Development and validation of a comprehensive genomic diagnostic tool for myeloid malignancies

et al.The diagnosis of hematologic malignancies relies on multidisciplinary workflows involving morphology, flow cytometry, cytogenetic, and molecular genetic analyses. Advances in cancer genomics have identified numerous recurrent mutations with clear prognostic and/or therapeutic significance to different cancers. In myeloid malignancies, there is a clinical imperative to test for such mutations in mainstreamdiagnosis; however, progress toward this has been slow and piecemeal. Here we describe Karyogene, an integrated targeted resequencing/analytical platform that detects nucleotide substitutions, insertions/deletions, chromosomal translocations, copy number abnormalities, and zygosity changes in a single assay. We validate the approach against 62 acute myeloid leukemia, 50 myelodysplastic syndrome, and 40 blood DNA samples from individuals without evidence of clonal blood disorders.Wedemonstrate robust detection of sequence changes in 49 genes, including difficult-to-detect mutations such as FLT3 internal-tandem and mixed-lineage leukemia (MLL) partial-tandem duplications, and clinically significant chromosomal rearrangements including MLL translocations to known and unknown partners, identifying the novel fusion gene MLL-DIAPH2 in the process. Additionally, we identify most significant chromosomal gains and losses, and several copy neutral loss-of-heterozygosity mutations at a genome-wide level, including previously unreported changes such as homozygosity for DNMT3A R882 mutations. Karyogene represents a dependable genomic diagnosis platform for translational research and for the clinical management of myeloid malignancies, which can be readily adapted for use in other cancers.This study was supported by a Wellcome Trust Clinician Scientist Fellowship (100678/Z/12/Z) (T. McKerrell), the Wellcome Trust Sanger Institute (WT098051), and an educational grant from Celgene (ref: 51261). G.S.V. is funded by a Wellcome Trust Senior Fellowship in Clinical Science (WT095663MA), and work in his laboratory is also funded by Bloodwise and the Kay Kendall Leukaemia Fund. A.J.W. is supported by a Specialist Programme from Bloodwise (12048) and by the Medical Research Council (MC_U105161083). I.V. is funded by the Spanish Ministerio de Economía y Competitividad subprograma Ramón y Cajal.Peer Reviewe