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Fatty acid synthase (FASN) is a key enzyme involved in fatty acid biosynthesis and serves an important role in breast cancer development. The aim of the present study was to investigate the effects of patuletin on the gene expression and activity of FASN in the human breast cancer SK-BR-3 cell line, and the apoptotic effects of patuletin to breast cancer cells. Quantitative reverse transcription polymerase chain reaction, western blotting and intracellular FASN activity assays were used to evaluate FASN gene expression, protein expression and activity in patuletin-treated SK-BR-3 cells. MTT assays and flow cytometry were used to measure cell growth and cell apoptosis, respectively, following patuletin treatment. As a result, it was demonstrated that patuletin dose-dependently reduces FASN expression and intracellular activity in human breast cancer cells, and induces apoptosis in FASN over-expressing SK-BR-3 cells. Notably, apoptosis is associated with the reduction of intracellular FASN activity. The present study demonstrates that patuletin may be considered as a novel natural inhibitor of FASN, may induce anti-proliferative and pro-apoptotic effects in certain human breast cancer cells and may be useful for preventing and/or treating human breast cancer.