Department of Biology, Faculty of Basic Sciences, University of Guilan, Rasht, Iran.
Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
PLoS One. 2024 May 2;19(5):e0300702. doi: 10.1371/journal.pone.0300702. eCollection 2024.
The objective of the current investigation was to evaluate the induction of heat shock proteins (HSPs) in SP2/0 transgenic cells and the effect of these proteins on the production of monoclonal antibodies (mAbs). The SP2/0 cell line expressing the PSG-026 antibody, a biosimilar candidate of golimumab, the culture parameters, and the target protein expression were not justified for industrial production and were used for the experiments. Paracetamol and heat shock were used as chemical and physical inducers of HSPs, respectively. The results showed that paracetamol and heat shock increased the expression of HSP70 and HSP27 at the mRNA and protein levels. The expression of HSPs was greater in paracetamol-treated cells than in heat shock-treated cells. Paracetamol treatment at concentrations above 0.5 mM significantly reduced cell viability and mAb expression. However, treatment with 0.25 mM paracetamol results in delayed cell death and increased mAb production. Heat shock treatment at 45°C for 30 minutes after enhanced mAb expression was applied after pre-treatment with paracetamol. In bioreactor cultures, pretreatment of cells with paracetamol improved cell viability and shortened the lag phase, resulting in increased cell density. The production of mAbs in paracetamol-treated cultures was markedly greater than that in the control. Analysis of protein quality and charge variants revealed no significant differences between paracetamol-treated and control cultures, indicating that the induction of HSPs did not affect protein aggregation or charge variants. These findings suggest that inducing and manipulating HSP expression can be a valuable strategy for improving recombinant protein production in biopharmaceutical processes.
本研究旨在评估热休克蛋白（HSPs）在 SP2/0 转基因细胞中的诱导作用，以及这些蛋白对单克隆抗体（mAbs）生产的影响。表达 PSG-026 抗体的 SP2/0 细胞系是一种类似于 golimumab 的生物类似候选药物，其培养参数和目的蛋白表达水平不适合工业生产，因此用于实验。对乙酰氨基酚和热休克分别作为 HSPs 的化学和物理诱导剂。结果表明，对乙酰氨基酚和热休克均能上调 HSP70 和 HSP27 的 mRNA 和蛋白表达水平。与热休克处理相比，对乙酰氨基酚处理能显著上调 HSP 的表达。对乙酰氨基酚处理浓度高于 0.5mM 时，会显著降低细胞活力和 mAb 表达。然而，用 0.25mM 对乙酰氨基酚处理时，会延迟细胞死亡，增加 mAb 产量。在增强 mAb 表达后，用 45°C 热休克处理 30 分钟，作为对乙酰氨基酚预处理的后续处理。在生物反应器培养中，细胞用对乙酰氨基酚预处理后可提高细胞活力，缩短迟滞期，从而提高细胞密度。对乙酰氨基酚处理培养物中的 mAb 产量明显高于对照组。对蛋白质量和电荷变异体的分析表明，对乙酰氨基酚处理组和对照组之间没有显著差异，表明 HSP 的诱导不会影响蛋白聚集或电荷变异体。这些发现表明，诱导和操纵 HSP 表达可以成为提高生物制药过程中重组蛋白生产的一种有价值的策略。
