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Combining iPSC and gene editing with cell therapy to develop the next generation of MSC-based therapeutics to counteract bone fragility in OI.

将 iPSC 和基因编辑与细胞疗法相结合，开发下一代基于 MSC 的疗法，以对抗成骨不全症中的骨脆性。

基本信息

批准号：
MR/S009256/1
负责人：
Pascale V GUILLOT
金额：
$ 111.67万
依托单位：
University College London
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2019
资助国家：
英国
起止时间：
2019 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=MR%2FS009256%2F1
关键词：
Combining iPSC gene editing cell

项目摘要

Bone loss and skeletal fragility impose a huge healthcare and economic burden, affecting millions of people in the world due to the ageing process, microgravity, prolonged immobilisation, or pathologies such as osteoporosis or the brittle bone disease osteogenesis imperfecta (OI). OI develops because a genetic defect causes the main component of the bone matrix (type I collagen) to be faulty, preventing the bone-forming cells (osteoblasts) from maturing, which causes the bones to become brittle. There is no cure for OI and patients need care all their lives. Thus, developing therapeutics to counteract bone fragility is of paramount importance. Using a pre-clinical model of OI, we have recently shown that transplantation of human mesenchymal stem cells (MSC) isolated from the pregnancy fluid surrounding healthy babies improved the quality of fragile bones, stimulated the maturation of resident osteoblasts and reduced the activity of resident bone resorbing cells (osteoclasts). However, there are two main hurdles to overcome before MSC can be routinely used in the clinic. First, MSC isolated from donor organs or pregnancy fluid progressively lose their regenerative potential whilst they are cultivated in the laboratory to reach sufficient numbers. Second, we need to understand how MSC work in vivo to improve transplantation protocols and develop tests to determine the repair potential of putative new donor cell samples. Luckily, a new source of MSC is rejuvenated cells, also called induced pluripotent stem cells (iPSC). Skin cells, which can be isolated from healthy or OI patients, can be rejuvenated in vitro and the genetic defect causing OI can be repaired through genome editing. Since iPSC do not change during culture, it is possible to obtain sufficient number of cells before converting them back into MSC. These cells are called induced MSC (iMSC). This raises the possibility to develop the next generation of cell therapy using iMSC derived from the patient's own cells. The aim of this project is to to test the potential of iMSC to counteract bone fragility and replace the use of primary MSC and also to understand how donor MSC ameliorate OI bones. This project will test whether transplantation of iMSC (derived from healthy iPSC or from OI-iPSC following correction of the genetic defect) into a pre-clinical model of OI improves bone quality and normalise osteoblast function. We will also establish whether the differentiation of donor MSC towards the osteoblast lineage directly contributes to bone formation and/or whether the tiny sacks (called exosomes) released by MSC function as activators to stimulate resident osteoblasts and/or inhibit osteoclasts. Finally, we will use ex vivo co-culture to assess how the OI genetic defect impedes the behaviour of OI human and mouse osteoblasts and determine whether the function of bone-forming and bone-resorbing cells can be modulated by human MSC through direct cell contact, or via exosomes or the soluble proteins secreted by human MSC. Our long-term goal is to develop personalized treatments for people suffering from bone loss and at risk of fractures using patients' own skin cells.

骨丢失和骨骼脆性造成了巨大的医疗和经济负担，由于老龄化过程、微重力、长时间静止或骨质疏松症或脆性骨病成骨不全(OI)等病理因素，世界上数百万人受到影响。OI的发生是因为遗传缺陷导致骨基质的主要成分(I型胶原)出现缺陷，阻止骨形成细胞(成骨细胞)成熟，从而导致骨骼变得脆弱。OI无法治愈，患者需要终生护理。因此，开发治疗方法来抵消骨脆性是至关重要的。利用OI的临床前模型，我们最近发现，从健康婴儿周围的孕液中分离出的人间充质干细胞(MSC)移植改善了脆性骨骼的质量，刺激了常驻成骨细胞的成熟，并降低了常驻骨吸收细胞(破骨细胞)的活性。然而，在MSC可以在临床上常规使用之前，有两个主要的障碍需要克服。首先，从捐献器官或妊娠液中分离出来的MSC在实验室培养到足够数量时会逐渐失去再生能力。其次，我们需要了解MSC如何在体内发挥作用，以改进移植方案，并开发测试来确定假定的新捐赠者细胞样本的修复潜力。幸运的是，MSC的新来源是再生细胞，也被称为诱导多能干细胞(IPSC)。可以从健康或OI患者身上分离的皮肤细胞可以在体外恢复活力，导致OI的遗传缺陷可以通过基因组编辑来修复。由于IPSC在培养过程中不会发生变化，因此在将其转化回MSC之前获得足够数量的细胞是可能的。这些细胞被称为诱导间充质干细胞(IMSC)。这增加了使用来自患者自身细胞的iMSC开发下一代细胞疗法的可能性。该项目的目的是测试iMSC在抵消骨脆性和取代原始MSC使用方面的潜力，并了解供体MSC如何改善OI骨。该项目将测试将iMSC(来源于健康的IPSC或纠正遗传缺陷后的OI-IPSC)移植到OI的临床前模型中是否可以改善骨骼质量并使成骨细胞功能正常化。我们还将确定供体MSC向成骨细胞谱系的分化是否直接有助于骨形成和/或由MSC释放的微小袋子(称为外切体)是否起到激活作用，刺激常驻成骨细胞和/或抑制破骨细胞。最后，我们将使用体外共培养来评估OI基因缺陷如何阻碍OI人和小鼠成骨细胞的行为，并确定人MSC是否可以通过直接细胞接触、或通过外体或人MSC分泌的可溶性蛋白来调节成骨细胞和骨吸收细胞的功能。我们的长期目标是利用患者自己的皮肤细胞为骨丢失和骨折风险的患者开发个性化的治疗方法。