Investigating immunophenotype and metabolism of TCR KO donor and third-party CD19-targeted chimeric antigen receptor T cells

摘要 复发是异基因造血细胞移植(allo-hct)后死亡的最重要原因，但 几十年来，研究进展甚微。针对CD19(CD19)的嵌合抗原受体 Cars)改变T细胞效应器功能，以消除表达CD19的白血病和淋巴瘤细胞。然而， 许多患者仍在复发。候选人有临床前数据表明使用基因组编辑的可行性 删除内源性T细胞受体(TCR)以降低捐赠者CD19 CAR T细胞的同种异体反应性，但它是 尚不清楚这些TCR基因敲除(KO)细胞在体内如何作为抗肿瘤药物发挥作用，在多大程度上移植- 将导致抗宿主病(GVHD)，或工程如何影响T细胞代谢。这一知识是 在为血液病患者创造现成的CAR T细胞方面取得进展 恶性肿瘤。拟议研究的总体目标是确定捐赠者和第三方TCR如何 KO CD19 CAR T细胞在临床前模型中影响免疫重建、移植物抗宿主病和移植物排斥反应 了解去除TCR如何影响这些细胞的免疫代谢。中心假设是 强大的抗肿瘤作用以及微不足道的移植物抗宿主病和移植物排斥反应可以在临床前得到证实。 通过在allo-HCT后使用TCR KO CD19 CAR T细胞(供者或第三方)，产生更好的 异基因红细胞移植后常规CD19 CAR T细胞的转归。这一假设将在拟议的 明确的目标。 这项工作将为候选人提供理想的培训，因为她正在为领导一个 研究细胞疗法和异基因血细胞移植的独立实验室。纪念斯隆·凯特琳癌症中心 有一个著名的免疫学项目，候选人的主要导师马塞尔·范登布林克博士是一位领导者 在免疫治疗研究中。她的共同导师和咨询委员会成员具有多样性和互补性 他们都有指导独立科学家的良好记录。候选人和她 指导团队制定了严格的培训计划，旨在增加她在以下方面的知识基础：1) 开发下一代CAR T细胞；2)生物信息学和编程；3)细胞代谢， CAR T细胞的代谢流量、线粒体功能和代谢分析；以及4)职业发展 技能。应聘者将通过课程作业、研讨会和指导来进行这些领域的培训。 这项研究项目和培训将为她未来的独立职业生涯奠定基础 医生兼科学家。拟议中的研究有望产生指导未来基因组的发现 CAR T细胞工程学。这位候选人的目标是启动一项独立的研究计划，设计“脱机”。 -货架“CAR T细胞，预计将为一系列 恶性血液病。

Abstract Relapse is the most important cause of mortality after allogeneic hematopoietic cell transplant (allo-HCT), but little research progress has been made in several decades. Chimeric antigen receptors targeting CD19 (CD19 CARs) redirect T cell effector functions to eliminate CD19-expressing leukemia and lymphoma cells. However, many patients still relapse. The candidate has preclinical data indicating the feasibility of using genome editing to delete the endogenous T cell receptor (TCR) to reduce the alloreactivity of donor CD19 CAR T cells, but it is unknown how these TCR knockout (KO) cells will function in vivo as anti-tumor agents, to what extent graft- versus-host-disease (GVHD) will result, or how engineering impacts T cell metabolism. This knowledge is essential for progress toward creating readily available “off-the-shelf” CAR T cells for patients with hematologic malignancies. The overall objectives of the proposed research are to determine how donor and third-party TCR KO CD19 CAR T cells impact immune reconstitution, GVHD, and graft rejection in preclinical models, as well as to understand how removing the TCR impacts the immunometabolism of these cells. The central hypothesis is that potent anti-tumor effects as well as negligible GVHD and graft rejection can be demonstrated preclinically by using TCR KO CD19 CAR T cells (either donor or third-party) following allo-HCT, to produce superior outcomes to conventional CD19 CAR T cells following allo-HCT. This hypothesis will be tested in the proposed Specific Aims. This work will provide ideal training for the candidate as she prepares for her long-term career goal to lead an independent laboratory studying cellular therapeutics and allo-HCT. Memorial Sloan Kettering Cancer Center has a renowned immunology program, and Dr. Marcel van den Brink, the candidate’s primary mentor, is a leader in immunotherapy research. Her co-mentor and advisory committee members have diverse and complementary expertise, and all have strong track records of mentoring independent scientists. The candidate and her mentoring team have developed a rigorous training plan designed to increase her knowledge base in: 1) development of next-generation CAR T cells; 2) bioinformatics and programming; 3) cellular metabolism, metabolic flux, mitochondrial function, and metabolic analyses in CAR T cells; and 4) professional development skills. The candidate will undertake training in these areas through coursework, workshops, and mentorship. This research project and training will provide the foundation to establish her future career as an independent physician-scientist. The proposed studies are expected to generate findings that will guide future genome engineering of CAR T cells. The candidate’s aim is to launch an independent research program designing “off- the-shelf” CAR T cells, which are expected to provide much improved therapeutic options for a range of hematologic malignancies.