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The roles of RNA 5-methylcytosine (RNA:m5C) and RNA:m5C methyltransferases (RCMTs) in lineage-associated chromatin organization and drug response/resistance are unclear. Here we demonstrate that the RCMTs, namely NSUN3 and DNMT2, directly bind hnRNPK, a conserved RNA-binding protein. hnRNPK interacts with the lineage-determining transcription factors (TFs), GATA1 and SPI1/PU.1, and with CDK9/P-TEFb to recruit RNA-polymerase-II at nascent RNA, leading to formation of 5-Azacitidine (5-AZA)-sensitive chromatin structure. In contrast, NSUN1 binds BRD4 and RNA-polymerase-II to form an active chromatin structure that is insensitive to 5-AZA, but hypersensitive to the BRD4 inhibitor JQ1 and to the downregulation of NSUN1 by siRNAs. Both 5-AZA-resistant leukaemia cell lines and clinically 5-AZA-resistant myelodysplastic syndrome and acute myeloid leukaemia specimens have a significant increase in RNA:m5C and NSUN1-/BRD4-associated active chromatin. This study reveals novel RNA:m5C/RCMT-mediated chromatin structures that modulate 5-AZA response/resistance in leukaemia cells, and hence provides a new insight into treatment of leukaemia.

Introduction Cutaneous composite lymphoma (CCL) is extremely rare. When 2 potentially distinct lymphoid lesions occur at one skin site, distinguishing between one neoplastic clone and a secondary reactionary lymphoid response versus a second neoplasm is difficult. In this study, we describe a unique case of CCL along with a review of reported cases in literature to identify clues and discuss issues that are relevant to the diagnosis of CCL. Design Review of a CCL case from our institution and a systematic review of reported cases of CCL in the literature. Results A total of 18 studies describing 22 cases and a case report from our institution are included. The mean age at diagnosis was 68 years. Most cases herein presented with multiple skin lesions (67%) and reported a history of immune suppression (76%). Nineteen cases (83%) had a combination of T-cell and B-cell neoplasms, whereas the remaining cases had 2 distinct B-cell clones. Clonal differentiation was confirmed based on morphology and immunohistochemistry in all cases, and by polymerase chain reaction studies in 19 cases. Complete remission was achieved in only one quarter of reported cases. Conclusion Diagnosing CCL can be challenging because accurate differentiation of 2 or more clonal populations at 1 site is tedious. A stepwise approach and integration of clinical, morphologic, immunohistochemistry, and molecular data along with an understanding of the prognosis of the lymphomas in question is essential for an accurate diagnosis and necessary because of therapeutic and prognostic implications.

The roles of RNA 5-methylcytosine (RNA:m5C) and RNA:m5C methyltransferases (RCMTs) in lineage-associated chromatin organization and drug response/resistance are unclear. Here we demonstrate that the RCMTs, namely NSUN3 and DNMT2, directly bind hnRNPK, a conserved RNA-binding protein. hnRNPK interacts with the lineage-determining transcription factors (TFs), GATA1 and SPI1/PU.1, and with CDK9/P-TEFb to recruit RNA-polymerase-II at nascent RNA, leading to formation of 5-Azacitidine (5-AZA)-sensitive chromatin structure. In contrast, NSUN1 binds BRD4 and RNA-polymerase-II to form an active chromatin structure that is insensitive to 5-AZA, but hypersensitive to the BRD4 inhibitor JQ1 and to the downregulation of NSUN1 by siRNAs. Both 5-AZA-resistant leukaemia cell lines and clinically 5-AZA-resistant myelodysplastic syndrome and acute myeloid leukaemia specimens have a significant increase in RNA:m5C and NSUN1-/BRD4-associated active chromatin. This study reveals novel RNA:m5C/RCMT-mediated chromatin structures that modulate 5-AZA response/resistance in leukaemia cells, and hence provides a new insight into treatment of leukaemia., Resistance to chemotherapy is a serious issue that can be influenced by RNA epigenetics and chromatin structure. Here, the authors show in leukaemia cells that RNA 5-methylcytosine (RNA:m5C) and RNA:m5C methyltransferases (RCMTs) mediate chromatin structures that can modulate 5-Azacitidine response and resistance.

