Improving Brain Recovery Through Glycoengineering

通过糖工程改善大脑恢复

• 批准号: 10666616
• 负责人: Xiaofeng Jia
• 金额: $ 48.23万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10666616
• 关键词: Adhesions; Affinity; Animal Model; Animals; Apoptosis; Behavior assessment; Behavioral; Biochemical; Biological; Biological Process; Biology; Brain; Brain Injuries; Cadherins; Carbohydrates; Cell Adhesion; Cell Communication; Cell surface; Cells; Cellular biology; Central Nervous System Diseases; Chemical Structure; Chemicals; Chemistry; Complex; Development; Dose; Electrophysiology (science); Engineering; Event; Extracellular Matrix; Foundations; Generations; Glycoconjugates; Glycoengineering; Gold; Healthcare; Heart Arrest; Human; Implant; In Vitro; Intercept; Ischemia; Kinetics; Lead; Malignant Neoplasms; Mass Spectrum Analysis; Mediator; Metabolic; Metabolism; Methodology; Modeling; Monosaccharides; N-acetylmannosamine; Nature; Nerve Regeneration; Neurites; Neuronal Differentiation; Neurons; Neurophysiology - biologic function; Organism; Pathway interactions; Pharmaceutical Preparations; Physiological; Polysaccharides; Preclinical Testing; Proteins; Rattus; Recovery; Recovery of Function; Reporting; Rodent Model; Safety; Sialic Acids; Signal Transduction; Spatial Distribution; Stimulus; Structure-Activity Relationship; Sulfhydryl Compounds; Surface; Synaptic plasticity; Techniques; Testing; Therapeutic; Time; Tissues; Translations; Transplantation; WNT Signaling Pathway; Wnt proteins; Work; analog; blastomere structure; cell motility; clinical application; clinical translation; conventional therapy; cost effectiveness; covalent bond; design; drug candidate; embryo cell; extracellular; functional group; functional improvement; glycoproteomics; healing; immune cell infiltrate; improved; in vivo; injured; injury recovery; innovation; intercellular communication; migration; nerve stem cell; neural; neuroregulation; novel; receptor; response; scaffold; stem cell biology; stem cell differentiation; stem cell fate; stem cell therapy; technology platform; therapeutic candidate; translational approach

Project Summary This project is based on recent advances in metabolic glycoengineering (MGE), a technology platform where non-natural monosaccharides intercept the biosynthetic pathways for cell surface-displayed glycans. As a result, chemical functionalities not naturally found in carbohydrates are installed in the glycalyx, which can alter cell adhesion, receptor activity, and downstream events (e.g., apoptosis, differentiation, and motility). In previous work, we developed the N-acetylmannosamine (ManNAc) analog “Ac5ManNTGc” to install thiol groups into sialic acids in human embryonic cells and found that – when the cells were grown on a “high affinity” surface (e.g., gold, which forms coordinate covalent bonds with thiols) – Wnt signaling was upregulated in the absence of extracellular Wnt proteins and neuronal differentiation was induced. In vivo translation of this approach, however, was hindered by the requirement for a non-degradable gold scaffold. We recently overcame this impediment by designing new ManNAc analogs with thiols presented on longer linkers, which extends this functional group further away from the core monosccharide and increases analog potency. Critically, the new analogs provide pro-neurogenic activity in the absence of a scaffold thereby simplifying in vivo translation. This project will explore analog mechanism in hNSCs in Specific Aim 1; this aim will define how the chemical structure, kinetics, and dose of thiol-modified ManNAc analogs (along with chemically inert size-matched controls) modulate cellular glycans in Aim 1a; evaluate changes to cell adhesion and motility in Aim 1b, and evaluate the differentiation of human neural stem cells (hNSCs) in Aim 1c. Next, in Specific Aim 2, we will apply the optimized analog-treatment conditions to improve neural regeneration in a rat cardiac arrest (CA) model of brain injury by transplanting MGE-modified into injured animals. We will compare hNSCs treated with our new thiol-modified analogs with appropriate controls on functional recovery after CA by evaluating survival, adhesion, distribution, and migration of transplanted hNSCs in rat brain. In Specific Aim 3, we will evaluate biochemical (Wnt signaling and cadherin involvement) and cellular (tissue infiltrating immune cells) level mechanisms we propose contribute to the healing effects of MGE in brain injury recovery (in Aim 3a). Finally, in Aim 3b we will characterize cell-wide “glycosites” by mass spectrometry and use glycobioinformatics analyses to identify unknown biochemical mediators of MGE. Specifically, we anticipate identifying new mediators of the beneficial effects of MGE in the implanted hNSCs as well as trans-acting host proteins. We hypothesize that thio-analogs modulate hNSC fate through a complex combination of receptor- specific effects on cell signaling and adhesion providing a pleiotropic suite of healing effects that cannot be achieved through conventional therapies. Accordingly, our innovative approach opens a new avenue to improve stem cell therapy with our new thiol-based MGE technique.

项目摘要 该项目基于代谢糖工程（MGE）的最新进展，该技术平台是一个技术平台 非天然的单糖拦截了细胞表面播种聚糖的生物合成途径。作为 结果，在碳水合物中没有自然发现的化学功能安装在Glycayx中，这可能会改变 细胞粘附，受体活性和下游事件（例如凋亡，分化和运动性）。 以前的工作，我们开发了N-乙酰基甘露胺（MANNAC）类似物“ AC5MANNTGC”来安装硫醇 将人类胚胎细胞中的唾液酸分组成盐酸，发现 - 当细胞在“高亲和力”上生长时 表面（例如黄金，形成坐标与硫醇的共价键） - Wnt信号在 诱发了细胞外Wnt蛋白和神经元分化的缺乏。体内翻译 然而，通过不可降解的金支架的要求阻碍了方法。我们最近 通过在更长的接头上呈现的硫醇设计新的Mannac类似物，克服了这种障碍 将这个功能组扩展到远离核心单糖的范围内，并增加了模拟效力。 至关重要的是，新的类似物在没有脚手架的情况下提供了促神经的活性，从而简化了 体内翻译。该项目将在特定目标1中探索HNSC中的模拟机制；这个目标将定义 化学结构，动力学和硫醇改性的甘露纳克类似物如何以及化学惰性 尺寸匹配的控件）在AIM 1A中调节细胞聚糖；评估细胞粘合剂和运动性的变化 AIM 1B，并评估AIM 1C中人神经干细胞（HNSC）的分化。接下来，以特定的目的 2，我们将应用优化的模拟处理条件来改善大鼠心脏的神经再生 通过将MGE修饰移植到受伤的动物中，抑制了脑损伤模型。我们将比较HNSCS 用我们的新硫醇修饰的类似物处理，并通过适当的对照在CA后进行功能恢复的适当控制 评估大鼠大脑中移植的HNSC的生存，粘附，分布和迁移。在特定的目标3中 我们将评估生化（Wnt信号传导和钙粘蛋白受累）和细胞（组织浸润免疫 我们提出的水平机制有助于MGE在脑损伤恢复中的愈合作用（在AIM中 3a）。最后，在AIM 3B中，我们将通过质谱法表征整个细胞的“糖材”并使用 糖菌信息学分析以鉴定MGE的未知生化介质。具体来说，我们期望 确定MGE在植入的HNSC中的有益作用的新调解人以及跨性宿主 蛋白质。我们假设硫代动物通过接收器的复杂组合调节HNSC命运 - 对细胞信号传导和粘合剂的具体影响，提供了一套多效性的愈合作用套件，无法是 通过常规疗法实现。根据每种创新方法， 通过我们新的基于硫醇的MGE技术改善干细胞疗法。