Design, Synthesis, and Biological Evaluation of Rapamycin Derivatives as the First Mammalian Target of Rapamycin and GLS1 Dual Inhibitors

The mammalian target of rapamycin (mTOR) and glutaminase 1 (GLS1) are key enzymes regulating metabolic reprogramming in breast cancer. The first generation of mTOR and GLS1 dual inhibitors was designed and synthesized on the basis of anticancer synergism. Compound <b>9d</b> showed selective and potent antiproliferative activity against all breast cancer cell lines and displayed potent inhibitory activity against both mTOR (mTORC1 and mTORC2) and GLS1. Mechanism studies revealed that <b>9d</b> effectively modulated the level of biomarkers and metabolites associated with mTOR and GLS1 inhibition and triggered sustained and massive reactive oxygen species generation, leading to cell death by autophagy, apoptosis, and ferroptosis. Moreover, <b>9d</b> inhibited the metastasis, invasion, and angiogenesis of breast cancer cells. In vivo experiments demonstrated that <b>9d</b> significantly inhibited tumor growth and metastasis, without observable toxicity. These findings proved mTOR/GLS1 dual inhibitors' great therapeutic potential for breast cancer.