Department of Biomolecular Engineering, The University of California at Santa Cruz, Santa Cruz, California, United States of America.
PLoS One. 2018 Aug 2;13(8):e0197656. doi: 10.1371/journal.pone.0197656. eCollection 2018.
The production of envelope glycoproteins (Envs) for use as HIV vaccines is challenging. The yield of Envs expressed in stable Chinese Hamster Ovary (CHO) cell lines is typically 10-100 fold lower than other glycoproteins of pharmaceutical interest. Moreover, Envs produced in CHO cells are typically enriched for sialic acid containing glycans compared to virus associated Envs that possess mainly high-mannose carbohydrates. This difference alters the net charge and biophysical properties of Envs and impacts their antigenic structure. Here we employ a novel robotic cell line selection strategy to address the problems of low expression. Additionally, we employed a novel gene-edited CHO cell line (MGAT1- CHO) to address the problems of high sialic acid content, and poor antigenic structure. We demonstrate that stable cell lines expressing high levels of gp120, potentially suitable for biopharmaceutical production can be created using the MGAT1- CHO cell line. Finally, we describe a MGAT1- CHO cell line expressing A244-rgp120 that exhibits improved binding of three major families of bN-mAbs compared to Envs produced in normal CHO cells. The new strategy described has the potential to eliminate the bottleneck in HIV vaccine development that has limited the field for more than 25 years.
包膜糖蛋白（Env）的生产用于 HIV 疫苗极具挑战性。稳定的中国仓鼠卵巢（CHO）细胞系中 Env 的表达产量通常比其他具有药物应用价值的糖蛋白低 10-100 倍。此外，与主要含有高甘露糖碳水化合物的病毒相关的 Env 相比，CHO 细胞中产生的 Env 通常富含含有唾液酸的聚糖。这种差异改变了 Env 的净电荷和生物物理特性，并影响了它们的抗原结构。在这里，我们采用了一种新的机器人细胞系选择策略来解决低表达的问题。此外，我们还采用了一种新的基因编辑 CHO 细胞系（MGAT1-CHO）来解决高唾液酸含量和抗原结构不良的问题。我们证明，使用 MGAT1-CHO 细胞系可以创建表达高水平 gp120 的稳定细胞系，这些细胞系可能适合生物制药生产。最后，我们描述了一种表达 A244-rgp120 的 MGAT1-CHO 细胞系，与在正常 CHO 细胞中产生的 Env 相比，该细胞系能够改善与三种主要 bN-mAb 家族的结合。所描述的新策略有可能消除 HIV 疫苗开发中超过 25 年来限制该领域发展的瓶颈。
