Extrachromosomal circular DNA drives oncogenic genome remodeling in neuroblastoma

Abstract

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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Fig. 1: A genome-wide map of circular DNA in neuroblastoma.
Fig. 2: Monoallelic large circular DNAs are an origin of oncogene amplification and overexpression in neuroblastoma.
Fig. 3: The majority of structural rearrangements involve sites of DNA circularization and form clustered rearrangement patterns in neuroblastoma.
Fig. 4: Rearrangement of circular DNAs drives transcriptional deregulation and dismal prognosis in neuroblastoma.

Data availability

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.

Code availability

The scripts used to analyze the sequencing data have been uploaded to www.github.com/henssenlab. Data on tree-shaped rearrangements can be accessed and visualized online (https://kons.shinyapps.io/trees/).

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Acknowledgements

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.

Author information

R.P.K., E.R.F., K.H., J.M., F.H., I.C.M., R.C., A.W., M.B., M.P., C. Röefzaad, T.T., H.D., R.F.S., C. Rosswog, J. Theissen, V.B., N.M.P., H.D.G., Y.B., A.S., N. Timme, K.K., S.F., N. Thiessen, E.B., K.S, A.K., P.H., J. Toedling, M.F., D.B., S.S., A.E., D.T., J.H.S. and A.G.H. contributed to the study design and the collection and interpretation of the data. R.P.K. performed the analysis of Circle-seq and WGS. E.R.F. performed the data analysis of the WGS data. I.C.M., R.C., N.M.P. and H.D.G. performed the circular DNA extraction and PCR-based validation. V.B., C. Röefzaad, A.W., P.H., K.S., M.F., A.S. and F.H. collected and prepared the patient samples. C. Rosswog. and J. Theissen performed and analyzed the FISH. M.B. and R.F.S. performed the haplotype phasing analyses. M.P. and J. Toedling. analyzed the tumor genome sequencing data. M.B. performed the allele-specific analysis of Circle-seq, WGS and RNA-seq. S.F., F.K., R.P.K, K.H. and J.M. performed the RNA-seq data analysis. P.H., H.D.G., N.M.P., A.S., D.B. and K.S. performed the experiments and analyzed the data. R.P.K., A.K., A.E. and J.H.S. contributed to the study design. R.P.K. and A.G.H. led the study design, performed the data analysis and wrote the manuscript. All authors contributed to manuscript drafting.

Correspondence to Richard P. Koche.

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Supplementary Figs. 1–12, Tables 1–4 and Note

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Source Data Fig. 1

Unprocessed Western blots for Supplementary Fig. 10f–h.

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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

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