| 引用本文: | 彭延杰,张林,吴正理,范加金,仲立军,曹强庚.HER2阳性乳腺癌重组免疫毒素的开发[J].中国现代应用药学,2023,40(7):881-887. |
| PENG Yanjie,ZHANG Lin,WU Zhengli,FAN Jiajin,ZHONG Lijun,CAO Qianggeng.Development of Recombinant Immunotoxin for HER2 Positive Breast Cancer[J].Chin J Mod Appl Pharm(中国现代应用药学),2023,40(7):881-887. |
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| 摘要: |
| 目的 制备一种人表皮生长因子受体2(human epidermal growth factor receptor 2,HER2)阳性乳腺癌的高活力重组免疫毒素TP25。方法 利用基因工程技术将曲妥珠单抗可变区与绿脓杆菌细胞毒素片段进行重组,经原核表达获得具有完整结构的高活性融合蛋白TP25,采用免疫荧光法与流式细胞技术检测TP25与不同乳腺癌细胞系的结合能力,通过3H-Thymidin掺入法检测其对肿瘤细胞的杀伤活力,通过测定单链DNA的产生研究TP25诱发肿瘤细胞发生凋亡。结果 经酶切与PCR验证成功构建融合蛋白TP25重组表达质粒,经原核包涵体表达制备出分子量为52 kDa的TP25制品,经验证TP25能与HER2高表达的乳腺癌细胞结合,具有活性空间构象,细胞增殖试验显示TP25的细胞毒性强弱与用量呈正相关,能引发HER2阳性乳腺癌细胞发生凋亡。结论 制备了HER2阳性乳腺癌靶向免疫毒素TP25。 |
| 关键词: 人表皮生长因子受体2 乳腺癌 免疫毒素 原核表达 细胞增殖 凋亡 |
| DOI：10.13748/j.cnki.issn1007-7693.20220389 |
| 分类号:R965.1 |
| 基金项目:山东省医药卫生科技发展计划项目(202002041518) |
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| Development of Recombinant Immunotoxin for HER2 Positive Breast Cancer |
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PENG Yanjie1, ZHANG Lin1, WU Zhengli2, FAN Jiajin1, ZHONG Lijun1, CAO Qianggeng1
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1.Shandong Provincial Maternal and Child Health Care Hospital, Jinan 250014, China;2.Southwest University, Chongqing 400715, China
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| Abstract: |
| OBJECTIVE To prepare a highly active recombinant immunotoxin TP25 for human epidermal growth factor receptor 2(HER2) positive breast cancer. METHODS The recombinant region of the trastuzumab variable fragment and cytotoxin was recombined by gene engineering technology, and a highly active fusion protein TP25 with intact structure was obtained by prokaryotic expression. The binding ability of TP25 to different breast cancer cell lines was detected by immunofluorescence and flow cytometry. The killing activity of TP25 was detected by 3H-Thymidin incorporation. The apoptosis of tumor cells induced by TP25 was studied by measuring the production of single stranded DNA. RESULTS The recombinant expression plasmid of TP25 was successfully constructed and verified by restriction enzyme digestion and PCR experiment. The TP25 product with a molecular weight of 52 kDa was prepared by prokaryotic expression. It was proved that TP25 could bind to breast cancer cells with high expression of HER2 and had an active spatial conformation. Cell proliferation assay showed that the cytotoxicity of TP25 was positively correlated with dosage and could induce apoptosis of HER2 positive breast cancer cell. CONCLUSION HER2 positive breast cancer targeting protein TP25 is prepared. |
| Key words: human epidermal growth factor receptor 2 breast cancer immunotoxin prokaryotic expression cell proliferation apoptosis |
