The Mechanism of Sijunzi Decoction Suppresses Gastric Cancer Metastasis via the m6A Methyltransferase METTL14 Based on Untargeted Metabolomics Studies and Network Pharmacology Analysis

Xiangnan Li,1,2,&ast; Linlin Zhao,3,&ast; Jiluan Wang,3 Tianchi Ma,1,2 Jing Zhou,1,2 Yue Bian,4 Junfu Guo1,2 1Teaching and Experiment Center, Liaoning University of Traditional Chinese Medicine, Shenyang, Liaoning, People’s Republic of China; 2Shenyang Key Laboratory for TCM Emotional Disorder, Liaoning University of Traditional Chinese Medicine, Shenyang, Liaoning, People’s Republic of China; 3College of Chinese Medicine Integrated with Western Medicine, Liaoning University of Traditional Chinese Medicine, Shenyang, Liaoning, People’s Republic of China; 4Department of Nursing, Shenyang Medical College, Shenyang, Liaoning, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Junfu Guo, Liaoning University of Traditional Chinese Medicine, No. 79 Chongshan Road, Huanggu Area, Shenyang, Liaoning Province, People’s Republic of China, Tel +86-024-31207185, Email [email protected]: Sijunzi Decoction (SJZ), a Traditional Chinese Medicine (TCM) formula, is renowned for its capacity to fortify Qi and enhance spleen function. However, additional research is necessary to comprehend the mechanisms beneath the therapeutic potential of SJZ in gastric cancer.Objective: This research endeavored to analyze how SJZ treats gastric cancer using network pharmacology and experimental validation.Methods: Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and network pharmacology were applied to systemically clarify the mechanism of SJZ against gastric cancer. We used a xenograft tumor model of gastric cancer and gastric cancer cell lines to explore the effect of SJZ on N6-methyladenosine (m6A) modification. Cell transfection, plate clone formation, scratch migration, and transwell assays were performed in gastric cancer cell lines. The expression levels of m6A enzymes and epithelial-mesenchymal transition (EMT) markers were assessed by Quantitative real-time reverse transcription (RT-qPCR) and Western blotting.Results: The results revealed 511 active components and 196 targets of SJZ, with 167 targets associated with gastric cancer therapy. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis disclosed notable enrichment of pathways related to cancer, metabolism, and immunity. The protein-protein interaction (PPI) network comprised 274 nodes and 2902 edges, whereas the herbal component-target protein-pathway-disease network included 107 nodes and 345 edges, identifying four components with more than 20 putative targets. Experimental assays demonstrated a significant decrease in METTL3 expression following SJZ treatment, whereas the expression level of METTL14 was markedly elevated in the SJZ group across both gastric cancer cell lines and gastric cancer tissues derived from a mouse model (P< 0.01, P< 0.001, or P< 0.05). SJZ inhibited clone formation, migration, and invasion of gastric cancer cells, and EGFR and Vimentin expression via METTL14 (P< 0.05, P< 0.01, or P< 0.001).Conclusion: METTL14 appears integral to the inhibition of EMT by SJZ as a treatment for gastric cancer.Keywords: Sijunzi decoction, gastric cancer, METTL14, network pharmacology