Collaborative Research: Advanced Biomanufacturing of Functional Bionanoparticles for Biomedical Engineering Applications

在过去的十年里，新型纳米材料的研究经历了爆炸式的增长。然而，从实验室规模向大规模生产过渡的主要挑战，如高通量制造工艺、统一性和监测大量产品质量的方法，一直是实现其巨大潜力的瓶颈。本提案的目标是设计一种先进的制造工艺来制造基因工程多功能生物纳米颗粒（bio-NPs），并检查和验证其在脑肿瘤非侵入性成像中的应用。如果成功，这将提供一个从nsf风格的基础科学到现实世界应用的极好示范。该项目将先进的生物制造和生物成像模块整合到研究生、本科生和高中生的教育和实验室培训中，将对他们的教育产生积极的影响。将为学生开设一门新的生物制造研究型课程。本项目旨在通过基因工程纳米囊泡形成大肠杆菌的发酵，合成以工程蛋白修饰的新型纳米多功能外膜囊泡，并将修饰后的外膜囊泡应用于脑肿瘤的无创生物成像。为了实现这一目标，重组DNA技术将首先用于设计新的基因工程蛋白多功能生物nps，用于生物成像的捕获和检测功能。生物nps是基于脂质的omv，具有均匀的大小，双分子层的外层被新型工程蛋白融合装饰，赋予多功能。omv，共同显示多个超活性NanoLuc荧光素酶拷贝（比传统萤火虫或Renilla荧光素酶活性高150倍），将包含(i)抗体结合结构域用于锚定感兴趣的抗体，以及（ii）热响应弹性蛋白样蛋白结构域，用于通过尺寸过滤简单纯化omv。然后，将设计一种集成两级尺寸过滤的发酵工艺，用于生产多功能omv。最后，该项目将验证这些omv在脑肿瘤高性能生物成像方面的功能。该研究将为生物制造提供新的视角，该产品将极大地促进全球公共卫生。这种新型的可扩展的基因工程制造平台可以推广到具有许多其他所需功能的omv，适用于广泛的应用，包括生物修复、生物催化剂、生物传感、生物质转化、疫苗和药物输送。

1604826/1604925:Lei/ChenResearch on new nanomaterials has undergone explosive growth in the past decade. However, the main challenges of the transition from laboratory-scale to mass production, such as high throughput manufacturing processes, uniformity, and methodology of monitoring the quality of large-quantity products have been the bottlenecks to realize their tremendous potential. The goal of this proposal is to design an advanced manufacturing process to manufacture genetically engineered multi-functional bio-nanoparticles (bio-NPs) and to examine and validate their utility for non-invasive imaging of brain tumor cancer. If successful, this will provide an excellent demonstration from NSF-style basic science to real-world applications. This project will positively impact education of graduate, undergraduate and high school students by integrating advanced biomanufacturing and bioimaging modules into their educational and laboratory training. A new research-oriented course in Biomanufacturing will be offered to students. This multidisciplinary project aims to synthesize novel nano-sized multi-functional outer membrane vesicles (OMVs)decorated with engineered proteins through fermentation of genetically engineered nano-vesicle-forming E. coli and then apply the decorated OMVs for non-invasive bioimaging of brain tumor. To accomplish this, recombinant DNA technology will first be used to design novel genetically engineered protein multi-functional bio-NPs for capture and detection functions for bioimaging. The bio-NPs are lipid-based OMVs with a uniform size and the outer leaflet of the bilayer is decorated with novel engineered protein fusion, endowing multi-functionality. The OMVs, co-displaying multiple copies of super-active NanoLuc luciferase enzyme (~150-fold more active than that of conventional firefly or Renilla luciferase), will contain (i) an antibody-binding domain for anchoring antibodies of interest, and (ii) a thermo-responsive elastin-like protein domain for simple purification of the OMVs via size filtration. A fermentation process integrated with two-stage size filtration will then be designed for production of multi-functional OMVs. Finally, the project will validate the functionality of these OMVs for high performance bioimaging of brain tumor. The proposed research will offer a new perspective to biomanufacturing while the product can greatly promote global public health. This novel scalable genetically-engineered manufacturing platform can be generalized to prepare the OMVs with many other desired functions suitable for a wide range of applications including bioremediation, biocatalysts, biosensing, biomass conversion, vaccines, and drug delivery.