Recent studies have implicated a role for midkine (MK) in cancer progression. This is based upon its structural homology with pleiotrophin, an angiogenic growth factor, and its ability to enhance fibrinolytic activity of bovine endothelial cells. To investigate whether MK plays a role in breast cancer, we examined MK mRNA expression in N-nitroso-N-methylurea-induced rat mammary tumors at various stages of tumor progression, including hormone independence and distant metastasis. Well-differentiated mammary adenocarcinomas showed levels of MK comparable to those of normal mammary gland. A 10- to 20-fold reduction in MK mRNA levels was observed in mammary tumors that had progressed to hormone independence and metastasis. The data suggest that loss of MK expression correlates with breast tumor progression. Treatment of rat mammary tumor cell lines with retinoic acid increased MK expression, decreased proliferation, and markedly reduced colony-forming efficiency in agar. This raises the possibility that agents that upregulate MK could have potential in prevention and therapy by causing breast cells to terminally differentiate.