MYCN is a novel oncogenic target in adult B-ALL that activates the Wnt/β-catenin pathway by suppressing DKK3

Abstract

Dickkopf-3 (DKK3) is frequently down-regulated by promoter hypermethylation and is closely associated with a poor prognosis in many cancers. Our previous studies have shown that miR-708 down-regulates DKK3 at the post-transcriptional level in B-ALL. However, whether transcriptional mechanisms lead to DKK3 silencing remains unclear. Here, we analysed the promoter regions of DKK3 by bioinformatics and found binding sites for MYCN. A dual-luciferase reporter gene assay and ChIP experiments revealed that MYCN negatively regulates DKK3 at the transcriptional level in B-ALL cell lines, and using bisulphite sequencing PCR, we affirmed that MYCN has no effect on the methylation of the DKK3 promoter. MYCN silencing in B-ALL cells resulted in reduced cell proliferation, increased apoptosis and G1 phase arrest. Treatment with MYCN siRNA or 5-aza-2'-deoxycytidine (5-AdC), a demethylating agent, significantly increased the levels of DKK3 mRNA and protein and decreased the protein levels of p-GSK3β and nuclear β-catenin, which indicates inhibition of the Wnt/β-catenin pathway in vitro. MYCN knockdown significantly decreased the tumorigenic capacity of Nalm6 cells, which restored DKK3 levels and inhibited the Wnt/β-catenin pathway in vivo. Our study provides an increased understanding of adult B-ALL pathogenesis, which may be beneficial to the development of effective prognostic markers or therapeutic targets.

Keywords: 5-AdC; DKK3; MYCN; Wnt/β-catenin; adult B-cell acute lymphoblastic leukaemia.