chronic myelomonocytic leukemia ( cmml ) is a clonal hematopoietic stem cell neoplasm with overlapping myelodysplastic and myeloproliferative features . while progression to acute myeloid leukemia ( aml ) is common , the driving molecular aberrations involved in transformation remain obscure . clonal cytogenetic abnormalities predict adverse prognosis and are evident in up to 40% of cases . next generation sequencing has identified molecular aberrations in approximately 90% of cmml patients with tet2 gene mutations ( tet2 + ) being particularly frequent . concomitant tet2 + and jak2 + have been described in chronic myeloproliferative disorders , but none of these are sufficient to drive blastic transformation . moreover , transformation of myeloproliferative disorders most often results in loss of jak2 + indicating the transformation to arise in a jak2-negative subclone . we here report a case of tet2 + cmml who acquired jak2 + and progressed into overt aml 12 years after initial diagnosis . array comparative genomic hybridization ( acgh ) revealed a novel del(8q)(q23.2q24.11 ) ( del(8q ) ) comprising genes with recently reported relevance in cancer , including aml . in 1998 , a 64-year old male presented with pleural effusion , pericarditis , and leukocytosis of 3810/l with 30% monocytes . a complete overview of the clinical course of disease is given in table 1 . lymph node enlargement emerged in 2007 and a biopsy showed infiltration of 50% fully differentiated monocytes . despite these various cmml - related manifestations , no cytopenias were evident . however , in 2010 bm failure developed with decreasing hemoglobin and thrombocyte counts and increasing leukocyte count . nine months later , in june 2011 at the age of 76 the patient was admitted with abdominal pain and severely deranged hematopoiesis and was diagnosed with aml . flow cytometry of peripheral blood showing 40% myeloblasts staining positive for cd34 , cd117 , hla - dr , and cd13 . a ct scan showed splenomegaly , and signs of malignant infiltration in the renal parenchyma and the soft tissue along the lower thoracic vertebras , suspicious of myelosarcomas . the patient received palliative care , but succumbed from progressive renal failure in july 2011 . routine molecular analysis at aml diagnosis showed a jak2 + allelic burden of 92% of mononuclear cells ( mncs ) from peripheral blood ( pb ) . since jak2 + is usually lost upon transformation this led us to retrospectively delineate the longitudinal evolution of molecular aberrations at critical time points , summarized in table 1 . in short , dna was extracted from diagnostic paraffin embedded bm biopsies ( 1998 ) , from thawed bm mncs from the time of disease progression ( 2010 ) , and from pb mncs at the time of overt aml ( 2011 ) . jak2 + was analyzed for by standard quantitative polymerase chain reaction ( qpcr ) . while the diagnostic cmml sample was negative for jak2 + , the allelic burden was above the 90% level both in the transformation- and aml phase of disease . the entire coding sequences and all splice sites of the tet2 gene ( exons 311 ) were scanned for mutations by pcr combined with denaturing gradient gel electrophoresis ( dgge ) as described in ref . . in all samples , a stable frameshift mutation in exon 6 was found ( c.3658_3658del a , thr1220fs . dna extracted from a benign skin biopsy from 1990 proved negative for tet2 mutations , hence excluding germline mutation ( fig . 1a ) . by multiplex fluorescent - labeled pcr followed by capillary gel electrophoresis we retrospectively analyzed mncs from the time of aml - diagnosis for recurrent aml related mutations , namely flt3-itd , flt3-d835 , ckit d816v , idh1 r132 , cebpa , npm1 exon 12 , and wt1 exon 7 of which all came out negative . standard g - band karyotyping was not performed at initial diagnosis in 1998 , but the karyotype was 46,xy two and a half years from diagnosis in stable phase of disease . however , in september 2010 the karyotype had changed to 46,xy , dup(9)(p21p21)/46,xy . since progressive bm failure was not evident until september 2010 , we considered this time point pivotal in aml transformation . consequently , we focused on identifying candidate aberrations of potential impact by applying acgh using human cancer cytochip at this time point . regions of gain or loss contained within copy number variable regions ( cnvs ) were discarded . high - resolution acgh analysis revealed an interstitial deletion in chromosome 8 at bands q23.2-q24.11 of maximal size 7,7  mb ( position 110.679.916118.334.826 ) ( fig . to longitudinally analyze for the del(8q ) , we performed fluorescence in situ hybridization ( fish ) with the following directly labeled probes : rp11 - 11a18 ( spectrumorange - labeled , bluegnome , cambridge , uk ) located at 8q23.3 ( position 113.491.837113.659.918 ) and centromere se 8 probe ( fitc - labeled , kreatech , amsterdam , the netherlands ) . the fish analysis confirmed the presence of del(8q ) in the progression- and aml phase and absence of del(8q ) in the stable phase of cmml in 2001 ( fig . array cgh did not detect the dup(9)(p21p21 ) found with standard g - banding karyotyping in september 2010 , since the fraction of dup(9 ) positive cells is below the sensitivity of acgh . we performed an ncbi gene database and pubmed search on each of these genes for their reported relevance in carcinogenesis . importantly , several of these are of notified relevance ; trps1 and eif3h are reported as candidate oncogenes with increased expression in breast cancer . in a study of non - small cell lung carcinomas the cub and sushi multiple domains 3 gene ( csmd3 ) was identified as the second most frequently mutated gene and loss of function cause increased proliferation of airway epithelial cells in vitro 
 . while the csmd gene family has recently been suggested tumor suppressor genes in colorectal cancer , the role of csmd3 in hematological cancers remains to be elucidated . strikingly , rad21 has been associated with aml as it encodes a protein in the cohesin complex , which is involved in chromosome segregation during cell division , regulation of transcription , and dna double strand break repair . recently , genes belonging to the cohesin complex were described as novel recurrent and mutually exclusive mutations in myeloid neoplasms . we have longitudinally described the molecular aberrations present at critical time points from initial diagnosis of cmml to overt aml applying a comprehensive panel of molecular analyses . firstly , the consistent and non - germ line tet2 + in all stages of disease verify that the aml stage indeed was transformed from cmml and not a de novo event . it can not be ruled out that the massive jak2 + allelic burden have contributed to genomic instability and , speculatively , also influenced the proliferative potential of the leukemic blasts . our results indicate that the leukemic transformation of an otherwise long - standing and stable cmml could have been driven by loss of gene function resulting from this exact deletion . of the deleted genes , rad21 and csmd3 are of particular interest , since their loss of function have recently been reported to play a role in carcinogenesis . finally , this case report emphasizes the relevance of acgh in the search for candidate genes involved in leukemogenesis , not least in heterogeneous neoplasms such as cmml . we wish to confirm that there are no known conflicts of interest associated with this publication . there has been no significant financial support for this work that could have influenced its outcome . we confirm that the manuscript has been read and approved by all named authors and that there are no other persons who satisfied the criteria for authorship . we further confirm that the order of authors listed in the manuscript has been approved by all of us . we confirm that we have given due consideration to the protection of intellectual property associated with this work and that there are no impediments to publication , including the timing of publication . , from the department of pathology , aarhus university hospital for helping with dna extraction from paraffin - embedded tissue and marcus celik hansen , m.sc . , for graphical support . the karen elise jensen foundation and the danish cancer society ( grant no .