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chronic myeloproliferative disorders. Br J Haematol 2007; 136: 745–751. Delhommeau F, Dupont S, Tonetti C, Masse A, Godin I, Le Couedic JP et al. Evidence that the JAK2 G1849T (V617F) mutation occurs in a lymphomyeloid progenitor in polycythemia vera and idiopathic myelofibrosis. Blood 2007; 109: 71–77. Pardanani A, Lasho TL, Finke C, Mesa RA, Hogan WJ, Ketterling RP et al. Extending Jak2V617F and MplW515 mutation analysis to single hematopoietic colonies and B and T lymphocytes. Stem Cells 2007; 25: 2358–2362. Bagasra O. Protocols for the in situ PCR-amplification and detection of mRNA and DNA sequences. Nat Protoc 2007; 2: 2782–2795. Gattenloehner S, Chuvpilo S, Langebrake C, Reinhardt D, MullerHermelink HK, Serfling E et al. Novel RUNX1 isoforms determine the fate of acute myeloid leukemia cells by controlling CD56 expression. Blood 2007; 110: 2027–2033. Schiller PI, Puchta U, Ogilvie AJ, Graf A, Kind P, Sander CA. [In situ PCR and PCR in situ hybridization of paraffin-embedded tissue. New diagnostic possibilities in pathology]. Pathologe 1998; 19: 313–317. Gattenlohner S, Peter C, Bonengel M, Einsele H, Bargou R, Muller-Hermelink HK et al. Detecting the JAK2 V617F mutation in fresh and ‘historic’ blood and bone marrow. Leukemia 2007; 21: 1599–1602. Baxter EJ, Scott LM, Campbell PJ, East C, Fourouclas N, Swanton S et al. Acquired mutation of the tyrosine kinase JAK2 in human myeloproliferative disorders. Lancet 2005; 365: 1054–1061.
10 Frantz C, Sekora DM, Henley DC, Huang CK, Pan Q, Quigley NB et al. Comparative evaluation of three JAK2V617F mutation detection methods. Am J Clin Pathol 2007; 128: 865–874. 11 Steensma DP. JAK2 V617F in myeloid disorders: molecular diagnostic techniques and their clinical utility: a paper from the 2005 William Beaumont Hospital Symposium on Molecular Pathology. J Mol Diagn 2006; 8: 397–411; quiz 526. 12 James C, Delhommeau F, Marzac C, Teyssandier I, Couedic JP, Giraudier S et al. Detection of JAK2 V617F as a first intention diagnostic test for erythrocytosis. Leukemia 2006; 20: 350–353. 13 Hussein K, Brakensiek K, Buesche G, Buhr T, Wiese B, Kreipe H et al. Different involvement of the megakaryocytic lineage by the JAK2 V617F mutation in polycythemia vera, essential thrombocythemia and chronic idiopathic myelofibrosis. Ann Hematol 2007; 86: 245–253. 14 Bonds LA, Barnes P, Foucar K, Sever CE. Acetic acid-zincformalin: a safe alternative to B-5 fixative. Am J Clin Pathol 2005; 124: 205–211. 15 Tbakhi A, Totos G, Hauser-Kronberger C, Pettay J, Baunoch D, Hacker GW et al. Fixation conditions for DNA and RNA in situ hybridization: a reassessment of molecular morphology dogma. Am J Pathol 1998; 152: 35–41. 16 Pellett PE, Spira TJ, Bagasra O, Boshoff C, Corey L, de Lellis L et al. Multicenter comparison of PCR assays for detection of human herpesvirus 8 DNA in semen. J Clin Microbiol 1999; 37: 1298–1301. 17 Piard F, Martin L, Chapusot C, Ponnelle T, Faivre J. [Genetic pathways in colorectal cancer: interest for the pathologist]. Ann Pathol 2002; 22: 277–288.
