Targeted non-genotoxic hematopoietic stem cell transplant conditioning approach

项目摘要 造血干细胞移植（HSCT）和基因治疗越来越多地用于确定性管理 影响淋巴造血系统的疾病条件反射疗法是 组件HSCT。目前使用的大多数预处理方案要么基于烷基化， 化疗，如白消安，或涉及不同剂量的全身照射（TBI）。这些试剂 遗传毒性和影响器官以外的造血隔室，导致无数的急性和长期 HSCT相关并发症。我们的首要目标是开发一种无遗传毒性的调理方案 靶向造血干细胞和祖细胞（HSPC）。我们实验室的初步数据表明MPL （血小板生成素受体），造血干细胞（HSC）的关键生存信号，可能是一个理想的 开发HSPC靶向HSCT预处理方法的候选者。限制性表达谱 在造血隔室内，与造血隔室相比，HSC中的表达更高。 祖细胞区室，使MPL成为一个非常有吸引力的目标。我们制定了两项战略， 表达MPL的HSPC隔室。首先，免疫毒素方法，通过缀合MPL抗体产生 核糖体毒素皂草素（MPL-ab-SAP），第二种是MPL受体结合血小板生成素， CAR-T细胞（TPO-CAR-T细胞）。我们假设，针对MPL受体的调节剂将导致 在选择性HSPC耗竭中，从而产生足够的生态位空间以允许供体植入， 在造血区室之外的细胞毒性风险最小。我们将确定机制， HSC区室中MPL-ab-SAP介导的细胞毒性的关键决定因素。然后，我们将确定 通过MPL-ab-SAP的HSPC消耗是否可用作HSCT的有效预处理策略， 自体造血干细胞基因治疗还将研究TPO-CAR-T细胞作为HSCT调节的作用。 K 08奖将用于保护我的研究时间，帮助拓宽我的研究技能和知识， 并为基础和转化HSCT和基因治疗的职业生涯提供出色的基础。这一指导 培训阶段将使我能够进一步整合和建立我的研究技能作为一个医生科学家。超过 在接下来的五年里，我将（1）获得开发治疗药物所需的分子方法方面的专业知识， 抗体和基于基因治疗的治疗方法，（2）在HSC生物学，移植 免疫学和小鼠HSCT，（3）获得细胞成像和生物信息学技能，（4）获得 翻译研究方法，grananisation和领导，（5）建立在我的出版记录， 为独立研究赠款提供研究思路。为了实现这些目标，我制定了一个 一个结构良好的培训计划，我已经组建了一个高素质的导师团队博士斯宾塞（专业知识 Bunting博士（HSC生物学），Waller博士（HSPC生物学和临床移植）， 博士拉森（移植免疫学）在埃默里大学。

Project Summary Hematopoietic stem cell transplant (HSCT) and gene therapy are increasingly utilized for definitive management of disorders affecting the lymphohematopoietic compartment. Conditioning regimens are one of the critical components HSCT. The majority of conditioning regimens used today are either based on alkylating chemotherapy, such as busulfan, or involve varying doses of total body irradiation (TBI). These agents are genotoxic and affect organs other than the hematopoietic compartment, leading to myriad acute and long-term HSCT associated complications. Our overarching goal is to develop a non-genotoxic conditioning regimen targeting hematopoietic stem and progenitor cells (HSPCs). Preliminary data from our lab indicates MPL (thrombopoietin receptor), a critical survival signal in hematopoietic stem cells (HSCs), could be an ideal candidate for development of HSPC-targeted HSCT conditioning approaches. Restricted expression profiles within the hematopoietic compartment, with a higher expression in HSC when compared to the hematopoietic progenitor compartment, make MPL a very attractive target. We have developed two strategies to target the MPL expressing HSPC compartment. First, an immunotoxin approach, generated by conjugating MPL antibody to a ribosomal toxin, saporin (MPL-ab-SAP), and the second, a MPL receptor engaging thrombopoietin based CAR-T cell (TPO-CAR-T cells). We hypothesize that a conditioning agent targeting the MPL receptor will result in selective HSPC depletion thereby creating adequate niche space to allow donor engraftment and will have minimal risk of cytotoxicity outside the hematopoietic compartment. We will determine the mechanism, and critical determinants of MPL-ab-SAP-mediated cytotoxicity in the HSC compartment. Then, we will determine whether HSPC depletion by MPL-ab-SAP could be utilized as an effective conditioning strategy for HSCT and autologous HSC-based gene therapy. The role of TPO-CAR-T cells as an HSCT conditioning will also be studied. The K08 Award will be used to protect my time for research, help broaden my research skills and knowledge, and provide an outstanding basis for a career in basic and translational HSCT and gene therapy. This mentored phase of training will allow me to further integrate and build on my research skills as a physician-scientist. Over the next five years, I will (1) gain expertise in molecular methods needed for development of therapeutic antibodies, and gene therapy based therapeutics, (2) gain further exposure in HSC biology, transplant immunology, and murine HSCT, (3) acquire skills in cell imaging and bioinformatics, (4) acquire skills in translational research methodology, grantsmanship and leadership, and (5) build on my publication record and develop research ideas for independent investigator grants. To accomplish these goals, I have developed a well-structured training plan, and I have assembled a highly-qualified mentorship team of Dr. Spencer (expertise in cell and gene therapy), Dr. Bunting (HSC biology), Dr. Waller (HSPC biology and clinical transplantation), and Dr. Larsen (transplant immunology) at Emory University.