iPSC-based blood regenerative therapies for AIDS

基于 iPSC 的艾滋病血液再生疗法

• 批准号: 9057122
• 负责人: Igor I. Slukvin
• 金额: $ 69.45万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9057122
• 关键词: AIDS therapy; Acquired Immunodeficiency Syndrome; Adult; Alleles; Antiviral Agents; Autologous; B-Lymphocytes; Basic Science; Benefits and Risks; Biological; Blood; Blood Cells; Bone Marrow Purging; CCR5 gene; CD34 gene; CD4 Positive T Lymphocytes; Cell Culture Techniques; Cell Therapy; Cells; Cellular biology; Clinic; Clinical; Collaborations; Cyclic GMP; Cytoprotection; Development; Endothelium; Engraftment; Evaluation; Frequencies; Future; Generations; Genes; Genetic; Genomics; Goals; HIV; HIV Infections; HIV resistance; HIV therapy; HIV-1; Health; Hematological Disease; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic Stem Cell Transplantation; Hematopoietic System; Hereditary Disease; Human; IL2RA gene; Immune System Diseases; Immune system; In Vitro; Individual; Knock-out; Latent Virus; Liver; Mediating; Methodology; Modification; Molecular; Molecular Profiling; Mus; Mutation; Myeloid Cells; National Heart, Lung, and Blood Institute; Null Lymphocytes; Patients; Population; Production; Protocols documentation; Regulatory Pathway; Research; Safety; Somatic Cell; Source; Staging; Stem cells; Stromal Cells; T-Cell Development; T-Lymphocyte; Technology; Testing; Therapeutic; Thymus Gland; Time; Translations; Transplantation; Viral; base; cancer genetics; clinical application; clinically relevant; fetal; hematopoietic repopulating cell; homologous recombination; human embryonic stem cell; humanized mouse; improved; in vivo; induced pluripotent stem cell; leukemia; mouse model; mutant; pluripotency; reconstitution; regenerative; regenerative therapy; standard of care

DESCRIPTION (provided by applicant): HSC transplantations have become a standard of care for treating otherwise incurable blood cancers and genetic diseases. The curing of HIV and leukemia by transplanting HSCs from HIV-resistant patients with a CCR5-D32 mutation has demonstrated the power of stem cell-based therapies for AIDS. However, difficulties in the genetic modifications of autologous HSCs and in finding HLA-compatible CCR5-D32 donors significantly hamper the widespread use of somatic HSC-based AIDS therapies in the clinical setting. Converting adult human cells to induced pluripotent stem cells (iPSCs) provides a unique opportunity to produce immunologically matched gene-edited therapeutic cells for diseases of the blood and immune system as iPSCs can be expanded indefinitely ex vivo, genetically modified using homologous recombination and differentiated into hematopoietic cells. However, transferring this approach to the clinic requires the improvement of iPSC- derived blood cell engraftment, development of robust cGMP-compatible protocols for blood production from iPSCs, and the bi-allelic CCR5 disruption to provide an anti-HIV effect. The proposed studies capitalize on our recent advances in identification of pre-HSC hemogenic endothelium (HE) stage in human ESC/iPSC cultures and progress in locus-specific gene editing in ESC/iPSCs using ZNF-mediated homologous recombination. The three related specific aims are directed at understanding the molecular mechanisms controlling development of HSCs from human PSCs through the HE stage, with the ultimate goal to develop clinically- relevant protocols for ex vivo production of CCR5-knockout autologous HSCs for AIDS therapies. In aim 1, we will identify the biological regulators guiding the formation of engraftabl hematopoietic cells from HE with a goal to improve production of blood cells with regenerative potential from human PSCs. In aim 2, we will develop homologous recombination-based technology for the bi-allelic CCR5 knockout in iPSCs and test the engraftability and safety of genetically corrected iPSC-derived blood cells following transplantation in NOD/SCD/IL2Rg-/- (NSG) mice. In aim 3, we will test whether iPSC-derived CCR5-null cells are protected from HIV-1 challenge in NSG mice. Successful completion of the studies will validate a methodology for generation of regenerative blood cells from iPSCs and their potential use for HIV therapies. The applications of the methodology proposed here will be also useful for basic research and for future clinical applications for modification of any genomic target in iPSCs.

描述（由申请人提供）：HSC移植已成为治疗原本无法治愈的血液癌和遗传疾病的护理标准。通过从HIV耐HIV患者移植CCR5-D32突变的HSC通过移植HSC来固化HIV和白血病，这证明了基于干细胞的艾滋病疗法的能力。但是，自体HSC的遗传修饰以及发现与HLA兼容的CCR5-D32供体的困难显着阻碍了在临床环境中基于体细胞HSC的艾滋病疗法的广泛使用。将成年人类细胞转换为诱导的多能干细胞（IPSC）提供了一个独特的机会，可以生产与血液和免疫系统疾病的免疫学匹配的基因编辑的治疗细胞，因为IPSC可以无限期地扩展，可以在体内扩展，使用同源性重组并分化为血液瘤细胞的基因修饰。但是，将这种方法转移到诊所需要改善IPSC衍生的血细胞植入，开发与IPSC的血液产生的可靠CGMP兼容方案以及BI-平行性CCR5破坏，以提供抗HIV效应。拟议的研究利用了我们在人类ESC/IPSC培养物中鉴定HSC前血解性内皮（HE）阶段的最新进展以及使用ZNF介导的同源重组在ESC/IPSC中的基因座特异性基因编辑中的进展。这三个相关的特定目的旨在了解控制HSC从人PSC到HE阶段开发的分子机制，其最终目标是开发用于临床相关的方案，以实现CCR5敲除自体HSC用于艾滋病疗法。在AIM 1中，我们将确定从HE中指导植入造血细胞形成的生物调节剂，目的是改善人类PSC的再生潜力的血细胞产生。在AIM 2中，我们将开发IPSC中BI-CRALLIC CCR5敲除的同源重组技术，并测试在NOD/SCD/IL2RG中移植后遗传校正的IPSC衍生的血细胞的植入性和安全性。在AIM 3中，我们将测试NSG小鼠中iPSC衍生的CCR5-NULL细胞免受HIV-1挑战的保护。成功完成研究将验证一种从IPSC产生再生血细胞及其对HIV疗法的潜在用途的方法。此处提出的方法的应用也将用于基础研究和将来的临床应用，以修改IPSC中任何基因组靶标。