Simultaneous occurrence of acute myeloid leukaemia with mutated nucleophosmin (NPM1) in the same family
Leukemia (2009) 23, 199–203; doi:10.1038/leu.2008.170; published online 3 July 2008
Acute myeloid leukaemia (AML) with mutated nucleophosmin (NPM1), accounting for about 30% of adult de novo AML1 and 7% of childhood AML,2 displays distinctive biological and clinical characteristics.3 We report for the first time on the clinical, pathological and molecular features of NPM1-mutated AML which developed almost simultaneously (at an interval of only 2 months) in a father and his daughter, and discuss the biological significance of this finding. The father (SG) presented in April 2006, at the age of 33 years, because of fever. Laboratory examination revealed anaemia (8.5 g per 100 ml), moderate thrombocytopenia (74 000 per mm3), a white blood cell (WBC) count of 9820 per mm3 and a few blast cells in the peripheral blood smear. Bone marrow aspirate and trephine showed diffuse infiltration by blast cells that accounted for about 80% of the bone marrow population. Morphology (Figures 1a and b), immunophenotype of leukaemic cells, that is expression of myeloid (myeloperoxidase, CD33) and monocyte markers (macrophage-restricted CD68, CD14), were diagnostic of acute myelomonocytic leukaemia, according to the WHO (World Health Organization) classification. Leukaemic cells were CD34-negative (Figure 1c). Cytogenetics showed a normal karyotype (46, XY). Initially, no molecular studies for mutations of FLT3, NPM1 and CEBPA genes were performed. The patient was treated according to the GIMEMA (Gruppo Italiano Malattie Ematologiche dell’ Adulto) LAM 99-P protocol (Supplementary Materials). Bone marrow examinations carried out after induction and consolidation therapy showed complete remission (CR). The patient
underwent allogeneic peripheral blood stem cell (PBSC) transplantation from his 40-year-old HLA-identical brother (Supplementary Materials). Currently (June 2008), the patient is in CR. His daughter (SV), aged 7 years, presented in June 2006, because routine blood tests showed Hb 8.9 g per 100 ml, platelets 195 000 per mm3, and a WBC count of 46 920 per mm3 with 90% blasts. The morphological features (Figure 2a) and positivity for myeloperoxidase of a fraction of blast cells (not shown) indicated an AML without maturation, according to the WHO classification. Leukaemic cells were CD34 negative (Figure 2b). Cytogenetics showed a normal karyotype (46, XX). Molecular studies revealed an NPM1 gene mutation of type B (insertion of 4 bp CATG after nucleotide 1018 of the reference sequence, GenBank accession number NM_002520), which is one of the most common NPM1 mutation variants in childhood AML.4 Further studies also revealed an internal tandem duplication of the FLT3 gene (FLT3-ITD). The patient was treated according to the high-risk arm of the Associazione Italiana Ematologia Oncologia Pediatrica protocol LAM 2002/ 01 (Supplementary Materials). Complete haematological remission was followed by autologous PBSC transplantation (December 2006; Supplementary Materials). Bone marrow biopsy (February 2007) showed about 15% leukaemic cells. Rescue therapy was immediately started according to the I-BFMSG protocol but only the first FLAG (fludarabine, cytarabine, granulocyte-colony stimulating factor) cycle was administered because of severe neutropenia. In April 2007, bone marrow biopsy showed massive infiltration by leukaemic cells exhibiting the original morphological and phenotypic characteristics (myeloperoxidase positivity, CD34 negativity). In May 2007, the child underwent a haploidentical PBSC transplant from her Leukemia
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Figure 1 Bone marrow at initial diagnosis of AML (acute myeloid leukaemia, father). (a) Blasts with myelomonocytic appearance in bone marrow smear (May–Grunwald–Giemsa; 1000). (b) The myelomonocytic blasts show irregular nuclei in a paraffin section from a trephine biopsy (hematoxylin–eosin; 800). (c) Leukaemic cells are CD34 negative; the arrow indicates a CD34-positive vessel. (d) Immunostaining with anti-NPM monoclonal antibody reveals aberrant expression of nucleophosmin in the cytoplasm of the leukaemic cells (single arrow); residual normal hemopoietic cells show nucleus-restricted NPM positivity. (c and d) Alkaline phosphatase anti-alkaline phosphatase (APAAP) technique; paraffin-embedded sections from bone marrow trephine; hematoxylin counterstaining ( 800).
