Jain Nina K, Barkowski-Clark Susan, Altman Richard, Johnson Krista, Sun Fang, Zmuda Jonathan, Liu Chao Yan, Kita Adriana, Schulz Ryan, Neill Alyssa, Ballinger Robert, Patel Rekha, Liu Jian, Mpanda Alinafe, Huta Brian, Chiou Henry, Voegtli Walter, Panavas Tadas
Discovery Research CT, Alexion Pharmaceuticals, 100 College Street, New Haven, CT 06510, United States.
Cell Biology, Life Sciences Solutions Group, Thermo Fisher, 7335 Executive Way, Frederick, MD 21704, United States.
Protein Expr Purif. 2017 Jun;134:38-46. doi: 10.1016/j.pep.2017.03.018. Epub 2017 Mar 23.
Chinese Hamster Ovary (CHO) cells are the principal mammalian host used for stable cell line generation and biotherapeutic protein production. Until recently, production of milligrams to grams of protein in CHO transient systems was challenging. As such, Human Embryonic Kidney (HEK293) cells are the most common mammalian cell type used for transient transfection. The post-translational modifications (PTMs) of a protein are dictated in part by the cell line used for expression, and changes in PTMs have been shown to affect both the activity and biophysical properties of proteins. Therefore, it is potentially advantageous to keep the host cell type consistent throughout drug discovery and development. To this end, we compared the ExpiCHO system, a high density CHO-S transient transfection system, to the Expi293 and FreeStyle MAX CHO transient systems. Fourteen proteins were expressed in both the Expi293 and ExpiCHO systems. For a majority of proteins tested, the protein titers observed with the ExpiCHO system were higher than those seen with both the FreeStyle MAX CHO and Expi293 systems. Antibodies expressed using the ExpiCHO system had glycosylation patterns more similar to antibodies produced in stable CHO cell lines than Expi293-derived antibodies. However, culture duration and temperature were found to affect protein titer, monodispersity, enzyme activity, and PTMs and should be carefully selected when using the ExpiCHO system. The ExpiCHO transient transfection systems allows for facile production of milligrams to grams of protein in CHO cells and de-risks the transition from transient to stable material during drug development.
中国仓鼠卵巢（CHO）细胞是用于稳定细胞系构建和生物治疗性蛋白生产的主要哺乳动物宿主。直到最近，在CHO瞬时表达系统中生产毫克至克级别的蛋白仍具有挑战性。因此，人胚肾（HEK293）细胞是用于瞬时转染的最常见哺乳动物细胞类型。蛋白质的翻译后修饰（PTM）部分取决于用于表达的细胞系，并且已表明PTM的变化会影响蛋白质的活性和生物物理特性。因此，在整个药物发现和开发过程中保持宿主细胞类型一致可能具有优势。为此，我们将高密度CHO-S瞬时转染系统ExpiCHO与Expi293和FreeStyle MAX CHO瞬时系统进行了比较。在Expi293和ExpiCHO系统中都表达了14种蛋白质。对于大多数测试的蛋白质，ExpiCHO系统观察到的蛋白产量高于FreeStyle MAX CHO和Expi293系统。使用ExpiCHO系统表达的抗体的糖基化模式比源自Expi293的抗体更类似于在稳定CHO细胞系中产生的抗体。然而，发现培养持续时间和温度会影响蛋白产量、单分散性、酶活性和PTM，在使用ExpiCHO系统时应仔细选择。ExpiCHO瞬时转染系统能够在CHO细胞中轻松生产毫克至克级别的蛋白质，并降低了药物开发过程中从瞬时表达材料过渡到稳定表达材料的风险。
