Nox4 through mutant p53 induces a pro-inflammatory and pro-migratory secretome in the tumor microenvironment

The tumor microenvironment (TME) is a complex environment of tumor cells and resident stromal cells that are surrounded by secreted factors and extracellular matrix (ECM) proteins. Recent studies suggestreactive oxygen species (ROS) plays an important role in tumor progression and stromal cell activation.Previously, we showed wild-type (WT) and mutant (mt) forms of p53 differentially regulate ROS generation by NADPH oxidase-4 (NOX4). We found that WT-p53 suppresses TGF-β-induced NOX4, ROS production, and cell migration, whereas tumor-associated mt-p53 proteins enhance NOX4 expression and cell migration by TGF-β/SMAD3-dependent mechanisms. In this study, we investigated the role of mutant p53-induced NOX4 on the cancer cell secretome and the effect NOX4-derived ROS has on the TME. We found that conditioned media collected from H1299 lung epithelial cells stably expressing either mutant p53-R248Q or R273H promotes the migration and invasion of naive H1299 cells in scratch-wound repair and trans-well invasion assays, respectively. Furthermore, conditioned media from cells co-transfected with mutant p53 and dominant negative NOX4 (Nox4-DN) elicited diminished migratory behavior by naive H1299 cells. We utilized a Western blot-based cytokine array analysis to identify inflammatory factors in the H1299 mutant p53 CM that promote cell migration and invasion. We found CCL5 was significantly reduced in CM from H1299 cells expressing p53-R248Q and NOX4-DN. Collectively, our findings provide further insight into NOX4-based regulation of communication between cells in the tumor microenvironment and its potential as a therapeutic target affecting inflammation-driven metastatic disease.