The ETV6-RUNX1 fusion gene, found in 25% of childhood acute lymphoblastic leukemia (ALL) cases, is acquired in utero but requires additional somatic mutations for overt leukemia. We used exome and low-coverage whole-genome sequencing to characterize secondary events associated with leukemic transformation. RAG-mediated deletions emerge as the dominant mutational process, characterized by recombination signal sequence motifs near breakpoints, incorporation of non-templated sequence at junctions, ∼30-fold enrichment at promoters and enhancers of genes actively transcribed in B cell development and an unexpectedly high ratio of recurrent to non-recurrent structural variants. Single-cell tracking shows that this mechanism is active throughout leukemic evolution, with evidence of localized clustering and reiterated deletions. Integration of data on point mutations and rearrangements identifies ATF7IP and MGA as two new tumor-suppressor genes in ALL. Thus, a remarkably parsimonious mutational process transforms ETV6-RUNX1-positive lymphoblasts, targeting the promoters, enhancers and first exons of genes that normally regulate B cell differentiation.

] Cancer Genome Project Wellcome Trust Sanger Institute Hinxton UK [2] Addenbrooke's National Health Service Foundation Trust Cambridge UK [3] Department of Haematology University of Cambridge Cambridge UK [4]

] Cancer Genome Project Wellcome Trust Sanger Institute Hinxton UK [2] Department of Human Genetics VIB and University of Leuven Leuven Belgium

] Institute for Cancer Research Sutton London UK [2]

] Institute for Cancer Research Sutton London UK [2] Northern Institute for Cancer Research University of Newcastle Newcastle upon Tyne UK

Cancer Genome Project Wellcome Trust Sanger Institute Hinxton UK

Centro Ricerca Tettamanti Hospital San Gerardo Monza Italy

Department of Laboratory Medicine University of California San Francisco San Francisco California USA

Department of Paediatric Haematology and Oncology 2nd Faculty of Medicine Charles University Prague and University Hospital Motol Prague Czech Republic

Hospital Universitario 12 de Octubre Madrid Spain

Institute for Cancer Research Sutton London UK

Paediatric Malignancy Unit Molecular Haematology and Cancer Biology Unit Camelia Botnar Laboratories Great Ormond Street Hospital for Children and University College London Institute of Child Health London UK

Komentář v

Nat Genet. 2014 Feb;46(2):96-8. doi: 10.1038/ng.2879. PubMed

Zobrazit více v PubMed

Bhojwani D, et al. ETV6-RUNX1-positive childhood acute lymphoblastic leukemia: improved outcome with contemporary therapy. Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K. 2012;26:265–270. PubMed PMC

Greaves MF, Wiemels J. Origins of chromosome translocations in childhood leukaemia. Nature reviews. Cancer. 2003;3:639–649. PubMed

Mori H, et al. Chromosome translocations and covert leukemic clones are generated during normal fetal development. Proceedings of the National Academy of Sciences of the United States of America. 2002;99:8242–8247. PubMed PMC

Bateman CM, et al. Acquisition of genome-wide copy number alterations in monozygotic twins with acute lymphoblastic leukemia. Blood. 2010;115:3553–3558. PubMed

Mullighan CG, et al. Genome-wide analysis of genetic alterations in acute lymphoblastic leukaemia. Nature. 2007;446:758–764. PubMed

Zhang M, Swanson PC. V(D)J recombinase binding and cleavage of cryptic recombination signal sequences identified from lymphoid malignancies. The Journal of biological chemistry. 2008;283:6717–6727. PubMed

Mullighan CG, et al. Genomic analysis of the clonal origins of relapsed acute lymphoblastic leukemia. Science. 2008;322:1377–1380. PubMed PMC

Raschke S, Balz V, Efferth T, Schulz WA, Florl AR. Homozygous deletions of CDKN2A caused by alternative mechanisms in various human cancer cell lines. Genes, chromosomes & cancer. 2005;42:58–67. PubMed

Waanders E, et al. The origin and nature of tightly clustered BTG1 deletions in precursor B-cell acute lymphoblastic leukemia support a model of multiclonal evolution. PLoS genetics. 2012;8:e1002533. PubMed PMC

Holmfeldt L, et al. The genomic landscape of hypodiploid acute lymphoblastic leukemia. Nature genetics. 2013 PubMed PMC

