EAGER: Peptide affinity membranes for binding influenza viruses: H1N1 and H5H1

0943424 Karim智力优点：EAGER方案是一个高风险高收益方案。最近H1N1流感的爆发和H5 N1禽流感病毒的全球传播及其对人类的感染引起了全球对这些病毒引起大流行的关注。根据世界卫生组织2009年5月20日的报告，41个国家已确诊10，243例H1N1流感病例，其中80人死亡。在美国，这两个数字分别为5，469和6。目前迫切需要一种针对H1N1和H5 N1的治疗方法或快速接种疫苗的方法。该提案解决了这一问题，提出了一个变革性的想法，即开发一种基于肽的膜系统，以便从细胞培养液或人类血液中捕获流感病毒。主要目标是开发一种基于细胞培养的疫苗生产方法，该方法使用基于肽的纯化步骤。该肽是利用关于流感病毒血凝素（HA）表面蛋白的可用基因组信息开发的。我们假设，对于给定的病毒，可以通过使用筛选工具选择具有高特异性的亲和肽。此外，通过亲和肽基团与膜表面的化学连接，这种亲和基质可用于重复结合和固定特定类型的病毒。我们进一步假设病毒表面蛋白在病毒结合过程中起着重要作用。因此，基于流感病毒的特定毒株设计的亲和膜可用于结合与该病毒具有高度同源性的其它病毒。本研究将有助于更好地了解病毒表面蛋白在亲和结合过程中的作用。“同质性”一词通常用于描述两种流感病毒株的相似程度。通过选择具有不同同质性的几种流感病毒，可以表征表面蛋白对病毒与亲和膜结合的影响。本研究将回答关于均一性对病毒与给定亲和膜结合的影响有多敏感的问题。由于微孔亲和膜结合了微滤和基于树脂的亲和层析的优点，可以显著简化病毒的生产和纯化过程。这特别适用于基于细胞培养的流感病毒疫苗生产工艺。通过仔细选择合适的膜孔径，可以在一个步骤中实现细胞去除和病毒纯化。细胞培养物将流过微孔亲和膜的一侧。在此期间，细胞和细胞碎片将被膜孔排斥，而病毒和蛋白质和/或DNA和其他小的生物组分将在膜孔内迁移。然后，病毒将被接枝在膜孔表面上的这些功能性肽基团特异性地结合，而蛋白质、DNA和其他生物组分将通过膜孔。洗涤后，可以从结合的膜上洗脱出高纯度的病毒。更广泛的影响：完成拟议的研究将对H1N1和H5 H1流感干预产生重大影响和可能的应用。提出的新型肽基膜将简化病毒纯化过程，并可能加快疫苗开发、商业化和生产过程。所制备的膜也可用于快速收集和纯化病毒样品的病毒诊断。此外，这种方法可以扩展到诊断其他病毒，如西尼罗河病毒。此外，可以开发一种可能的临床装置，其可以特异性地从人血液中去除病毒。通过使人类血液循环通过含有针对特定流感病毒的亲和膜的膜装置，可以使这些病毒结合到膜表面，从而减少血液中的总体病毒群体;然后可以将该装置与一些其他干预方法结合，例如使用TAMIFLU。该项目将涉及来自化学工程和TTU健康科学中心的多学科研究人员。该项目的教育目标之一是培养一名博士后和一名博士生，从事疫苗和肽基膜技术的前沿研究。一名本科生将从TTU广为宣传的推广计划中选出，例如工程推广中心和麦克奈尔学者计划。然后，UG学生将被鼓励去研究生院，并在生物相关领域的职业生涯。PI在吸引女性和少数民族候选人加入其研究小组方面拥有丰富的经验。对当地学校的推广将通过资助的方案进行，如科学是女孩的事情，大学预科工程，教师培训讲习班和工程推广导师。

0943424KarimIntellectual Merit: This EAGER proposal is a high risk high gain proposal. Recent outbreak of H1N1 flu and the worldwide spread of H5N1 avian influenza virus and its infection to human beings has raised a global concern about these viruses to cause a pandemic. According to the World Health Organization report of May 20, 2009, 41 countries have confirmed 10,243 cases of H1N1 flu with 80 fatalities. In the US, these numbers are 5,469 and 6, respectively. A treatment or a rapid vaccination process for H1N1 and H5N1 is urgently needed. The proposal addresses this issue in this transformative idea to develop a peptide-based membrane system so that flu viruses can be captured from either a cell culture broth or from human blood. The main goal is to develop a cell culture-based vaccine production method, which uses a peptide-based purification step. The peptide is developed using the available genomic information regarding hemagglutinin (HA) surface protein of flu viruses. We hypothesize that for a given virus, an affinity peptide with a high specificity could be selected by using a screening tool. Moreover, with the chemical attachment of the affinity peptide group to a membrane surface, this affinity matrix could be used to repeatedly bind and elute the specific type of virus. We further hypothesize that the virus surface protein plays a significant role on the virus binding process. Therefore, an affinity membrane designed based on a specific strain of influenza virus may be used to bind other viruses which have high homogeneity to this virus. This research will be helpful in better understanding the role of the virus surface protein on the affinity binding process. A term of 'homogeneity' is usually used to describe how similar two influenza virus strains are. By choosing several influenza viruses with different homogeneity, the effect of the surface protein on the virus binding to an affinity membrane could be characterized. This research will answer the question regarding how sensitive the effect of the homogeneity is on the binding of a virus to a given affinity membrane.Since microporous affinity membrane combines both the advantages of the microfiltration and resin based affinity chromatography, the virus production and purification process could be significantly simplified. This is specifically suitable for the cell culture based influenza virus vaccine production process. By carefully choosing an appropriate membrane pore size, cells removal and virus purification could be achieved within one step. Cell cultures will be flown through one side of a microporous affinity membrane. During this period, cells and cell debris will be rejected by the membrane pores while viruses and proteins and/or DNA and other small biological components will migrate inside the membrane pores. Viruses will then be specifically bound by these functional peptide groups grafted on the membrane pore surface while protein, DNA and other biological components will pass through the membrane pores. After washing, viruses with high purity can be eluted from the bounded membranes. This simplification of the purification process may speed up the H1N1 and H5N1 vaccine development, commercialization and production process.Broader Impact: Completion of the proposed studies will have significant impact and possible applications in H1N1 and H5H1 flu intervention. The proposed new peptide-based membrane will simplify the virus purification process and it may speed up the vaccine development, commercialization and production process. The prepared membrane may also be used in the virus diagnosis by quick collection and purification of virus samples. Further, this method may be extended to diagnose other viruses, such as the West Nile virus. Further, a possible clinical device may be developed which can specifically remove viruses from human blood. By circulating human blood through a membrane device containing affinity membranes for a specific flu virus, these viruses can be made to bind to the membrane surfaces thus reducing the overall virus population in the blood; this device may be then combined with some other intervention methods such as using TAMIFLU. This project will involve multidisciplinary researchers from Chemical Engineering and TTU Health Sciences Center. One of the educational goals of the project is to train one post-doc and one PhD student in the cutting edge research of vaccine and peptide-based membrane technology. An undergraduate student will be selected from well publicized outreach programs at TTU, e.g. Engineering Outreach Center, and McNair Scholars Program. The UG student will then be encouraged to go to graduate school and have a career in bio related field. The PI has significant experience in attracting female and minority candidates to his research group. Outreach to local schools will be done through funded programs such as Science it's a Girl's Thing, Pre-college Engineering, Teacher Training Workshop, and Engineering Outreach Mentor.