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Defective Epstein–Barr virus in chronic active infection and haematological malignancy

Abstract

Epstein–Barr virus (EBV) infection is highly prevalent in humans and is implicated in various diseases, including cancer1,2. Chronic active EBV infection (CAEBV) is an intractable disease classified as a lymphoproliferative disorder in the 2016 World Health Organization lymphoma classification1,2. CAEBV is characterized by EBV-infected T/natural killer (NK) cells and recurrent/persistent infectious mononucleosis-like symptoms3. Here, we show that CAEBV originates from an EBV-infected lymphoid progenitor that acquires DDX3X and other mutations, causing clonal evolution comprising multiple cell lineages. Conspicuously, the EBV genome in CAEBV patients harboured frequent intragenic deletions (27/77) that were also common in various EBV-associated neoplastic disorders (28/61), including extranodal NK/T-cell lymphoma and EBV-positive diffuse large B-cell lymphoma, but were not detected in infectious mononucleosis or post-transplant lymphoproliferative disorders (0/47), which suggests a unique role of these mutations in neoplastic proliferation of EBV-infected cells. These deletions frequently affected BamHI A rightward transcript microRNA clusters (31 cases) and several genes that are essential for producing viral particles (20 cases). The deletions observed in our study are thought to reactivate the lytic cycle by upregulating the expression of two immediate early genes, BZLF1 and BRLF14,5,6,7, while averting viral production and subsequent cell lysis. In fact, the deletion of one of the essential genes, BALF5, resulted in upregulation of the lytic cycle and the promotion of lymphomagenesis in a xenograft model. Our findings highlight a pathogenic link between intragenic EBV deletions and EBV-associated neoplastic proliferations.

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Code availability

Custom codes used for the simulation of EBV deletions are available from GitHub repository (https://github.com/yusukeokuno/ebvsimulation) with no restriction on access.

Data availability

Sequence data has been deposited at the European Genome-phenome Archive (EGA), which is hosted by the EBI and the CRG, under accession number EGAS00001003159. Further information about EGA can be found on https://ega-archive.org and ref. 49. We also registered assembled sequences of the EBV genomes in DNA Data Bank of Japan (DDBJ; https://www.ddbj.nig.ac.jp/) under BioProject accession number PRJDB7503. The sequences can be individually via getentry (http://getentry.ddbj.nig.ac.jp/) using the accession numbers AP019012–AP019188.

Additional information

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Acknowledgements

The authors would like to thank all of the subjects and families for participating in this study. The authors would like to thank N. Yoshida, M. Okada, H. Moriuchi, K. Yamamoto, S. Kamimura, Y. Horikoshi and T. Kinoshita for providing samples. The authors would also like to thank Y. Miura, Y. Imanishi, F. Ando, S. Kumagai, T. Kunogi, H. Matsuda, H.M.A. Masud and H. Namizaki for their valuable assistance. The authors acknowledge M. Nakatochi for valuable comments. The authors acknowledge the Division for Medical Research Engineering, Nagoya University Graduate School of Medicine for technical support of cell sorting and next-generation sequencing. The authors acknowledge the Human Genome Center, the Institute of Medical Science, the University of Tokyo (http://sc.hgc.jp/shirokane.html) for providing super-computing resources. This work was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) to H.K. (grant nos 25293109 and 17H04081), grants from Japan Agency for Medical Research and Development to H.K. (grant no. JP17ek0109286) and T.M. (grant no. JP17fm0208016), a grant from the Hori Sciences and Arts Foundation and from the Ministry of Health, Labor, and Welfare of Japan to H.K. (grant no. H29-Nanchi-016).

Author information

Y.O., T.M., Y.Sato and H.M. designed and performed the research, analysed the data and wrote the paper. T.O., N.M., K.Y., Y.Shiraishi, K.C., H.T. and S.Miyano performed bioinformatics analyses of the sequencing data. Y.N., M.Y., T.W. and F.G. performed the research and analysed the data. Y.Ito, A.S., M.I., K.K., M.S., K.O., J.K., T.N., H.Kiyoi, S.Kato, S.N., S.Morishima, T.Y., S.F., N.S., Y.Isobe, M.N., A.K., K.I. and Y.T. collected specimens and clinical data. S.Kojima, S.O. and H.Kimura led the entire project and wrote the paper. All authors critically reviewed the manuscript for its content.

Competing interests

S.Kojima was supported by a grant from Sanofi K.K. All other authors declare no competing interests.

Correspondence to Hiroshi Kimura.

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    Supplementary Figures 1–12, Supplementary Tables 1–9 and Supplementary Discussion.

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About this article

Fig. 1: Mutational analysis of CAEBV.
Fig. 2: Clonal evolution of CAEBV.
Fig. 3: Cell origin of CAEBV.
Fig. 4: Mutational analysis of the EBV genome.
Fig. 5: Lymphomagenesis of LCLs carrying EBV with an essential gene deletion.