| Background and PurposeIn China, the incidence of gastric cancer has increased year by year, and mortality rate ranks the first. While the early gastric cancer lacks serious symptoms, so about 80%-90% of patients are already at advanced stage, thus leading to poor prognosis. At present, chemotherapy is the primary therapy for advanced carcinoma of stomach, while traditional chemotherapy drugs have considerable toxicity and are easy to produce drug resistance. As the molecular biology is inverstigated molecular targeting therapy is a new therapeutic strategy for gastric carcinoma. Nowdays, several molecule-targeting agents that are mainly EGFR and VEGF as the target, such as bevacizumab, showing anti-tumor activity in clinical researches. From the follow-up data in recent 30 years, 5 year survival f advanced gastric cancer is only 5%-17%. Therefore, how to improve the effect of treatment of gastric cancer is a big challenge in the clinical trials. Recent studies show that Neuropilin-1(NRP-1), as co-receptor of Semaphorin3, vascular endothelial growth factor(VEGF) and and others, is involved in vascular generation, growth and metastasis of tumor. So it is expected to become a new target for cancer therapy. Currently, some studies about NRP-1 at home and abroad, but uncommon in gastric cancer. This article is designed to detect influence of anti-NRP-1m Ab produced by our laboratory on the gastric cancer cell lines, which could lay a foundation for the development of a new anti-tumor strategy targeting NRP-1.MethodsOur laboratory has successfully produced NRP-1 monoclonal antibody(NRP-1m Ab) with high purity and sconcentration. The NRP-1 expression in gastric cancer cells were measured by Western blotting, and choose the high expression NRP-1 of a plant of cell for follow-up experiments. Meanwhile, the affinity and the location detection of NRP-1 were detected by flow cytometry and immunofluorescence techniques. Flow cytometry was set aside for observe cell apoptosis. Transwell detected migration and invasion of cells. In vivo, the NRP-1 m Ab anti-gastric effect was achieved on BGC-823 xenografts nude mice,and obtained the rate of tumor inhibition. Tumor tissue morphology was observed by hematoxylin eosin(HE). Finally, immunohistochemistry tested the expression of vascular endothelial growth factor(VEGF), microvessel density(MVD) and NRP-1 in tumor tissues.ResultsWestern blotting indicated that the NRP-1 was situated on the membrane of gastric cancer cell lines(BGC-823, MKN-28, SGC-7901), ecpecially in BGC-823 cells. MTT and cell colony formation inhibition experiment showed that NRP-1m Ab have inhibitory action on BGC-823 cell. Flow cytometry showed that NRP-1m Ab could induce led to BGC-823 cell apoptosis. Transwell results show that NRP-1m Ab can inhibit migration and invasion of gsatric cancer cell, and the inhibition of growth were showed dose dependent. In vivo, NRP-1m Ab can inhibit the growth of transplanted tumor. HE results showed that apoptosis drug group compared with the control group, the cell is more obvious. HE indicted that the apoptosis of cell were more obvious than the control group. Immunohistochemistry showed that expression of VEGF, MVD, and NRP-1 in tumor tissue were lower than the contrast group.Conclusions1、 NRP-1mAb can specially bind to the NRP-1 of the gastric cancer cell lines, especially in BGC-823 cells.2、 NRP-1mAb have inhibitory effect on the gastric cancer cells in dose-dependent method and time-dependent method.3、 NRP-1mAb can inhibit migration and invasion of gsatric cancer cells in dose dependent.4、 NRP-1mAb can inhibit the growth of transplanted tumor, and may be phase cooperation with VEGF and MVD. |
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