A Mixture of Baicalein, Wogonin, and Oroxylin-A Inhibits EMT in the A549 Cell Line via the PI3K/AKT-TWIST1-Glycolysis Pathway.

Hui-Juan Cao,Xiao-Le Xian,Fu-Ling Tian,Pei-Jie Ding,Ting-Ting Fu,Chun-Yu Tian,Shu Zhao,Jian-Ye Dai,Chun-Hua Jiang,Wei Zhou,Shu-Jun Sun

doi:10.3389/fphar.2021.821485

Abstract

Non-small cell lung cancer (NSCLC) is a worldwide disease with a high morbidity and mortality rate, which is most derived from its metastasis. Some studies show that the epithelial–mesenchymal transition (EMT) process promotes lung cancer cell migration and invasion, leading to NSCLC metastasis. Total flavonoid aglycones extract (TFAE) isolated from Scutellaria baicalensis was reported to inhibit tumor growth and induce apoptosis. In this study, we found that baicalein, wogonin, and oroxylin-A were the active compounds of TFAE. After reconstructing with these three compounds [baicalein (65.8%), wogonin (21.2%), and oroxylin-A (13.0%)], the reconstructed TFAE (reTFAE) inhibited the EMT process of A549 cells. Then, bioinformatic technology was employed to elucidate the potential pharmacodynamic mechanism network of reTFAE. We identified the relationship between reTFAE and PI3K/Akt signaling pathways, with TWIST1 as the key protein. LY294002, the inhibitor of the PI3K/Akt signaling pathway, and knock-down TWIST1 could significantly enhance the efficacy of reTFAE, with increasing expression of epithelial markers and decreasing expression of mesenchymal markers in A549 cells at the same time. Furthermore, stable isotope dimethyl-labeled proteomics technology was conducted to complement the follow-up mechanism that the EMT-inhibition process may be realized through the glycolysis pathway. In conclusion, we claim that TWIST1-targeted flavonoids could provide a new strategy to inhibit EMT progress for the treatment of NSCLC.

Highlights

Lung cancer is a worldwide disease with a high morbidity and mortality rate
To explore reconstructed TFAE (reTFAE)’s effects in the epithelial–mesenchymal transition (EMT) process of lung cancer, A549 cells were treated with reTFAE (2, 4, and 8 μg/ml) for 24 h
The 0th hour, the wound was dose-dependently healed by reTFAE at the 48th hour (Figures 1E,F), which means that reTFAE inhibited the migration of A549 cells

Summary

Introduction

Lung cancer is a worldwide disease with a high morbidity and mortality rate. According to the World Health Organization (2020), there were 2.207 million new lung cancer patients worldwide in 2020, only less than breast cancer, and lung cancer ranked first in the global causes of cancer-related death (World Health Statistics, 2020; Thai et al, 2021). NSCLC microenvironment has limited nutrition, so cancer cells reTFAE Suppresses EMT of A549. The metabolic change leads to further tumor cell growth, aggravation of epithelial–mesenchymal transition (EMT) process, and resistance to treatment. During the treatment of NSCLC, 40% of patients have metastases (Dwyer-Nield et al, 2017), and the main biological process is EMT (Tulchinsky et al, 2018). Cellular EMT process is related to TGFR, Wnt/β-Catenin, PI3K/Akt, Ras, MAPK, NF-κB pathways, and the transcription factors involved are TWIST (TWIST1 and TWIST2), Slug, ZEB (ZEB1 and ZEB2), and Snail (Shaw and Solomon, 2011; Otsuki et al, 2018). Inhibition of TWIST1 was known to induce growth inhibition and apoptosis of EGFR-mutant NSCLC cells (Yochum et al, 2019)

Methods

Results

Conclusion