Nanolipidoids-Conjugated MicroRNA Enhance Oral and Cranial Bone Regeneration

 描述（由申请人提供）：虽然美国每年有超过500，000例矫正骨畸形和严重尺寸缺损的手术，但50%的手术以失败告终。因此，利用分子工程方法使骨缺损再生，使其具有正常的解剖外形、物理性质和生物学功能，具有重要的意义。microRNAs（miRNAs）是一种非编码RNA，正逐渐成为新的、强有力的药物靶点。目前有157个基于miRNA的治疗方法的临床前和临床试验。我们已经鉴定了一种新的miRNA，miR-335- 5 p，其靶向Wnt信号拮抗剂DKK 1的3 '-UTR序列，并下调DKK 1的表达。我们还发现miR-335- 5 p是成骨分化的重要调节因子。我们进一步产生了转基因小鼠系，其中miR-335- 5 p基因在骨组织中过表达，显示标记基因的表达增强和骨量增加。基于这些有趣的发现，我们计划使用miR-335- 5 p作为治疗靶点，使用我们新合成和表征的类胡萝卜素-miRNA纳米颗粒作为新型递送载体来增强口腔和颅面骨再生。 目标1.我们将使用转基因动物模型确定miR-335- 5 p过表达在骨再生和创伤修复中的调节和成骨作用。我们的假设是，由于miR-335- 5 p转基因被整合到小鼠基因组中，其超物理转录水平将提高靶细胞中的骨形成能力。 目标2.我们将开发两种治疗策略，使用纳米颗粒类胡萝卜素系统在体内递送miR-335- 5 p分子用于口腔和颅面骨再生。 首先，我们将miR-335- 5 p修饰的骨髓基质细胞移植到骨创伤部位以促进骨形成。我们的假设是，miR-335- 5 p特异性抑制DKK 1并激活Wnt/β-Catenin信号，从而进一步触发成骨级联反应，导致BMSC向成骨细胞谱系分化并增加骨形成能力。 其次，我们将确定miR-335- 5 p在原位促进骨形成细胞和增强伤口愈合和组织再生中的作用。我们的假设是，在伤口部位局部施用miR-335- 5 p直接导致间充质干细胞分化为骨形成细胞，加速伤口愈合和骨再生。 这项转化研究将探索miR-335- 5 p作为促进骨再生和伤口愈合的治疗剂的潜力。这种创新的基于miRNA的疗法具有临床治疗其他骨疾病的潜力，包括骨质疏松症、骨折、牙周炎和骨/关节植入物的骨整合受损。

 DESCRIPTION (provided by applicant): While over 500,000 surgeries correcting bone deformities and critical size defects occur each year in the US, 50% of the procedures end in failure. Therefore, using molecular engineering approaches to regenerate lost bone with normal anatomic contour, physical properties and biological function is of enormous importance. MicroRNAs (miRNAs) are non-coding RNAs that are emerging as new and powerful drug targets. Currently there are 157 preclinical and clinical trials of miRNA-based therapeutics. We have identified a novel miRNA, miR-335-5p, which targets the 3'-UTR sequence of DKK1, a Wnt signal antagonist, and down-regulates DKK1 expression. We also revealed that miR-335-5p is an important regulator for osteogenic differentiation. We have further generated a transgenic mouse line in which miR-335-5p gene is overexpressed in bone tissue showing enhanced expression of marker genes and increased bone mass accrual. Based on these interesting findings we plan to use miR-335-5p as a therapeutic target to enhance oral and craniofacial bone regeneration using our newly synthesized and characterized lipidoid-miRNA nanoparticles as a novel delivery vehicle. Aim 1. We will determine regulatory and osteogenic effects of miR-335-5p overexpression in bone regeneration and wound repairing using a transgenic animal model. Our hypothesis is that since miR-335-5p transgene is incorporated into mouse genome its super-physical transcript levels will boost the bone forming capacity in target cells. Aim 2. We will develop two therapeutic strategies with a nanoparticle lipidoid system to deliver miR-335-5p molecules in vivo for oral and craniofacial bone regeneration. Firstly, we will transplant miR-335-5p-modified bone marrow stromal cells into bone wound site to enhance bone formation. Our hypothesis is that miR-335-5p specifically suppresses DKK1 and activates Wnt/β−Catenin signals, which further triggers an osteogenic cascade leading BMSC differentiation towards osteoblastic lineage and an increased bone formation capability. Secondly, we will determine the effect of miR-335-5p in promoting bone-forming cells in situ and enhancing wound healing and tissue regeneration. Our hypothesis is that local administration of miR-335-5p in wound site directly leads mesenchymal stem cells differentiation into bone forming cells, accelerating wound healing and bone regeneration. This translational research will explore the potential of miR-335-5p as a therapeutic agent for promoting bone regeneration and wound healing. This innovative miRNA-based therapy has potential for the clinical treatment of other bone disorders including osteoporosis, fracture, periodontitis and impaired osseointegration of bone/joint implants.