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;但它受到器官可用性的限制。再生医学可以提供一种替代疗法，使用细胞或实验室培养的人造组织来再生或替换受损器官。这种方法特别适用于胆管疾病，这是儿童和成人肝移植的主要原因，没有其他治疗。胆管是将称为胆汁的有毒消化液从肝脏转移到肠道的管道。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，我在实验室里培养了人类导管细胞，在3D结构中称为类器官。然后，我用我的类器官制作了人造管道，将它们移植到小鼠体内，并展示了它们的治疗潜力。我的目标是利用类器官进行再生医学的首次人体研究。我将首先关注胆道疾病，但开发的技术和诀窍将高度转移到其他器官。为了实现这一目标，我将应对三个主要挑战。1.优化在实验室中生长的类器官用于人体移植移植的细胞应与它们所替代的组织相匹配，并符合监管标准。我将与Bilitech LTD和英国细胞和基因治疗弹射器（CGTC）合作，生产符合人类移植要求的类器官。我将用一种称为单细胞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并最大限度地减少差异。2.开发针对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和太小的胆管，以至于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射到导管中的类器官来达到和再生，为实验室生长的组织提供了一种有价值的治疗替代方案。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挽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类器官不能在临床试验之外移植到人体器官中，大型动物实验是此类研究的先决条件。在与Bilitech LTD和CGTC一起获得UKRI可行性奖的背景下，我已经将我的人工胆管从小鼠尺寸升级到人类尺寸。我将在成年猪中移植这些人工管道，并对动物进行6个月的随访，以证明安全性和有效性。同时，我将使用我开发的新系统探索细胞在人体器官中移植的能力。我将捕获不用于移植的肝脏移植物，通过使含氧血液在肝脏中循环来维持它们在体外（离体），并将我的类器官注射到这些器官中以修复它们的胆管。这项工作将推进使用类器官的再生医学从原理证明到首次人体研究，这与英国再生医学战略（再生医学专家组报告）直接一致。它将解决紧迫的临床需求;提供肝移植的替代方案，肝移植是胆管疾病的唯一治疗方法;并减轻移植计划的压力，使所有其他肝脏疾病受益。它将产生可转移的成果，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的合作将建立学术界和工业界之间的联系。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.