EVALUATING THE ENGINEERING OF CYTOMEGALOVIRUS-SPECIFIC CD8+ T CELL CLONES

评估巨细胞病毒特异性 CD8 T 细胞克隆的工程设计

• 批准号: 8172786
• 负责人: STANLEY R. RIDDELL
• 金额: $ 15.51万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2011-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8172786
• 关键词: Adoptive Immunotherapy; Adoptive Transfer; Adrenal Cortex Hormones; Allogenic; Antigens; Autologous; CD8B1 gene; Cell Therapy; Clone Cells; Cloning; Computer Retrieval of Information on Scientific Projects Database; Cytomegalovirus; Engineering; Exhibits; Funding; Glucocorticoid Receptor; Graft Rejection; Grant; Hematopoietic stem cells; Immunity; Immunocompetent; Immunosuppressive Agents; Immunotherapy; In Vitro; Institution; Macaca; Mature T-Lymphocyte; Modeling; Morbidity - disease rate; Organ Transplantation; Patients; Receptor Gene; Receptor Signaling; Research; Research Personnel; Resistance; Resources; Safety; Solid; Source; T cell response; T-Lymphocyte; United States National Institutes of Health; Virus Diseases; Zinc Fingers; endonuclease; graft vs host disease; immunosuppressed; innovation; mortality; peripheral blood; receptor; vector

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Reactivation of cytomegalovirus (CMV) represents a major cause of morbidity and mortality in immunosuppressed recipients of allogeneic hematopoietic stem cell and solid organ transplant as a consequence of deficient CD8+ T cell immunity. Adoptive immunotherapy with antigen-specific T cell clones has been shown to safely correct quantitative or qualitative deficiencies of T cells that permit progression of viral infection. However, T cell therapy has failed to restore protective T cell immunity in the subset of patients that receive immunosuppressive corticosteroids to treat graft versus host disease or graft rejection, respectively. To overcome this obstacle, we have investigated an innovative strategy for selectively interfering with glucocorticoid receptor (GR) signaling using zinc-finger endonucleases (ZFNs) for targeted deletion of the GR gene in mature T cells. We have developed a macaque model for adoptive transfer of CMV-specific T cell clones and employ this model to investigate the safety and efficacy of immunotherapy with CMV-specific T cells that lack the GR gene (GR-/-) after ZFN targeted deletion. We identified immunocompetent macaques with detectable CMV-specific CD8+ T cell responses in the peripheral blood and used a replication defective Ad5/35 vector to deliver ZFNs that target the GR gene in macaque T cells. CMV-specific GR-/- CD8+ T cell clones were then isolated by limiting dilution cloning. The GR-/- T cell clones exhibited CMV-specific cytolytic reactivity comparable to CMV-specific GRwt clones. However, in contrast to GRwt T cells, the GR-/- T cells were resistant to lymphotoxic effects of corticosteroids in vitro, consistent with disruption of the GR gene and lack of a functional GR protein. We are currently generating autologous GR-/- CMV-specific macaque CD8+ T cell clones from 2 macaques that can be used for adoptive transfer studies to evaluate the safety and efficacy of adoptively transferred CD8+ CMV-specific T cell clones with an edited GR gene.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 巨细胞病毒（CMV）的重新激活代表了同种异体造血干细胞和固体器官移植的免疫抑制受体发病率和死亡率的主要原因，这是由于CD8+ T细胞免疫力不足。用抗原特异性T细胞克隆的收养免疫疗法已被证明可以安全地纠正允许病毒感染进展的T细胞的定量或定性缺陷。然而，T细胞疗法未能恢复接受免疫抑制皮质类固醇的患者子集中的保护性T细胞免疫，分别治疗移植物与宿主疾病或移植物排斥。为了克服这一障碍，我们研究了一种创新的策略，用于选择性地干扰糖皮质激素受体（GR）信号传导，使用锌指内切核酸内切酶（ZFN）在成熟T细胞中靶向缺失GR基因的靶向缺失。我们开发了一种猕猴模型，用于通过CMV特异性T细胞克隆的过继转移，并采用该模型研究了在ZFN靶向缺失后缺乏GR基因（GR - / - ）的CMV特异性T细胞免疫疗法的安全性和功效。我们在外周血中鉴定出具有可检测到的CMV特异性CD8+ T细胞反应的免疫能力猕猴，并使用了复制有缺陷的AD5/35载体来提供靶向猕猴T细胞中GRENE的ZFN。然后通过限制稀释克隆分离CMV特异性的GR - / - CD8+ T细胞克隆。 GR - / - T细胞克隆表现出与CMV特异性GRWT克隆相当的CMV特异性细胞溶解反应性。然而，与GRWT T细胞相反，GR - / - T细胞在体外具有抗皮质类固醇的淋巴细胞毒性作用，与GR基因的破坏和缺乏功能性GR蛋白有关。我们目前正在从2个猕猴中生成自体GR - / - CMV特异性猕猴CD8+ T细胞克隆，这些猕猴可用于继承转移研究，以评估具有编辑GR基因的过继转移CD8+ CMV特异性T细胞克隆的安全性和功效。