Nature Communications 3: Article number: 688 (2012); Published: 21 February 2012; Updated: 10 April 2018 The original HTML version of this Article had an incorrect volume number of 2; it should have been 3. This has now been corrected in the HTML; the PDF version of the Article was correct from the time of publication.

Acute myeloid leukemia and myelodysplastic syndrome with inv(3)(q21q26.2)/t(3;3)(q21;q26.2) have a poor prognosis. Indeed, the inv(3)(q21q26.2)/t(3;3)(q21;q26.2) has been recognized as a poor risk karyotype in the revised International Prognostic Scoring System. However, inv(3)(q21q26.2)/t(3;3)(q21;q26.2) is not among the cytogenetic abnormalities pathognomonic for diagnosis of acute myeloid leukemia irrespective of blast percentage in the 2008 WHO classification. This multicenter study evaluated the clinico-pathological features of acute myeloid leukemia/myelodysplastic syndrome patients with inv(3)(q21q26.2)/t(3;3)(q21;q26.2) and applied the revised International Prognostic Scoring System to myelodysplastic syndrome patients with inv(3)(q21q26.2)/t(3;3)(q21;q26.2). A total of 103 inv(3)(q21q26.2)/t(3;3)(q21;q26.2) patients were reviewed and had a median bone marrow blast count of 4% in myelodysplastic syndrome (n=40) and 52% in acute myeloid leukemia (n=63) (P

In this study we investigated the distribution of IgG4+ plasma cells and regulatory T (TREG) cells, a major regulator of IgG4 production, in nodal and extranodal Rosai-Dorfman disease (RDD). Twenty-six specimens (15 nodal, 11 extranodal) were examined, with reactive lymph nodes and site-matched extranodal specimens as controls. Overall, 84.6% (22/26) of the specimens showed various degrees of sclerosis (7 mild, 8 moderate, and 7 severe). Nineteen cases (73.1%) exhibited more than 10 IgG4+ cells/0.060 mm2 (photographed area at ×40), and 8 cases (30.8%) showed more than 40% of IgG+ cells being IgG4+. Only 1 control case exhibited more than 10 IgG4+ cells/0.060 mm2 (P < .05). The number of TREG cells was comparable between nodal RDD and controls, whereas extranodal RDD exhibited significantly higher numbers of TREG cells than controls. These findings demonstrate that a subset of RDD shows features of IgG4-related disease and indicate an overlap between certain aspects of the 2 diseases.

We have identified a family in which three members developed B-cell lymphoproliferative disorders within a nine month period. The 33 year old proband and his mother have hairy cell leukemia, and his 37 year old brother developed a large cell lymphoma. Chromosomal fragile site analysis of peripheral blood lymphocytes of the three patients as well as two healthy family members was performed. The mean number of rare fragile sites present per cell analyzed was not significantly different from that observed in a group of healthy adults used as controls. However, the level of expression of the common fragile sites detected in each of the study patients was significantly elevated compared to the control population. Although the relationship between the level of expression of common fragile sites and the subsequent development of a malignant process is unknown, many of the agents that induce these sites are known mutagens, and the level of their expression may reflect a genetic susceptibility to mutagenic damage. Thus, it is possible that common exposure to an environmental mutagen may have contributed to the temporal clustering of malignancy in this family.

