Bio-nanomanufacturing of Protein Therapeutics Using Membrane Microfluidics

使用膜微流体的蛋白质治疗药物的生物纳米制造

• 批准号: 1728049
• 负责人: Susan Daniel
• 金额: $ 37.44万
• 依托单位: Cornell University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1728049&HistoricalAwards=false
• 关键词: Bio; nanomanufacturing; Protein; Therapeutics; Using

The market for therapeutic proteins is valued near $140 billion annually. Many of these therapeutics are glycoproteins, which require the addition of specific sugars, called 'glycans' at an exact position on the protein through a process called protein glycosylation. The glycan affects protein folding and function and ensures it retains its therapeutic potency. In nature, glycoproteins are produced through a series of sequential reactions inside a cell. Making therapeutic glycoproteins within cells is challenging for a variety of reasons, and extensive and costly purification steps are required to harvest the therapeutic material. With this award, a cell-free glycosylation network will be constructed in a microfluidic device that separates reactions in space and time, giving supreme flexibility in optimizing individual reactions and constructing glycans with high specificity. The benefits of this manufacturing paradigm to society are reducing the cost of these drugs and providing scientists an avenue to design and develop synthetic drug compounds that may or may not exist in nature to treat disease. The related education plan creates a hands-on bio-nanomanufacturing activity for a high school girls organized by the PIs and their student trainees, so that young women will understand the power of biotechnology and be inspired to pursue these career paths. Cell-free protein synthesis holds great promise for producing high-value, biotherapeutic nanomaterials without cell culture and benefitting from chemical manufacturing know-how. Here, raw materials and biological enzymes are mixed to produce biological products. Shortcomings of this approach are competing reactions, side products, and low yields. Cells avoid these shortcomings by localizing reactions within subcellular compartments and orchestrating the reaction sequences. The biocatalysts that give the final molecule its essential posttranslational features are compartmentalized in membranes. Handling enzymes outside of their native lipid environment can drastically reduce their activity. Thus, in vitro, sequential, bio-enzymatic reactions have never been achieved in a cell-free manner. The research objective is to mimic the elegant compartmentalization strategies used by cells in a microfluidic biomembrane device that organizes biological reactions in proper spatial and temporal sequence. These devices will generate authentically glycosylated proteins. Through assessment of nanostructure product architectures, this work will advance understanding of nanoscale phenomena and processes for nanomaterials manufacture and discovery. This cell-free device concept will enable facile optimization of glycosylated protein production, and provide a framework for understanding how experimental conditions affect product yield and quality that is broadly applicable to the bio-nanomanufacturing of virtually any posttranslationally-modified protein.

治疗蛋白的市场每年价值接近1400亿美元。这些治疗剂中的许多是糖蛋白，它需要添加特定的糖，即通过称为蛋白质糖基化的过程在蛋白质上的精确位置称为“聚糖”。聚糖会影响蛋白质折叠和功能，并确保其保留其治疗效力。在自然界中，糖蛋白是通过细胞内的一系列顺序反应产生的。由于各种原因，在细胞内制造治疗性糖蛋白是具有挑战性的，并且需要广泛且昂贵的纯化步骤来收集治疗材料。通过此奖项，将在微流体设备中构建一个无细胞的糖基化网络，该机构将空间和时间的反应分开，从而在优化单个反应和构造具有高特异性的聚糖方面具有至高的灵活性。这种制造范式对社会的好处是降低了这些药物的成本，并为科学家提供了设计和开发合成药物化合物的途径，这些途径在自然界中可能存在或可能不存在以治疗疾病。相关的教育计划为PIS及其学生学员组织的高中女生创造了动手的生物制造活动，因此年轻女性将了解生物技术的力量，并受到启发来追求这些职业道路。无细胞的蛋白质合成对生产没有细胞培养的高价值，生物治疗纳米材料并从化学制造知识中受益。在这里，将原材料和生物酶混合在一起以生产生物产品。这种方法的缺点是竞争反应，副产品和低收率。细胞通过将反应定位在亚细胞隔室中并策划反应序列来避免这些缺点。赋予最终分子的生物催化剂其基本的翻译后特征在膜中被划分。处理本机脂质环境外的酶可以大大减少其活性。因此，在体外，顺序，生物酶反应从未以无细胞的方式实现。研究的目标是模仿细胞在微流体生物膜装置中使用的优雅分隔策略，该策略在适当的空间和时间序列中组织生物反应。这些设备将生成真实的糖基化蛋白。通过评估纳米结构架构，这项工作将提高对纳米级现象的理解和纳米材料制造和发现的过程。这种无细胞的装置概念将使糖基化蛋白质的产生可轻松优化，并提供一个框架，以了解实验条件如何影响产品产量和质量，这些产品几乎适用于几乎任何翻译后修饰的蛋白质的生物 - 纳米制造。

项目成果

Synthetic Glycobiology: Parts, Systems, and Applications

• DOI: 10.1021/acssynbio.0c00210
• 发表时间: 2020-07-17
• 期刊: ACS SYNTHETIC BIOLOGY
• 影响因子: 4.7
• 作者: Kightlinger, Weston Y.;Warfel, Katherine F.;Jewett, Michael C.
• 通讯作者: Jewett, Michael C.

Cell-Free Synthetic Glycobiology: Designing and Engineering Glycomolecules Outside of Living Cells

• DOI: 10.3389/fchem.2020.00645
• 发表时间: 2020-07-29
• 期刊: FRONTIERS IN CHEMISTRY
• 影响因子: 5.5
• 作者: Jaroentomeechai，Thapakorn;Taw，May N.;DeLisa，Matthew P.
• 通讯作者: DeLisa，Matthew P.