Detachment-activated FAK-STAT3-NNMT inhibits anoikis by enhancing fatty acid oxidation in breast cancer

Abstract

Purpose: Breast cancer metastasis claims the majority of breast cancer-related deaths. Anoikis resistance is a key prerequisite for tumor metastasis. Possible mechanism of evading anoikis in breast cancer remains unclear. Our previous research demonstrated that Nicotinamide N-methyltransferase (NNMT) has essential functions in cancer development and metastasis. However, the spercific functional mechanisms of NNMT in breast cancer anoikisp remain to be explored Method: Immunofluorescence staining, confocal microscopy analysis, immunoblotting and qPCR were used to analyze NNMT expression. Flow cytometry and soft agar assay were used to analyze the proportion of breast cancer cells undergoing anoikis. ChIP assay and detached cell model were used to confirm the presence of FAK-STAT3-NNMT in detached breast cancer cells. BODIPY staining was used to visualize lipid deposit in the cytoplasm, and Seahorse analysis was used to directly measure FAO rate of attached and detached breast cancer cells. Results: Detachment of breast cancer cells activates the FAK-STAT3 axis to increase the transcription of NNMT, which then protects breast cancer cells from anoikis by enhancing fatty acid oxidation (FAO). Mechanistically, NNMT promotes the expression of CPT1A, a FAO rate-limiting gene, by suppressing PP2A methylation. Furthermore, NNMT-induced FAO accelerates the clearance of reactive oxygen species (ROS), which triggers anoikis and hinders cancer metastasis, by maintaining NADP+/NADPH balance. In vivo experiments show that NNMT-knockdown breast cancer cells colonize the lung much less than that of NNMT high expression cells. Additionally, inhibitors of NNMT or FAO suppressed breast cancer cell metastasis, suggesting that NNMT-mediated FAO enhances the metastatic potential of breast cancer. Conclusion: Our study identifies that NNMT, an enzyme upregulated during detachment via the FAK-STAT3 axis, activates the FAO pathway to resist anoikis of breast cancer cells.