HIF-1α and HIF-2α Are Differentially Regulated In vivo in Neuroblastoma: High HIF-1α Correlates Negatively to Advanced Clinical Stage and Tumor Vascularization

Purpose: Hypoxia is considered to be a major driving force behind tumor angiogenesis. The stabilization and activation at hypoxia of the hypoxia-inclucible factors HIF-1 alpha and HIF-2 alpha and the concomitant induction of expression of vascular endothelial growth factor (VEGF) and other proangiogenic factors provide a molecular frame for hypoxiadriven tumor angiogenesis. This study has investigated how HIF and VEGF protein levels relate to each other with regard to vascularization, tumor stage, and overall survival in neuroblastoma. Experimental Design: Tissue cores taken from tumor specimens representing 93 children with neuroblastoma were arranged on a microarray and stained for HIF-1 alpha, HIF-2 alpha, VEGF, and CD31 proteins. Both fraction of positive cells and staining intensity were evaluated and protein levels were correlated with each other and with clinical variables. Results: Although high levels of both HIF-1 alpha (P < 0.001) and HIF-2 alpha (P < 0.001) correlated positively to VEGF expression, they did not fully correlate with each other. Moreover, HIF-1 alpha (P = 0.002) and VEGF (P < 0.001), but not HIF-2 alpha, correlated negatively to vascularization as determined by CD31 staining abundance. VEGF expression or degree of vascularization did not correlate with tumor stage or overall survival. High HIF-1 alpha levels correlated with low tumor stage (P < 0.001) and were associated with a favorable patient prognosis (P = 0.08). Conclusions: The discordant results on expression of HIF-1 alpha and HIF-2 alpha suggest that these two proteins are differentially regulated in vivo, thus reflecting distinctive protein expression/stabilization mechanisms. The association between HIF-1 alpha and favorable outcome stresses the importance of discriminating HIF-2 alpha from HIF-1 alpha expression and has implications for using HIFs as treatment targets. (Clin Cancer Res 2009; 15(23):7130-6)