Abstract A33: EphA2 Receptor: Evidence for a proto-oncogenic role in melanoma

Abstract

Abstract In nature, the cells are under constant physiological stress imposed by local microenvironment as well as by endogenous oxidative processes. In order to counteract these insults, the cells have evolved a complex network of signaling molecules to elicit appropriate responses. Ultraviolet Radiation (UVR) is a most ubiquitous source of environmental stress on the human skin and the most lethal consequence of this exposure is malignant melanoma. The propensity for the exposed cell to become an oncogene depends on how quickly it would respond to the stress (early events) and how the response would lead to oncogenic induction (late events). In our previous study, we set out to identify UVR-responsive genes in pigment cells that may function as candidate oncogenes or tumor suppressor genes in melanomas. We discovered that a tyrosine kinase receptor, EphA2 upregulated by UVR preferentially over other related family of Ephrin receptors. We also found that EphA2 acts as a pro-apoptotic gene in a p53 independent, MAPK dependent mechanism in normal human melanocytes. Interestingly, the role of EphA2 in tumor physiology albeit well described is highly complex with increasing evidence pointing to its role as a proto-oncogene. Hence, the balance between the oncogenic and apoptotic effects of EphA2 likely depends on the cellular context and the unique pathways that are activated. On account of this dual behavior exhibited by EphA2, we hypothesized that EphA2 might be a key modulator during the transformation stage of the tumor cells. Hence, we set out to address our hypothesis through a set of molecular genetic and functional approaches to understand the mechanistic role of EphA2 in melanoma tumorigenesis. We selected a set of melanoma cell lines with high and low levels of endogenous EphA2. We overexpressed EphA2 in the low expresser cell lines using eukaryotic expression plasmid which was stably selected for high EphA2 expression. The phenotypic changes were evaluated through proliferation assay, migration and ability to form colonies in three dimensional cultures. In parallel, we knocked down EphA2 expression in high EphA2 expresser cell lines through Lentiviral shRNA system. The phenotypic changes are evaluated by migration and apoptotic assays. In the study presented here, we have shown evidence for EphA2's role as a proto-oncogene in the induction of melanoma tumorigenesis. When EphA2 was ectopically and stably overexpressed in low EphA2-expressing cell line, we observed an increase in proliferation rate, increase in migration and also exhibited increased invasive phenotype in three dimensional organotypic cultures. When EphA2 expression is knocked-down by shRNA approach in high EphA2-expressing line, we saw an increase in apoptotic response, and a significant decrease in the migration potential of the knocked-down cells. Also, our preliminary evidence suggests that EphA2 regulatory role is mediated through FAK signaling in high EphA2 expressing melanoma cells. These studies strongly suggest that EphA2 is a proto-oncogene in melanoma tumorigenesis. Our further ongoing attempt to underpin the mechanistic pathways underlying these observed phenotypic changes, in complement with our previous findings on EphA2 as UV inducible gene, would provide additional pathways for therapeutic intervention to tackle the challenges in the treatment of metastatic melanoma. Citation Information: Cancer Res 2009;69(23 Suppl):A33.