The World Health Organization (WHO) classification of tumors of the hematopoietic and lymphoid tissues was last updated in 2008. Since then, there have been numerous advances in the identification of unique biomarkers associated with some myeloid neoplasms and acute leukemias, largely derived from gene expression analysis and next-generation sequencing that can significantly improve the diagnostic criteria as well as the prognostic relevance of entities currently included in the WHO classification and that also suggest new entities that should be added. Therefore, there is a clear need for a revision to the current classification. The revisions to the categories of myeloid neoplasms and acute leukemia will be published in a monograph in 2016 and reflect a consensus of opinion of hematopathologists, hematologists, oncologists, and geneticists. The 2016 edition represents a revision of the prior classification rather than an entirely new classification and attempts to incorporate new clinical, prognostic, morphologic, immunophenotypic, and genetic data that have emerged since the last edition. The major changes in the classification and their rationale are presented here.

BACKGROUND: Cancer and Leukemia Group B (CALGB) Study 19802, a phase 2 study, evaluated whether dose intensification of daunorubicin and cytarabine could improve disease-free survival (DFS) in adults with acute lymphoblastic leukemia (ALL) and whether high-dose systemic and intrathecal methotrexate could replace cranial radiotherapy for central nervous system (CNS) prophylaxis. METHODS: One hundred sixty-one eligible, previously untreated patients ages 16 to 82 years (median age, 40 years) were enrolled, and 33 (20%) were aged ≥60 years. RESULTS: One hundred twenty-eight patients (80%) achieved complete remission (CR). Dose intensification of daunorubicin and cytarabine was feasible. At a median follow-up of 10.4 years for surviving patients, the 5-year DFS rate was 25% (95% confidence interval, 18%-33%), and the overall survival (OS) rate was 30% (95% confidence interval, 23%-37%). Patients aged

The myelodysplastic/myeloproliferative neoplasms (MDS/MPN) include clonal myeloid neoplasms that overlap the MDS and MPN categories and at the time of initial diagnosis exhibit some clinical, laboratory, or morphologic features supporting the diagnosis of myelodysplastic syndrome (MDS) and at the same time show proliferative features in keeping with the diagnosis of a myeloproliferative neoplasm (MPN). Although the clinical, morphologic, and laboratory findings vary along a continuum from MDS to MPN, distinctive features are usually present that allow assignment of most of the cases to 1 of 3 distinct subtypes recognized by the 2008 World Health Organization (WHO) classification: chronic myelomonocytic leukemia (CMML), atypical chronic myeloid leukemia, BCR-ABL - (aCML, BCR-ABL1 - ), and juvenile myelomonocytic leukemia (JMML). The WHO classification also recognizes a provisional category of the MDS/MPN, unclassifiable (MDS/MPN, U), including the provisional entity of refractory anemia with ring sideroblasts and thrombocytosis (RARS-T). In the past 2 to 3 years since the publication of the WHO classification in 2008, dynamic progress in array technologies and next-generation amplicon deep sequencing has provided new insights into the molecular pathogenesis of MDS/MPN, especially CMML and JMML. In this review we will give an overview of these neoplasms and focus on adult MDS/MPN, especially CMML. We will give only brief updates for aCML and RARS-T; JMML will be discussed in a separate article.

Cancer and Leukemia Group B 19808 (CALGB 19808) is the only randomized trial of a second-generation P-glycoprotein (Pgp) modulator in untreated patients with acute myeloid leukemia (AML) younger than age 60 years. We randomly assigned 302 patients to receive induction chemotherapy regimens consisting of cytosine arabinoside (Ara-C; A), daunorubicin (D), and etoposide (E), without (ADE) or with (ADEP) PSC-833 (P). The incidence of complete remission was 75% with both regimens. Reversible grade 3 and 4 liver and mucosal toxicities were significantly more common with ADEP. Therapy-related mortality was 7% and did not differ by induction arm. Excess cardiotoxicity was not seen with high doses of D in ADE. The median disease-free survival was 1.34 years in the ADE arm and 1.09 years in the ADEP arm (P = .74, log-rank test); the median overall survival was 1.86 years in the ADE arm and 1.69 years in the ADEP arm (P = .82). There was no evidence of a treatment difference within any identifiable patient subgroup. Inhibition of Pgp-mediated drug efflux by PSC-833 did not improve clinical outcomes in younger patients with untreated AML. This trial was registered at www.clinicaltrials.gov as #NCT00006363.

