Microcarrier culture system for human influenza vaccine

描述（由申请人提供）：拟议研究的目标是开发一种优化的微载体-生物反应器细胞培养系统，以实现快速、高效、大规模、低成本的人流感疫苗生产。大多数人类流感病毒疫苗是在胚胎蛋中生产的。联邦政府和其他国际组织正在寻求生产流感疫苗的替代细胞培养方法，因为蛋源疫苗存在固有的困难。在正常情况下，从(a)确定每个流感季节可能出现的特定病毒株到(b)获得针对这些病毒株的适当疫苗之间需要很长时间。微载体-生物反应器细胞培养技术有可能降低生产成本，并将生产9300万剂单价疫苗所需的时间缩短到几个月。在流感大流行的情况下，这种迅速实现大规模的潜力将是无价的，并可能应对大流行或生物恐怖主义行为。为了实现研究目标，将优化影响Vero和MDCK细胞病毒产量的微载体生物反应器培养工艺参数，并制定适用于经济商业应用的方案。我们将(i)向流感疫苗的大型生产商许可生产协议，（ii）销售用于这些协议的Hillex微载体。

DESCRIPTION (provided by applicant): The goal of the proposed research is to develop an optimized microcarrier-bioreactor cell culture system that enables the rapid, efficient, large scale, low cost production of human influenza vaccine. Most human influenza virus vaccine is produced in embryonated eggs. The Federal government and other worldwide organizations are seeking alternative cell-culture approaches to influenza vaccine production since there are inherent difficulties with the egg-derived vaccines. Under normal conditions there are long lead times between (a) the identification of particular virus strains likely in each flu season and (b) the availability of appropriate vaccines against these virus strains. Microcarrier-bioreactor cell-culture technology has the potential to lower production costs and reduce the time required to produce 93 million doses of monovalent vaccine to a few months. This potential to rapidly achieve large scale, would be invaluable in case of an influenza pandemic and possibly could counter a pandemic or bioterrorist acts. To achieve the research goals, five specific aims will optimize microcarrier-bioreactor culture process parameters that affect viral yield with Vero and MDCK cells and develop protocols applicable to economic commercial use. We will (i) license manufacturing protocols to large producers of flu vaccine and (ii) sell Hillex microcarriers for those protocols. Specific Aim I. To identify and integrate critical cell culture and viral growth parameters for optimal production of human influenza virus in Vero cells grown in Hillex microcarrier-suspension culture. Specific Aim II. To extend the range of viral growth parameters (as identified in Specific Aim I) to include six additional strains of influenza virus and to compare the virus production in direct "head-to-head" competition between embryonated eggs and Vero cells grown in Hillex microcarrier-suspension culture. Specific Aim III. To assess production of human influenza virus in Vero cells grown in a microcarrier-bioreactor culture at the one liter scale. Specific Aim IV: To develop a process to expand and scale-up Vero cells from a working seed ampoule to a five liter culture and to demonstrate that the passaged cells will support high influenza virus concentrations at a larger scale. Specific Aim V: To identify and integrate critical cell culture parameters for optimal production of human influenza virus (A/Wuhan/395/95) in MDCK cells grown in Hillex microcarrier-bioreactor culture.