Formestane (4-hydroxy-4-androstene-3,17-dione) is an anti-estrogenic drug used on the treatment of breast cancer. It is a type I aromatase inhibitor acting as a “suicide substrate”. In women it suppresses the estrogens formation and then can be used therapeutically in those pathologies that are estrogen dependent (i.e. breast cancer). In humans, estrogens are strong pituitary inhibitors of gonadotrophins releasing factors. The inhibition of the estrogens synthesis produces an increase of luteinizing hormone (LH) and then a net increase of testosterone production is observed. In addition the combined administration with testosterone and/or its precursors (i.e. androstenedione) will reduce the side effects linked to aromatization like gynecomastia. For these reasons, anti-estrogenic substances including aromatase inhibitors were included in 2004 in the World Anti Doping Agency (WADA) List of Prohibited substances in sports. The basic analytical methodologies developed so far are based on GC/MS or LC/MS, targeting formestane itself. It has been demonstrated that traces of formestane can be produced endogenously and found in urine samples in concentrations as low as 0.5-20 ng/mL and thus, since 2011, it is mandatory according to the WADA rules to perform a confirmation based on GC/C/IRMS in order to assess the synthetic origin of formestane for samples below 150 ng/mL before releasing an adverse analytical finding. Different IRMS methods have been already described in the literature, generally requiring two consecutive liquid chromatographic purifications (HPLC) before obtaining an extract of adequate purity for the subsequent IRMS analysis. The metabolism of formestane has been described in a single male volunteer. It appears that among the high number of metabolites described, 3b,4a-dihydroxy-5a-androstan-17-one (4a-hydroxy-epiandrosterone; 4OH-EA) has a longer detection window. We propose a global mass spectrometric approach for the confirmation of the exogenous origin of formestane in urine samples. Data obtained by the in house GC/C/IRMS and GC/MS/MS developed methods will be discussed.