Tang Qing, Onitsuka Masayoshi, Tabata Atsushi, Tomoyasu Toshifumi, Nagamune Hideaki
Department of Biological Science and Technology, College of Life and Materials Systems Engineering, Graduate School of Advanced Technology and Science, Tokushima University, Tokushima, Japan.
Department of Bioengineering, Division of Bioscience and Bioindustry, Graduate School of Technology, Industrial and Social Sciences, Tokushima University, Tokushima, Japan.
Anticancer Res. 2018 Jul;38(7):4319-4325. doi: 10.21873/anticanres.12731.
BACKGROUND/AIM: Recombinant antibodies have been investigated and used in applications such as targeting modules of drug-delivery systems (DDS) against cancers. This study aimed to prepare recombinant antibodies against HER2, containing sortase A (SrtA) recognition sequence, that are applicable as targeting modules in DDS after linkage with the drug-carrier containing oligoglycine-acceptor peptide by SrtA transpeptidation.
The recombinant trastuzumab fragment antibodies (scFvs and Fab) with the SrtA-recognition motif (LPXTG) at their C-terminal were constructed and expressed in Escherichia coli and Chinese hamster ovary (CHO) cells, respectively. The reactivity of the purified recombinant antibodies towards HER2-expressing cells was also evaluated via immunofluorescent staining.
Fab demonstrated higher yield and purity and better reactivity towards HER2-expressing cells (HCT-15 and HeLa) when compared to scFvs.
The CHO expression system possesses superior yield and purity when compared to the E. coli expression system with respect to the preparation of recombinant antibodies applicable in targeting modules for DDS (DDS-TM). Moreover, a Fab variant prepared in this study demonstrated the potential to be a DDS-TM against HER2-expressing cancer cells.
背景/目的：重组抗体已被研究并应用于诸如针对癌症的药物递送系统（DDS）靶向模块等应用中。本研究旨在制备针对HER2的重组抗体，其含有分选酶A（SrtA）识别序列，通过SrtA转肽作用与含寡甘氨酸受体肽的药物载体连接后，可作为DDS中的靶向模块。
构建了在其C末端带有SrtA识别基序（LPXTG）的重组曲妥珠单抗片段抗体（单链抗体片段和Fab片段），并分别在大肠杆菌和中国仓鼠卵巢（CHO）细胞中表达。还通过免疫荧光染色评估了纯化的重组抗体对HER2表达细胞的反应性。
与单链抗体片段相比，Fab片段表现出更高的产量和纯度，以及对HER2表达细胞（HCT - 15和HeLa）更好的反应性。
在制备适用于DDS靶向模块（DDS - TM）的重组抗体方面，与大肠杆菌表达系统相比，CHO表达系统具有更高的产量和纯度。此外，本研究制备的Fab片段变体显示出成为针对HER2表达癌细胞的DDS - TM的潜力。