mother at the Division of Pediatric Oncology and Haematology, and Haematopoietic Stem Cell transplantation Unit of the University of Perugia, Italy (Supplementary Materials). A CR was sustained from June to November 2007. As bone marrow biopsy in December 2007 indicated AML had relapsed, the patient was treated with a donor lymphocyte infusion (5 105 per kg). In February 2008, bone marrow biopsy showed massive leukaemic infiltration. Molecular analyses confirmed the presence of NPM1 and FLT3-ITD mutations. In March 2008, the child received unmanipulated bone marrow stem cells from the same donor after conditioning with treosulfan, thiotepa and clofarabine. She developed grade II acute graft-versus-host disease and achieved a CR with a full-donor type chimerism. Finding that the daughter’s leukaemic cells harboured a NPM1 mutation and both father’s and daughter’s showed normal karyotype and CD34 negativity (which are distinguishing features of AML with mutated NPM11), prompted us to retrospectively investigate the father’s leukaemic cells, which had been frozen at diagnosis of AML in April 2006. NPM1 gene sequencing revealed a 4 bp (TCTG) insertion after nucleotide 1018 of the reference sequence (GenBank accession number NM_002520). This corresponds to NPM1 mutation A,1 the most common type of NPM1 mutation in adult AML, which is found in 75–80% of cases.3 The father’s leukaemic cells, unlike his daughter’s, carried no FLT3-ITD mutation. In conclusion, the daughter’s AML cells were NPM1 mutated (type B) and FLT3-ITD positive whereas her father’s were NPM1 mutated (type A) and harboured two FLT3 gene alleles in germline configuration. These different molecular profiles of leukaemic cells in father and daughter suggested that NPM1 mutations were not inherited. Immunostaining with a mouse monoclonal anti-NPM antibody of bone marrow at diagnosis (in father and daughter) and relapse (daughter) showed aberrant nucleophosmin expression in the cytoplasm of leukaemic cells (Figures 1d and 2c). This Leukemia
expression pattern is predictive of a mutated NPM1 gene.5 In contrast, normal residual cells (haemopoietic precursors, vessel endothelial cells, fibroblasts and osteoblasts) showed a nucleusrestricted nucleophosmin expression (Figures 1d, 2c and d) which is predictive of NPM1 gene in germline configuration.5 Nuclear NPM positivity was also found in all normal cells during CR (not shown). Immunohistochemical results were confirmed by molecular studies that detected an NPM1 mutation in leukaemic cells but not in bone marrow mononuclear cells taken during CR (Figure 3). None of the other family members we studied (I,1FI,2FII,1FIII,2) showed NPM1 or FLT3-ITD mutations (Figure 3), proving the NPM1 mutation was somatic in origin in father and daughter. Thus, the mutation pattern in our patients is quite different from what is observed in familial leukaemia associated with heterozygous mutations of genes encoding the runt-related transcription factor 1 (RUNX1) or CEBPA, both of which are of germline type and linked to familial susceptibility to AML.6,7 To further clarify the significance of our findings, we used the Affymetrix GeneChip 250K SNP (single-nucleotide polymorphism) mapping array technology and CNAG (v3) software8 to perform an high-throughput and genome-wide profiling of chromosomal alterations and loss of heterozygosity (LOH) events in several family members. In the father’s cells we observed an altered status of chromosome 3p (II,2), which had not been detected by routine cytogenetics. A hemizygous deletion from 3p14.1 to 3p12.3 (from 67 525 645 to 79 347 258 bp of Human May 2004 Assembly, 11.8 Mb in length) was found at diagnosis but not at remission (Figure 4), which suggested it was a tumour-specific alteration. Moreover, a large LOH region, spanning from 3p23 to 3p22.1 (from 32 328 557 to 43 502 256 bp, 11 Mb in length), was present at diagnosis and maintained during remission (Figure 4). This germline LOH event was associated with combined deletion of one allele and duplication of the other, thus preserving a diploid
Letters to the Editor
201 genes, which could be affected by mutations, or they could contain loci related to disease predisposing events. None of the other family members presented this peculiar combination of chromosome 3 alterations. No recurrent-specific DNA aberrations that might suggest familial predisposing genetic factors were identified. Moreover, there was no history of leukaemias or other malignant diseases in the family tree.
copy number status (defined as copy number neutral LOH). Stretches of homozygosity have been found through the genome of normal individuals.9 These traits of homozygosity can originate from uniparental dysomy, but most probably they represent autozygosity traits found in region of broad linkage disequilibrium and low frequency of recombination, due to presence of a common ancestor and consanguinity. These regions might include
Figure 2 Bone marrow at first relapse of AML (acute myeloid leukaemia, daughter). (a) Diffuse marrow infiltration by myeloid blasts without maturation (paraffin section from bone marrow trephine; hematoxylin–eosin; 800). (b) Leukaemic cells are CD34 negative; the arrow indicates a CD34-positive vessel. (c and d) Immunostaining with anti-NPM monoclonal antibody reveals aberrant expression of nucleophosmin in the cytoplasm of leukaemic cells (single arrows); normal endothelial cells in a vessel (c; double arrows) and residual normal haemopoietic cells, including macrophage (d; double arrows) show nucleus-restricted NPM positivity. (c and d) Alkaline phosphatase anti-alkaline phosphatase (APAAP) technique; paraffin-embedded sections from bone marrow trephine; hematoxylin counterstaining ( 800).
