Humans and wildlife are continuously and simultaneously exposed to various pesticides that have been identified as endocrine disruptors which interfere with regulations of sexual differentiation and fertility. Low-dose effects of combined exposure from mixtures of pesticides have been extensively reported and need to be addressed in the context of human health risk assessment. The objective of the study is to assess the individual and combined anti-androgenic effects of twelve widely used pesticides in MDA-kb2 cells. The order of potency for seven pesticides with moderate anti-androgenic activities was in the order: fenitrothion > dimethomorph > difenoconazole > bromopropylate > prochloraz > imazalil > endosulfan, which was induced by the androgen receptor (AR) antagonism rather than cytotoxicity (with the exception of endosulfan which exhibited the highest cytotoxicity). The other five pesticides exhibited lower anti-androgenic activities. At 10% of AR antagonistic effect, three mixtures comprised of the seven pesticides (Mix-EC10, Mix-EC20, and Mix-EC25) at equi-effect concentrations showed summed concentrations of 6.75E-11, 17.63 and 25.21 μM, respectively. The combined effects were essentially close to the predicted of concentration addition (CA) at realistically low concentrations. In addition, molecular docking simulation indicated that hydrophobic interaction and polar functional groups of the pesticides contributed to the binding energy, which might be responsible for the AR antagonism. Our findings provide a basis for defining similarly acting antagonists in the context of cumulative risk assessment for pesticides in foods.