Bottom-up, high-throughput prototyping of extracellular vesicle mimetics using cell-free synthetic biology

使用无细胞合成生物学对细胞外囊泡模拟物进行自下而上的高通量原型设计

基本信息

批准号：
10638114
负责人：
Randy Carney
金额：
$ 57.06万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-01 至 2027-02-27
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10638114
关键词：
Address Animals Autoimmune Responses Benchmarking Biodistribution Biological Biological Assay Biological Models Cell secretion Cells Cellular Assay Chemicals Clinic Complex Coupled Disease Dose Drug Delivery Systems Engineering Evaluation Exhibits Experimental Autoimmune Encephalomyelitis Future Half-Life Heterogeneity Image Immune In Vitro Integral Membrane Protein Lead Lipids Liposomes Mass Spectrum Analysis Measurement Membrane Membrane Proteins Mesenchymal Mesenchymal Stem Cells Methods MicroRNAs Molecular Multiple Sclerosis Neurons Nucleic Acids Organ Performance Physiologic pulse Play Printing Production Protein Biosynthesis Proteins Proteomics Quality Control RNA Recombinants Reproducibility Research Resolution Role Set protein Standardization Stromal Cells System Technology Testing Therapeutic Therapeutic Effect Toxic effect Translating Untranslated RNA Vesicle Wild Type Mouse Work analytical method angiogenesis cell type clinical application delivery vehicle design experimental study extracellular vesicles high throughput screening immunoregulation in vivo in vivo Model insight intercellular communication interest mimetics mouse model nanolitre nanoparticle nervous system disorder neuroprotection pharmacokinetics and pharmacodynamics prevent prototype public health relevance reconstitution regenerative stem superresolution microscopy synthetic biology transcriptome sequencing unilamellar vesicle uptake

项目摘要

PROJECT SUMMARY Bottom-up, high-throughput prototyping of extracellular vesicle mimetics using cell-free synthetic biology Cells secrete extracellular vesicles (EVs) that function as primary messengers of intercellular communication and are studied as promising drug-delivery vehicles and therapeutics. However, the clinical application of native EVs has been hindered by their low production yield, impurity, and inherent heterogeneity. Native EVs contain many biologically active components, such as RNAs and proteins, spread out over numerous subpopulations. This biological complexity is both the strength and the Achilles’ heel of native EVs. While various features of this complexity enable the beneficial therapeutic effects of EVs, it is not clear which plays a dominant role. However, the complex set of proteins and RNAs results in heterogeneous EVs that are challenging to study and use as a standardized treatment. Therefore, separating out and defining the critical biomolecular features from the overall heterogeneous set will allow us to perform quality control of EVs and to reproducibly produce or study EVs. A major bottleneck in finding the critical molecular parts of EVs is the lack of high-throughput methods. To overcome this difficulty, our team will create a synthetic biology-based, cell-free high-throughput discovery platform. The platform will be able to synthesize EV mimetics using a cell-free synthesis approach (Aim 1), coupled with high-throughput examination of EV mimetic potency in vitro (Aim 2). Select EV mimetics will also be investigated using an in vivo model system of neuroprotection and immune modulation (Aim 3). Throughout the study, we will use native mesenchymal stem/stromal cell EVs and neurological diseases as our model system to evaluate the platform. Our work will enable the high-throughput study of EVs for any disease and biological questions of interest. In addition, we will unveil new insights into EVs that address key debated topics in the EV field.

项目概要使用无细胞合成生物学对细胞外囊泡模拟物进行自下而上的高通量原型设计细胞分泌细胞外囊泡 (EV)，充当细胞间通讯的主要信使并被研究为有前途的药物输送载体和治疗方法。然而，天然药物的临床应用。电动汽车因其低产量、杂质和固有的异质性而受到阻碍。许多生物活性成分，例如 RNA 和蛋白质，分布在众多亚群中。这种生物复杂性既是原生电动汽车的优势，也是其致命弱点。复杂性使得 EV 具有有益的治疗效果，但尚不清楚哪个发挥主导作用。复杂的蛋白质和 RNA 组会产生异质 EV，这些 EV 的研究和使用具有挑战性因此，从整体中分离并定义关键的生物分子特征。异构集将使我们能够对电动汽车进行质量控制并可重复地生产或研究电动汽车 A。寻找EV关键分子部分的主要瓶颈是缺乏高通量方法。克服这个困难，我们的团队将创造一个基于合成生物学的、无细胞的高通量发现该平台将能够使用无细胞合成方法合成 EV 模拟物（目标 1），加上体外 EV 模拟效力的高通量检查（目标 2）。使用神经保护和免疫调节的体内模型系统（目标 3）进行研究。在这项研究中，我们将使用天然间充质干/基质细胞 EV 和神经系统疾病作为我们的模型系统我们的工作将使电动汽车针对任何疾病和生物的高通量研究成为可能。此外，我们还将推出有关电动汽车的新见解，解决电动汽车中的关键争议话题。场地。