Effect of Gubenyiliu formula II and its disassembled prescriptions on cell autophagy in breast cancer through PI3K/AKT/mTOR pathway

Abstract

The aim of this study is to reveal the mechanism of Gubenyiliu II (GYII) inhibiting autophagy in breast cancer and the effect of its disassembled prescriptions, Quxie (QX) and Fuzheng (FZ), which cause autophagy difference on tumor growth. After a breast cancer in situ tumor model was established, mice were randomly distributed into different groups: model, GYII, QX, FZ and tamoxifen groups, and treated correspondingly. Then, the tumor volumes and weights were monitored. Immunohistochemistry detected the contents of microtubule-associated protein light chain 3 (LC3), phosphorylated phosphatidylinositol 3-kinase (p-PI3K), phosphorylated protein kinase B (p-AKT) and phosphorylated mammalian target of rapamycin (p-mTOR) in tumor tissues. Furthermore, 4T1 cells were administrated with the 20% contained serum. Cell proliferation, migration and invasion were measured using cell counting kit-8 and transwell assays. Electron microscopy and flow cytometry detected autophagy and apoptosis. The content of LC3 was measured by immunofluorescence. Western blot detected the protein levels of LC3, Beclin1, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR in tumor tissues and 4T1 cells. GYII, QX and FZ treatment significantly reduced the tumor volumes and weights in breast cancer tumor-bearing mice. The cell proliferation, migration and invasion were restrained, and cell apoptosis and autophagy were promoted in GYII, QX and FZ groups. Moreover, GYII, QX and FZ increased the expression of LC3 in 4T1 cells and tumor tissues and decreased the phosphorylation levels of PI3K, AKT and mTOR in tumor tissues. The protein levels of LC3 and Beclin1 were upregulated, and p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR were downregulated in tumor tissues and 4T1 cells of treatment groups. Our study confirmed that GYII could treat breast cancer by restraining the PI3K/AKT/mTOR signaling pathway-mediated autophagy. While QX focuses on inhibiting tumor growth, FZ acts on inhibiting tumor metastasis.