16α-Hydroxydehydrotrametenolic acid attenuates MDA-MB-231 triple-negative breast cancer xenograft growth and modulates integrin β1/c-Met signaling and glycolytic metabolism.

BACKGROUND: Abnormal glycolysis, enhanced metastatic potential, and dysregulated oncogenic signaling, including integrin β1 and c-Met pathways, are key drivers of triple-negative breast cancer (TNBC) progression and contribute to its poor prognosis and limited therapeutic options. 16α-Hydroxydehydrotrametenolic acid (HDTRA) is a candidate therapeutic agent, and our previous work suggested its potential antitumor activity. PURPOSE: We aimed to evaluate the therapeutic potential of HDTRA in MDA-MB-231 TNBC cells by characterizing its modulatory effects on integrin β1 and c-Met signaling and clarifying the associated molecular mechanisms, including inhibition of cell migration and invasion in vitro, induction of apoptosis, and cell cycle arrest, and disruption of abnormal glycolysis. These findings warrant further preclinical investigation of HDTRA in the MDA-MB-231 model. STUDY DESIGN: This research combined in vitro MDA-MB-231 functional assays, in vivo subcutaneous xenograft models, quantitative proteomic screening, target-rescue validation, and computational molecular simulation. Different concentrations of HDTRA were applied in cellular and animal models to assess antitumor efficacy, followed by multi-omics pathway analysis, target agonist rescue experiments, molecular docking, and molecular dynamics simulations to explore the underlying mechanism. METHODS: MDA-MB-231 TNBC cells and xenograft models were used to evaluate the in vitro and in vivo efficacy of HDTRA. Quantitative proteomic analysis was performed to identify global HDTRA-induced proteomic changes, followed by multi-dimensional functional annotation and weighted gene co-expression network analysis to define core modulated pathways. Functional assays were conducted to confirm the biological effects of HDTRA. Rescue experiments using specific agonists of integrin β1 and c-Met were performed to verify target involvement. Molecular docking and molecular dynamics simulations were applied to predict potential interactions between HDTRA and the two proteins. RESULTS: HDTRA exerted potent antitumor activity in vitro and in vivo, significantly inhibiting MDA-MB-231 cell proliferation, migration, and invasion, while inducing apoptosis and G2/M cell cycle arrest, with no overt systemic toxicity observed under the tested conditions. Quantitative proteomics suggested HDTRA may modulate a subset of cancer-related pathways, associated with altered glycolytic metabolism and time-dependent modulation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression. Mechanistically, HDTRA suppressed integrin β1 and c-Met signaling, thereby inhibiting downstream FAK/PI3K/AKT and c-Met-associated cascades. Rescue experiments further showed that restoring integrin β1 or c-Met activity significantly reversed the antitumor effects of HDTRA. CONCLUSION: This study identifies a multi-mechanistic antitumor action of HDTRA in MDA-MB-231 TNBC through suppression of integrin β1 and c-Met-mediated signaling, inhibition of migration and invasion in vitro, induction of apoptosis and cell cycle arrest, and association with altered glycolytic metabolism. These findings indicate antitumor activity of HDTRA in the MDA-MB-231 TNBC model and warrant further preclinical investigation.