Cancer is a complex disease that requires multidisciplinary treatment. One of the most common cancers is breast cancer, and these cancers are usually estrogen receptor-positive. Today, chemotherapy, as well as surgery, has an important place in the treatment of cancer. For this reason, serious research is carried out on the search for new chemotherapeutic drugs. Some of the drugs currently used in breast cancer are targeted at the estrogen receptor. As a result of the preliminary analyses carried out within our study's scope, five new diphenylurea derivative compounds that we predict will be targeted to the estrogen receptor were determined and synthesized. Chemical analyzes of the synthesized compounds were performed. In addition, in vitro analyzes were performed for their biological effects. In this context, it was determined that DPU2, one of the compounds applied to the estrogen receptor (+) breast cancer cell line, MCF-7, showed the cytotoxic effect. The IC50 value was determined, and it was determined that although it showed a cytotoxic effect on MCF-7 cells, it did not show severe cytotoxicity in healthy fibroblast cells. Then, LC3B, one of the autophagy indicators, and IL-1β, a proinflammatory cytokine, were examined immunohistochemically. Both LC3B and IL-1β expression were lower in the DPU2 applied group than in the control group. These findings supported that DPU2 has antitumor activity on MCF-7. In conclusion, in the light of in silico and in vitro analyses conducted within the scope of our study, it was determined that DPU2, one of the five phenyl urea derivative compounds I synthesized, has the potential to be a drug in the treatment of estrogen receptor (+) breast cancer. However, the study needs to be supported by in vivo and clinical studies.