Background: Triple-negative breast cancer (TNBC) is clinically characterized by the absence of therapeutic targets and limited treatment options after surgery, leading to poor therapeutic outcomes and prognosis. Shuyu Pills (SYP), a classical traditional Chinese medicine (TCM) formula for treating deficiency syndromes with pathogenic excess, has been widely applied in the treatment of TNBC. However, its mechanisms of action against TNBC remain unclear. Purpose: This study aimed to investigate the anti-tumor effect and the underlying mechanisms of SYP in combination with the mTOR inhibitor everolimus in TNBC both in vitro and in vivo experiments. Methods: Network pharmacology was used to predict the potential signaling pathways targeted by SYP. The pharmacological effects of SYP and its combination with everolimus were evaluated through in vitro and in vivo experiments, including CCK-8 assay, transwell migration, wound healing assay, flow cytometry, Immunohistochemistry staining and Western blot analysis. Results: The combination of SYP and everolimus exhibits synergistic effects in inhibiting TNBC. Synergistic therapy suppressed TNBC cell proliferation and colony formation, cell migration and invasion, induced cell cycle arrest and apoptosis and reduced tumor growth in TNBC-bearing mice. The addition of SYP significantly promoted apoptosis of TNBC cells via regulated apoptosis-related protein including Bax, Caspase3, Bcl-2, CDK1 and Cyclin B. Further, synergistic treatment reduced TNBC tumor growth and Ki67 expression. Furthermore, the combined therapy effectively suppresses the expression of PI3K, p-PI3K, AKT, p-AKT, mTOR, and p-mTOR proteins. Conclusion: The present study aimed to investigate the pharmacological mechanism of the SYP. SYP can inhibit the proliferation, migration, and invasion of TNBC cells by arresting the cell cycle and inducing apoptosis. This mechanism may involve downregulation of the PI3K/AKT/mTOR signaling pathway. SYP can cooperate with everolimus to inhibit TNBC. Overall, these findings provide valuable evidence for the potential clinical application of SYP. Keywords: triple-negative breast cancer, cell cycle, apoptosis, pi3k/akt/mtor signaling pathway, shuyu pills, everolimus
Zhao et al. (Sun,) studied this question.