Specific molecular events, characteristic of each cell-cycle phase may have direct effect to the functionality of nuclear receptors. Based on this understanding, the evaluation of lipophilic chemicals at the different cell-cycle phases is significant and should be considered. In order to achieve the aim of performing large-scale dose-response analysis on the effects of lipophilic chemicals at the different cell-cycle phases, a stable, sensitive and highly selective human progesterone receptor (hPR) expressing HeLa reporter cell line, hPRLuc-20, was established. Upon the establishment of the hPRLuc-20 cells, they were synchronized to the G(1), S and G(2) phases and treated with progesterone (PROG) and promegestone (R5020). The cells successfully showed that at the different cell-cycle phase, both agonists resulted in different cellular responses. The differences in response supports that hPR expressed within the hPRLuc-20 cells do respond in a cell-cycle dependent manner, thus showing the cells' compatibility in large-scale dose-response analyses of chemicals. It is hopeful that the advanced application of the hPRLuc-20 cells could contribute to provide fundamental hints to further understand the function of hPR, and provide key observations to elucidate the nature of these chemicals with hPR, its corresponding co-regulators and transcription factors.