Extrachromosomal circularization of DNA is an important genomic feature in cancer. However, the structure, composition and genome-wide frequency of extrachromosomal circular DNA have not yet been profiled extensively. Here, we combine genomic and transcriptomic approaches to describe the landscape of extrachromosomal circular DNA in neuroblastoma, a tumor arising in childhood from primitive cells of the sympathetic nervous system. Our analysis identifies and characterizes a wide catalog of somatically acquired and undescribed extrachromosomal circular DNAs. Moreover, we 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. Cancer-causing lesions can emerge out of circle-derived rearrangements and are associated with adverse clinical outcome. It is highly probable that circle-derived rearrangements represent an ongoing mutagenic process. Thus, extrachromosomal circular DNAs represent a multihit mutagenic process, with important functional and clinical implications for the origins of genomic remodeling in cancer.
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The WGS and RNA-seq data that support the findings of this study have been deposited with the European Genome-phenome Archive (https://www.ebi.ac.uk/ega/) under accession nos. EGAS00001001308 and EGAS00001004022. The Circle-seq data that support the findings of this study are available from the corresponding author upon request. Source data for Fig. 1 are available online.
Turner, K. M. et al. Extrachromosomal oncogene amplification drives tumour evolution and genetic heterogeneity. Nature 543, 122–125 (2017).
Møller, H. D. et al. Circular DNA elements of chromosomal origin are common in healthy human somatic tissue. Nat. Commun. 9, 1069 (2018).
Shibata, Y. et al. Extrachromosomal microDNAs and chromosomal microdeletions in normal tissues. Science 336, 82–86 (2012).
Pennisi, E. Circular DNA throws biologists for a loop. Science 356, 996 (2017).
Verhaak, R. G. W., Bafna, V. & Mischel, P. S. Extrachromosomal oncogene amplification in tumour pathogenesis and evolution.Nat. Rev. Cancer 19, 283–288 (2019).
Møller, H. D., Parsons, L., Jørgensen, T. S., Botstein, D. & Regenberg, B. Extrachromosomal circular DNA is common in yeast. Proc. Natl Acad. Sci. USA 112, E3114–E3122 (2015).
Tjio, J. H. & Levan, A. The chromosome number of man. Hereditas 42, 1–6 (1956).
Garsed, D. W. et al. The architecture and evolution of cancer neochromosomes. Cancer Cell 26, 653–667 (2014).
Rausch, T. et al. Genome sequencing of pediatric medulloblastoma links catastrophic DNA rearrangements with TP53 mutations. Cell 148, 59–71 (2012).
Kohl, N. E. et al. Transposition and amplification of oncogene-related sequences in human neuroblastomas. Cell 35, 359–367 (1983).
deCarvalho, A. C. et al. Discordant inheritance of chromosomal and extrachromosomal DNA elements contributes to dynamic disease evolution in glioblastoma. Nat. Genet. 50, 708–717 (2018).
Nikolaev, S. et al. Extrachromosomal driver mutations in glioblastoma and low-grade glioma. Nat. Commun. 5, 5690 (2014).
Schwab, M. et al. Amplified DNA with limited homology to myc cellular oncogene is shared by human neuroblastoma cell lines and a neuroblastoma tumour. Nature 305, 245–248 (1983).
Balaban-Malenbaum, G. & Gilbert, F. Double minute chromosomes and the homogeneously staining regions in chromosomes of a human neuroblastoma cell line. Science 198, 739–741 (1977).
Cox, D., Yuncken, C. & Spriggs, A. I. Minute chromatin bodies in malignant tumours of childhood. Lancet 1, 55–58 (1965).
Sanborn, J. Z. et al. Double minute chromosomes in glioblastoma multiforme are revealed by precise reconstruction of oncogenic amplicons. Cancer Res. 73, 6036–6045 (2013).
Deshpande, V. et al. Exploring the landscape of focal amplifications in cancer using AmpliconArchitect. Nat. Commun. 10, 392 (2019).
Avet-Loiseau, H. et al. Morphologic and molecular cytogenetics in neuroblastoma. Cancer 75, 1694–1699 (1995).
Dillon, L. W. et al. Production of extrachromosomal microDNAs is linked to mismatch repair pathways and transcriptional activity. Cell Rep. 11, 1749–1759 (2015).
Forbes, S. A. et al. COSMIC: somatic cancer genetics at high-resolution. Nucleic Acids Res. 45, D777–D783 (2017).
Bouzas-Rodriguez, J. et al. Neurotrophin-3 production promotes human neuroblastoma cell survival by inhibiting TrkC-induced apoptosis. J. Clin. Invest. 120, 850–858 (2010).
