Tumor suppressor ARF inhibits HER-2/neu-mediated oncogenic growth.

HER2/neu, a receptor tyrosine kinase oncogene, promotes mitogenic growth and antiapoptotic activity in cancer cells. Strong expression of HER2/neu in cancers has been associated with poor prognosis. Alternative reading frame protein (ARF), a tumor suppressor protein encoded by a gene located in the Ink4a/ARF gene locus, is frequently inactivated in human cancers. Little is known about the tumor suppressor role of ARF in HER2/neu-overexpressing cancers. Here, we applied the ARF gene as a tumor-suppressive agent for HER2/neu-overexpressing cells under the control of a tetracycline (tet)-regulated gene expression system. We found that ARF antagonized protein kinase B (PKB)/Akt-mediated p27Kip1 phosphorylation and increased p27 stability in HER2/neu-overexpressing cells. ARF expression also led to decreased levels of Cul1 and Skp2, two proteins involved in p27 degradation. We also found that ARF caused apoptosis in HER2/neu-overexpressing cells, and sensitized cells to apoptosis induced by the chemotherapeutic agents taxol and 2-methoxyestradiol. Most significantly for clinical application, we found that ARF inhibited HER2/neu-mediated cell growth, transformation, and tumorigenesis. These findings indicate that modulation of ARF activity may be a useful therapeutic intervention in HER2-overexpressing cancers.