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)可以通过CCR5基因的突变或缺失而对艾滋病毒感染产生抵抗力。这些细胞 移植回自体捐献者将防止艾滋病毒复制并起到“治愈”的作用。第一，提纯的人类 来自未感染个体的CD34干细胞或外周血单个核细胞将转化为 通过各种策略诱导多能干细胞(IPS)，重点是非整合载体。 然后，这些iPS细胞将被基因改造，要么缺乏CCR5基因，要么具有自然突变 CCR5.将天然缺乏CCR5表达的细胞注射给没有艾滋病毒证据的“柏林病人” 几年后感染。 IPS来源的CCR5突变细胞将被分化为CD34细胞(即iPS来源的HSPC)，然后 分化为造血子代细胞。这些细胞将被评估为正常的细胞功能和 在细胞培养和移植到人源化小鼠后对艾滋病毒感染的抵抗力。同样的程序 将对来自艾滋病毒感染者的CD34细胞进行研究。 这些研究旨在优化我们向艾滋病毒感染者提供iPS-HSPC的方法，以 防止艾滋病毒疾病的发展，并有可能建立一种“功能疗法”。

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 or deletion in the CCR5 gene. These cells transplanted back to autologous donors will prevent HIV replication and effect a "cure." First, purified human CD34+ stem cells or peripheral blood mononuclear cells from uninfected individuals will be converted into induced pluripotent stem (iPS) cells by a variety of strategies with emphasis given to non-integrating vectors. These iPS cells will then be genetically modified to either lack the CCR5 gene or have the natural mutation of CCR5. 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 differentiated into CD34+ cells (i.e. iPS-derived HSPC) and then differentiated into hematopoietic progeny cells. These cells will be evaluated for normal cell function and resistance to HIV infection in cell culture and after transplantation into humanized mice. The same procedures will be undertaken with CD34+ cells from HIV-infected individuals. These studies are directed at optimizing our approaches for providing iPS-HSPC to HIV-infected individuals to prevent HIV disease progression and potentially establish a "functional cure."