Regenerative medicine applications of cholangiocyte organoids

移植仍然是所有终末期疾病的唯一治疗方法；但它受到器官供应的限制。再生医学可以提供一种治疗选择，使用细胞或实验室培养的人造组织来再生或替换受损的器官。这种方法尤其适用于胆管疾病，胆管疾病是儿童和成人在没有其他治疗的情况下进行肝移植的主要原因。胆管是将一种叫做胆汁的有毒消化液从肝脏输送到肠道的管道。在疾病中，胆管破裂，胆汁损害周围组织，导致肝功能衰竭。为了修复这种损伤，我在实验室里培育了人类导管细胞，形成了一种叫做类器官的3D结构。然后，我用我的类器官制造了人造导管，将它们移植到老鼠身上，并显示出它们的治疗潜力。我的目标是利用类器官推进再生医学的首次人体研究。我将首先关注胆道疾病，但所开发的技术和诀窍将高度可转移到其他器官。为实现这一目标，我将应对三大挑战。1. 优化实验室培养的用于人体移植的类器官移植细胞应与其所替代的组织相匹配，并符合监管标准。我将与Bilitech LTD和UK Cell and Gene Therapy Catapult （CGTC）合作，生成符合人体移植要求的类器官。我将用一种叫做单细胞RNA测序的高分辨率方法将这些类器官与人类胆管进行比较，并利用这些信息进一步改善我的细胞，尽量减少差异。2. 发展适合不同疾病适应症的类器官移植技术某些疾病可能会影响太多或太小的胆管，因此无法用人工组织进行手术矫正。然而，这些小分支可以通过直接注入导管的类器官来达到和再生，这为实验室培养的组织提供了一种有价值的治疗选择。为了确定这种方法的安全性和有效性，我将尝试使用类器官注射来拯救广泛导管损伤的小动物模型。在大型动物模型和离体人体器官中验证我的类器官的安全性和有效性在临床试验之外，类器官不能移植到人体器官中，而大型动物实验是此类研究的先决条件。在Bilitech LTD和CGTC的UKRI可行性奖的背景下，我已经将我的人工胆管从老鼠的大小扩大到人类的大小。我将在成年猪身上移植这些人工导管，并对动物进行6个月的随访，以证明其安全性和有效性。与此同时，我将利用我开发的新系统探索细胞植入人体器官的能力。我将捕获不用于移植的肝移植物，通过肝脏循环含氧血液在体外（离体）维持它们，并将我的类器官注射到这些器官中，以修复它们的胆管。这项工作将推动使用类器官的再生医学从原理验证到首次人体研究，这与英国再生医学战略（再生医学专家组报告）直接一致。它将解决迫切的临床需求；提供一种替代肝移植的方法，肝移植是胆管疾病的唯一治疗方法；并减少对其他肝脏疾病有益的移植项目的压力。它将产生可转移的产出，影响更广泛的领域，如组织工程，类器官生物学和生物信息学，这可以指导我未来的研究。与Bilitech LTD和CGTC的合作将加强学术界和工业界的联系。所开发的专业知识和技术将适用于其他组织和器官，并将为英国再生和药物制造技术做出重大贡献。

Transplantation remains the only treatment for all end-stage diseases; but it is limited by organ availability. Regenerative medicine could provide a therapeutic alternative, using cells or lab-grown artificial tissue to regenerate or replace damaged organs. This approach is particularly relevant in bile duct diseases, which are a leading cause for liver transplantation in children and adults with no other treatment. The bile ducts are tubes transferring a toxic digestive fluid, called bile, from the liver to the intestine. In disease, bile ducts break down and bile damages surrounding tissue causing liver failure. To repair this damage, I grew human duct cells in the lab, in 3D structures called organoids. I then made artificial ducts from my organoids, transplanted them in mice and showed their therapeutic potential. My goal is to progress regenerative medicine using organoids to first-in-man studies. I will first focus on biliary disease, but the technology and know-how developed will be highly transferable to other organs. To achieve this, I will address three main challenges. 1. Optimise organoids grown in the lab for human transplantationTransplanted cells should match the tissue they are replacing and meet regulatory standards. I will collaborate with Bilitech LTD and the UK Cell and Gene Therapy Catapult (CGTC) to generate organoids complying with requirements for human transplantation. I will compare these organoids to human bile ducts with a high-resolution method called Single Cell RNA sequencing and use this information to further improve my cells and minimize differences. 2. Develop organoid transplantation techniques tailored to different disease indicationsCertain diseases may affect too many and too small bile ducts to be amenable to surgical correction with artificial tissue. However, these small branches may be reached and regenerated by organoids injected directly in the ducts, providing a valuable therapeutic alternative to lab-grown tissue. To establish the safety and efficacy of this approach, I will attempt to rescue small animal models of widespread duct damage using organoid injection.3. Validate the safety and efficacy of my organoids in large animal models and ex-vivo human organsOrganoids can't be transplanted in human organs outside clinical trials, and large animal experiments are a pre-requisite for such studies. In the context of a UKRI feasibility award with Bilitech LTD and CGTC, I have upscaled my artificial bile ducts from mouse to human size. I will transplant these artificial ducts in adult pigs and follow up the animals for 6 months to demonstrate safety and efficacy. In parallel, I will explore the cells' capacity to engraft in human organs using a new system I developed. I will capture liver grafts not used for transplantation, maintain them outside the body (ex-vivo) by circulating oxygenated blood through the liver, and inject my organoids in these organs to repair their bile ducts. This work will advance regenerative medicine using organoids from proof-of-principle to first-in-human studies, which is in direct alignment with the UK strategy on Regenerative Medicine (Regenerative Medicine Expert Group report). It will address a pressing clinical need; provide an alternative to liver transplantation, which is the only treatment for bile ducts disorders; and reduce pressure on the transplantation program benefiting all other liver diseases. It will generate transferrable outputs impacting on broader fields, such as tissue engineering, organoid biology and bioinformatics, which can instruct my future research. The collaboration with Bilitech LTD and CGTC will forge ties between academia and industry. The expertise and technology developed will be applicable to other tissues and organs and will make a significant contribution to UK regenerative and medicines manufacturing know-how.