The World Health Organization (WHO) classification of myeloid and lymphoid neoplasms utilizes morphology, immunophenotype, genetics and clinical features to define disease entities of clinical significance. It is a consensus classification in which a number of experts have agreed on the classification and diagnostic criteria. In general, the classification stratifies neoplasms according to their lineage (myeloid, lymphoid, histiocytic/dendritic) and distinguishes neoplasms of precursor cells from those comprised of functionally mature cells. Lymphoid neoplasms are derived from cells that frequently have features that recapitulate stages of normal B-, T-, and NK-cell differentiation and function, so to some extent they can be classified according to the corresponding normal counterpart, although additional features, such as genotype, clinical features and even location of the tumor figure into the final classification listing as well. Five major subgroups of myeloid neoplasms are recognized based mainly on their degree of maturation and biologic properties: myeloproliferative neoplasms (MPNs) which are comprised primarily of mature cells with effective proliferation; myeloid (and lymphoid) neoplasms with eosinophilia and abnormalities of PDGFRA, PDGFRB and FGFR1, defined largely by the finding of significant eosinophilia and specific genetic abnormalities; myelodysplastic/myeloproliferative neoplasms (MDS/MPN), comprised mainly of mature cells with both effective and ineffective proliferation of various lineages; myelodysplastic syndromes (MDS), in which immature and mature cells are found with abnormal, dysplastic and ineffective maturation, and acute myeloid leukemia (AML), comprised of precursor cells with impaired maturation. Genetic abnormalities play an important role as diagnostic criteria for further sub-classification of some myeloid neoplasms, particularly of AML. Although therapy-related MDS and AML (t-MDS/AML) often have genetic defects identical to those found in de novo AML and de novo MDS, they are classified separately from de novo AML and MDS in order to emphasize their unique clinical and biologic properties.

Purpose Thalidomide and lenalidomide can alleviate anemia in myelofibrosis. However, their value is undermined by their respective potential to cause peripheral neuropathy and myelosuppression. We therefore evaluated the safety and therapeutic activity of another immunomodulatory drug, pomalidomide. Methods In a phase II randomized, multicenter, double-blind, adaptive design study, four treatment arms were evaluated: pomalidomide (2 mg/d) plus placebo, pomalidomide (2 mg/d) plus prednisone, pomalidomide (0.5 mg/d) plus prednisone, and prednisone plus placebo. Pomalidomide was administered for up to 12 28-day treatment cycles. Prednisone (30 mg/d) was given in a tapering dose schedule during the first three cycles. Response was assessed by International Working Group criteria. Results Eighty-four patients with myelofibrosis-associated anemia were randomly assigned to the aforementioned treatment arms: 22, 19, 22, and 21, respectively. Response in anemia was documented in 20 patients, including 15 who became transfusion independent. Response rates in the four treatment arms were 23% (95% CI, 5% to 41%), 16% (95% CI, 0% to 33%), 36% (95% CI, 16% to 56%), and 19% (95% CI, 2% to 36%). The corresponding figures for patients receiving ≥ 3 cycles of treatment (n = 62) were 38%, 23%, 40%, and 25%. Response to pomalidomide with or without prednisone was durable (range, 3.2 to 16.9+ months) and significantly better in the absence of leukocytosis (37% v 8%; P = .01); JAK2V617F or cytogenetic status did not affect response. Grade ≥ 3 toxicities were infrequent and included (in each treatment arm) neutropenia (9%; 16%; 5%; 5%), thrombocytopenia (14%; 16%; 9%; 5%), and thrombosis (9%; 5%; 0%; 0%). Conclusion Pomalidomide therapy at 0.5 or 2 mg/d with or without an abbreviated course of prednisone is well tolerated in patients with myelofibrosis and active in the treatment of anemia.

