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Use of Chemically Modified RNA to Enhance Bone Healing

使用化学修饰的 RNA 来增强骨愈合

基本信息

批准号：
10673114
负责人：
CHRISTOPHER Howard EVANS
金额：
$ 45.38万
依托单位：
MAYO CLINIC ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10673114
关键词：
5&apos Untranslated Regions Acceleration Address Allografting Animal Model Animals Autologous Autologous Transplantation BMP2 gene BMP7 gene Biochemical Process Biology Blood Bone Matrix Bone Morphogenetic Proteins Cells Chemicals Clinical Clinical Trials Cloning Collaborations Collagen Complementary DNA Congenital Abnormality Cytosine DNA Data Defect Development Disadvantaged Excision Failure Female Femur Funding Gene Delivery Gene Expression Genetic Transcription Goals Growth Factor Harvest Hour Human Implant In Vitro Inflammatory Injury Insertional Mutagenesis Laboratories Location Marketing Measures Messenger RNA Methods Modeling Modification Morbidity - disease rate Natural regeneration Operating Rooms Operative Surgical Procedures Osteogenesis Outcome Performance Pharmaceutical Preparations Physiological Pilot Projects Porifera Production Productivity Property Proteins Protocols documentation Published Comment Pyrimidines RNA Rattus Receptor Activation Recombinant Proteins Recombinants Resources Rodent Model Safety Sheep Site Speed Tail Technology Testing Time Tissue Transplantation Toll-like receptors Transcript Transfection Translations Universities Uracil Viral Vector Virus Work bone bone healing collagen scaffold cost demineralization design experimental study gene therapy healing immunoreaction implantation improved in vivo in vivo imaging interest invention male member osteogenic quantum recombinant human bone morphogenetic protein-2 response scaffold side effect therapeutic gene tumor vector

项目摘要

PROJECT SUMMARY/ABSTRACT This proposal addresses the problem of bone healing in large segmental osseous defects that occur as a result of injury, tumor resection or the correction of congenital deformities. They present important clinical problems because they heal poorly and there are no clinically expedient ways to regenerate the lost bone. Much interest was aroused by the cloning of bone morphogenetic proteins (BMPs), which have potent osteogenic properties in animal models. Recombinant BMP-2 and BMP-7 advanced to clinical use, but they have improved the field only incrementally. Their disappointing clinical performance is thought to reflect delivery problems. The proteins are mixed with a collagen scaffold and surgically implanted into a bone defect in the operating room. Most of the BMP leaves the defect within hours. To address this, massively supraphysiological amounts of the BMP are implanted, leading to major side-effects and greatly increasing the cost. Attempts to improve the delivery of the BMPs have focused on the development of scaffolds that incorporate BMPs and release them slowly after implantation, and gene therapy, which enables cells within and around the osseous defect to synthesize their own BMP endogenously. Recent data from the latter approach demonstrate that, when the BMP is synthesized endogenously, expression of the BMP needs to be neither prolonged nor high for effective bone healing. This raises the possibility of delivery mRNA rather than DNA to enable local, endogenous synthesis of the BMP. While theoretically satisfying, this approach is restricted because mRNA is unstable, toxic to cells and inflammatory. This project will explore the use of chemically modified RNA (cmRNA) that lacks these disadvantages. Among the chemical modifications are the inclusion of iodo-substituted pyrimidines uracil and cytosine; the introduction of specific 5’-untranslated regions (UTRs); and the inclusion of an extended polyA tail. Chemically modified RNA technology was invented and pioneered at the Technical University of Munich (TUM), which has produced a cmRNA encoding BMP-2. Pilot studies in the PI’s laboratory in collaboration with TUM have confirmed that BMP-2-cmRNA heals critical size defects in the rat femur with impressive speed and reliability. Funds are thus requested by this collaboration to develop the work further with the goal of eventual human trials. This proposal seeks to establish the central premise that healing of critical size defects is effective when BMP-2 is expressed locally and transiently from cmRNA molecules. A rat, femoral critical sized defect will be used in both male and female animals. Specific Aim 1 will determine the location, duration and level of cmRNA expression. Specific Aim 2 will study the biology of healing, particularly with regard to identifying the cells that express and respond to BMP-2. Because large animal studies are a necessary prelude to contemplating human trials, Specific Aim 3 will use a sheep drill hole model to establish whether BMP-2-cmRNA is osteogenic in sheep.

项目摘要/摘要该建议解决了由此导致的大段骨缺损处的骨愈合问题。用于损伤、肿瘤切除或矫正先天畸形。它们带来了重要的临床问题。因为它们愈合得很差，而且没有临床上方便的方法来再生丢失的骨骼。很感兴趣是由骨形态发生蛋白(BMPs)的克隆引起的，BMPs具有强大的成骨特性在动物模型中。重组BMP-2和BMP-7已进入临床应用，但它们改善了这一领域只是循序渐进。他们令人失望的临床表现被认为反映了分娩问题。这个蛋白质与胶原蛋白支架混合，通过手术植入手术室的骨缺损处。大多数BMP在几个小时内就会出现缺陷。为了解决这个问题，大量的超生理数量的植入骨形态发生蛋白，导致较大的副作用，大大增加了成本。试图改善 BMP的交付专注于开发包含BMP并释放它们的支架植入后缓慢，以及基因治疗，使骨缺损内和周围的细胞能够内源性合成自己的骨形态发生蛋白。后一种方法的最新数据表明，当 BMP是内源性合成的，BMP的表达既不需要延长也不需要高表达才能有效骨愈合。这增加了传递mRNA而不是DNA的可能性，从而使本地的、内源性的骨形成蛋白的合成。虽然理论上令人满意，但这种方法受到限制，因为mRNA是不稳定的。对细胞有毒性，并有炎症作用。该项目将探索化学修饰的RNA(CmRNA)的使用，缺乏这些缺点。在化学修饰中包括了碘取代的嘧啶类化合物。尿嘧啶和胞嘧啶；引入特定的5‘-非翻译区；以及纳入加长的Polya尾巴。化学修饰的RNA技术是在技术会议上发明并开创的慕尼黑大学(TUM)，它已经产生了编码BMP-2的cmRNA。国际和平研究所的试点研究与TUM合作的实验室已经证实BMP-2-cmRNA可以修复大鼠临界大小的缺陷股骨的速度和可靠性令人印象深刻。因此，这种合作需要资金来发展这项工作更进一步的目标是最终进行人体试验。这项提议试图建立一个中心前提，即治愈当BMP-2从cmRNA分子局部瞬时表达时，临界大小缺陷的表达是有效的。一个雄性和雌性动物都将使用大鼠、股骨临界大小的缺损区。具体目标1将决定 CmRNA表达的部位、持续时间和水平。《特殊目标2》将学习愈合的生物学，特别是关于识别表达BMP-2并对其做出反应的细胞。因为大型动物研究是一项酝酿人体试验的必要前奏，《特定目标3》将利用绵羊钻孔模型建立 BMP-2-cmRNA在绵羊体内是否具有成骨作用。