MCPB-21 targets GYG2 to regulate fatty acid oxidation and promote ferroptosis of breast cancer

This study systematically evaluated the anti-breast cancer potential and mechanisms of the cannabidiol (CBD) derivative MCPB-21. The structure of MCPB-21 was confirmed via nuclear magnetic resonance (NMR) following chemical synthesis. Differential gene analysis of breast cancer was conducted using public databases, and molecular docking demonstrated MCPB-21's binding affinity to the GYG2 protein. Experiments on MDA-MB-231 and MCF-7 breast cancer cells assessed MCPB-21's impact on apoptosis, invasion, and lipid changes through flow cytometry, Transwell invasion assays, cell proliferation, and Oil Red O staining. Western blotting (WB) was employed to examine expression changes in proteins related to fatty acid β-oxidation and ferroptosis, including ACOX1, ABCD3, ABCD4, EHHADH, CPT1α, GPX4, SLC7A11, and ACSL4. The role of fatty acid oxidation in ferroptosis was further analyzed using the ACOX1 inhibitor 10,12-Tricosadiynoic acid. Additionally, the effects of MCPB-21 on fatty acid oxidation and ferroptosis were evaluated by interfering with GYG2 expression. Finally, the anti-tumor efficacy of various doses of MCPB-21 was compared to the control drug CBD. In vitro experimental results showed that MCPB-21 can affect the behavior of breast cancer cells by inducing cancer cell apoptosis, increasing reactive oxygen species (ROS) levels, and promoting lipid accumulation. At the same time, Western blot detection showed that MCPB-21 could downregulate key enzymes of fatty acid β-oxidation (such as ACOX1, ABCD3, ABCD4, EHHADH, CPT1α) and antioxidant factors (GPX4, SLC7A11), and upregulate the enzyme ACSL4 that promotes lipid peroxidation. Mechanistic studies further showed that MCPB-21 affects the expression of ACOX1 by regulating GYG2, inhibits fatty acid β-oxidation, and induces ferroptosis. At the same time, the combined use of ACOX1 inhibitors enhanced lipid accumulation and ROS levels, verifying its role in regulating fatty acid oxidation. In animal experiments, MCPB-21 (10 and 40 mg/kg) significantly inhibited the growth of nude mouse transplanted tumors, caused tumor tissue necrosis, inhibited the proliferation marker Ki67, and regulated the expression of ferroptosis-related proteins (GPX4 and SLC7A11 decreased, and ACSL4 increased). Immunohistochemical analysis showed that MCPB-21 had a stronger anti-tumor effect than CBD, mainly by regulating the fatty acid β-oxidation pathway to promote ferroptosis. In summary, MCPB-21 exhibits excellent anti-breast cancer potential. Its mechanism of action is mainly to achieve anti-tumor effects by inhibiting fatty acid β-oxidation and activating ferroptosis, which provides a theoretical basis and potential therapeutic strategy for the development of new anti-breast cancer drugs.