Recently the World Health Organization (WHO), in collaboration with the European Association for Haematopathology and the Society for Hematopathology, published a revised and updated edition of the WHO Classification of Tumors of the Hematopoietic and Lymphoid Tissues. The 4th edition of the WHO classification incorporates new information that has emerged from scientific and clinical studies in the interval since the publication of the 3rd edition in 2001, and includes new criteria for the recognition of some previously described neoplasms as well as clarification and refinement of the defining criteria for others. It also adds entities—some defined principally by genetic features—that have only recently been characterized. In this paper, the classification of myeloid neoplasms and acute leukemia is highlighted with the aim of familiarizing hematologists, clinical scientists, and hematopathologists not only with the major changes in the classification but also with the rationale for those changes.

BACKGROUD: The Cancer and Leukemia Group B evaluated oral topotecan administered at 2 schedules and doses for myelodysplastic syndrome (MDS). METHODS: Patients with previously untreated primary or therapy-related MDS were eligible. Patients with refractory anemia (RA), RA with ringed sideroblasts, or refractory cytopenia with multilineage dysplasia (RCMD) were eligible only if they were dependent on erythrocyte transfusion, had a platelet count

We performed a retrospective comparison of presenting features, planned treatment, complete remission (CR) rate, and outcome of 321 adolescents and young adults (AYAs) ages 16 to 20 years with newly diagnosed acute lymphoblastic leukemia (ALL) who were treated on consecutive trials in either the Children's Cancer Group (CCG) or the Cancer and Leukemia Group B (CALGB) from 1988 to 2001. CR rates were identical, 90% for both CALGB and CCG AYAs. CCG AYAs had a 63% event-free survival (EFS) and 67% overall survival (OS) at 7 years in contrast to the CALGB AYAs, in which 7-year EFS was only 34% (P < .001; relative hazard rate [RHR] = 2.2) and OS was 46% (P < .001; RHR = 1.9). While CALGB AYAs aged 16 to 17 years achieved similar outcomes to all CCG AYAs with a 7-year EFS of 55%, the EFS for 18- to 20-year-old CALGB patients was only 29%. Comparison of the regimens showed that CCG AYAs received earlier and more intensive central nervous system prophylaxis and higher cumulative doses of nonmyelosuppressive agents. There were no differences in outcomes of those who reached maintenance therapy on time compared with those who were delayed. Based on these observations, a prospective study for AYAs with ALL using the more successful approach of the CCG has been initiated.

KIT mutations have been identified in several malignancies, including acute myeloid leukemia (AML) and systemic mastocytosis (SM). Mast cell leukemia (MCL) is the most aggressive mast cell neoplasm, but has not been well studied due to its rarity. We identified novel KIT transcripts in two patients with MCL and two patients with SM with an associated hematological disorder, but not from two patients with SM. Similar novel KIT transcripts were also observed in normal CD34+ cells from bone marrow and umbilical cord blood, suggesting that altered KIT isoforms may be specific to the blast stage of hematopoietic precursors. The novel KIT proteins lack several domains including the ATP binding site, and one was inactive in a functional test for autophosphorylation. Our discovery of novel KIT transcripts underscores the importance of analysing entire protein encoding regions when studying genes of interest.

