The Wnt/β-catenin signaling pathway plays an important role in regulating mammary organogenesis and oncogenesis. However, therapeutic methods targeting the Wnt pathway against breast cancer have been limited. To address this challenge, we investigate the function of cyclin-dependent kinase 14 (CDK14), a member of the Wnt signaling pathway, in mammary development and breast cancer progression. We show that CDK14 is expressed in the mammary basal layer and elevated in triple negative breast cancer (TNBC). CDK14 knockdown reduces the colony-formation ability and regeneration capacity of mammary basal cells and inhibits the progression of murine MMTV-Wnt-1 basal-like mammary tumor. CDK14 knockdown or pharmacological inhibition by FMF-04-159-2 suppresses the progression and metastasis of TNBC. Mechanistically, CDK14 inhibition inhibits mammary regeneration and TNBC progression by attenuating Wnt/β-catenin signaling. These findings highlight the significance of CDK14 in mammary development and TNBC progression, shedding light on CDK14 as a promising therapeutic target for TNBC.Keywords: CDK14; CP: Cancer; Wnt/β-catenin signal; mammary stem cell; triple negative breast cancer.