Cascading effect in bioprocessing-The impact of mild hypothermia on CHO cell behavior and host cell protein composition
Cascading effect in bioprocessing-The impact of mild hypothermia on CHO cell behavior and host cell protein composition
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DOI：
10.1002/bit.26437
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发表时间：
2017-12-01
影响因子：
3.8
通讯作者：
Kontoravdi, Cleo
中科院分区：
文献类型：
--
作者：
Goey, Cher H.;Tsang, Joshua M. H.;Kontoravdi, Cleo
单克隆抗体 (mAb) 下游纯化的一个主要挑战是宿主细胞蛋白 (HCP) 的去除。先前的研究表明，细胞培养条件显着影响收获时的 HCP 含量。然而，目前尚不清楚过程条件如何影响宿主细胞群的生理变化，以及这些变化如何依次级联以改变 HCP 谱。我们通过细胞行为变化的角度，研究了温度下降 (TDS) 至轻度低温如何影响关键上游性能指标，即抗体滴度、HCP 浓度和 HCP 种类，跨越细胞培养衰退期和收获时。与维持在标准生理温度的生物反应器相比，在补料分批中国仓鼠卵巢 (CHO) 细胞生物反应器的第 5 天引入的温和低温条件导致细胞增殖率较低，但在细胞培养衰退阶段健康细胞的百分比较高。此外，在轻度低温培养中，细胞凋亡的发生不太明显。因此，轻度低温培养需要额外 5 天才能达到与标准生理温度下对照相似的完整活细胞浓度 (IVCC) 和抗体产量。当细胞活力降至 80% 以下时，轻度低温细胞培养物的 HCP 种类减少了 36%，其中蛋白酶和分子伴侣分别减少了约 44% 和 27%，尽管 HCP 浓度相似。这项研究表明，TDS 可能是一个很好的策略，可以通过减少 HCP 的种类来提供更清洁的下游原料，并通过减少蛋白酶和分子伴侣的数量来保持产品的完整性。
A major challenge in downstream purification of monoclonal antibodies (mAb) is the removal of host cell proteins (HCPs). Previous studies have shown that cell culture conditions significantly impact the HCP content at harvest. However, it is currently unclear how process conditions affect physiological changes in the host cell population, and how these changes, in turn, cascade down to change the HCP profile. We examined how temperature downshift (TDS) to mild hypothermia affects key upstream performance indicators, that is antibody titre, HCP concentration and HCP species, across the cell culture decline phase and at harvest through the lens of changes in cellular behavior. Mild hypothermic conditions introduced on day 5 of fed-batch Chinese hamster ovary (CHO) cell bioreactors resulted in a lower cell proliferation rate but larger percentages of healthier cells across the cell culture decline phase compared to bioreactors maintained at standard physiological temperature. Moreover, the onset of apoptosis was less evident in mild hypothermic cultures. Consequently, mild hypothermic cultures took an extra 5 days to reach an integral viable cell concentration (IVCC) and antibody yield similar to that of the control at standard physiological temperature. When cell viability dropped below 80%, mild hypothermic cell cultures had a reduced variety of HCP species by 36%, including approximately 44% and 27% lower proteases and chaperones, respectively, despite having similar HCP concentration. This study suggests that TDS may be a good strategy to provide cleaner downstream feedstocks by reducing the variety of HCPs and to maintain product integrity by reducing the number of proteases and chaperones.