Holstein Melissa, Jang Dongyoun, Urrea Christine, Botta Lakshmi Sirisha, Grimm William, Ghose Sanchayita, Li Zheng Jian
Biologics Process Development, Global Product Development and Supply, Bristol-Myers Squibb Co., Devens, Massachusetts, USA.
Biotechnol Prog. 2021 Jan;37(1):e3086. doi: 10.1002/btpr.3086. Epub 2020 Oct 16.
Beta-glucans are polysaccharides of D-glucose monomers linked by (1-3) beta-glycosidic bonds, are found to have a potential immunogenicity risk in biotherapeutic products, and are labeled as process contaminants. A common source of beta-glucans is from the cellulose found in traditional depth filter media. Typically, beta-glucan impurities that leach into the product from the primary clarification depth filters can be removed by the subsequent bind-and-elute affinity chromatography capture step. Beta-glucans can also be removed by a bind-and-elute cation exchange chromatography step, which is useful for removing beta-glucans introduced by a post-Protein A depth filtration step. However, the increasing prevalence of flowthrough polishing chromatography poses a challenge for beta-glucan removal due to the lack of any bind-and-elute chromatography steps after the post-Protein A depth filter. In this work, a depth filter flush strategy was developed to control beta-glucan leaching into the product pool. Different loading conditions for the depth filtration and subsequent chromatography steps were evaluated to determine the robustness of the optimized flush strategy. Carry through runs demonstrated greater than two-fold reduction in beta-glucan levels using the optimized wash as compared to standard filter flush conditions.
β-葡聚糖是由D-葡萄糖单体通过(1-3)β-糖苷键连接而成的多糖，在生物治疗产品中被发现具有潜在的免疫原性风险，并被标记为工艺污染物。β-葡聚糖的常见来源是传统深层过滤介质中的纤维素。通常，从初级澄清深层过滤器中渗入产品的β-葡聚糖杂质可通过后续的结合-洗脱亲和层析捕获步骤去除。β-葡聚糖也可通过结合-洗脱阳离子交换层析步骤去除，这对于去除蛋白A深度过滤步骤后引入的β-葡聚糖很有用。然而，由于蛋白A深度过滤器之后缺乏任何结合-洗脱层析步骤，流通式精制层析的日益普及对β-葡聚糖的去除构成了挑战。在这项工作中，开发了一种深层过滤器冲洗策略来控制β-葡聚糖渗入产品池。评估了深层过滤和后续层析步骤的不同负载条件，以确定优化冲洗策略的稳健性。与标准过滤器冲洗条件相比，采用优化冲洗的连续运行显示β-葡聚糖水平降低了两倍以上。
