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Improving product quality and productivity of bispecific molecules through the application of continuous perfusion principles

机译：通过应用连续灌注原理，提高双特异性分子的产品质量和生产率

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Bispecific protein scaffolds can be more complex than traditional monoclonal antibodies (MAbs) because two different sites/domains for epitope binding are needed. Because of this increased molecular complexity, bispecific molecules are difficult to express and can be more prone to physical and chemical degradation compared to MAbs, leading to higher levels of protein aggregates, clipped species, or modified residues in cell culture. In this study, we investigated cell culture performance for the production of three types of bispecific molecules developed at Amgen. In particular, we cultured a total of six CHO cell lines in both an approximately 12-day fed-batch process and an approximately 40-day high-density perfusion process. Harvested cell culture fluid from each process was purified and analyzed for product quality attributes including aggregate levels, clipped species, charge variants, individual amino acid modifications and host cell protein (HCP) content. Our studies showed that in average, the intensified perfusion process increased 15-fold the integrated viable cell density and the total harvested product (and fivefold the daily volumetric productivity) compared to fed-batch. Furthermore, bispecific product quality improved in perfusion culture (as analyzed in affinity-capture pools) with reduction in levels of aggregates (up to 72% decrease), clipped species (up to 75% decrease), acidic variants (up to 76% decrease), deamidated/isomerized species in complementarity-determining regions, and HCP (up to 84% decrease). In summary, the intensified perfusion process exhibited better productivity and product quality, highlighting the potential to use it as part of a continuous manufacturing process for bispecific scaffolds.
机译：双特异性蛋白质支架可以比传统的单克隆抗体（mAb）更复杂，因为需要两种不同的表位结合的位点/结构域。由于这种增加的分子复杂性，与mAb相比，双特异性分子难以表达并且可以更容易发生物理和化学降解，导致细胞培养物中更高水平的蛋白质聚集体，剪裁物种或改性残基。在这项研究中，我们研究了在AMGEN中产生的三种类型的双特异性分子的细胞培养性能。特别是，我们在大约12天的喂食批处理和大约40天的高密度灌注过程中培养了总共六种Cho细胞系。纯化来自每种方法的收获细胞培养物，并分析包括骨料水平，剪裁物种，电荷变体，单个氨基酸修饰和宿主细胞蛋白（HCP）含量的产品质量属性。我们的研究表明，平均而言，与FED批次相比，增强的灌注过程增加了15倍的综合活细胞密度和总收获的产品（和每日体积生产率的5倍）。此外，在灌注培养中改善的双特异性产品质量（如在亲和捕获池中分析），减少骨料水平（降低72％），剪裁物种（降低高达75％），酸性变体（高达76％降低） ），循环/异构化物种在互补确定区域，HCP（降低高达84％）。总之，加强的灌注过程表现出更好的生产率和产品质量，突出了将其用作双特异性支架的连续制造过程的一部分。

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