Effects of a novel long noncoding RNA, lncUSMycN, on N-Myc expression and neuroblastoma progression.

Abstract

Patients with neuroblastoma due to the amplification of a 130-kb genomic DNA region containing the MYCN oncogene have poor prognoses. Bioinformatics data were used to discover a novel long noncoding RNA, lncUSMycN, at the 130-kb amplicon. RNA-protein pull-down assays were used to identify proteins bound to lncUSMycN RNA. Kaplan-Meier survival analysis, multivariable Cox regression, and two-sided log-rank test were used to examine the prognostic value of lncUSMycN and NonO expression in three cohorts of neuroblastoma patients (n = 47, 88, and 476, respectively). Neuroblastoma-bearing mice were treated with antisense oligonucleotides targeting lncUSMycN (n = 12) or mismatch sequence (n = 13), and results were analyzed by multiple comparison two-way analysis of variance. All statistical tests were two-sided. Bioinformatics data predicted lncUSMycN gene and RNA, and reverse-transcription polymerase chain reaction confirmed its three exons and two introns. The lncUSMycN gene was coamplified with MYCN in 88 of 341 human neuroblastoma tissues. lncUSMycN RNA bound to the RNA-binding protein NonO, leading to N-Myc RNA upregulation and neuroblastoma cell proliferation. High levels of lncUSMycN and NonO expression in human neuroblastoma tissues independently predicted poor patient prognoses (lncUSMycN: hazard ratio [HR] = 1.87, 95% confidence interval [CI] = 1.06 to 3.28, P = .03; NonO: HR = 2.48, 95% CI = 1.34 to 4.57, P = .004). Treatment with antisense oligonucleotides targeting lncUSMycN in neuroblastoma-bearing mice statistically significantly hindered tumor progression (P < .001). Our data demonstrate the important roles of lncUSMycN and NonO in regulating N-Myc expression and neuroblastoma oncogenesis and provide the first evidence that amplification of long noncoding RNA genes can contribute to tumorigenesis.