Rapid Optimization of Protein Freeze-Drying Formulations Using Ultra Scale-Down and Factorial Design of Experiment in Microplates
Rapid Optimization of Protein Freeze-Drying Formulations Using Ultra Scale-Down and Factorial Design of Experiment in Microplates
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DOI：
10.1002/bit.22448
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发表时间：
2009-12-01
影响因子：
3.8
通讯作者：
Dalby, Paul A.
中科院分区：
文献类型：
--
作者：
Grant, Yitzchak;Matejtschuk, Paul;Dalby, Paul A.
保持生物制药蛋白的稳定状态对其安全性和功效至关重要，也是优化最终产品的主要因素。冻干制剂提供了一种提高稳定性的途径。目前，冷冻干燥配方的优化是一个经验过程，需要大量耗时的实验，并且使用大量的产品材料。在这里，我们描述了一个用于快速识别和优化配方赋形剂的通用框架，以防止冻干过程中蛋白质活性的损失。使用因子实验设计 (DOE) 方法与微孔板冻干相结合，快速确定了一系列最佳配方，可在冷冻干燥过程中稳定乳糖脱氢酶（源自莱克曼乳杆菌）。本文概述的程序作为初始筛选，以确定关键赋形剂和潜在的相互作用，然后进行中心复合面设计的优化实验。聚乙二醇 (PEG) 和乳糖在筛选阶段对维持蛋白质稳定性具有显着影响，优化产生了用于绘制操作窗口的准确模型。还描述了冷冻干燥期间微孔板中发生的冷冻温度和升华速率的变化。然后将最佳配方在带塞的小瓶中冷冻干燥，以验证微尺度数据与较大中试规模观察到的效果相关。这项工作提供了一种生物制药制剂筛选的通用方法，可以筛选可能的赋形剂的单一效应和相互作用效应，从而提高通量，同时降低时间和材料方面的成本。生物技术。生物工程。 2009；104：957-964。 (C) 2009 年 Wiley 期刊公司。
Retaining biopharmaceutical proteins in a stable form is critical to their safety and efficacy, and is a major factor for optimizing the final product. Freeze-dried formulations offer one route for improved stability. Currently the optimization of formulations for freeze-drying is an empirical process that requires many time-consuming experiments and also uses large quantities of product material. Here we describe a generic framework for the rapid identification and optimization of formulation excipients to prevent loss of protein activity during a lyophilization process. Using factorial design of experiment (DOE) methods combined with lyophilization in microplates a range of optimum formulations were rapidly identified that stabilized lactose dehydrogenase (derived from Lactobacillus leichmanii) during freeze-drying. The procedure outline herein-as initial screen to identify key excipients and potential interactions followed by a central composite face designed optimization experiment. Polyethylene glycol (PEG) and a lactose were shown to have significant effects on maintaining protein stability at the screening stage and optimization resulted in an accurate model that was used to plot a window of operation. The variation of freezing temperatures and rates of sublimation that occur across a microplate during freeze-drying have been characterized also. The optimum formulation was then freeze-dried in stoppered vials to verify that the microscale data was relevant to the effects observed at larger pilot scales. This work provides a generic approach to biopharmaceutical formulation screening where possible excipients can be screened for single and interactive effects thereby increasing throughput while reducing costs in terms of time and materials. Biotechnol. Bioeng. 2009;104: 957-964. (C) 2009 Wiley Periodicals, Inc.