Refined detection and phasing of structural aberrations in pediatric acute lymphoblastic leukemia by linked-read whole-genome sequencing

Structural chromosomal rearrangements that may lead to in-frame gene-fusions represent a leading source of information for diagnosis, risk stratification, and prognosis in pediatric acute lymphoblastic leukemia (ALL). However, short-read whole genome sequencing (WGS) technologies struggle to accurately identify and phase such large-scale chromosomal aberrations in cancer genomes. We therefore evaluated linked-read WGS for detection of chromosomal rearrangements in an ALL cell line (REH) and primary samples of varying DNA quality from 12 patients diagnosed with ALL. We assessed the effect of input DNA quality on phased haplotype block size and the detectability of copy number aberrations (CNAs) and structural variants (SVs). Biobanked DNA isolated by standard column-based extraction methods was sufficient to detect chromosomal rearrangements even at low 10x sequencing coverage. Linked-read WGS enabled precise, allele-specific, digital karyotyping at a base-pair resolution for a wide range of structural variants including complex rearrangements and aneuploidy assessment. With use of haplotype information from the linked-reads, we also identified additional structural variants, such as a compound heterozygous deletion of ERG in a patient with the DUX4-IGH fusion gene. Thus, linked-read WGS allows detection of important pathogenic variants in ALL genomes at a resolution beyond that of traditional karyotyping or short-read WGS.

[1]  Bertil Johansson,et al.  High hyperdiploid childhood acute lymphoblastic leukemia , 2009, Genes, chromosomes & cancer.

[2]  Cheng Cheng,et al.  Genomic Profiling of Adult and Pediatric B-cell Acute Lymphoblastic Leukemia , 2016, EBioMedicine.

[3]  A. Nordgren,et al.  High-resolution detection of chromosomal rearrangements in leukemias through mate pair whole genome sequencing , 2018, PloS one.

[4]  Victor Guryev,et al.  Dense and accurate whole-chromosome haplotyping of individual genomes , 2017, Nature Communications.

[5]  C. Harrison,et al.  Genes commonly deleted in childhood B-cell precursor acute lymphoblastic leukemia: association with cytogenetics and clinical features , 2013, Haematologica.

[6]  C. Mullighan,et al.  Genetic Basis of Acute Lymphoblastic Leukemia. , 2017, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[7]  Hanlee P. Ji,et al.  Haplotyping germline and cancer genomes using high-throughput linked-read sequencing , 2015, Nature Biotechnology.

[8]  M. Greaves,et al.  Phenotypic characterisation of a unique non-T, non-B acute lymphoblastic leukaemia cell line , 1977, Nature.

[9]  P. Kwok,et al.  A Hybrid Approach for de novo Human Genome Sequence Assembly and Phasing , 2016, Nature Methods.

[10]  A. Syvänen,et al.  PAX5-ESRRB is a recurrent fusion gene in B-cell precursor pediatric acute lymphoblastic leukemia , 2016, Haematologica.

[11]  Michael C. Heinold,et al.  The landscape of genomic alterations across childhood cancers , 2018, Nature.

[12]  M. Gerstein,et al.  CNVnator: an approach to discover, genotype, and characterize typical and atypical CNVs from family and population genome sequencing. , 2011, Genome research.

[13]  N. Weisenfeld,et al.  Direct determination of diploid genome sequences , 2016, bioRxiv.

[14]  Rolf Larsson,et al.  DNA methylation-based subtype prediction for pediatric acute lymphoblastic leukemia , 2014, Clinical Epigenetics.

[15]  B. Johansson,et al.  Identification of ETV6-RUNX1-like and DUX4-rearranged subtypes in paediatric B-cell precursor acute lymphoblastic leukaemia , 2016, Nature Communications.

[16]  Nour-al-dain Marzouka,et al.  CopyNumber450kCancer: baseline correction for accurate copy number calling from the 450k methylation array , 2015, Bioinform..

