CD19-Specific CB T-cell Therapy for Patients with B-cell Malignancies

项目3解决了以下问题：（i）机会性病毒感染和（ii）脐带血移植（UCBT）后B系恶性肿瘤复发。我们假设，病毒感染和白血病复发的发生率后，异基因UCBT可以减少过继转移供体来源的多病毒特异性细胞毒性T淋巴细胞（CTL），遗传修饰（转导）的白血病细胞表达的CD 19分子的特异性。为了巩固UCBT，我们设计了嵌合抗原受体（CAR），以将T细胞的抗原特异性重定向至B细胞谱系限制的细胞表面分子CD 19。来自项目2的病毒特异性CTL将被转导以表达该CAR，通过嵌合CD 28和CD 3-zeta导致MHC非依赖性和CAR依赖性活化，以裂解淋巴细胞，上调IL-2，和抗凋亡基因。具体目标#1中提出的研究将评价UCB衍生的多病毒特异性CTL是否可对GDI 9具有特异性，以及HSV-1胸苷激酶转基因是否可在免疫缺陷小鼠中表达以通过正电子发射断层扫描（PET）成像。使用非侵入性生物发光成像（BLI）和微型PET的连续免疫疗法将纵向评估输注细胞的持久性和抗肿瘤效果。具体目标#2将评估在UCBT后输注经修饰以表达CAR CD 19特异性的多病毒特异性CTL的可行性、安全性和持久性。相关研究将描述过继转移的CTL的（i）持续性的幅度和持续时间，（ii）运输，和（iii）抗病毒和抗白血病作用。在为人类PET T细胞成像做准备时，在具体目标#3中，我们将开发非人灵长类动物模型。将用汽车和TK对猴源性T细胞进行遗传修饰，并输注。使用[18F] FEAU（由胸苷激酶（TK）代谢）的纵向PET成像将用于评价这些过继转移的猕猴T细胞的分布。总之，研究结果将促进使用病毒和CD 19特异性CTL靶向UCBT后MRD的进展，以提高B细胞恶性肿瘤患者的无病生存期。

Project 3 addresses the problems of (i) opportunistic viral infection and (ii) relapse of B-lineage malignancies after umbilical cord blood transplantation (UCBT). We hypothesize that the incidence of viral infection and leukemia relapse following allogeneic UCBT can be reduced by adoptively transferred donor-derived multi virus-specific cytotoxic T lymphocytes (CTL), genetically modified (transduced) to be specific for the CD19 molecule expressed by leukemic cells. To consolidate UCBT we have designed a chimeric antigen receptor (CAR) to redirect antigen specificity of T cells to the B cell lineage-restricted cell-surface molecule CD19 The virus-specific CTLs from Project 2 will be transduced to express this CAR, resulting in MHC-independent and CAR-dependent activation through chimeric CD28 and CD3-zeta to lyse lymphoid cells, upregulate IL-2, and anti-apoptotic genes in response to CD19. The studies proposed in Specific Aim #1 will evaluate whether UCB-derived multivirus-specific CTLs can be rendered specific for GDI 9 and whether HSV-1 thymidine kinase transgene can be expressed for imaging by positron emission tomography (PET) in immunodeficient mice. Adoptive immunotherapy using non-invasive bioluminescent imaging (BLI) and micro PET will longitudinally asses the persistence of the infused cells, and the anti-tumor effect. Specific Aim #2 will evaluate the feasibility, safety, and persistence of infusing multivirus-specific CTLs modified to express CAR CD19-specific after UCBT. The correlative studies will delineate the (i) magnitude and duration of persistence, (ii) trafficking, and (iii) anti-viral and anti-leukemic effects of adoptively transferred CTLs. In preparation for human PET T-cell imaging, in Specific Aim #3, we will develop a nonhuman primate model Monkey-derived T cells will be genetically modified with CARs and TK, and infused. Longitudinal PET imaging using [18FJFEAU, metabolized by thymidine kinase (TK) will be used to evaluate the distribution of these adoptively transferred macaque T cells. In aggregate, the results of the studies will facilitate the evolution of targeting post-UCBT MRD with viral- and CD19-specific CTLs for enhanced disease-free survival of patients with B cell malignancies.