Hubner S, et al. High incidence and unique features of antigen receptor gene rearrangements in TEL-AML1-positive leukemias. Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K. 2004;18:84–91. PubMed

Schatz DG, Swanson PC. V(D)J recombination: mechanisms of initiation. Annual review of genetics. 2011;45:167–202. PubMed

Fugmann SD, Lee AI, Shockett PE, Villey IJ, Schatz DG. The RAG proteins and V(D)J recombination: complexes, ends, and transposition. Annual review of immunology. 2000;18:495–527. PubMed

Komori T, Okada A, Stewart V, Alt FW. Lack of N regions in antigen receptor variable region genes of TdT-deficient lymphocytes. Science. 1993;261:1171–1175. PubMed

Raghavan SC, Swanson PC, Ma Y, Lieber MR. Double-strand break formation by the RAG complex at the bcl-2 major breakpoint region and at other non-B DNA structures in vitro. Molecular and cellular biology. 2005;25:5904–5919. PubMed PMC

Tsai AG, et al. Human chromosomal translocations at CpG sites and a theoretical basis for their lineage and stage specificity. Cell. 2008;135:1130–1142. PubMed PMC

Hesse JE, Lieber MR, Mizuuchi K, Gellert M. V(D)J recombination: a functional definition of the joining signals. Genes & development. 1989;3:1053–1061. PubMed

Mullighan CG, et al. BCR-ABL1 lymphoblastic leukaemia is characterized by the deletion of Ikaros. Nature. 2008;453:110–114. PubMed

Zhang J, et al. The genetic basis of early T-cell precursor acute lymphoblastic leukaemia. Nature. 2012;481:157–163. PubMed PMC

Nik-Zainal S, et al. Mutational processes molding the genomes of 21 breast cancers. Cell. 2012;149:979–993. PubMed PMC

Campbell PJ, et al. The patterns and dynamics of genomic instability in metastatic pancreatic cancer. Nature. 2010;467:1109–1113. PubMed PMC

Baca SC, et al. Punctuated evolution of prostate cancer genomes. Cell. 2013;153:666–677. PubMed PMC

Bailey TL, Williams N, Misleh C, Li WW. MEME: discovering and analyzing DNA and protein sequence motifs. Nucleic acids research. 2006;34:W369–373. PubMed PMC

Stephens PJ, et al. Complex landscapes of somatic rearrangement in human breast cancer genomes. Nature. 2009;462:1005–1010. PubMed PMC

Berger MF, et al. The genomic complexity of primary human prostate cancer. Nature. 2011;470:214–220. PubMed PMC

Dunham I, et al. An integrated encyclopedia of DNA elements in the human genome. Nature. 2012;489:57–74. PubMed PMC

Stephens PJ, et al. Massive genomic rearrangement acquired in a single catastrophic event during cancer development. Cell. 2011;144:27–40. PubMed PMC

Shimazaki N, Tsai AG, Lieber MR. H3K4me3 stimulates the V(D)J RAG complex for both nicking and hairpinning in trans in addition to tethering in cis: implications for translocations. Molecular cell. 2009;34:535–544. PubMed PMC

Anderson K, et al. Genetic variegation of clonal architecture and propagating cells in leukaemia. Nature. 2011;469:356–361. PubMed

Potter NE, et al. Single cell mutational profiling and clonal phylogeny in cancer. Genome research. 2013 PubMed PMC

Familiades J, et al. PAX5 mutations occur frequently in adult B-cell progenitor acute lymphoblastic leukemia and PAX5 haploinsufficiency is associated with BCR-ABL1 and TCF3-PBX1 fusion genes: a GRAALL study. Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K. 2009;23:1989–1998. PubMed

Kempski H, et al. An investigation of the t(12;21) rearrangement in children with B-precursor acute lymphoblastic leukaemia using cytogenetic and molecular methods. British journal of haematology. 1999;105:684–689. PubMed

van Delft FW, et al. Clonal origins of relapse in ETV6-RUNX1 acute lymphoblastic leukemia. Blood. 2011;117:6247–6254. PubMed

Hurlin PJ, Steingrimsson E, Copeland NG, Jenkins NA, Eisenman RN. Mga, a dual-specificity transcription factor that interacts with Max and contains a T-domain DNA-binding motif. The EMBO journal. 1999;18:7019–7028. PubMed PMC

Cancer Genome Atlas Research N. Genomic and epigenomic landscapes of adult de novo acute myeloid leukemia. The New England journal of medicine. 2013;368:2059–2074. PubMed PMC