Severe congenital neutropenia (SCN) is an inborn disorder of granulopoiesis. Like most other bone marrow failure syndromes, it is associated with a marked propensity to transform into a myelodysplastic syndrome (MDS) or acute leukemia, with a cumulative rate of transformation to MDS/leukemia that exceeds 20%. The genetic (and/or epigenetic) changes that contribute to malignant transformation in SCN are largely unknown. In this study, we performed mutational profiling of 14 genes previously implicated in leukemogenesis using 14 MDS/leukemia samples from patients with SCN. We used high-throughput exon-based resequencing of whole-genome–amplified genomic DNA with a semiautomated method to detect mutations. The sensitivity and specificity of the sequencing pipeline was validated by determining the frequency of mutations in these 14 genes using 188 de novo AML samples. As expected, mutations of tyrosine kinase genes (FLT3, KIT, and JAK2) were common in de novo AML, with a cumulative frequency of 30%. In contrast, no mutations in these genes were detected in the SCN samples; instead, mutations of CSF3R, encoding the G-CSF receptor, were common. These data support the hypothesis that mutations of CSF3R may provide the “activated tyrosine kinase signal” that is thought to be important for leukemogenesis.

Purpose of review The sighting of the Philadelphia chromosome in 1960, later shown to harbor the BCR-ABL mutation in chronic myeloid leukemia, is arguably the most seminal contribution to molecular oncology. In the decades that followed, other cytogenetic and molecular disease markers have been described and effectively incorporated into routine diagnostic tests. This review discusses how this process is unfolding in myeloproliferative disorders. Recent findings In 2003, a karyotypically-occult FIP1L1-PDGFRA was reported in a subset of patients with blood eosinophilia and bone marrow mastocytosis; this mutation has since joined several other molecular markers for eosinophilic (e.g. PDGFRbeta- and FGFR1-rearrangements) and mast cell (e.g. KITD816V) disorders. In 2005, JAK2V617F was described in polycythemia vera and other BCR-ABL myeloproliferative disorders; the particular discovery has already had a major impact on current diagnostic approaches in polycythemia vera. These remarkable molecular discoveries are both redefining and reinforcing the diagnostic role of bone marrow histopathology. Summary Recent progress in the molecular pathogenesis of myeloproliferative disorders calls for a paradigm shift in traditional diagnostics, which is based on subjective technologies or assignment to a 'consensus'-based ever-changing list of inclusionary and exclusionary criteria. Routine clinical practice might be better served by diagnostic algorithms that incorporate molecular disease markers, which complement histological impression.

In practice, cases of therapy-related myelodysplastic syndrome (t-MDS) are often classified according to morphologic schemes used for de novo MDS. However, there are few data addressing the appropriateness of such classification. We studied 155 patients with therapy-related acute myeloid leukemia (t-AML)/t-MDS to determine whether subclassification by the World Health Organization (WHO) criteria for de novo MDS provides prognostic information in t-MDS. In addition, we assessed whether cytogenetic stratification by the International Prognostic Scoring System (IPSS) guidelines or karyotypic complexity was prognostically important. We found no differences in median survival times among patients classified into the different WHO subgroup of MDS or according to their bone marrow blast percentage; our results indicate a uniformly poor outcome in t-MDS regardless of morphologic classification. However, significant survival differences correlated with cytogenetic stratification according to IPSS guidelines and/or karyotypic complexity. We found only a borderline difference in median survival of patients with an initial t-MDS diagnosis compared with patients with an initial t-AML diagnosis.

Objectives: Acute myeloid leukemia with myelodysplasia-related changes (AML-MRC) is a heterogeneous disorder defined by morphologic, genetic, or clinical features. Genetic abnormalities associated with AML-MRC are often associated with adverse prognostic features, and many cases are preceded by a myelodysplastic syndrome (MDS) or a myelodysplastic/myeloproliferative neoplasm. Methods: Although the criteria of 20% or more blasts in blood or bone marrow and multilineage dysplasia affecting 50% or more of cells in two or more of the myeloid lineages seem straightforward for AML-MRC, identification of morphologic dysplasia among observers is not always consistent, and there is morphologic overlap with other leukemic disorders such as acute erythroleukemia. Results: Session 3 of the workshop cases displayed heterogeneity as expected within AML-MRC, yet several cases suggested that recently recognized entities may exist within this category, such as familial MDS/AML predisposition syndromes and rare cases of high-risk AML associated with the cryptic t(5;11)(q35;p15); NUP98-NSD1 that may masquerade as a del(5q). However, most cases of AML-MRC were usually associated with adverse genetic abnormalities, particularly −5/del(5q), −7/del(7q), and/or complex karyotypes. Conclusions: Whole-genome sequencing and array studies may identify genetic abnormalities, such as those affecting TP53, which may provide prognostic information.

