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LED red light has been reported to have many health benefits. The present study was conducted to characterise anti-proliferation properties of four LED red light wavelengths (615, 630, 660 and 730 nm) against non-triple negative (MCF-7) and triple negative (MDA-MB-231) breast cancer-origin cell lines. It has been shown by MTT assay that at 24 h post-exposure time point, only LED red light with wavelength 660 nm possessed anti-proliferative effects against both cell lines with 40% reduction of cell viability. The morphology of LED 660 nm irradiated cells was found flatten with enlarged cell size, typical characteristic of cell senescent. Indications of autophagy activities following the irradiation have been provided by acridine orange staining, showing high presence of acidic vesicle organelles (AVOs). In addition, high LC3-II/LC3-I to LC3 ratio has been observed qualitatively in Western blot analysis indicating an increase number of autophagosomes formation in LED 660 nm irradiated cells compared to control cells. Electron dense bodies observed in these cells under TEM micrographs provided additional support to the above observations, leading to the conclusion that LED 660 nm irradiation promoted anti-proliferative activities through autophagy in breast cancer-origin cells. These findings have suggested that LED 660 nm might be developed and be employed as an alternative cancer treatment method in future.