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Neuroblastoma Study Reveals Rearrangements Originating From Extrachromosomal Circular DNA

NEW YORK – New research suggests circularized DNA falling outside of linear chromosomes may serve as a recurrent source of somatic rearrangements in neuroblastoma, a pediatric cancer affecting immature cells in the sympathetic nervous system.

"[W]e find that extrachromosomal circular DNAs are an unanticipated major source of somatic rearrangements, contributing to oncogenic remodeling through chimeric circularization and reintegration of circular DNA into the linear genome" co-senior and corresponding author Anton Henssen, a pediatric oncology and hematology researcher affiliated with the Charité Berlin University of Medicine, the Berlin Institute of Health, and the German Cancer Research Center, and his co-authors reported online today in Nature Genetics.

While past research has pointed to a role for circularized, extrachromosomal MYCN oncogene sequences in neuroblastoma, the team explained, the full suite and the frequency of somatic mutations involving small or large stretches of circularized extrachromosomal DNA amplifications had not been fully explored.

With that in mind, Henssen and colleagues from Germany, the US, and Spain profiled matched tumor and normal blood samples from 93 neuroblastoma patients using whole-genome sequencing and an algorithm that uncovers circularized DNA based on paired read orientation, uncovering preliminary evidence for complex and relatively frequent ecDNAs in neuroblastoma.

That finding prompted the team to take a closer look at these sequences using a modified version of circle sequencing (Circle-seq) in 21 of the neuroblastoma tumors, making it possible to enrich for circularized DNA. The circularized sequences were mapped back to their original sites in the genome using additional long-read and single-molecule real-time sequences, the investigators explained, and they validated candidate DNA circles with PCR and Sanger sequencing.

Together, these approaches uncovered almost 5,700 small extrachromosomal circular DNAs per tumor, on average, and an average of 0.82 large, copy number-amplified extrachromosomal circular DNA sequences, the authors reported, noting that "DNA circularization is not restricted to proto-oncogenes, but also affects various coding and non-coding regions with yet unknown functional consequences."

Even so, the team's follow-up analyses — including RNA sequencing experiments — indicated that rearrangements stemming from extrachromosomal circular DNA from MYCN and other genes may be a recurrent and ongoing source of new mutations through a multi-hit model in neuroblastoma.

"We demonstrate that the majority of genomic rearrangements in neuroblastoma involve circular DNA, challenging our current understanding about cancer genome remodeling," the authors wrote. "We envision that our findings extend to other cancers and that further detailed analyses of circle-derived rearrangements will shed new insights into our understanding of cancer genome remodeling."