Inhibitory Effect Of 5-FU/EGCG Combination On The Growth Of Gastric Carcinoma Cells

Increasing studies suggest that chemotherapeutics in combination with natural products can not only increase the chemosensitivity of tumor cells,but also reduce the toxic and side effects of drugs at high doses,demonstrating a promising strategy in cancer chemotherapy.Gastric cancer contributes significantly to cancer-related deaths worldwide,and is ranked second in cancer incidence and mortality in China.5-Fluorouracil(5-FU)is a basic medicine for treatment of gastrointestinal malignant tumors,but some patients have shown severe adverse effects.Epigallocatechin gallate(EGCG)is one of the main phenolic contents of Camellia sinensis and has shown strong antioxidiant,anti-tumor and other activities.To evaluate the synergistic anti-tumor effect of 5-FU and EGCG combination,we studied here the effects of 5-FU,EGCG and 5-FU/EGCG combination on proliferation,apoptosis,cell cycle and related signaling pathways using human gastric cancer cell lines AGS and SGC-7901.Then we analyzed the molecular network of lncRNA and mRNA using gene chips and bioinformatics.We show that both 5-FU and EGCG were able to inhibit the proliferation of both gastric cancer cell lines in a dose-dependent manner albeit AGS was more sensitive than SGC-7901 to 5-FU.Compared with 5-FU or EGCG alone,5-FU/EGCG combination was more effective to inhibit the proliferation of the gastric cancer cells and to induce cell apoptosis.As revealed by quantitative real-time PCR and Western blotting,the expression level of CHOP,a specific marker of endoplasmic reticulum stress response,was significantly increased in the cells treated with 5-FU/EGCG combination,while that of the inflammation marker COX-2 was dramatically reduced.To further investigate the anti-tumor mechanisms of 5-FU/EGCG combination in gastric cancer cells,gene chip analysis was used to study IncRNA and mRNA expression profiles using the SGC-7901 cells.In total,28,320 lncRNAs and 28,416 mRNAs were found differentially expressed in cells treated with 5-FU/EGCG combination.Meanwhile,candidate genes involved in the development of gastric cancer were unravelled using bioinformatic approaches against TCGA database.Results from the experimental data as well as from the database unravelling reveal that 85 coding genes and 11 lncRNAs related to the development of gastric cancer were affected by the treatment of 5-FU/EGCG combination.GO gene function enrichment analysis suggests that the biological processes involving the function of 5-FU/EGCG combination include nucleic acid metabolism,cytoskeleton,actin filament polymerization and proteolysis,while KEGG pathway enrichment analysis reveals that the main pathways involved were PI3K-Akt,MAPK and mTOR signaling pathways.To predict the target genes associated with the identified coding genes and IncRNA,an lncRNA-miRNA-Gene regulatory network map was constructed,which reveals that lncRNA NR2F1-AS1 was either directly or indirectly related to multiple target genes and miRNAs,implying that NR2F1-AS 1-linked molecular networks may play an important role in the inhibitory effect of 5-FU/EGCG on gastric cancer cells.A regulatory relationship diagram between drugs and genes was also constructed,showing that both 5-FU and EGCG may have direct effects on IGF1R and SERPINE1.In conclusion,this study reveal that 5-FU/EGCG combination was effective on inhibition of cell proliferation and induction of apoptosis in gastric cancer cells,and its functions were associated with nucleic acid metabolism,cytoskeleton,proteolysis and other biological processes as well as with PI3K-Akt,MAPK and mTOR signaling pathways.These results provide an important insight into the potentials of 5-FU/EGCG combination as a high-efficiency and low-toxicity chemotherapeutic strategy.