Utrophin (UTRN), known as a tumor suppressor, potentially regulates tumor development and the immune microenvironment. However, its impact on breast cancer's development and treatment remains unstudied. We conducted a thorough examination of UTRN using both bioinformatic and in vitro experiments in this study. We discovered UTRN expression decreased in breast cancer compared to standard samples. High UTRN expression correlated with better prognosis. Drug sensitivity tests and RT-qPCR assays revealed UTRN's pivotal role in tamoxifen resistance. Furthermore, the Kruskal-Wallis rank test indicated UTRN's potential as a valuable diagnostic biomarker for breast cancer and its utility in detecting T stage of breast cancer. Additionally, our results demonstrated UTRN's close association with immune cells, inhibitors, stimulators, receptors, and chemokines in breast cancer (BRCA). This research provides a novel perspective on UTRN's role in breast cancer's prognostic and therapeutic value. Low UTRN expression may contribute to tamoxifen resistance and a poor prognosis. Specifically, UTRN can improve clinical decision-making and raise the diagnosis accuracy of breast cancer.