Abstract
The epithelial-mesenchymal transition (EMT) is an important factor in lung cancer metastasis, and targeting EMT is a potential therapeutic strategy. Fucosyltransferase IV (FUT4) and its synthetic cancer sugar antigen Lewis Y (LeY) was abnormally elevated in many cancers. In this study, a traditional Chinese medicine ginsenoside Rg3 was used to investigate whether its inhibition to EMT and invasion of lung cancer is by the glycobiology mechanism. We found that Rg3 treatment (25, 50, 100 μg/ml) inhibited cell migration and invasion by wound-healing and transwell assays. Rg3 could significantly alter EMT marker proteins with increased E-cadherin, but decreased Snail, N-cadherin and Vimentin expression. Rg3 also down-regulated FUT4 gene and protein expression in lung cancer cells by qPCR, Western blot and immunofluorescence. After FUT4 down-regulated with shFUT4, EMT was obviously inhibited. Furthermore, the activation of EGFR through decreased LeY biosynthesis was inhibited, which blocked the downstream MAPK and NF-κB signal pathways. In addition, Rg3 reduced tumor volume and weight in xenograft mouse model, and significantly decreased tumor metastasis nodules in lung tissues by tail vein injection. In conclusion, Rg3 inhibits EMT and invasion of lung cancer by down-regulating FUT4 mediated EGFR inactivation and blocking MAPK and NF-κB signal pathways. Rg3 may be a potentially effective agent for the treatment of lung cancer.
Highlights
RESULTSWe examined epithelial-mesenchymal transition (EMT) marker proteins, and found that E-cadherin expression was increased and N-cadherin expression was decreased by Quantitative real-time PCR (qPCR) (Figure 4F), Western blot (Figure 4G) and immunofluorescent staining (Figure 4H) in shFUT4 transfected cells compared with the untreated control and mock transfected cells
The addition of anti-Lewis Y (LeY) antibody, EGFR inhibitor (AG1478) or NF-κB inhibitor (BAY11-7082) significantly inhibited the activation of these molecules (Figure 5E), indicating that there is a crosstalk in the EGFR activated MAPK and NF-κB signal pathways. These results suggest that downregulating FUT4 expression decreased LeY biosynthesis which leads to the inhibition of EGFR activation, the downstream MAPK and NF-κB signal pathways in lung cancer cells
Cisplatin exhibits an anti-tumor effect by impairing the structure and function of DNA, whereas the action of pemetrexed is through regulating intracellular folatedependent metabolism and inhibiting cancer cell replication and proliferation
Summary
We examined EMT marker proteins, and found that E-cadherin expression was increased and N-cadherin expression was decreased by qPCR (Figure 4F), Western blot (Figure 4G) and immunofluorescent staining (Figure 4H) in shFUT4 transfected cells compared with the untreated control and mock transfected cells These results suggest that down-regulating FUT4 expression can inhibit migration, invasion and EMT in lung cancer cells. EMT marker proteins were examined, and the Rg3 treatment group had a higher expression of E-cadherin and a lower expression of Snail, N-cadherin, and Vimentin by Western blot (Figure 6F), and a stronger staining of E-cadherin and a weaker staining of N-cadherin by immunohistochemical staining (Figure 6G) compared with the control group These data showed that Rg3 significantly inhibited EMT in vivo. Tumor metastasis nodules in Lung tissues of Rg3 group were less and smaller than that control group after treatment for 2 months (Figure 6H)
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