5-Hydroxymethylcytosine Profiles Are Prognostic of Outcome in Neuroblastoma and Reveal Transcriptional Networks That Correlate With Tumor Phenotype

Abstract

PURPOSE Whole-genome profiles of the epigenetic modification 5-hydroxymethylcytosine (5-hmC) are robust diagnostic biomarkers in adult patients with cancer. We investigated if 5-hmC profiles would serve as novel prognostic markers in neuroblastoma, a clinically heterogeneous pediatric cancer. Because this DNA modification facilitates active gene expression, we hypothesized that 5-hmC profiles would identify transcriptomic networks driving the clinical behavior of neuroblastoma. PATIENTS AND METHODS Nano-hmC-Seal sequencing was performed on DNA from Discovery (n = 51), Validation (n = 38), and Children’s Oncology Group (n = 20) cohorts of neuroblastoma tumors. RNA was isolated from 48 tumors for RNA sequencing. Genes with differential 5-hmC or expression between clusters were identified using DESeq2. A 5-hmC model predicting outcome in high-risk patients was established using linear discriminant analysis. RESULTS Comparison of low- versus high-risk tumors in the Discovery cohort revealed 577 genes with differential 5-hmC. Hierarchical clustering of tumors from the Discovery and Validation cohorts using these genes identified two main clusters highly associated with established prognostic markers, clinical risk group, and outcome. Genes with increased 5-hmC and expression in the favorable cluster were enriched for pathways of neuronal differentiation and KRAS activation, whereas genes involved in inflammation and the PRC2 complex were identified in the unfavorable cluster. The linear discriminant analysis model trained on high-risk Discovery cohort tumors was prognostic of outcome when applied to high-risk tumors from the Validation and Children’s Oncology Group cohorts (hazard ratio, 3.8). CONCLUSION 5-hmC profiles may be optimal DNA-based biomarkers in neuroblastoma. Analysis of transcriptional networks regulated by these epigenomic modifications may lead to a deeper understanding of drivers of neuroblastoma phenotype.

5-Hydroxymethylcytosine profiles of cfDNA are highly predictive of R-CHOP treatment response in diffuse large B cell lymphoma patients

“…2 d) was highly enriched in hydroxymethylation for non-responders ( p = 0.00033). In addition, for the top 205 DhMRs, the most significant enrichment was found in intronic, intergenic, and promoter regions, consistent with previous studies [ 45 , 46 ] (Fig. 2 e).…”

“…In addition, for the top 205 DhMRs, the most significant enrichment was found in intronic, intergenic, and promoter regions, consistent with previous studies [45,46] ( Fig. 2e).…”

A human tissue map of 5-hydroxymethylcytosines exhibits tissue specificity through gene and enhancer modulation

“…Previous studies have shown that 5hmC-modified loci can serve as excellent biomarkers for the diagnosis and prognosis of human diseases including cancer [19][20][21][23][24][25][26] . These data suggest that 5hmC may mark specific genes or promoter/enhancers in disease samples that undergo concerted changes representative of the disease pathogenesis in question.…”

“…Moreover, 5hmC modifications co-localize with gene bodies and enhancers are known to mark for transcription activation 6,17,18 . Notably, in several recent studies, the 5hmC-modified loci have been shown to serve as informative biomarkers for a variety of human cancers and other complex diseases [19][20][21][22][23][24][25][26] . Interestingly, two early studies that investigated the 5hmC in circulating cell-free DNA from patients with solid tumors suggested that 5hmC had the potential to be specific biomarkers for human cancers arising from different tissue origins 19,20 .…”

Fragmentation patterns of circulating cell-free DNA demonstrate biomarker potential for human cancers

“…Thirdly, by focusing on the fragmentation patterns, the DELFI approach understandably missed opportunities to utilize other important molecular features of cfDNA (e.g., epigenetic markers). For example, considering recent progress in exploiting novel epigenetic markers in cfDNA, such as 5-hydroxymethylcytosines, in biomarker discovery for human cancers (2, 20, 21), integrating both the fragmentation patterns and epigenetic features of cfDNA could possibly enhance our understanding of the underlying mechanisms of molecular alterations in the cancer genome and epigenome, as well as improve the detection performance. In summary, Cristiano et al provided exciting new progress in this emerging area with potentially tremendous impact on clinical oncology.…”