Brower Kevin P, Ryakala Venkat K, Bird Ryan, Godawat Rahul, Riske Frank J, Konstantinov Konstantin, Warikoo Veena, Gamble Jean
Late Stage Process Development, Genzyme - A Sanofi Company, 45 New York Avenue, Framingham, MA, 01701.
Biotechnol Prog. 2014 May-Jun;30(3):708-17. doi: 10.1002/btpr.1870. Epub 2014 Feb 5.
Downstream sample purification for quality attribute analysis is a significant bottleneck in process development for non-antibody biologics. Multi-step chromatography process train purifications are typically required prior to many critical analytical tests. This prerequisite leads to limited throughput, long lead times to obtain purified product, and significant resource requirements. In this work, immunoaffinity purification technology has been leveraged to achieve single-step affinity purification of two different enzyme biotherapeutics (Fabrazyme® [agalsidase beta] and Enzyme 2) with polyclonal and monoclonal antibodies, respectively, as ligands. Target molecules were rapidly isolated from cell culture harvest in sufficient purity to enable analysis of critical quality attributes (CQAs). Most importantly, this is the first study that demonstrates the application of predictive analytics techniques to predict critical quality attributes of a commercial biologic. The data obtained using the affinity columns were used to generate appropriate models to predict quality attributes that would be obtained after traditional multi-step purification trains. These models empower process development decision-making with drug substance-equivalent product quality information without generation of actual drug substance. Optimization was performed to ensure maximum target recovery and minimal target protein degradation. The methodologies developed for Fabrazyme were successfully reapplied for Enzyme 2, indicating platform opportunities. The impact of the technology is significant, including reductions in time and personnel requirements, rapid product purification, and substantially increased throughput. Applications are discussed, including upstream and downstream process development support to achieve the principles of Quality by Design (QbD) as well as integration with bioprocesses as a process analytical technology (PAT).
用于质量属性分析的下游样品纯化是非抗体生物制品工艺开发中的一个重大瓶颈。在许多关键分析测试之前，通常需要多步色谱工艺序列纯化。这一前提导致通量有限、获得纯化产品的前置时间长以及资源需求巨大。在这项工作中，免疫亲和纯化技术已被用于分别以多克隆抗体和单克隆抗体作为配体，对两种不同的酶生物治疗药物（法布赞 [阿加糖酶β] 和酶2）进行单步亲和纯化。从细胞培养收获物中快速分离出目标分子，其纯度足以进行关键质量属性（CQA）分析。最重要的是，这是第一项证明应用预测分析技术来预测商业生物制品关键质量属性的研究。使用亲和柱获得的数据被用于生成适当的模型，以预测在传统多步纯化序列之后将获得的质量属性。这些模型利用与原料药等效的产品质量信息为工艺开发决策提供支持，而无需生成实际原料药。进行了优化以确保最大程度的目标物回收和最小程度的目标蛋白降解。为法布赞开发的方法成功地重新应用于酶2，表明了该平台的机会。该技术的影响是巨大的，包括减少时间和人员需求、快速产品纯化以及大幅提高通量。讨论了其应用，包括上游和下游工艺开发支持，以实现设计质量（QbD）原则以及作为过程分析技术（PAT）与生物工艺的整合。
