Nanowired human isogenic cardiac organoids to treat acute myocardial ischemia/reperfusion injuries

项目摘要：在美国，每年有超过735，000例心肌梗塞（MI）。而 经皮冠状动脉介入治疗（PCI）显著降低了急性不良反应， 缺血/再灌注（I/R）后患者的预后仍然很差。由于有限的再生能力， 在人类心脏中，人类多能干细胞衍生的心肌细胞（hPSC-CM）已经接受了显著的 由于它们被证明在移植到受损心脏后恢复收缩功能的能力， 各种哺乳动物模型，导致多个正在进行的临床试验。目前，移植 方法主要依赖于解离的hPSC-CM，导致低细胞存活，适度的功能改善， 易出错的风险和较差的可扩展性。为了应对这些挑战，我们的实验室开发了一种新的， 由hPSC-CM组成的血管化人心脏类器官，人原代心脏成纤维细胞， 内皮细胞、基质细胞和导电硅纳米线（e-SiNW）。内皮细胞是 用于诱导类器官内的脉管系统形成，并且添加e-SiNW以产生电连接。 导电微环境，以促进hPSC-CM收缩发育和它们与 宿主心肌我们的初步体内数据显示，有毛类器官显示了强大的hPSC- CM植入和上级功能恢复。其临床翻译的主要障碍包括：1） 在细胞和类器官培养中使用动物蛋白，以及2）在细胞和类器官培养中缺乏功能益处证明， 大型动物模型。用等基因hPSC衍生的细胞替代人原代细胞用于类器官制造 通过主要组织相容性降低批次间差异并增强免疫相容性 图2示出了将hPSC供体与人受体匹配的MHC复合物（MHC）。此外，虽然目前的hPSC-CM 植入策略一直集中在心肌内注射，开发了一种有效的方法， 类器官的冠状动脉内递送将加速它们的临床转化。本提案的目的是 开发临床级hPSC心脏类器官，并证明其在大型动物中的功能益处 模型，以生成IND提交的启用数据。这一提议的核心假设是： 同基因hPSC心脏类器官提供了可扩展系统以高效且有效地植入hPSC-CM 用于心脏修复该提议是创新的，因为我们将1）在无异种中衍生等基因hPSC衍生细胞， 化学定义的条件，以开发用于植入的临床级心脏类器官，以及2）利用 大小和类器官中的内皮管腔样结构，以开发有效的冠状动脉内递送 战略因此，我们将追求以下两个目标：1）制造和表征头发的人 - 心脏类器官，其在无异种、化学限定的条件下使用源自hPSC的同基因心脏细胞， 和2）用猪I/R测定经纯化的同基因hPSC心脏类器官的治疗功效 （缺血/再灌注）模型。

Project Summary: In the U.S., there are more than 735,000 myocardial infarctions (MI) each year. While percutaneous coronary intervention (PCI) has significantly reduced acute adverse repones, the long-term prognosis for post-ischemia/reperfusion (I/R) patients remains poor. Due to the limited regenerative capacity of human hearts, human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have received significant attention due to their proven capacity to restore contractile function upon transplantation to injured hearts in various mammalian models, leading to multiple ongoing clinical trials. However, the current transplantation approach mainly relies on dissociated hPSC-CMs, leading to low cell survival, moderate functional improvement, arrhythmogenic risk, and poor scalability. To address these challenges, our lab developed nanowired, pre- vascularized human cardiac organoids composed of hPSC-CMs, human primary cardiac fibroblasts, endothelial cells, stromal cells, and electrically conductive silicon nanowires (e-SiNWs). Endothelial cells are used to induce vasculature formation within the organoids, and e-SiNWs are added to create an electrically conductive microenvironment to facilitate hPSC-CM contractile development and their electrical integration with the host myocardium. Our preliminary in vivo data showed that nanowired organoids illustrated robust hPSC- CM engraftment and superior functional recovery. The major barriers in their clinical translation include: 1) the use of animal proteins in the cell and organoid culture and 2) the lack of functional benefit demonstration in a large animal model. Replacing human primary cells with isogenic hPSC-derived cells for organoid fabrication would reduce batch-to-batch variations and enhance immune compatibility through Major Histocompatibility Complex (MHC) matching hPSC donors with human recipients. In addition, while the current hPSC-CM implantation strategy has been focused on intramyocardial injection, developing an effective approach for intracoronary delivery of the organoids will accelerate their clinical translation. The goal of this proposal is to develop clinical-grade hPSC cardiac organoids and demonstrate their functional benefits with a large animal model to generate enabling data for IND submission. The central hypothesis of this proposal is the nanowired isogenic hPSC cardiac organoids provide a scalable system to both efficiently and effectively implant hPSC-CMs for cardiac repair. The proposal is innovative in that we will 1) derive isogenic hPSC-derived cells in xeno-free, chemically defined conditions to develop clinical-grade cardiac organoids for implantation and 2) leverage the size and the endothelial lumen-like structures in the organoids to develop an effective intracoronary delivery strategy. Accordingly, we will pursue the following 2 aims: 1) Fabricate and characterize nanowired human cardiac organoids using isogenic cardiac cells derived from hPSCs in xeno-free, chemically defined conditions, and 2) Determine the therapeutic efficacy of the nanowired isogenic hPSC cardiac organoids with a porcine I/R (ischemia/reperfusion) model.