全人源抗VEGF165单克隆抗体培养工艺的研究

李昕然; 刘睿; 刘方堃; 刘煜

doi:10.11665/j.issn.1000-5048.20150617

全人源抗VEGF165单克隆抗体培养工艺的研究

1.
中国药科大学天然药物活性组分与药效国家重点实验室，生命科学与技术学院
2.
江苏先声药业有限公司

基金项目: 国家高技术研究发展计划(863计划)资助项目(No.2012AA02A303)；国家“重大新药创制”科技重大专项资助项目(No.2013ZX09301303-004)

计量
- 文章访问数: 1206
- HTML全文浏览量: 7
- PDF下载量: 1891
出版历程
- 刊出日期: 2015-12-24

Culture process for fully human anti-VEGF165 monoclonal antibody

摘要

摘要: 为研究一种表达全人源抗VEGF165单克隆抗体(HVAB)的重组DG44细胞的培养工艺。在摇瓶培养阶段，利用常见的流加和分阶段温度控制培养的方法，初步确定培养工艺。在生物反应器培养阶段，研究了不同pH范围对DG44细胞生长和单克隆抗体表达的影响。结果表明，通过补料可使HVAB表达量从101 mg/L提高到654 mg/L；通过在生长中期的适当降温，使细胞终活性保持在80%以上；pH控制范围6.4~7.4更适合DG44细胞的生长和HVAB表达。反应器中的抗体表达量为526 mg/L，与对照摇瓶相比降低了11%，为之后的中试研究奠定了工艺基础。
- DG44细胞 /
- 生物反应器 /
- 全人源单克隆抗体 /
- 悬浮培养 /
- 工艺优化
Abstract: This study was focus on the optimizing cell culture process of recombinant DG44 cells which expressing novel fully human anti-VEGF165 monoclonal antibody(HVAB). Feed-batch culture and two-phase temperature control were studied for optimizing the HVAB productivity in shaken flasks. The influences of pH changes were determined to study the growth of DG44 cell and expression of HVAB in bioreactors. The HVAB productivity was rose from 101 mg/L to 654 mg/L after feed-batch culture in shaken flask, and cell viability maintained above 80% after reduce the temperature in middle growth phase. It is also suggested that pH range at 6. 4-7. 4 is beneficial to DG44 cells growth and HVAB expression. The productivity in bioreactor is 526 mg/L decreased 11% compare with in shaken flask, which laid an foundation for further pilot scale production.
- DG44 cells /
- bioreactor /
- fully human monoclonal antibody /
- suspension culture /
- process optimization

HTML全文

参考文献(15)

[1]	Ferrara N. Vascular endothelial growth factor[J]. Arterioscler Thromb Vasc Biol,2009,29(6):789-791.
[2]	Chen G,Li J,Cui WY,et al.Construction of eukaryotic expression vector for mouse-human chimeric antibody against vascular endothelial growth factor in CHO DG44 cells[J].Chin J Biologicals(中国生物制品学杂志),2014,27(11):1394-1399.
[3]	Dai F, Shu L, Bian Y, et al. Safety of bevacizumab in treating metastatic colorectal cancer:a systematic review and meta-analysis of all randomized clinical trals[J].Clin Drug Investig,2013,33(11):779-788.
[4]	Wang XN,Wang LH,Gao P.The advantages,characteristics and experimental study for animal cells in serum-free culture medium[J].Asia-Pacific Tradit Med,2009,5(2):21-23.
[5]	Xu J,Wang D,Liu Y,et al.Serum-free medium optimization based on trial design and support vector regression[J].Biomed Res Int,2014,2014:269305.doi: 10.1155/2014/269305.
[6]	Wang D.The effect of silkworm 30kc6 gene for CHO cell expressing VEGF mAb(家蚕30kc6基因对CHO细胞表达VEGF单克隆抗体影响的研究)[D].Nanjing:China Pharmaceutical University,2014.
[7]	Guo JY.Development and optimization of the serm-free medium for CHO DG44 stable cell line(CHO DG44稳定细胞株无血清培养基的研发与优化)[D].Xiamen:Xiamen University,2014.
[8]	Liu Y, Yun J, Chen Q, et al. Isolation and characterization of human anti-VEGF165 monoclonal antibody with anti-tumor efficacy from transgenic mice expressing human immunoglobulin loci[J].Cancer Lett,2009,273(1):28-34.
[9]	Yang JD,Lu C,Stasny B,et al.Fed-batch bioreactor process scale-up from 3-L to 2,500-L scale for monoclonal antibody production from cell culture[J].Biotechnol Bioeng,2007,98(1):141-154.
[10]	Duan XJ,Ren T,Luo HY,et al.Review of animal cell medium for antibody drug production[J].Chin Med Biotechnol,2014,9(1):53-57.
[11]	Arden N,Betenbaugh MJ.Life and death in mammalian cell culture:strategies for apoptosis inhibition[J].Trends Biotechnol,2004,22(4):174-180.
[12]	Meuwlya F,Webera U,Zieglerb T,et al.Conversion of a CHO cell culture process from perfusion to fed-batch technology without altering product quality[J].J Biotechnol,2006,123(1):106-116.
[13]	Zeng AP. Quantitative assessment of cell density effect on the metabolism and antibody production rate of hybridoma cells at high cell density[J].Biotechnol,1996,45:243-251.
[14]	Mei JG,Zhuang JQ,Wang JL,et al.Advance of large-scale animal cell culture technology[J].China Biotechnol(中国生物工程杂志),2012,32(7):127-132.
[15]	Duan XJ.Problems and solutions of bioreactor scale-up for antibody production[J].Chin Med Biotechnol,2013,8(6):466-469.

施引文献

资源附件(0)

计量

文章访问数: 1206
HTML全文浏览量: 7
PDF下载量: 1891
被引次数: 0

全人源抗VEGF165单克隆抗体培养工艺的研究

计量

出版历程

Culture process for fully human anti-VEGF165 monoclonal antibody

计量

出版历程

目录