Expression and characterization of recombinant humanized anti-HER2 single-chain antibody in Pichia pastoris for targeted cancer therapy

详细信息    查看全文

• 作者：Xiaodan Cao ; Haijun Yu ; Chao Chen ; Jia Wei ; Ping Wang
• 关键词：Anti ; HER2 ; Breast cancer cell line SKBR3 ; Cancer therapy ; Heterogeneous expression ; Humanized single ; chain variable fragment ; Pichia pastoris
• 刊名：Biotechnology Letters
• 出版年：2015
• 出版时间：July 2015
• 年：2015
• 卷：37
• 期：7
• 页码：1347-1354
• 全文大小：849 KB
• 参考文献：Afshar S, Olafsen T, Wu AM, Morrison SL (2009) Characterization of an engineered human purine nucleoside phosphorylase fused to an anti-her2/neu single chain Fv for use in ADEPT. J Exp Clin Cancer Res 28:147PubMed Central PubMed View Article
  Ahmad M, Hirz M, Pichler H, Schwab H (2014) Protein expression in Pichia pastoris: recent achievements and perspectives for heterologous protein production. Appl Microbiol Biotechnol 98:5301-317PubMed Central PubMed View Article
  Athmaram TN, Saraswat S, Singh AK et al (2012) Influence of copy number on the expression levels of pandemic influenza hemagglutinin recombinant protein in methylotrophic yeast Pichia pastoris. Virus Genes 45:440-51PubMed View Article
  Cheng LS, Liu AP, Yang JH et al (2003) Construction, expression and characterization of the engineered antibody against tumor surface antigen, p185(c-erbB-2). Cell Res 13:35-8PubMed View Article
  Colombo M, Corsi F, Foschi D et al (2010) HER2 targeting as a two-sided strategy for breast cancer diagnosis and treatment: outlook and recent implications in nanomedical approaches. Pharmacol Res 62:150-65PubMed View Article
  Galeffi P, Lombardi A, Donato MD et al (2005) Expression of single-chain antibodies in transgenic plants. Vaccine 23:1823-827PubMed View Article
  Galeffi P, Lombardi A, Pietraforte I et al (2006) Functional expression of a single-chain antibody to ErbB-2 in plants and cell-free systems. J Transl Med 4:39PubMed Central PubMed View Article
  Gerber DE (2008) Targeted therapies: a new generation of cancer treatments. Am Fam Physician 77:311-19PubMed
  Hohenblum H, Gasser B, Maurer M et al (2004) Effects of gene dosage, promoters, and substrates on unfolded protein stress of recombinant Pichia pastoris. Biotechnol Bioeng 85:367-75PubMed View Article
  Hu S, Li L, Qiao J et al (2006) Codon optimization, expression, and characterization of an internalizing anti-ErbB2 single-chain antibody in Pichia pastoris. Protein Expr Purif 47:249-57PubMed View Article
  Inan M, Aryasomayajula D, Sinha J, Meagher MM (2006) Enhancement of protein secretion in Pichia pastoris by overexpression of protein disulfide isomerase. Biotechnol Bioeng 93:771-78PubMed View Article
  Liu D, Wang C, Li C et al (2011) Production and characterization of a humanized single-chain antibody against human integrin αvβ3 protein. J Biol Chem 286:24500-4507PubMed Central PubMed View Article
  Lombardi A, Sperandei M, Cantale C et al (2005) Functional expression of a single-chain antibody specific for the HER2 human oncogene in a bacterial reducing environment. Protein Expr Purif 44:10-5PubMed View Article
  Mack M, Wannemacher M, Hobl B et al (2009) Comparison of two expression platforms in respect to protein yield and quality: Pichia pastoris versus Pichia angusta. Protein Expr Purif 66:165-71PubMed View Article
  Mansur M, Cabello C, Hernández L et al (2005) Multiple gene copy number enhances insulin precursor secretion in the yeast Pichia pastoris. Biotechnol Lett 27:339-45PubMed View Article
  Ménard S, Casalini P, Campiglio M et al (2001) HER2 overexpression in various tumor types, focussing on its relationship to the development of invasive breast cancer. Ann Oncol 12(Suppl 1):S15–S19PubMed View Article
  Mitri Z, Constantine T, O’Regan R (2012) The HER2 receptor in breast cancer: pathophysiology, clinical use, and new advances in therapy. Chemother Res Pract 2012:743193PubMed Central PubMed
  Monnier PP, Vigouroux RJ, Tassew NG (2013) In vivo applications of single chain Fv (variable domain) (scFv) fragments. Antibodies 2:193-08View Article
  Muzard J, Bouabdelli M, Zahid M et al (2009) Design and humanization of a murine scFv that blocks human platelet glycoprotein VI in vitro. FEBS J 276:4207-222PubMed View Article
  Nejatollahi F, Jaberipour M, Asgharpour M (2014) Triple blockade of HER2 by a cocktail of anti-HER2 scFv antibodies induces high antiproliferative effects in breast cancer cells. Tumour Biol 35:7887-895PubMed View Article
  Nordén K, Agemark M, Danielson J?H et al (2011) Increasing gene dosage greatly enhances recombinant expression of aquaporins in Pichia pastoris. BMC Biotechnol 11:47PubMed Central PubMed View Article
  Pavlinkova G, Colcher D, Booth BJ et al (2001) Effects of humanization and gene shuffling on immunogenicity and antigen binding of anti-TAG-72 single-chain Fvs. Int J Cancer 94:717-26PubMed View Article
  Qiu MZ, Xu RH (2013) The progress of targeted therapy in advanced gastric cancer. Biomark Res 1:32PubMed Central PubMed View Article
  Reynolds JG, Geretti E, Hendriks BS et al (2012) HER2-targeted liposomal doxorubicin displays enhanced anti-tumorigenic effects without associated cardiotoxicity. Toxicol Appl Pharmacol 262:1-0PubMed View Article
  Sinclair G, Choy FYM (2002) Synonymous codon usage bias and the expression of human glucocerebrosidase in the methylotrophic yeast, Pichia pastoris. Protein Expr Purif 26:96-05PubMed V
• 作者单位：Xiaodan Cao (1)
  Haijun Yu (1)
  Chao Chen (1)
  Jia Wei (1)
  Ping Wang (1)
  
  1. Biomedical Nanotechnology Center, State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, 200237, People’s Republic of China
  
• 刊物类别：Biomedical and Life Sciences
• 刊物主题：Life Sciences
  Microbiology
  Biotechnology
  Applied Microbiology
  Biochemistry
  
• 出版者：Springer Netherlands
• ISSN：1573-6776

文摘

Objectives The availability of self-targeting and low immunogenic therapeutic agents is critical to efficient cancer therapy. Therefore, the development of humanized therapeutic antibodies is particularly appealing. Results A humanized single-chain variable fragment (scFv) antibody that can target human epidermal growth factor receptor-2 (HER2)-overexpressing cancer cells was designed and produced via expression in Pichia pastoris. The expression gave a high yield of 8?mg protein/l (with a purity of 92?%) using shake-flask cultures. Functional studies also revealed that the purified recombinant anti-HER2 scFv exhibited anti-proliferative activity and could bind efficiently to HER2-overexpressing human breast cancer cell line SKBR3. Conclusion The recombinant scFv offers promising therapeutic and binding efficiencies that are desirable for targeted cancer therapies.