Ye Jianxin, Kober Vanessa, Tellers Melanie, Naji Zubia, Salmon Peter, Markusen Julia F
Merck & Co., Inc., Bioprocess Research & Development, Rahway, New Jersey 07065, USA.
Biotechnol Bioeng. 2009 Jun 15;103(3):542-51. doi: 10.1002/bit.22265.
Chinese hamster ovary cells (CHO) have been extensively utilized as the production platform for therapeutic proteins including monoclonal antibodies in pharmaceutical industry. For early development, it would be advantageous to rapidly produce large amounts of protein in the same cell line; therefore, development of a CHO transient transfection platform with high protein expression level is highly desirable. Here, we describe the development of such a platform in CHO cells. Polyethylenimine (PEI) was used as the transfection reagent. Different media were screened for the best transfection and expression performance, and UltraCHO was chosen as the best performer. DMSO and lithium acetate (LiAc) were discovered to improve CHO transient transfection expression levels significantly. A 14-day fed-batch process was successfully developed to further increase production yield. With an optimized transient transfection process, we were able to express monoclonal antibody (Mab) in CHO cells at a high level, averaging 80 mg/L. The process was successfully scaled up to 10 L working volume in a 20 L wave bioreactor. As expected, the Mabs had similar glycosylation patterns in comparison to the Mabs produced from a stably transfected CHO cell line, while in contrast Mabs expressed transiently from HEK293EBNA cells differed.
中国仓鼠卵巢细胞（CHO）已被广泛用作制药行业生产包括单克隆抗体在内的治疗性蛋白质的平台。在早期开发中，能够在同一细胞系中快速大量生产蛋白质将具有优势；因此，非常需要开发一种具有高蛋白表达水平的CHO瞬时转染平台。在此，我们描述了在CHO细胞中开发这样一个平台的过程。聚乙烯亚胺（PEI）用作转染试剂。筛选了不同的培养基以获得最佳的转染和表达性能，UltraCHO被选为表现最佳的培养基。发现二甲基亚砜（DMSO）和醋酸锂（LiAc）可显著提高CHO瞬时转染的表达水平。成功开发了一个14天的补料分批培养过程以进一步提高产量。通过优化的瞬时转染过程，我们能够在CHO细胞中高水平表达单克隆抗体（Mab），平均水平为80 mg/L。该过程成功放大至20 L波浪生物反应器中的10 L工作体积。正如预期的那样，与从稳定转染的CHO细胞系产生的单克隆抗体相比，这些单克隆抗体具有相似的糖基化模式，而相比之下，从HEK293EBNA细胞瞬时表达的单克隆抗体则有所不同。
