PBX1 Is a Favorable Prognostic Biomarker as It Modulates 13-cis Retinoic Acid–Mediated Differentiation in Neuroblastoma

Abstract

Purpose Neuroblastoma is an embryonic childhood cancer with high mortality. 13-cis retinoic acid (13-cisRA) improves survival for some patients, but many recur, suggesting clinical resistance. The mechanism of resistance, and the normal differentiation pathway, are poorly understood. Three-Amino-acid Loop Extension (TALE) family genes are master regulators of differentiation. Since retinoids promote differentiation in neuroblastoma, we evaluated TALE family gene expression in neuroblastoma. Experimental Design We evaluated expression of TALE family genes in RA-sensitive and -resistant neuroblastoma cell lines, with and without 13cis-RA treatment, identifying genes whose expression correlate with retinoid sensitivity. We evaluated the roles of one gene, PBX1, in neuroblastoma cell lines, including proliferation and differentiation. We evaluated PBX1 expression in primary human neuroblastoma samples by RT-qPCR, and three independent clinical cohort microarray datasets. Results We confirmed induction of PBX1 expression, and no other TALE family genes, was associated with 13-cisRA responsiveness in NB cell lines. Exogenous PBX1 expression in neuroblastoma cell lines, mimicking induced PBX1 expression, significantly impaired proliferation and anchorage-independent growth, and promoted RA-dependent and -independent differentiation. Reduced PBX1 protein levels produced an aggressive growth phenotype and RA resistance. PBX1 expression correlated with histological neuroblastoma subtypes, with highest expression in benign ganglioneuromas and lowest in high-risk neuroblastomas. High PBX1 expression is prognostic of survival, including in multivariate analysis, in the three clinical cohorts. Conclusions PBX1 is an essential regulator of differentiation in neuroblastoma and potentiates retinoid-induced differentiation. Neuroblastoma cells and tumors with low PBX1 expression have an immature phenotype with poorer prognosis, independent of other risk factors.

Preclinical Evaluation of a Novel RXR Agonist for the Treatment of Neuroblastoma

“…2A, closed arrows ). The UAB30-induced neurite outgrowth was quantified by counting the number of neurite outgrowths per cell [11] and reporting as fold change, and UAB30 led to significantly increased neurite outgrowths in all cell lines (Fig. 2B).…”

“…Pictures [Photometrics CoolSNAP HQ2 CCD camera (Tucson, AZ) attached to a Nikon Eclipse Ti microscope (Tokyo, Japan)] were obtained and the number of neurite outgrowths per cell were counted and reported as fold change neurite outgrowths [11]. …”

Achaete-Scute Homolog 1 Expression Controls Cellular Differentiation of Neuroblastoma

“…Despite the negative correlation between hASH1 and neuron differentiation observed in neuroblastoma patients, future in vivo studies aimed at evaluating the potential prognostic usefulness of hASH1 in predicting the responsiveness of these tumors to RA-based therapies are needed. Although other factors have been described to modulate RA differentiation in neuroblastoma, hASH1 represents a unique target and a possible biomarker for all neuroblastoma, independent of MYCN amplification status (Shah et al, 2014; Cimmino et al, 2015; Heynen et al, 2016). Future studies will focus on defining the mechanism of action of hASH1 during the early phase of RA-differentiation, which may reveal novel cellular targets of importance to retinoid therapy.…”

UAB30, a novel RXR agonist, decreases tumorigenesis and leptomeningeal disease in group 3 medulloblastoma patient-derived xenografts

“…Pictures [Photometrics CoolSNAP HQ2 CCD camera (Tucson, AZ) attached to a Nikon Eclipse Ti microscope (Tokyo, Japan)] of the cells were obtained and number of neurite outgrowths per cell were counted using the image software SPOT Basic 5.2 (Diagnostic Instruments Inc., Sterling Heights, MI) and reported as mean neurite outgrowths [26]. …”