[17]  A. Syvänen,et al.  Transcriptome sequencing in pediatric acute lymphoblastic leukemia identifies fusion genes associated with distinct DNA methylation profiles , 2017, Journal of Hematology & Oncology.

[18]  F. Sigaux,et al.  An intragenic ERG deletion is a marker of an oncogenic subtype of B-cell precursor acute lymphoblastic leukemia with a favorable outcome despite frequent IKZF1 deletions , 2014, Leukemia.

[19]  Paul Shannon,et al.  VariantAnnotation: a Bioconductor package for exploration and annotation of genetic variants , 2014, Bioinform..

[20]  J. Downing,et al.  Childhood Acute Lymphoblastic Leukemia: Progress Through Collaboration. , 2015, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[21]  Bernhard Hiller,et al.  CyDAS: a cytogenetic data analysis system , 2005, Bioinform..

[22]  Anders Isaksson,et al.  Allele-specific copy number analysis of tumor samples with aneuploidy and tumor heterogeneity , 2011, Genome Biology.

[23]  Hanlee P. Ji,et al.  Linked read sequencing resolves complex genomic rearrangements in gastric cancer metastases , 2017, Genome Medicine.

[24]  Behrooz Torabi Moghadam,et al.  The Mutational Landscape in Pediatric Acute Lymphoblastic Leukemia Deciphered by Whole Genome Sequencing , 2014, Human mutation.

[25]  Heather L. Mulder,et al.  Deregulation of DUX4 and ERG in acute lymphoblastic leukemia , 2016, Nature Genetics.

[26]  Mats G Gustafsson,et al.  DNA methylation for subtype classification and prediction of treatment outcome in patients with childhood acute lymphoblastic leukemia. , 2010, Blood.

[27]  N. Lindeman,et al.  Clinical and Technical Aspects of Genomic Diagnostics for Precision Oncology. , 2017, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[28]  M. Harris,et al.  Future of clinical genomics in pediatric oncology. , 2013, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[29]  Christofer L. Bäcklin,et al.  Genome-wide signatures of differential DNA methylation in pediatric acute lymphoblastic leukemia , 2013, Genome Biology.

[30]  O. Kallioniemi,et al.  FusionCatcher – a tool for finding somatic fusion genes in paired-end RNA-sequencing data , 2014, bioRxiv.

[31]  G. Lucchini,et al.  Imatinib after induction for treatment of children and adolescents with Philadelphia-chromosome-positive acute lymphoblastic leukaemia (EsPhALL): a randomised, open-label, intergroup study , 2012, The Lancet. Oncology.

[32]  R. Foà,et al.  RNA sequencing unravels the genetics of refractory/relapsed T-cell acute lymphoblastic leukemia. Prognostic and therapeutic implications , 2016, Haematologica.

[33]  A. Moorman.  The clinical relevance of chromosomal and genomic abnormalities in B-cell precursor acute lymphoblastic leukaemia. , 2012, Blood reviews.

[34]  Robert Huether,et al.  The genomic landscape of hypodiploid acute lymphoblastic leukemia , 2013, Nature Genetics.

[35]  R. Wade,et al.  A novel integrated cytogenetic and genomic classification refines risk stratification in pediatric acute lymphoblastic leukemia. , 2014, Blood.

[36]  Kevin K Dobbin,et al.  Identification of novel cluster groups in pediatric high-risk B-precursor acute lymphoblastic leukemia with gene expression profiling: correlation with genome-wide DNA copy number alterations, clinical characteristics, and outcome. , 2010, Blood.

[37]  Shinichi Morishita,et al.  Integrative analysis of genomic alterations in triple-negative breast cancer in association with homologous recombination deficiency , 2017, PLoS genetics.

[38]  K. Schmiegelow,et al.  Long-term results of NOPHO ALL-92 and ALL-2000 studies of childhood acute lymphoblastic leukemia , 2010, Leukemia.

[39]  Shinichi Morishita,et al.  Recurrent DUX4 fusions in B cell acute lymphoblastic leukemia of adolescents and young adults , 2016, Nature Genetics.