Codon engineering for improved antibody expression in mammalian cells
Codon engineering for improved antibody expression in mammalian cells
复制标题
DOI：
10.1016/j.pep.2007.05.017
复制
发表时间：
2007-10-01
影响因子：
1.6
通讯作者：
Sweet, Ray
中科院分区：
文献类型：
--
作者：
Carton, Jill M.;Sauerwald, Tina;Sweet, Ray
虽然在细菌宿主中已经很好地建立，但在哺乳动物系统中编码序列变异对蛋白质表达的影响在病毒蛋白质或来自远缘系统发生家族的蛋白质之外的特征很差。考虑到哺乳动物表达系统在蛋白质生物治疗药物的研究和生产中的广泛使用，潜在影响是巨大的。我们正在研究密码子工程对重组抗体表达的影响，重点是开发生产细胞系。CNTO 888是与MorphoSys AG合作通过抗体噬菌体展示获得的人MCP-1特异性mAb。抗体的分离的DNA序列偏向细菌密码子，反映了用于在大肠杆菌中噬菌体展示表达的Fat文库的工程化。我们比较了含有亲本V区序列的CNTO 888与两种工程编码变体的表达。在天然密码子交换（NCE）变体中，V区密码子被天然来源的人抗体基因中使用的密码子替换。在人密码子优化（HCO）变体中，V区密码子是基于人密码子使用表以最高频率使用的那些。测量来自哺乳动物宿主细胞中稳定转染的抗体表达水平。CNTO 888的HCO密码子变体产生了最高表达的细胞系和筛选群体的最高平均表达。这对生成CNTO 888生产细胞系的过程具有显著的积极影响，并表明通过密码子工程改造改善哺乳动物表达系统中抗体表达的潜力。(C)2007年爱思唯尔公司All rights reserved.
While well established in bacterial hosts, the effect of coding sequence variation on protein expression in mammalian systems is poorly characterized outside of viral proteins or proteins from distant phylogenetic families. The potential impact is substantial given the extensive use of mammalian expression systems in research and manufacturing of protein biotherapeutics. We are studying the effect of codon engineering on expression of recombinant antibodies with an emphasis on developing manufacturing cell lines. CNTO 888, a human mAb specific for human MCP-1, was obtained by antibody phage display in collaboration with MorphoSys AG. The isolated DNA sequence of the antibody was biased towards bacterial codons, reflecting the engineering of the Fat) library for phage display expression in Escherichia coli. We compared the expression of CNTO 888 containing the parental V-region sequences with two engineered coding variants. In the native codon exchanged (NCE) variant, the V-region codons were replaced with those used in naturally derived human antibody genes. In the human codon optimized (HCO) variant the V-region codons were those used at the highest frequency based on a human codon usage table. The antibody expression levels from stable transfections in mammalian host cells were measured. The HCO codon variant of CNTO 888 yielded the highest expressing cell lines and the highest average expression for the screened populations. This had a significant positive effect on the process to generate a CNTO 888 production cell line and indicates the potential to improve antibody expression in mammalian expression systems by codon engineering. (C) 2007 Elsevier Inc. All rights reserved.