一种抗肝细胞癌的杀肿瘤单链抗体的构建及其高细胞质表达

Construction and high cytoplasmic expression of a tumoricidal single-chain antibody against hepatocellular carcinoma.

作者信息

Sandee Duanpen, Tungpradabkul Sumalee, Tsukio Manae, Imanaka Tadayuki, Takagi Masahiro

机构信息

Department of Biotechnology, Graduate school of Engineering, Osaka University, Suita, Osaka 565-0871, Japan.

出版信息

BMC Biotechnol. 2002 Sep 12;2:16. doi: 10.1186/1472-6750-2-16.

DOI:10.1186/1472-6750-2-16

PMID:12227831

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC128817/

Abstract

BACKGROUND

Hep27 monoclonal (Hep27 Mab) is an antibody against hepatocellular carcinoma. Hep27 Mab itself can inhibit the growth of a hepatocellular carcinoma cell line (HCC-S102). We attempted to produce a single-chain fragment (scFv), a small fragment containing an antigen-binding site of Hep27 Mab, by using DNA-recombinant techniques.

RESULTS

The sequences encoding the variable regions of heavy (VH) and light (VL) chains of a murine Hep27 Mab were linked together by a linker peptide (Gly4Ser)3 and tagged with a hexa-histidine at the C-terminal; the resultant DNA construct was expressed in E. coli as an insoluble protein. The denatured scFv was refolded and purified by immobilized metal ion affinity chromatography (12 mg/l with a molecular weight of 27 kDa). Hep27scFv exhibited a tumoricidal activity against the HCC-S102 cell as its parental antibody (Hep27 Mab).

CONCLUSION

This scFv may be a potential candidate for a targeting agent in HCC immunodiagnosis or immunotherapy.

摘要

背景

Hep27单克隆抗体（Hep27 Mab）是一种针对肝细胞癌的抗体。Hep27 Mab自身能够抑制一种肝癌细胞系（HCC-S102）的生长。我们试图利用DNA重组技术制备一种单链片段（scFv），即包含Hep27 Mab抗原结合位点的小片段。

结果

将小鼠Hep27 Mab重链（VH）和轻链（VL）可变区的编码序列通过连接肽（Gly4Ser）3连接在一起，并在C末端标记六组氨酸；所得DNA构建体在大肠杆菌中表达为不溶性蛋白。变性的scFv经固定化金属离子亲和层析复性并纯化（产量为12 mg/l，分子量为27 kDa）。Hep27scFv与其亲本抗体（Hep27 Mab）一样，对HCC-S102细胞具有杀瘤活性。