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Figure 3 NPM1 mutation detection in family members. Exon 12 and flanking intron sequences of the NPM1 gene were screened for mutations by DHPLC (Wave System; MD Transgenomic Inc., Omaha, NE, USA) and direct sequencing (data not shown) in all family members, including diagnostic and remission samples of the affected patients, II,2 and III,1. Electropherograms from patients’ samples were compared with normal sequenced controls. Chromatograms obtained with DHPLC technology are shown beside each family member. Sequencing results are indicated on the bottom of each chromatogram. Nucleotide numbers are referred to the GenBank NPM1 wild-type sequence NM_002520. Leukemia
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202 Chromosome 3
II,2. Diagnosis
CNN-LOH; (3)(p22.1p23) del(3)(p12.3p14.1) II,2. Remission
Figure 4 Map of DNA alterations on chromosome 3 in father’s cells at diagnosis and remission. DNA from father’s cells at diagnosis (II,2 diagnosis) and remission (II,2 remission) were analysed on Affymetrix GeneChip 250K Nsp SNP Mapping Arrays. Whole-genome DNA profiles were assembled using Copy Number Analyzer for Affymetrix GeneChip (CNAG, v3) software and comparing each sample to a pool of normal controls (48 HapMap samples available on Affymetrix website). Chromosome 3 map is shown from p to q end (from right to left) in both samples. The upper two graphs represent single SNP copy number signals on log scale (red dots) and total copy number values averaged on adjacent 10 SNPs (blue lines), whereas copy number values for each allele (red and green lines) are shown below. Green and pink bars in the middle represent heterozygous genotype calls and homozygous calls between each father’s sample and normal controls, respectively. The two bars at the bottom represent the colour-coded visualisation of total copy number status (yellow, diploidy; pink, amplification) and LOH (loss of heterozygosity; blue, significant LOH; yellow, no LOH). Deletion at 3p14.1–12.3 (from 67 525 645 to 79 347 258 bp, according to UCSC Genome Browser, Human Assembly May 2004) observed in II,2-diagnostic sample and CNN-LOH at 3p23–22.1 (from 32 328 557 to 43 502 256 bp) observed both in II,2diagnosis and II,2-remission samples are shown.
The apparent absence of genetic predisposing factors concurs with the observation that the NPM1 mutation seems to be a founder genetic lesion rather than a secondary event, as it is specific for AML,1,10 does not usually associate with secondary AML,1,11 is mutually exclusive of other recurrent genetic abnormalities (except for rare cases bearing NPM1 and CEPBA mutations),12 is stable over the course of the disease13 and is associated with distinctive gene expression profile3 and microRNA signature.14 However, although the SNP array did not identify genetic predisposing factors, we cannot exclude that our patients carried leukaemia susceptibility genes whose alterations could not be detected by SNP arrays (as in case of balanced translocations, inversions or point mutations) or that might have been affected by epigenetic changes. An alternative hypothesis of the almost simultaneous occurrence of AML in father and daughter is exposure to a common environmental agent. Investigation into family occupations and workplace revealed that father and mother managed a cafeteria within a gasoline station for a period of about 10 years, where their daughter used to spend time almost every day after school. As exposure to benzene is related to higher risk of leukaemia,15 it could be speculated that low but continuous exposure to benzene from gasoline might have played a role in the development of AML in our patients. On the other hand, although extremely improbable, disease onset in father and daughter might just have been coincidental. Thus, despite extensive investigation, the almost simultaneous occurrence of AML with mutated NPM1 in the two family members remains unexplained. Father and daughter both underwent allogeneic PBSC transplantation but with different outcomes. The father’s Leukemia
favourable outcome was expected as leukaemic cells carried an NPM1 mutation in the absence of FLT3-ITD.3 Assessment of minimal residual disease by quantitative PCR after allogeneic transplant revealed no NPM1 mutant copies, which is indicative of complete molecular remission and may be predictive of long-term survival.16 In contrast, AML in the daughter was characterized by an initial response to therapy followed by early relapses, even after haploidentical PBSC transplantation. Her unfavourable prognosis was probably due to co-existence of an FLT3-ITD (documented at diagnosis and during relapses), which is known to counteract the favourable prognostic impact of NPM1 mutations.3
Acknowledgements Supported by the Associazione Italiana per la Ricerca sul Cancro (AIRC), MIUR PRIN and Fondazione Cariplo (to GC and AB). LLN was supported by IBISCUS. This work was supported by MIUR (Ministero Universita` e Ricerca) through a FIRB grant RBLA03ER38 and funds to Interdisciplinary Center for Biomolecular Studies and Industrial Applications (CISI) and Department of Biomedical Sciences and Technologies, University of Milan, Italy. We thank Dr GA Boyd for editorial assistance. B Falini has applied for a patent on the clinical use of NPM mutants.
