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：
10251257
负责人：
CHRISTOPHER Howard EVANS
金额：
$ 50.77万
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/10251257
关键词：
5&apos Untranslated Regions 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 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 Translations Universities Uracil Viral Vector Virus Work base bone bone healing cost demineralization design experimental study gene therapy healing immunoreaction implantation improved in vivo in vivo imaging interest 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.

项目概要/摘要该提案解决了由此产生的大节段性骨缺损的骨愈合问题损伤、肿瘤切除或先天畸形的矫正它们提出了重要的临床问题。因为它们愈合不良并且临床上没有有效的方法来再生丢失的骨头。是由具有强大成骨特性的骨形态发生蛋白（BMP）的克隆引起的重组 BMP-2 和 BMP-7 已进入临床应用，但它们已经改善了该领域。他们令人失望的临床表现被认为反映了分娩问题。蛋白质与胶原蛋白支架混合，并在手术室通过手术植入骨缺损处。大多数 BMP 在数小时内就会留下缺陷。为了解决这个问题，需要大量的超生理量。植入BMP会产生较大的副作用并大大增加成本。 BMP 的交付重点是开发结合 BMP 并释放它们的支架植入后缓慢进行，并进行基因治疗，使骨缺损内部和周围的细胞能够后一种方法的最新数据表明，当 BMP 是内源合成的，BMP 的表达既不需要太长也不需要太高才能有效这提高了传递 mRNA 而不是 DNA 来实现局部、内源性愈合的可能性。虽然理论上令人满意，但这种方法受到限制，因为 mRNA 不稳定，该项目将探索化学修饰的 RNA (cmRNA) 的用途。化学修饰没有这些缺点，其中包括碘取代的嘧啶。尿嘧啶和胞嘧啶；引入特定的 5'-非翻译区 (UTR)；化学修饰 RNA 技术是由技术部发明并首创的。慕尼黑大学 (TUM) 在 PI 的试点研究中生产了编码 BMP-2 的 cmRNA。与 TUM 合作的实验室证实 BMP-2-cmRNA 可以治愈大鼠的关键尺寸缺陷因此，此次合作需要资金以令人印象深刻的速度和可靠性来开展这项工作。该提案旨在进一步实现最终人体试验的目标，即建立治愈的中心前提。当 BMP-2 从 cmRNA A 局部瞬时表达时，临界尺寸缺陷的修复是有效的。对于大鼠，股骨临界尺寸缺损将用于雄性和雌性动物，具体目标 1 将确定。 cmRNA 表达的位置、持续时间和水平具体目标 2 将研究愈合生物学，特别是。关于鉴定表达 BMP-2 并对其作出反应的细胞，因为大型动物研究是一项研究。作为考虑人体试验的必要前奏，Specific Aim 3 将使用绵羊钻孔模型来建立 BMP-2-cmRNA 在绵羊中是否具有成骨作用。