Xu, K. Structure and evolution of double minutes in diagnosis and relapse brain tumors. Acta Neuropathol. 137, 123–137 (2019).
Storlazzi, C. T. et al. Gene amplification as double minutes or homogeneously staining regions in solid tumors: origin and structure. Genome Res. 20, 1198–1206 (2010).
Villamón, E. et al. Genetic instability and intratumoral heterogeneity in neuroblastoma with MYCN amplification plus 11q deletion. PLoS ONE 8, e53740 (2013).
Marrano, P., Irwin, M. S. & Thorner, P. S. Heterogeneity of MYCN amplification in neuroblastoma at diagnosis, treatment, relapse, and metastasis. Genes Chromosomes Cancer 56, 28–41 (2017).
Gröbner, S. N. et al. The landscape of genomic alterations across childhood cancers. Nature 555, 321–327 (2018).
Northcott, P. A. et al. Enhancer hijacking activates GFI1 family oncogenes in medulloblastoma. Nature 511, 428–434 (2014).
Henssen, A. G. et al. PGBD5 promotes site-specific oncogenic mutations in human tumors. Nat. Genet. 49, 1005–1014 (2017).
Henssen, A. G. et al. Genomic DNA transposition induced by human PGBD5. eLife 4, e10565 (2015).
Henssen, A. G. et al. Forward genetic screen of human transposase genomic rearrangements. BMC Genomics 17, 548 (2016).
Guzmán, C., Bagga, M., Kaur, A., Westermarck, J. & Abankwa, D. ColonyArea: an ImageJ plugin to automatically quantify colony formation in clonogenic assays. PLoS ONE 9, e92444 (2014).
Henssen, A. G., MacArthur, I., Koche, R. & Dorado-García, H. Purification and sequencing of large circular DNA from human cells. Protoc Exch (2019); https://doi.org/10.1038/protex.2019.006
Boeva, V. et al. Control-FREEC: a tool for assessing copy number and allelic content using next-generation sequencing data. Bioinformatics 28, 423–425 (2012).
Van Loo, P. et al. Allele-specific copy number analysis of tumors. Proc. Natl Acad. Sci. USA 107, 16910–16915 (2010).
Chong, Z. et al. novoBreak: local assembly for breakpoint detection in cancer genomes. Nat. Methods 14, 65–67 (2017).
Wala, J. A. et al. SvABA: genome-wide detection of structural variants and indels by local assembly. Genome Res. 28, 581–591 (2018).
Rausch, T. et al. DELLY: structural variant discovery by integrated paired-end and split-read analysis. Bioinformatics 28, i333–i339 (2012).
Moncunill, V. et al. Comprehensive characterization of complex structural variations in cancer by directly comparing genome sequence reads. Nat. Biotechnol. 32, 1106–1112 (2014).
Helmsauer, K. Tree-shaped Rearrangement Patterns in Pediatric Cancer Genomes (2019); https://kons.shinyapps.io/trees
Mootha, V. K. et al. PGC-1α-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat. Genet. 34, 267–273 (2003).
Subramanian, A. et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc. Natl Acad. Sci. USA 102, 15545–15550 (2005).
We thank A. Kentsis, S. Armstrong, B. Regenberg, F. Speleman, S. Perner, U. Ohler and N. Hübener for critical discussions, and K. Astrahantseff for editorial advice. We thank D. Sunaga-Franze, C. Quedenau, M. Sohn, K. Richter and C. Langnick for technical support. A.G.H. is supported by the Deutsche Forschungsgemeinschaft (German Research Foundation; grant no. 398299703) and the Wilhelm Sander Stiftung (2018.011.1). A.G.H., A.K. and S.F. are participants in the Berlin Institute of Health-Charité Clinical Scientist Program funded by the Charité-Universitätsmedizin Berlin and the Berlin Institute of Health. A.G.H., S.F., K.H. and V.B. are supported by Berliner Krebsgesellschaft e.V. K.H. is supported by Boehringer Ingelheim Fonds. This work was also supported by the TransTumVar project (project no. PN013600). This project was supported by the Berlin Institute of Health within the collaborative research project TERMINATE-NB (CRG04). We thank the patients and their parents for granting access to the tumor specimens and clinical information that were analyzed in this study. We thank B. Hero, H. Düren and N. Hemstedt of the Neuroblastoma Biobank and Neuroblastoma Trial Registry (University Children’s Hospital Cologne) of the GPOH for providing samples and clinical data.
The authors declare no competing interests.
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Koche, R.P., Rodriguez-Fos, E., Helmsauer, K. et al. Extrachromosomal circular DNA drives oncogenic genome remodeling in neuroblastoma. Nat Genet 52, 29–34 (2020). https://doi.org/10.1038/s41588-019-0547-z