State Key Laboratory of Cancer Biology & Xijing Hospital of Digestive Disease, the Fourth Military Medical University, 15 West Chang-Le Road, Xi'an 710032, PR China.
Gut. 2010 Mar;59(3):292-9. doi: 10.1136/gut.2008.155226. Epub 2009 Nov 30.
HER2, an oncogene, has been found to be over-expressed in 10-40% of human gastric carcinomas. The aims of this study were to investigate if a fusion protein consisting of anti-HER2 sFv and constitutively active caspase-3 was capable of inducing apoptosis in HER2-expressing human gastric cancer cells and blocking the growth of human gastric cancer xenografts in nude mice.
NIH3T3 cells stably transduced with the pcDNA3.1-HER-PE-CP3 recombinant plasmid containing a secretion signal, a single-chain anti-HER2 monoclonal antibody fragment, a Pseudomonas exotoxin A translocation domain and a constitutively active caspase-3 molecule were used to induce apoptosis in human gastric cancer cells both in vitro and in vivo. Immunofluorescence staining and western blotting were used to examine the expression of the recombinant protein HER-PE-CP3. Apoptosis was determined by flow cytometry and TUNEL assay.
Co-cultivation of HER-PE-CP3/ NIH3T3 with human gastric cancer cells led to internalisation of HER-PE-CP3 and apoptosis in HER2-expressing human gastric cancer cells but not in HER2-negative cancer cells. Inoculation of HER-PE-CP3/NIH3T3 in nude mice resulted in potent inhibition of human gastric cancer xenografts and much prolonged survival time of the tumour-bearing mice compared with the control. Significantly more apoptotic cells were detected in xenografts in mice receiving HER-PE-CP3/NIH3T3 than in control mice.
The HER-PE-CP3 chimeric molecule could induce selective apoptosis and potent growth inhibition of HER2-positive human gastric cancer cells and might represent a novel HER2-directed treatment option for human gastric cancer.
HER2 是一种癌基因，在 10-40%的人类胃癌中过表达。本研究的目的是探讨由抗 HER2 scFv 和组成性激活的 caspase-3 组成的融合蛋白是否能够诱导 HER2 表达的人类胃癌细胞凋亡，并阻断裸鼠中人类胃癌异种移植物的生长。
使用稳定转染 pcDNA3.1-HER-PE-CP3 重组质粒的 NIH3T3 细胞，该质粒包含分泌信号、单链抗 HER2 单克隆抗体片段、铜绿假单胞菌外毒素 A 易位结构域和组成性激活的 caspase-3 分子，在体外和体内诱导人类胃癌细胞凋亡。免疫荧光染色和 Western blot 用于检测重组蛋白 HER-PE-CP3 的表达。通过流式细胞术和 TUNEL 测定法确定细胞凋亡。
HER-PE-CP3/NIH3T3 与人类胃癌细胞共培养导致 HER-PE-CP3 内化，并导致 HER2 表达的人类胃癌细胞凋亡，但不导致 HER2 阴性癌细胞凋亡。将 HER-PE-CP3/NIH3T3 接种于裸鼠导致人类胃癌异种移植物的强烈抑制，并且与对照组相比，荷瘤小鼠的存活时间明显延长。与对照组相比，接受 HER-PE-CP3/NIH3T3 的小鼠异种移植物中检测到更多的凋亡细胞。
HER-PE-CP3 嵌合分子可以诱导 HER2 阳性人类胃癌细胞的选择性凋亡和强大的生长抑制，并且可能代表人类胃癌的一种新的 HER2 导向治疗选择。
