Derivation and Transplantation of Histocompatible Pluripotent Stem Cells

组织相容性多能干细胞的衍生和移植

• 批准号: 7924653
• 负责人: David W Russell
• 金额: $ 31.2万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7924653
• 关键词: Affect; Allogenic; American; Autologous; Behavior; CD34 gene; Cell Line; Cell Therapy; Cell Transplants; Cells; Chimeric Proteins; Chromosomes, Human, Pair 6; Clinic; Clinical; Clinical Protocols; Clinical Trials; Dependovirus; Derivation procedure; Development; Developmental Gene; Embryo; Engineering; Engraftment; Ensure; Fibroblasts; Ganciclovir; Gene Expression; Gene Expression Profiling; Gene Targeting; Genes; Genome; Globin; Goals; HLA-A gene; Haplotypes; Hematopoietic; Histocompatibility Antigens; Histocompatibility Testing; Human; Human Chromosomes; Immunosuppression; In Vitro; Individual; Investments; Karyotype determination procedure; Long Terminal Repeats; Macaca; Macaca nemestrina; Mesenchymal Stem Cells; Mitotic; Mitotic Recombination; Modeling; Mus; Patients; Phenotype; Pluripotent Stem Cells; Population; Preclinical Testing; Problem Solving; Procedures; Proliferating; Recombinants; Regenerative Medicine; Resistance; Siblings; Site; Solutions; Source; Spumavirus; Stem cells; Teratoma; Testing; Therapeutic; Time; Transgenes; Transplantation; United States; arm; blastomere structure; cell type; erythroid differentiation; human disease; human embryonic stem cell; hygromycin A; improved; in vivo; induced pluripotent stem cell; nonhuman primate; recombinase; research study; stem cell therapy; telomere; therapeutic cloning; vector

Human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) can only be used effectively in recipients with the same (or nearly the same) histocompatibility type, to avoid rejection or toxic immunosuppression. To overcome this, various strategies have been proposed to derive patient-specific stem cells. These often involve controversial procedures such as therapeutic cloning, and they require a major investment in time and money for every patient that is impractical for routine clinical use. The goal of this proposal is to solve this problem and create a panel of pluripotent stem cells that is histocompatible with a significant percentage of the U.S. population. This panel will consist of cells that are homozygous at the HLA locus, so only one set of major histocompatibility antigens is expressed. By choosing the most common haplotypes, 5-10 HLA-homozygous lines should match over 20-25% of Americans at HLA-A, B and DR loci, and 50 lines should match over 70% of the population. To accomplish this, we will derive a set of iPSCs with common HLA haplotypes from different types of non-embryonic cells. Integrating vectors will be used to deliver reprogramming transgenes, then they will be removed from the cells to create transgene-free iPSCs. Adeno-associated virus (AAV) gene targeting vectors will be used to insert a plus/minus selectable marker next to the HLA locus on human chromosome 6, then mitotic recombinants will be isolated that lost the marker gene and converted the short arm of chromosome 6 to homozygosity, including the entire HLA locus (an approach that has already succeeded in our pilot experiments). The parental iPSCs and their HLA-homozygous derivatives will be characterized extensively by SNP-chip analysis, karyotyping, and differentiation potential. This project will establish a bank of "patient-specific" HLA/MHC-homozygous stem cells that can be used for all types of regenerative medicine, avoiding the problems of graft rejection and immunosuppression, with significant potential for improving the health of Americans.

人胚胎干细胞（ESC）和诱导多能干细胞（iPSC） 只能有效地用于具有相同（或几乎相同） 组织相容性类型，以避免排斥或毒性免疫抑制。克服 为此，已经提出了各种策略来获得患者特异性干细胞。这些 往往涉及有争议的程序，如治疗性克隆，他们需要一个 对每一个病人的时间和金钱的重大投资是不切实际的常规 临床应用。该提案的目标是解决这一问题，并建立一个小组， 多能干细胞，这是组织相容性的一个显着百分比的美国。 人口这组细胞将由HLA基因座纯合的细胞组成， 仅表达一组主要组织相容性抗原。通过选择最 常见的单倍型，5-10个HLA纯合系应匹配超过20-25%的 美国人在HLA-A，B和DR位点，和50行应匹配超过70%的 人口为了实现这一点，我们将获得一组具有共同HLA的iPSC。 来自不同类型的非胚胎细胞的单倍型。将使用积分矢量 来传递重新编程的转基因，然后将它们从细胞中移除， 无转基因的iPSCs将使用腺相关病毒（AAV）基因靶向载体 在人类染色体上的HLA基因座旁边插入正/负选择标记 6，那么将分离丢失标记基因并将标记基因转化为有丝分裂的重组体。 6号染色体短臂的纯合性，包括整个HLA基因座（一个 我们的试验已经取得了成功）。父母的iPSC和 他们的HLA纯合衍生物将被SNP芯片广泛表征， 分析、核型分析和分化潜能。该项目将建立一个银行， “患者特异性”HLA/MHC纯合干细胞，可用于所有类型的肿瘤。 再生医学，避免移植排斥和免疫抑制的问题， 对改善美国人的健康有着巨大的潜力。