Chen S, et al. Novel non-TCR chromosome translocations t(3;11)(q25;p13) and t(X;11)(q25;p13) activating LMO2 by juxtaposition with MBNL1 and STAG2. Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K. 2011;25:1632–1635. PubMed

Jaffe JD, et al. Global chromatin profiling reveals NSD2 mutations in pediatric acute lymphoblastic leukemia. Nature genetics. 2013;45:1386–1391. PubMed PMC

Harris RS, Petersen-Mahrt SK, Neuberger MS. RNA editing enzyme APOBEC1 and some of its homologs can act as DNA mutators. Molecular cell. 2002;10:1247–1253. PubMed

Neuberger MS, Rada C. Somatic hypermutation: activation-induced deaminase for C/G followed by polymerase eta for A/T. The Journal of experimental medicine. 2007;204:7–10. PubMed PMC

Alexandrov LB, et al. Signatures of mutational processes in human cancer. Nature. 2013;500:415–421. PubMed PMC

Roberts SA, et al. Clustered mutations in yeast and in human cancers can arise from damaged long single-strand DNA regions. Molecular cell. 2012;46:424–435. PubMed PMC

Roberts KG, et al. Genetic alterations activating kinase and cytokine receptor signaling in high-risk acute lymphoblastic leukemia. Cancer cell. 2012;22:153–166. PubMed PMC

Tsai AG, Lieber MR. RAGs found “not guilty”: cleared by DNA evidence. Blood. 2008;111:1750. PubMed

Boubakour-Azzouz I, Bertrand P, Claes A, Lopez BS, Rougeon F. Terminal deoxynucleotidyl transferase requires KU80 and XRCC4 to promote N-addition at non-V(D)J chromosomal breaks in non-lymphoid cells. Nucleic acids research. 2012;40:8381–8391. PubMed PMC

Hong D, et al. Initiating and cancer-propagating cells in TEL-AML1-associated childhood leukemia. Science. 2008;319:336–339. PubMed

Tsuzuki S, Seto M, Greaves M, Enver T. Modeling first-hit functions of the t(12;21) TEL-AML1 translocation in mice. Proceedings of the National Academy of Sciences of the United States of America. 2004;101:8443–8448. PubMed PMC

Ross ME, et al. Classification of pediatric acute lymphoblastic leukemia by gene expression profiling. Blood. 2003;102:2951–2959. PubMed

Kozarewa I, et al. Amplification-free Illumina sequencing-library preparation facilitates improved mapping and assembly of (G+C)-biased genomes. Nature methods. 2009;6:291–295. PubMed PMC

Li H, Durbin R. Fast and accurate long-read alignment with Burrows-Wheeler transform. Bioinformatics. 2010;26:589–595. PubMed PMC

Varela I, et al. Exome sequencing identifies frequent mutation of the SWI/SNF complex gene PBRM1 in renal carcinoma. Nature. 2011;469:539–542. PubMed PMC

Ye K, Schulz MH, Long Q, Apweiler R, Ning Z. Pindel: a pattern growth approach to detect break points of large deletions and medium sized insertions from paired-end short reads. Bioinformatics. 2009 PubMed PMC

Van Loo P, et al. Allele-specific copy number analysis of tumors. Proceedings of the National Academy of Sciences of the United States of America. 2010;107:16910–16915. PubMed PMC

Alexandrov LB, Nik-Zainal S, Wedge DC, Campbell PJ, Stratton MR. Deciphering signatures of mutational processes operative in human cancer. Cell reports. 2013;3:246–259. PubMed PMC

Greenman C, Wooster R, Futreal PA, Stratton MR, Easton DF. Statistical analysis of pathogenicity of somatic mutations in cancer. Genetics. 2006;173:2187–2198. PubMed PMC

Klipper-Aurbach Y, et al. Mathematical formulae for the prediction of the residual beta cell function during the first two years of disease in children and adolescents with insulin-dependent diabetes mellitus. Medical hypotheses. 1995;45:486–490. PubMed

van Dongen JJ, et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K. 2003;17:2257–2317. PubMed

Prognostic value of rare IKZF1 deletion in childhood B-cell precursor acute lymphoblastic leukemia: an international collaborative study

Genomics and drug profiling of fatal TCF3-HLF-positive acute lymphoblastic leukemia identifies recurrent mutation patterns and therapeutic options