The primary objective was to identify the relationship between tramadol and breast cancer through the evaluation of EMT-associated biomarkers [hypoxia-inducible factor 1 alpha (HIF-1α)] and to examine whether tramadol treatment, in a normoxic or hypoxic microenvironment, affects the expression of HIF-1α, stress-induced reactive oxygen species and EMT- and cyclin-related proteins in the human breast adenocarcinoma cell lines MDA-MB-231 and MCF-7. Here, HIF1A is linked to breast adenocarcinoma.