Methylation analysis of DCR2 gene using tumor and serum DNA of neuroblastoma patients

Abstract

11067 Background: Since more than half of patients with high-risk neuroblastoma (NB) do not have a known, poor prognostic marker; MYCN amplification (MNA), it is crucial to find a novel predictor for the high-risk group in non-NMA NB patients to improve their outcome with the risk-adopted therapy.Rapidly progressing neuroblastomas have recently been associated with aberrant hypermethylation of the Tumor Necrosing Factor Related Apoptosis Inducing Ligand (TRAIL) decoy receptor 2 gene (DCR2) promoter. We developed a method to evaluate DCR2 methylation status in tumor and serum DNA, and assessed its utility as a prognostic factor and indicator of therapeutic efficacy in NB patients. Methods: Using DNA-based real-time PCR, we evaluated the ratio of a methylated-DCR2 specific sequence (M) and a reference sequence (R) located in a part of the DCR2 promoter that is unaffected by methylation in 5 NB cell lines and 53 NBs. Of these patients, 13 had MNA NB, and 40 had non-MNA NB. The association between DCR2-methylation and stage was evaluated by Fisher’s exact test, and differences of event-free survival (EFS) were evaluated by the Kaplan-Meier method. Results: DCR2 aberrant methylation was detected in all 5 NB cell lines and 16 of the 53 tumor samples. DCR2 methylation was associated with stage in both the NB group (n=53; p<0.001) and non-MNA group, (n=40; p<0.001), and patients with DCR2 methylation showed significantly poorer 5-year EFS in both the NB group (43% vs. 83%; p=0.002) and non-MNA group (34% vs. 96%; p<0.001). In 10 patients for which data was available, a strong correlation was observed between the M/R ratios in tumor and serum (r=0.825; p=0.006). Among 5 DCR2-methylated patients whose clinical courses were followed, the M/R ratios in serum fell to undetectable levels in the patients that experienced remission (n=2), whereas they increased to high levels in the relapsed patients (n=3). Conclusions: Our study indicates that DCR2 hypermethylation is a useful biomarker for predicting a poor prognosis in NB, especially in non-MNA NB patients. Furthermore, detection of methylated DCR2 in serum DNA might be an indicator of therapeutic efficacy in DCR2-methylated NB patients. No significant financial relationships to disclose.