The prostanoid EP4 receptor belongs to the family of G protein-coupled receptors and it has been reported that the signaling pathway mediated by the EP4 receptor contributes to the cancer development. The cytoplasmic region of the EP4 receptor has several characteristic sites that are significantly different from other prostanoid receptors. In this study, we focused on one amino acid and examined the effects of point mutation on EP4 receptor signaling pathway. Thus, the point mutation-introduced EP4 receptor-stably expressing HEK cell line was prepared, and comparative experiment were conducted with wild-type EP4 receptor-expressing cell line. As a result, constitutive cAMP production and ligand-independent cell proliferation retardation were observed in the mutant EP4 receptor-expressing cell line when compared with wild-type EP4 receptor expressing cell line. These results suggested that the introducing only one mutation of the specific amino acid of the EP4 receptor would have a possibility to suppress the EP4 receptor-initiated cancer development by delaying the growth of cells.