Background: Fanconi anemia (FA) is caused by mutations in one of seventeen genes that make up the FA DNA double strand break (DSB) repair pathway. Recently, two individuals with biallelic germline BRCA1 mutations, each consisting of one null and one hypomorphic mutation, were identified and noted to have features consistent with FA, including congenital anomalies and increased chromosomal breakage of lymphocytes on exposure to diepoxybutane (Domchek et al. Cancer Discov. 2013 Apr; (4):399-405; and Sawyer et al. Cancer Discov. Epub. 2014 Dec 3.), adding BRCA1 as the newest FA gene. However, neither patient developed bone marrow failure (BMF), making the bone marrow effects of BRCA1 deficiency still unclear. Methods: To test the hypothesis that Brca1 is also essential in hematopoiesis, we developed a conditional mouse model with Mx-1 Cre-mediated Brca1 deletion and examined the effects of Brca1 deficiency on hematopoiesis in this model. Results: At baseline, Brca1-/- mice have macrocytic anemia and leukopenia. Further, by 6 months of age, 30% and 50% of the Brca1-/- mice develop spontaneous BMF or hematologic malignancies (HM), respectively. Brca1-/- mice develop a diverse range of HM, including T-cell lymphomas and acute myeloid leukemias, suggesting a defect in an early hematopoietic progenitor population. Methylcellulose colony forming assays also demonstrate a defect in progenitor cell function with Brca1-/- bone marrow cells forming fewer colonies (44.4±31.9) than Brca1+/+ cells (200.3±30.5, p=0.004) at baseline, and show FA-like hypersensitivity to the DNA cross-linking agent, Mitomycin C (MMC) (mean colony survival % at 10 nM MMC 40% versus 82% and at 50 nM 1% vs 56%). Spectral karyotyping of bone marrow cells from mice that developed BMF demonstrated chromatid exchanges and breaks. Similarly, multiple chromosomal translocations were seen in the myeloid leukemia cells, implicating genomic instability in the pathogenesis of these disorders. Conclusions: Taken together, our results show that loss of Brca1 in murine bone marrow causes hematopoietic defects and MMC sensitivity similar to that seen in humans with FA, providing strong evidence that Brca1 is critical for normal hematopoiesis and that Brca1 is a bona fide FA gene. This novel mouse model provides the opportunity to gain functional insight into the key stage(s) of hematopoiesis that require Brca1 and the effects of Brca1 haploinsufficiency, as seen in humans, on hematopoiesis. Further, as nearly all of the single gene FA mouse models to-date have failed to recapitulate the bone marrow phenotype of human FA, this model will be critical for deepening our understanding of the pathogenesis of FA. Disclosures No relevant conflicts of interest to declare.

Patients with acute myeloid leukemia (AML) and normal karyotype are classified in an intermediate-risk group, albeit this subset is heterogeneous for clinical outcome. A recent complementary DNA microarray study identified a gene-expression signature that--when used to cluster normal karyotype patients--separated them into 2 prognostically relevant subgroups. We sought the first independent validation of the prognostic value of this signature. Using oligonucleotide microarrays to measure gene expression in samples from uniformly treated adults with karyotypically normal AML, we performed cluster analysis based on the previously identified signature. We also developed a well-defined classification rule using the signature to predict outcome for individual patients. Cluster analysis confirmed the prognostic utility of the signature: patient clusters differed in overall (P = .001) and disease-free (P = .001) survival. The signature-based classifier identified groups with differences in overall (P = .02) and disease-free (P = .05) survival. A strong association of the outcome classifier with the prognostically adverse FLT3 internal tandem duplication (FLT3 ITD) potentially explained the prognostic significance of the signature. However, in the subgroup of patients without FLT3 ITD there was a moderate difference in survival for the classifier-derived groups. Our analysis confirms the applicability of the gene-expression profiling strategy for outcome prediction in cytogenetically normal AML.

