Nanoparticle-based synthetic transcription factor to induce stem cell myogenesis

项目摘要 干细胞生物学的最新进展在开发治疗肿瘤的新方法方面具有巨大潜力。 许多毁灭性的疾病，包括肌肉骨骼疾病（MSD）。干细胞治疗 再生功能性肌细胞和恢复受损骨骼肌的肌肉功能， 对于肌肉骨骼疾病和病症的治疗进展的发展至关重要。这种方法， 然而，这需要产生功能性肌原细胞的可移植细胞来源和更好地控制 干细胞肌原性分化的有效，选择性和安全的方式。 为此，这一提案的主要目标是开发一种可以复制结构的生物启发平台 并在称为转染因子（TF; MyoD和Myogenin）的内源性蛋白质上发挥作用，这些蛋白质是特异性的 肌肉特异性基因，负责协调整个干细胞分化为肌肉细胞。我们 一个名为NanoScript的生物启发平台，是一种基于纳米颗粒的转录因子， 就像天然TF蛋白一样。这个NanoScript平台被设计为基因特异性的，可以有效地激活 靶向基因表达（例如MyoD、肌细胞生成素和相关内源性基因）， 病毒的方式。我们的中心假设，基于最近的成就和初步数据，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产生功能性肌肉细胞- 来源的间充质干细胞（AMDSC），其是干细胞的丰富来源，具有患者特异性， 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：Aim 1：设计和合成肌细胞特异性转录因子（MRF）和表观遗传调节剂， 构建增强的肌细胞特异性NanoScripts-MRF [NanoScripts-MRF]。目标2：利用NanoScript 激活ADMSC中的肌肉特异性基因以产生肌肉细胞。 所提出的研究是创新的，因为这种通过集成两个 多学科的方法（化学生物学和纳米医学）到一个单一的纳米平台的非病毒基因 在干细胞中的调节尚未被开发。这项研究意义重大，因为我们将开发一种 创新的技术平台，我们的NanoScript是一个易于调整和强大的平台，它可以进一步 开发出与表观遗传调节剂或其他增效剂联合收割机，用于有效和选择性诱导 功能性肌肉细胞总的来说，在拟议的研究成功完成后，我们的期望是 NanoScript-MRF将激活包含其同源TF的肌肉特异性基因的转录 共有DNA结合域，这将导致增强的干细胞分化为肌肉细胞。因为 NanoScript是无毒和非病毒的，产生的肌肉细胞将被考虑用于体内翻译 未来的动物研究。

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.