Metabolic reprogramming underlying the synergistic effect of energy metabolism inhibitors and chemotherapeutics in TNBC

Triple-negative breast cancer (TNBC) is characterized by aggressive behavior and metabolic plasticity, limiting the efficacy of standard chemotherapy. Targeting metabolic vulnerabilities presents a promising strategy to overcome chemoresistance. This study investigates the synergistic efficacy and underlying mechanisms of combining energy metabolism inhibitors with chemotherapeutic agents in MDA–MB–231, a representative TNBC cell line. Through cell viability screening, we identified two potent synergistic regimens: gemcitabine with metformin and 5-fluorouracil with 3-bromopyruvate. Real-time bioenergetic profiling demonstrated that these combinations significantly disrupted the energy metabolic phenotypes of these cells. To elucidate the molecular mechanisms, we employed Ultra-Performance Liquid Chromatography-Quadrupole Time-of-Flight tandem Mass Spectrometry (UPLC-Q-TOF-MS/MS) metabolomics coupled with stable isotope-labeled metabolite quantification. Our data revealed that metabolic inhibition sensitized cells to chemotherapy by inducing profound reprogramming of amino acid and lipid metabolism, tightly linked to energy deprivation. Pathway analysis confirmed that the therapeutic efficacy relies on the specific modulation of amino acid pathways associated with central carbon metabolism. Collectively, this study provides a mechanistic rationale for integrating energy metabolism inhibitors with chemotherapy, offering novel therapeutic strategies to enhance clinical outcomes in TNBC.