G Cazzaniga1,8, L Lo Nigro2,8, I Cifola3, G Milone4, S Schnittger5, T Haferlach5, E Mirabile2, F Costantino6, MP Martelli7, E Mastrodicasa7, F Di Raimondo4, F Aversa7, A Biondi1 and B Falini7 1 Centro Ricerca Tettamanti, Clinica Pediatrica, Ospedale San Gerardo, Universita` di Milano-Bicocca, Monza, Italy;
Letters to the Editor 2
Center of Pediatric Haematology Oncology, University of Catania, Catania, Italy; 3 Institute of Biomedical Technologies (ITB), National Research Council (CNR), Milan, Italy; 4 Institute of Haematology, Ospedale Ferrarotto, University of Catania, Catania, Italy; 5 MLL Munich Leukemia Laboratory, Munich, Germany; 6 Ospedale Cannizzaro, Catania, Italy and 7 Section of Haematology and Immunology, IBit Foundation, University of Perugia, Perugia, Italy 8 These authors contributed equally to this work. E-mail:
[email protected] References 1 Falini B, Mecucci C, Tiacci E, Alcalay M, Rosati R, Pasqualucci L et al. Cytoplasmic nucleophosmin in acute myelogenous leukemia with a normal karyotype. N Engl J Med 2005; 352: 254–266. 2 Cazzaniga G, Dell’Oro MG, Mecucci C, Giarin E, Masetti R, Rossi V et al. Nucleophosmin mutations in childhood acute myelogenous leukemia with normal karyotype. Blood 2005; 106: 1419–1422. 3 Falini B, Nicoletti I, Martelli MF, Mecucci C. Acute myeloid leukemia carrying cytoplasmic/mutated nucleophosmin (NPMc+ AML): biologic and clinical features. Blood 2007; 109: 874–885. 4 Thiede C, Creutzig E, Reinhardt D, Ehninger G, Creutzig U. Different types of NPM1 mutations in children and adults: evidence for an effect of patient age on the prevalence of the TCTG-tandem duplication in NPM1-exon 12. Leukemia 2007; 21: 366–367. 5 Falini B, Martelli MP, Bolli N, Bonasso R, Ghia E, Pallotta MT et al. Immunohistochemistry predicts nucleophosmin (NPM) mutations in acute myeloid leukemia. Blood 2006; 108: 1999–2005. 6 Song WJ, Sullivan MG, Legare RD, Hutchings S, Tan X, Kufrin D et al. Haploinsufficiency of CBFA2 causes familial thrombocytopenia with propensity to develop acute myelogenous leukaemia. Nat Genet 1999; 23: 166–175. 7 Smith ML, Cavenagh JD, Lister TA, Fitzgibbon J. Mutation of CEBPA in familial acute myeloid leukemia. N Engl J Med 2004; 351: 2403–2407.
