DETECTION OF VIRUSES IN CELL LINES USED TO MANUFACTURE BIOLOGICAL PRODUCTS

CBER感兴趣的是用于产品生产的细胞系是否 含有病毒或病毒样颗粒或病毒核酸序列，或 支持对人类具有传染性的病毒的生长。 在以前的研究中， 我们用电子显微镜检查内源性逆转录病毒样颗粒， 在中国卵巢癌（CHO）细胞系中发现， CHO细胞中制备的几种重组产物的底物。 CHO细胞 含有电子显微镜可见的内源性逆转录病毒样颗粒 我们试图确定是否生长因子或细胞因子（可能 在CHO细胞中生产）可能会影响这些内源性颗粒。 的 结果表明，在CH处理后，颗粒增加了30- 100 细胞与几种细胞因子（GM-CSF，IFN，TNF）或与佛波酯 TPA。描述这些发现的手稿已发表在《体外》杂志上 毒理学（1991年）。 我们最近研究了另一种细胞基质 问题. 由于许多新的重组生物制品是使用 杆状病毒表达系统并使用称为Sf 9昆虫细胞系; 对其他能感染Sf 9的病毒知之甚少 细胞，特别是那些与人类宿主范围，我们最近发起了一个 一项研究，以确定任何对人类致病的病毒是否可以感染Sf 9 细胞我们已经检测了一系列病毒感染Sf 9的能力 细胞，以及用于评估Sf 9细胞是否不含病毒的方案。 污染和适用于生产生物制品 已经被开发出来了。 我们评估了Sf 9细胞被感染的能力， 15种病毒代表了几种虫媒病毒， 以及其他几种宿主范围广的病毒。圣刘易斯 脑炎病毒（SLE）是一种黄病毒，可感染SF 9 细胞 SLE引起Sf 9细胞（10-20%巨细胞）的CPE，也可引起B细胞 免疫荧光和电子显微镜检测SF 9细胞。一 手稿在J. Gen. Virol出版。我们测试的许多其他病毒 显然可以在细胞培养中存活很长一段时间， 明显的CPE，几个可以在单独的培养基中存活长达8周。 这可能 存在Sf 9细胞中生产的产品的污染风险， 没有进行适当的测试。

It is of interest to CBER whether cell lines used for product production contain virus or virus-like particles or viral nucleic acid sequences or support the growth of viruses infectious for humans. In previous studies, we used electron microscopy to examine endogenous retroviral-like particles found in the Chinese Hamster Ovary (CHO) cell line which is used as the cel substrate for several recombinant products made in CHO cells. CHO cells contain endogenous retroviral-like particles visible by electron microscopy We attempted to determine whether growth factors or cytokines (which might be manufactured in CHO cells) might affect these endogenous particles. The results indicated increases in particles from 30-100% after treatment of CH cells with several cytokines (GM-CSF, IFN, TNF) or with the phorbol ester TPA. A manuscript describing these findings has been published in In Vitro Toxicology (1991). We have recently approached another cell substrate issue. Since many new recombinant biological products are produced using the Baculovirus expression system and use an insect cell line called Sf9; and relatively little is known about other viruses which can infect Sf9 cells, particularly those with human host range, we recently initiated a study to determine whether any viruses pathogenic for humans can infect Sf9 cells. We have examined a series of viruses for their ability to infect Sf9 cells, and protocols for evaluating whether Sf9 cells are free of viral contamination and suitable for use for production of biological products have been developed. We evaluated the ability of Sf9 cells to be infected by fifteen viruses chosen to represent several classes of arboviruses as well as several additional viruses with wide host range. St Louis Encephalitis virus (SLE) a flavivirus, can productively infect the SF9 cells. SLE causes CPE in the Sf9 cells (10-20% giant cells) and can also b detected in SF9 cells by immunofluorescence and electron microscopy. A manuscript is in press in J. Gen. virol. Many of the other viruses we teste apparently can survive for long periods in cell culture without causing apparent CPE, several can survive up to 8 weeks in media alone. This may present risks of contamination for products produced in Sf9 cells if appropriate testing is not performed.