Astragalus mongholicus Bunge and Curcuma aromatica Salisb. inhibits liver metastasis of colon cancer by regulating EMT via the CXCL8/CXCR2 axis and PI3K/AKT/mTOR signaling pathway
Authors: Liu Fuyan , Liang Yan , Sun Ruolan , Yang Weicheng , Liang Zhongqing , Gu Junfei , Zhao Fan , Tang Decai 
Published: August 2022

Abstract
Background
One of the most challenging aspects of colon cancer (CC) prognosis and treatment is liver-tropic metastasis. Astragalus mongholicus Bunge—Curcuma aromatica Salisb. (AC) is a typical medication combination for the therapy of many malignancies. Our previous studies found that AC intervention inhibits liver metastasis of colon cancer (LMCC). Nevertheless, the comprehensive anti-metastasis mechanisms of AC have not been uncovered.

Methods
In bioinformatics analysis, RNA-seq data of CC and LMCC patients were collected from TCGA and GEO databases, and differentially expressed genes (DEGs) were identified. The biological processes and signaling pathways involved in DEGs were enriched by GO and KEGG. The protein–protein interaction (PPI) network of DEGs was established and visualized using the Cytocape software, followed by screening Hub genes in the PPI network using Degree value as the criterion. Subsequently, the expression and survival relevance of Hub gene in COAD patients were verified. In the experimental study, the effects of AC on the inhibition of colon cancer growth and liver metastasis were comprehensively evaluated by cellular and animal models. Finally, based on the results of bioinformatics analysis, the possible mechanisms of AC inhibition of colon cancer EMT and liver metastasis were explored by in vivo and in vitro pharmacological experiments.

Results
In this study, we obtained 2386 DEGs relevant to LMCC from the COAD (colon adenocarcinoma) and GSE38174 datasets. Results of GO gene function and KEGG signaling pathway enrichment analysis suggested that cellular EMT (Epithelial-mesenchymal transition) biological processes, Cytokine-cytokine receptor interaction and PI3K/Akt signaling pathways might be closely related to LMCC mechanism. We then screened for CXCL8, the core hub gene with the highest centrality within the PPI network of DEGs, and discovered that CXCL8 expression was negatively correlated with the prognosis of COAD patients. In vitro and in vivo experimental evidence presented that AC significantly inhibited colon cancer cell proliferation, migration and invasion ability, and suppressed tumor growth and liver metastasis in colon cancer orthotopic transplantation mice models. Concomitantly, AC significantly reduced CXCL8 expression levels in cell supernatants and serum. Moreover, AC reduced the expression and transcription of genes related to the PI3K/AKT pathway while suppressing the EMT process in colon cancer cells and model mice.

Conclusions
In summary, our research predicted the potential targets and pathways of LMCC, and experimentally demonstrated that AC might inhibit the growth and liver metastasis in colon cancer by regulating EMT via the CXCL8/CXCR2 axis and PI3K/AKT/mTOR signaling pathway, which may facilitate the discovery of mechanisms and new therapeutic strategies for LMCC.