The role of BCL11B in T lineage fate during human thymopoiesis and pluripotent stem cell differentiation

BCL11B 在人类胸腺生成和多能干细胞分化过程中 T 谱系命运中的作用

• 批准号: 10639378
• 负责人: Gay M Crooks
• 金额: $ 84.6万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10639378
• 关键词: Affect; Autologous; Binding; CHD4 gene; Cell Lineage; Cells; Cellular Indexing of Transcriptomes and Epitopes by Sequencing; Chromatin; Data; Development; Epigenetic Process; Event; Generations; Genes; Genetic Engineering; Genetic Transcription; Goals; Hematopoiesis; Hematopoietic stem cells; Heterogeneity; Human; Immunotherapy; Infection; Knowledge; Leukocytes; Lymphoid; Lymphoid Cell; Mature T-Lymphocyte; Mediating; Methods; Molecular; Mus; NOTCH1 gene; NuRD complex; Organoids; Output; Pathway interactions; Patients; Pattern; Pluripotent Stem Cells; Population; Process; Production; Publishing; Reporter; Repression; Role; Series; Signal Transduction; Sorting; Source; Specific qualifier value; System; T cell differentiation; T cell therapy; T-Cell Development; T-Lymphocyte; Therapeutic; Thymus Gland; Umbilical Cord Blood; Variant; anti-cancer; cancer cell; candidate identification; chimeric antigen receptor; chromatin remodeling; design; fighting; histone modification; human pluripotent stem cell; improved; in vitro Model; indexing; insight; knock-down; loss of function; manufacture; mouse model; notch protein; novel; overexpression; progenitor; programs; recruit; self-renewal; single-cell RNA sequencing; stem cell differentiation; stem cells; transcription factor

ABSTRACT/SUMMARY The functional limitations and logistical challenges of using patient-derived (autologous) products for adoptive T cell therapy has prompted the exploration of a universal source of “off-the-shelf” T cells generated from self- renewing PSCs which can be readily genetically engineered to enhance function and expanded without limit. However current PSC differentiation systems are characterized by low T cell output and concurrent production of innate lymphoid cells (ILCs). Our preliminary studies suggest that the earliest stages of T cell specification and commitment seen during PSC differentiation do not fully recapitulate either normal human thymopoiesis or in vitro models that use definitive hematopoietic stem and progenitor cells (HSPC) to initiate T cell development. The goal of this proposal is to understand the cellular and molecular differences between normal and PSC-derived T cell development, with a focus on the role of the transcription factor BCL11B. T cells are generated in the thymus after notch signaling from the microenvironment triggers a series of transcriptional events that initiate the T-lineage program in HSPCs; these events first produce early thymic progenitors (ETPs) (T lineage specification) and then extinguish alternative (non-T) lineage programs in multipotent ETPs (T lineage commitment). BCL11B is a critical regulator of both of these processes. Our published and preliminary data show that, in contrast to the mouse model, BCL11B is essential for T cell specification during human thymopoiesis and initiates the expression of several T-cell genes. Moreover, when BCL11B is overexpressed in cord blood HSPCs, the T cell program is launched more rapidly and efficiently, even in the absence of notch signaling. Surprisingly little is known about how the T cell lineage is generated from PSCs. Through scRNA-Seq analysis we have identified candidate ETPs and their immediate progeny as they emerge from PSC-derived hematopoiesis. We hypothesize that the rare PSC-derived ETPs in which the T cell program is launched are functionally and transcriptionally different from ETPs in the thymus, and that these intrinsic differences are detrimental for the generation of conventional T cells from PSCs. Further, we propose that chromatin remodeling induced by BCL11B mediates both T lineage specification and the fate decisions between the conventional T cell and innate lymphoid pathways. Specifically we will: 1. Define the earliest T lineage progenitors generated during PSC differentiation; 2. Determine the epigenetic underpinnings of T-cell specification in PSC-ATOs and in primary thymopoiesis; and 3. Define how BCL11B affects conventional T and innate lineage fate choices. These studies will yield new mechanistic insights about T-cell differentiation that are critical for the development of PSC-derived T-cell immunotherapies.

摘要/总结 使用患者来源（自体）产品进行过继 T 的功能限制和后勤挑战 细胞疗法促使人们探索一种通用来源的“现成”T 细胞，这种细胞是由自体产生的。 更新的 PSC 可以很容易地进行基因改造，以增强功能并无限扩展。 然而，目前的 PSC 分化系统的特点是 T 细胞产量低且同时生产 先天淋巴细胞（ILC）。我们的初步研究表明 T 细胞分化的最早阶段 PSC 分化过程中看到的承诺并不能完全重现正常的人类胸腺生成或 使用定型造血干细胞和祖细胞 (HSPC) 启动 T 细胞发育的体外模型。 该提案的目标是了解正常和正常之间的细胞和分子差异 PSC 衍生的 T 细胞发育，重点关注转录因子 BCL11B 的作用。 T 细胞是 来自微环境的Notch信号触发一系列转录后在胸腺中产生 在 HSPC 中启动 T 谱系计划的事件；这些事件首先产生早期胸腺祖细胞（ETP） （T 谱系规范），然后消除多能 ETP 中的替代（非 T）谱系程序（T 谱系 承诺）。 BCL11B 是这两个过程的关键调节因子。我们公布的初步数据显示 与小鼠模型相比，BCL11B 对于人类胸腺生成过程中的 T 细胞规范至关重要 启动多个 T 细胞基因的表达。此外，当BCL11B在脐带血HSPC中过度表达时， 即使在没有 Notch 信号传导的情况下，T 细胞程序也能更快、更有效地启动。出奇 关于 T 细胞谱系是如何从 PSC 产生的，人们知之甚少。通过 scRNA-Seq 分析，我们得到 确定了候选 ETP 及其直接后代，因为它们是从 PSC 衍生的造血过程中产生的。我们 假设启动 T 细胞程序的罕见 PSC 衍生 ETP 在功能上和 转录上与胸腺中的 ETP 不同，并且这些内在差异是有害的 用于从 PSC 生成常规 T 细胞。此外，我们提出染色质重塑诱导 BCL11B 介导 T 谱系规范以及传统 T 细胞和 T 细胞之间的命运决定。 先天淋巴通路。具体来说，我们将： 1. 定义 PSC 期间产生的最早的 T 谱系祖细胞 差异化； 2. 确定 PSC-ATO 和原代 T 细胞规范的表观遗传基础 胸腺生成； 3. 定义 BCL11B 如何影响传统 T 和先天谱系命运选择。这些研究 将产生关于 T 细胞分化的新机制见解，这对于 PSC 衍生的细胞的开发至关重要 T 细胞免疫疗法。