203 8 Yamamoto G, Nannya Y, Kato M, Sanada M, Levine RL, Kawamata N et al. Highly sensitive method for genomewide detection of allelic composition in nonpaired, primary tumor specimens by use of affymetrix single-nucleotide-polymorphism genotyping microarrays. Am J Hum Genet 2007; 81: 114–126. 9 Simon-Sanchez J, Scholz S, Fung HC, Matarin M, Hernandez D, Gibbs JR et al. Genome-wide SNP assay reveals structural genomic variation, extended homozygosity and cell-line induced alterations in normal individuals. Hum Mol Genet 2007; 16: 1–14. 10 Liso A, Bogliolo A, Freschi V, Martelli MP, Pileri SA, Santodirocco M et al. In human genome, generation of a nuclear export signal through duplication appears unique to nucleophosmin (NPM1) mutations and is restricted to AML. Leukemia 2008; 22: 1285–1289. 11 Pasqualucci L, Li S, Meloni G, Schnittger S, Gattenlohner S, Liso A et al. NPM1-mutated acute myeloid leukaemia occurring in JAK2-V617F+ primary myelofibrosis: de-novo origin? Leukemia advance online publication, 17 January 2008; doi:10.1038/ sj.leu.2405093. 12 Falini B, Mecucci C, Saglio G, Lo Coco F, Diverio D, Brown P et al. NPM1 mutations and cytoplasmic nucleophosmin are mutually exclusive of recurrent genetic abnormalities: a comparative analysis of 2562 patients with acute myeloid leukemia. Haematologica 2008; 93: 439–442. 13 Falini B, Martelli MP, Mecucci C, Liso A, Bolli N, Bigerna B et al. Cytoplasmic mutated nucleophosmin is stable in primary leukemic cells and in a xenotransplant model of NPMc+ acute myeloid leukemia in SCID mice. Haematologica 2008; 93: 775–779. 14 Garzon R, Garofalo M, Martelli MP, Briesewitz R, Wang L, Fernandez-Cymering C et al. Distinctive microRNA signature of acute myeloid leukemia bearing cytoplasmic mutated nucleophosmin. Proc Natl Acad Sci USA 2008; 105: 3945–3950. 15 Glass DC, Gray CN, Jolley DJ, Gibbons C, Sim MR, Fritschi L et al. Leukemia risk associated with low-level benzene exposure. Epidemiology 2003; 14: 569–577. 16 Chou WC, Tang JL, Wu SJ, Tsay W, Yao M, Huang SY et al. Clinical implications of minimal residual disease monitoring by quantitative polymerase chain reaction in acute myeloid leukemia patients bearing nucleophosmin (NPM1) mutations. Leukemia 2007; 21: 998–1004.
Supplementary Information accompanies the paper on the Leukemia website (http://www.nature.com/leu)
TP53 gene mutation is frequent in patients with acute myeloid leukemia and complex karyotype, and is associated with very poor prognosis
Leukemia (2009) 23, 203–206; doi:10.1038/leu.2008.173; published online 3 July 2008
Risk-adapted therapy is now standard practice in acute myeloid leukemia (AML), with cytogenetics the main determinant of risk. Patients with good risk cytogenetics, t(15;17), t(8;21), inv(16), do not usually receive a stem cell transplant in first complete remission (CR), whereas consolidation with an allogeneic transplantation is often recommended for younger patients with intermediate and adverse cytogenetics.1 A number of mutations have also been described in genes such as fms-like tyrosine kinase-3 (FLT3) and nucleophosmin 1 (NPM1), which have prognostic significance and enable substratification of patients in the intermediate cytogenetic risk group.2 They are, however, rare in adverse risk cases, which include 7, 5, 5q, abnormalities of 3q and those with a complex karyotype. Several studies have shown that TP53 mutations are frequent in cases of therapy-related AML and myelodysplasia, and are particularly associated with loss of 5q and a complex karyotype.3
Loss of chromosome 17 material, including the region of the TP53 gene, has been observed in a number of cases. A high incidence of TP53 mutations in cases of primary AML with a complex karyotype has also been reported4 but no large cohort has been reported to date with comprehensive outcome data. In this study, we examined the incidence and prognostic impact of TP53 mutations in a cohort of patients with adverse cytogenetics and those with losses on chromosome 17p who were entered into the UK Medical Research Council adult AML trials. We also included cases of 7q, as patients with this abnormality have an outcome at the border between the adverse and intermediate risk groups. We studied 166 patients with cytogenetic abnormalities of chromosomes 5, 7 or 17, or a complex karyotype (5 or more abnormalities) who had been entered into Medical Research Council AML clinical trials and had diagnostic DNA. Of these, 148 had adverse, 11 had intermediate and 7 had favorable risk cytogenetics by Medical Research Council classification. Details of chemotherapy schedules have been published previously5,6 and the majority of patients received intensive chemotherapy. Patient Leukemia
