Inhibition of Pim Kinase Supports the Neural Stem Cell Phenotype in Neuroblastoma

Abstract

INTRODUCTION: Relapse rates for high-risk neuroblastoma exceed 50%. Expression of the serine-threonine protein kinase, PIM, is associated with cancer progression and is negatively correlated with neural stemness markers in neural progenitor cells. Because promoting cellular differentiation is a component of neuroblastoma therapy, and PIM kinase negatively affects neural stemness, we aimed to determine whether PIM affected the neural stem cell phenotype in neuroblastoma. METHODS: We queried the Kocak (R2) gene database to compare PIM1 and the neural stemness marker, Nanog, in neuroblastoma. For in vitro studies, SK-N-BE(2), human neuroblastoma cells and human neuroblastoma patient–derived xenograft (PDX06) cells were treated with siRNA to achieve PIM1 kinase knockdown or with AZD1208 for PIM inhibition. Abundance of mRNA for the neural stemness marker, Nanog, was detected with quantitative reverse transcription polymerase chain reaction, and protein expression was confirmed with immunoblotting. Flow cytometry detected cell surface expression of CD133 and CXCR4, 2 additional neural stemness markers. Data were reported as mean ± SEM, with p ≤ 0.05 considered significant. RESULTS: R2 database revealed a negative correlation between PIM1 and Nanog (Fig. 1A). Protein expression of Nanog was increased following siRNA knockdown of PIM1 (Fig. 1B). AZD1208 increased the abundance of Nanog mRNA and resultant protein expression (Fig. 1C). AZD1208 treatment significantly increased cell surface expression of CD133 and CXCR4 (Fig. 1D).Figure 1CONCLUSION: Neuroblastoma cells treated with PIM inhibition dedifferentiated to a more neuronal stem cell phenotype as demonstrated by increased expression of neuronal stemness markers. These findings support further studies exploring the clinical implications of PIM kinase inhibition as an adjunct to neuroblastoma therapy.