Streefland Mathieu, van de Waterbeemd Bas, Kint Joeri, van der Pol Leo A, Beuvery E Coen, Tramper Johannes, Martens Dirk E
Netherlands Vaccine Institute (NVI), Unit Research and Development, PO Box 457, 3720 AL Bilthoven, The Netherlands.
Biotechnol Bioeng. 2009 Jan 1;102(1):161-7. doi: 10.1002/bit.22030.
Modern (bio)pharmaceutical process development requires thorough investigation of all process parameters that are critical to product quality. The impact of a disturbance of such a parameter during processing needs to be known so that a rational decision can be made about the release of the product. In cultivation processes the dissolved oxygen (DO) concentration is generally accepted as being a critical parameter. In this article the impact of a 90 min period of oxygen limitation during the cultivation of the strictly aerobic Bordetella pertussis bacterium is investigated. The cultivation is the most important process step for the manufacturing of a vaccine against whooping cough disease. Samples were taken immediately before and after oxygen limitation and at the end of cultivation of four oxygen limited and three control cultivations. DNA microarray analysis of the full transcriptome of the B. pertussis bacterium revealed that a 90 min period of oxygen limitation has a substantial effect on overall gene expression patterns. In total 104 genes were identified as a significant hit at any of the sample points, of which 58 were directly related to oxygen limitation. The other genes were mainly affected towards the end of cultivation. Of all genes involved in oxygen limitation none were identified to show a significant difference between the oxygen limited and control cultivations at the end of the batch. This indicates a fully reversible effect of oxygen limitation on gene expression. This finding has implications for the risk assessment of dissolved oxygen concentration as a critical process parameter.
现代（生物）制药工艺开发需要对所有影响产品质量的关键工艺参数进行全面研究。必须了解加工过程中此类参数受到干扰时所产生的影响，以便能够就产品放行做出合理决策。在培养过程中，溶解氧（DO）浓度通常被视为一个关键参数。本文研究了在严格需氧的百日咳博德特氏菌培养过程中，90分钟的氧气限制期所产生的影响。培养是生产百日咳疫苗最重要的工艺步骤。在四次氧气受限培养和三次对照培养的氧气限制期前后以及培养结束时立即取样。对百日咳博德特氏菌全转录组进行的DNA微阵列分析表明，90分钟的氧气限制期对整体基因表达模式有重大影响。总共鉴定出104个基因在任何一个取样点都有显著变化，其中58个基因与氧气限制直接相关。其他基因主要在培养结束时受到影响。在所有与氧气限制相关的基因中，没有一个在批次结束时在氧气受限培养和对照培养之间表现出显著差异。这表明氧气限制对基因表达具有完全可逆的影响。这一发现对将溶解氧浓度作为关键工艺参数的风险评估具有重要意义。
