染者的转基因CD34+细胞进行。 这些研究旨在优化我们为HIV提供iPS来源的CD34+HSPC的方法。 以防止艾滋病毒疾病的发展，并有可能建立一种“功能治愈”。

Abstract The objective of this proposal is to develop an effective method for providing a "functional cure" for HIV- infected individuals. The approach is based on the observation that subjects lacking CCR5 expression can be highly resistant to HIV infection. Our hypothesis is that hematopoietic CD34+ stem and progenitor cells (HSPC) can be made resistant to HIV infection via mutation in the CCR5 gene. These cells transplanted back to autologous donors will prevent HIV replication and effect a "cure." First, purified human CD34+ cells or peripheral blood mononuclear cells from uninfected individuals will be converted into induced pluripotent stem (iPS) cells by strategies that emphasize non-integrating vectors. These iPS cells will then be genetically modified to have the ¿32bp natural mutation of CCR5 associated with the absence of this receptor expression on the cell surface. Notably, cells naturally lacking CCR5 expression were given to the "Berlin patient" who has no evidence of HIV infection after several years. The iPS-derived CCR5-mutated cells will be converted into CD34+ cells (i.e. iPS-derived CD34+ HSPC) and then differentiated into hematopoietic progeny cells. These cells will be evaluated for resistance to HIV infection and normal cell function in cell culture and after transplantation into humanized mice. The same procedures will be undertaken with genetically modified CD34+ cells from HIV-infected individuals. These studies are directed at optimizing our approaches for providing iPS- derived CD34+ HSPC to HIV- infected individuals to prevent HIV disease progression and potentially establish a "functional cure."