EAGER: Biomanufacturing: Engineering Cell-Intrinsic Control of Cardiomyocyte Differentiation in Human Pluripotent Stem Cells

主要研究者：Palecek，Sean P.提案编号：1547225人类多能干细胞（hPSC）发现的最新科学进展以及将hPSC定向为特定细胞类型的能力，为人类发育和疾病、药物筛选和安全性分析以及基于细胞的再生疗法的研究带来了革命性的潜力。 然而，为了实现hPSC的这一前景，需要高效的生物制造平台来稳健地产生大量高质量的hPSC及其衍生物。 传统的生物制造策略采用生物反应器来控制细胞的环境，提供指导细胞命运的外在信号。 在这里，研究人员提出了一种新的内在范式来控制hPSC的命运，利用分子生物学的最新进展来编程hPSC在没有外部线索的情况下发育成心肌细胞。 该项目的成功完成将为hPSC如何决定转化为更特化的细胞提供更深入的基本理解，并为通过工程细胞从干细胞生物制造特化细胞提供新的策略。 该项目将开发一种新的范式，用于控制人类多能干细胞（hPSC）分化，利用细胞内在的细胞状态传感和发育信号通路的控制。 研究人员将利用最新进展来了解经典Wnt信号传导如何驱动hPSC向中胚层的承诺以及随后向心脏中胚层和功能性心肌细胞的特异性，以设计感知其分化状态并驱动信号传导以引发所需分化轨迹的细胞。 研究人员将鉴定中胚层和心脏中胚层特异性启动子元件，并使用这些元件来激活Wnt通路调节剂的表达，产生在没有外源信号的情况下分化为功能性心肌细胞的hPSC系。 然后，研究人员将测试这样的假设，即细胞内在的分化控制比外在控制通过在高通量筛选应用的规模缩小和适用于临床评价的细胞生产的规模扩大期间应用小分子实现更稳健和可扩展的心肌细胞生物制造。 该项目的更广泛影响包括为工程化hPSC系提供蓝图，用于心肌细胞以外的体细胞谱系的生物制造。教育和外展活动也将在研究生和本科生水平的干细胞工程师，并将提供外展K-12学生，K-12教师，本科生，和一般公众对干细胞科学和生物制造方面。

PI: Palecek, Sean P. Proposal Number: 1547225Recent scientific advances in the discovery of human pluripotent stem cells (hPSCs) and the ability to direct hPSCs to specialized cell types offer the potential to revolutionize the study of human development and disease, drug screening and safety profiling, and cell-based regenerative therapies. To realize this promise of hPSCs, however, efficient biomanufacturing platforms are needed to robustly generate large quantities of high quality hPSCs and their derivatives. Traditional biomanufacturing strategies employ bioreactors to control the environment of the cell, providing extrinsic signals that direct cell fates. Here, the investigators propose a novel intrinsic paradigm for control of hPSC fate, using recent advances in molecular biology to program an hPSC to develop into a heart muscle cell in the absence of external cues. Successful completion of this project will provide a deeper fundamental understanding of how hPSCs make decisions to turn into more specialized cells and provide a new strategy to biomanufacture specialized cells from stem cells by engineering the cell. This project will develop a novel paradigm for controlling human pluripotent stem cell (hPSC) differentiation, using cell intrinsic sensing of cell state and control of developmental signaling pathways. The investigators will employ recent advances in understanding how canonical Wnt signaling drives hPSC commitment to mesoderm and subsequent specification to cardiac mesoderm and functional cardiomyocytes to design cells that sense their differentiation status and drive signaling to elicit desired differentiation trajectories. The investigators will identify mesoderm and cardiac mesoderm-specific promoter elements and use these to activate expression of Wnt pathway modulators, generating hPSC lines that differentiate to functional cardiomyocytes in the absence of extrinsic signaling. Then, the investigators will test the hypothesis that cell intrinsic control of differentiation enables more robust and scalable biomanufacturing of cardiomyocytes than extrinsic control via application of small molecules during scale down for high throughput screening application and scale up for production of cells suitable for clinical evaluation. The broader impacts of this project include providing a blueprint for engineering hPSC lines for biomanufacturing of somatic lineages beyond cardiomyocytes. Education and outreach activities will also stem cell engineers at the graduate and undergraduate levels, and will provide outreach to K-12 students, K-12 teachers, undergraduate students, and the general public on aspects of stem cell science and biomanufacturing.