Nanoparticle-based synthetic transcription factor to induce stem cell myogenesis

项目总结 干细胞生物学的最新进展在开发新的治疗方法方面具有巨大的潜力。 许多毁灭性的疾病，包括肌肉骨骼疾病(MSD)。基于干细胞的治疗方法 再生功能性肌肉细胞和恢复受损骨骼肌的肌肉功能可以是 对发展肌肉骨骼疾病和紊乱的治疗进展至关重要。这样的方法， 然而，需要生成具有功能的肌源性细胞的可移植细胞来源并更好地控制 以有效、选择性和安全的方式进行干细胞肌源性分化。 为此，这项提议的主要目标是开发一个受生物启发的平台，可以复制这种结构 并作用于内源性蛋白质，称为转导因子(TF；MyoD和Mygenin)，它们是特异的 肌肉特异性基因，并负责协调整个干细胞分化为肌肉细胞。我们的 生物启发平台名为NanoScript，是一种基于纳米颗粒的转录因子，其行为和功能 就像天然的转铁蛋白一样。该NanoScrip平台被设计为特定于基因，并可以有效地激活 靶基因(如MyoD、Mygenin和相关内源基因)在无毒和无毒的 病毒式的方式。根据最近的成果和初步数据，我们的核心假设是，我们提出的 NanoScript平台可以有效地从人类患者来源的脂肪中生成功能肌肉细胞- 来源的间充质干细胞(AMDSCs)是干细胞的丰富来源，具有患者特有的 干细胞治疗的可能性。 我们建议通过解决以下具体问题来检验我们的中心假设并实现我们的目标 目的：AIM1：设计和合成肌细胞特异性转录因子(MRF)和表观遗传调节剂 增强型肌肉细胞特异性纳米脚本的构建[纳米脚本-MRF]AIM2：利用NanoScript来 激活ADMSCs中的肌肉特异性基因以生成肌肉细胞。 提出的研究是创新的，因为这种通过集成两个组件来开发TF仿真器的概念 多学科方法(化学生物学和纳米医学)建立一个单一的非病毒基因纳米平台 对干细胞的调控还没有发展起来。这项拟议的研究具有重要意义，因为我们将开发一种 创新的技术平台和我们的NanoScrip是一个易于调整和强大的平台，它可以进一步 开发成与表观遗传调节剂或其他增效剂结合，以有效和选择性地诱导 功能性肌肉细胞。总的来说，在成功完成建议的研究后，我们的期望是 NanoScrip-MRF将激活包含其同源TF的肌肉特异性基因的转录 一致的DNA结合域，这将导致增强干细胞分化为肌肉细胞。因为 纳米脚本是无毒和无病毒的，产生的肌肉细胞将被考虑转化为体内 未来研究中的动物研究。

PROJECT SUMMARY Recent advances in stem cell biology hold great potential in developing new approaches for the treatment of many devastating diseases, including musculoskeletal disorders (MSDs). Stem cell-based therapies for regenerating functional muscle cells and restoring muscular functions to damaged skeletal muscles can be critical for the development of therapeutic advances in musculoskeletal disease and disorder. Such approaches, however, require the generation of engraftable cell sources of functional myogenic cells and better control of stem cell myogenic differentiation in an effective, selective, and safe manner. To this end, the main goal of this proposal is to develop a bio-inspired platform that can replicate the structure and function on endogenous proteins called transfection factors (TFs; MyoD and Myogenin), which are specific for muscle-specific genes and responsible for orchestrating overall stem differentiation into muscle cells. Our bio-inspired platform called NanoScript, is a nanoparticle-based transcription factor that behaves and function just like natural TF proteins. This NanoScript platform is designed to be gene-specific and can effectively activate targeted gene expressions (e.g. MyoD, Myogenin and the related endogenous genes) in a non-toxic and non- viral manner. Out central hypothesis, based upon recent achievement and preliminary data, is that our proposed NanoScript platform can effectively generate functional muscle cells from human patient-derived adipose- derived mesenchymal stem cells (AMDSCs), which are an abundant source of stem cells, with patient-specific stem cells treatment possibilities. We propose to test our central hypothesis and achieve our objectives by addressing the following specific aims: Aim1: Design and synthesize muscle cell-specific TFs (MRF) and epigenetic modulators for the construction of enhanced muscle cell-specific NanoScripts [NanoScripts-MRF]. Aim2: Utilize NanoScript to activate muscle-specific genes in ADMSCs for generating muscle cells. The proposed research is innovative, as this concept of developing a TF emulator by integrating two multidisciplinary approaches (chemical biology and nanomedicine) onto a single nano-platform for non-viral gene regulation in stem cells has not been developed. The proposed research is significant, since we will develop an innovative technology platform and our NanoScript is an easily tunable and robust platform, it can be further developed to combine with epigenetic modulators or other synergists for effective and selective induction of functional muscle cells. Collectively, upon successful completion of the proposed study, our expectations are that NanoScript-MRF will activate transcription of the muscular-specific genes containing their cognate TF consensus DNA binding domain, which will lead to an enhanced stem differentiation into muscle cells. Because NanoScript is non-toxic and non-viral, the generated muscle cells will be considered for translation into in vivo animal studies in future studies.