The role of the p38-FOXM1 pathway in the regulation of cancer cell invasion

Abstract

Head and neck cancer is the sixth most common cancer worldwide and associated with a poor clinical prognosis, due to development of recurrent tumors and metastasis. Tumor recurrence and low patient survival are strongly linked with the ability of tumor cells to invade and infiltrate the surrounding tissue. Stress-activated protein kinases (SAPK), particularly p38, are known to regulate a wide range of cellular phenotypes, including cell invasion via the activity of secreted proteases. The proliferation-associated Forkhead box protein M1 (FOXM1) transcription factor, a p38 downstream target, plays a role in the development and growth of many cancer types. However, only very little is known about the role of p38 and FOXM1 in invasive processes of head and neck cancer and the exact mechanism underlying this process. In this work we examined the downstream events of p38 signaling primarily focusing on the role of FOXM1 transcription factor in regulation of the urokinase-type plasminogen activator (uPA) gene and invasion of head and neck squamous cell carcinoma (HNSCC) cells. Using different HNSCC cell lines, we confirm that p38 regulates FOXM1 expression and provide evidence that p38 signaling driven in vitro invasion of HNSCC cells requires FOXM1 expression. Furthermore, siRNA-mediated FOXM1 knockdown is sufficient to inhibit the invasive behavior of HNSCC cells in vitro. By using reporter gene assays, bioinformatical analysis of the publically available ChIP-Seq data, chromatin immunoprecipitation assays, and transplantation-based mouse model of oral cancer, we identified the molecular mechanism of FOXM1-mediated invasion of HNSCC cells. FOXM1 controls the uPA-dependent invasion via activation of c-Fos and thus drives AP-1 activity on the uPA promoter, which enhances its expression and proteolytic activity. Further, an activated Ras signaling is necessary for a potent FOXM1-mediated uPA activity and tumor formation. The data are supported by a bioinformatical study, demonstrating concomitant up-regulation of FOXM1 and uPA in oral dysplasia and SCCs of head and neck, oesophagus, lung and cervix. In the mouse model of oral cancer we show that uPA expression is upregulated in recurrent tumors compared to primary tumors, giving further evidence for a crucial role of the p38-FOXM1-uPA axis in the development of recurrent tumors. Taken together, we conclude that the stress signalling cascade requires a FOXM1-dependent intermediate step preceding the activation of AP-1 transcription factor to enhance invasive behaviour of tumor cells. This novel mechanism promotes invasion of HNSCC and may provide a potential target for the adjuvant therapy of these highly invasive cancers.