T-cell Therapy for B-lineage Acute Lymphoblastic Leukemia

描述（由申请人提供）：此修订的R 01资助解决了异基因造血干细胞移植（HSCT）后B系急性淋巴细胞白血病（B-ALL）复发的问题。我们假设，通过采用过继转移的供体来源的T细胞（经遗传修饰，对CD 19具有特异性）靶向移植后B-ALL微小残留病（MRD），可以降低同种异体HSCT后癌症复发的发生率。为了巩固HSCT，我们设计了下一代嵌合抗原受体（CAR），命名为CD 19 RCD 28，将T细胞的特异性重定向到B细胞谱系限制的细胞表面分子CD 19，而不依赖于主要组织相容性复合体（MHC）。通过嵌合CD 28和CD 3 -6裂解B-ALL激活的基因修饰的CD 19 RCD 28 + T细胞以CAR调节的方式上调IL-2和抗凋亡基因的产生。睡美人（SB）系统已经与电穿孔组合以引入CAR以及共表达HSV-1胸苷激酶（TK）用于通过正电子发射断层扫描（PET）成像。目标#1中的研究现在将评估是否可以开发全人CD 19特异性CAR（hCD 19 RCD 28），其提供完全有能力的激活信号，如通过SB转座遗传修饰的T细胞中的CD 19依赖性杀伤、细胞因子产生和持续增殖所确定的。将使用播散性B系肿瘤的异种小鼠模型来确定过继治疗的可行性和安全性，所述过继治疗使用非侵入性生物发光成像（BLI）和PET来纵向评估输注的CAR+TK+细胞的持久性和抗肿瘤作用。目的#2将评价在用于高危CD 19 + B-ALL的同种异体HSCT后输注递增剂量的供体来源的hCD 19 RCD 28 + T细胞（有/无TK表达）的安全性、可行性和持久性。表达TK的T细胞将通过PET成像。如有必要，在发生严重毒性的情况下，将给予更昔洛韦（GCV）进行TK+细胞的条件性消融。目标#3中的相关研究将使用载体特异性Q-PCR和TCR谱分析对连续采集的标本描述规定T细胞剂量水平下转移T细胞的数量和持久性。其他相关研究将评估过继转移的T细胞向取样骨髓（BM）的运输以及转移的T细胞在MRD解剖部位的功能状态。使用输注的CAR+ T细胞中共表达的TK代谢/捕获的2 '-脱氧-20-[18 F]氟-5-乙基-1-2-D-阿拉伯呋喃糖基尿嘧啶（[18 F]-FEAU）的人PET成像将用于评价过继转移T细胞的分布。总的来说，研究结果将促进HSCT后MRD与供体来源的CD 19特异性T细胞的靶向进展，以提高B-ALL患者的无病生存期。概要：我们将在移植后输注CD 19特异性T细胞，以提高急性淋巴细胞白血病患者的生存率。

DESCRIPTION (provided by applicant): This revised R01 grant addresses the problem of relapse of B-lineage acute lymphoblastic leukemia (B-ALL) after allogeneic hematopoietic stem-cell transplantation (HSCT). We hypothesize that the incidence of cancer relapse following allogeneic HSCT can be reduced by targeting post-transplant B-ALL minimal residual disease (MRD) with adoptively transferred donor-derived T cells genetically modified to be specific for CD19. To consolidate HSCT, we have designed a next-generation chimeric antigen receptor (CAR), designated CD19RCD28, to redirect specificity of T cells to the B-cell lineage-restricted cell-surface molecule CD19 independent of major histocompatibility complex (MHC). Genetically modified CD19RCD28+ T cells activated through chimeric CD28 and CD3-6 lyse B-ALL, upregulate production of IL-2 and anti-apoptotic genes, in a CAR-regulated manner. The Sleeping Beauty (SB) system has been combined with electroporation to introduce the CAR as well as co-express HSV-1 thymidine kinase (TK) for imaging by positron emission tomography (PET). The studies in Aim #1 will now evaluate whether an all-human CD19-specific CAR can be developed (hCD19RCD28) that provides a fully-competent activation signal as determined by CD19-dependent killing, cytokine production, and sustained proliferation in T cells that have been genetically modified by SB transposition. A xenogeneic mouse model of disseminated B-lineage tumor will be used to ascertain the feasibility and safety of adoptive therapy using non-invasive bioluminescent imaging (BLI) and <PET to longitudinally asses the persistence of the infused CAR+TK+ cells and the anti-tumor effect. Aim #2 will evaluate the safety, feasibility and persistence, of infusing escalating doses of donor-derived hCD19RCD28+ T cells with/without TK expression, after allogeneic HSCT for high-risk CD19+ B-ALL. T cells expressing TK will be imaged by PET. If necessary, ganciclovir (GCV) will be given for conditional ablation of TK+ cells in the event of serious toxicity. Correlative studies in Aim #3 will delineate the magnitude and persistence of transferred T cells at the prescribed T-cell Dose Levels using vector-specific Q-PCR and TCR spectratyping analyses on serially acquired specimens. Other correlative studies will evaluate the trafficking to sampled bone marrow (BM) of adoptively transferred T cells and the functional status of transferred T cells in this anatomic site of MRD. Human PET imaging using 2'-Deoxy-20-[18F]fluoro-5-ethyl-1-2-D-arabinofuranosyluracil ([18F]-FEAU) metabolized/trapped by TK co-expressed in infused CAR+ T cells, will be used to evaluate the distribution of adoptively transferred T cells. In aggregate, the results of the studies will facilitate the evolution of targeting post-HSCT MRD with donor-derived CD19-specific T cells for enhanced disease-free survival of patients with B-ALL. LAY SUMMARY: We will infuse CD19-specific T cells after transplantation to improve survival for patients with acute lymphoblastic leukemia.