Myelofibrosis with myeloid metaplasia (MMM) is a clinicopathologic entity characterized by stem cell-derived clonal myeloproliferation, ineffective erythropoiesis, extramedullary hematopoiesis, and bone marrow fibrosis and osteosclerosis. Patients with MMM have shortened survival and their quality of life is compromised by progressive anemia, marked hepatosplenomegaly, and severe constitutional symptoms including cachexia. After decades of frustration with ineffective therapy, patients are now being served by promising treatment approaches that include allogeneic hematopoietic stem cell transplantation and immunomodulatory drugs. Recent information regarding disease pathogenesis, including a contribution to the myeloproliferative disorder phenotype by a gain-of-function JAK2 mutation (JAK2(V617F)), has revived the prospect of targeted therapeutics as well as molecular monitoring of treatment response. Such progress calls for standardization of response criteria to accurately assess the value of new treatment modalities, to allow accurate comparison between studies, and to ensure that the definition of response reflects meaningful health outcome. Accordingly, an international panel of experts recently convened and delineated 3 response categories: complete remission (CR), partial remission (PR), and clinical improvement (CI). Bone marrow histologic and hematologic remissions characterize CR and CR/PR, respectively. The panel agreed that the CI response category is applicable only to patients with moderate to severe cytopenia or splenomegaly.

Although the diagnosis and classification of most cases of the myelodysplastic syndromes (MDS) is usually accomplished without difficulty, a minority of cases may pose diagnostic problems. In many cases the diagnostic dilemma can be solved by adhering to basic guidelines recommended for evaluation of patients suspected of having MDS, and in particular to the quality of the blood and bone marrow specimens submitted for morphologic, immunophenotypic and genetic studies. In other cases, such as patients who have hypocellular MDS or MDS with fibrosis, the criteria for making a diagnosis may be difficult if not impossible to apply, and in still others the diagnostic uncertainty is because the minimal criteria necessary to establish the diagnosis of MDS are not always clearly stated. In this review, some of these diagnostic problems are addressed and some general guidelines for resolving them are suggested. In addition, data are presented that illustrate that the WHO classification offers a valuable tool in the diagnosis and classification of MDS.

Purpose Because both t(8;21) and inv(16) disrupt core binding factor (CBF) in acute myeloid leukemia (AML) and confer relatively favorable prognoses, these cytogenetic groups are often treated similarly. Recent studies, however, have shown different gene profiling for the two groups, underscoring potential biologic differences. Therefore, we sought to determine whether these two cytogenetic groups should also be considered separate entities from a clinical standpoint. Patients and Methods We analyzed 144 consecutive adults with t(8;21) and 168 with inv(16) treated on Cancer and Leukemia Group B front-line studies. We compared pretreatment features, probability of achieving complete remission (CR), overall survival (OS) and cumulative incidence of relapse (CIR) between the two groups. Results With a median follow-up of 6.4 years, for CBF AML as a whole, the CR rate was 88%, 5-year OS was 50% and CIR was 53%. After adjusting for covariates, patients with t(8;21) had shorter OS (hazard ratio [HR] = 1.5; P = .045) and survival after first relapse (HR = 1.7; P = .009) than patients with inv(16). Unexpectedly, race was an important predictor for t(8;21) AML, in that nonwhites failed induction more often (odds ratio = 5.7; P = .006) and had shorter OS than whites when certain secondary cytogenetic abnormalities were present. In patients with t(8;21) younger than 60 years, type of induction also correlated with relapse risk. For inv(16) AML, secondary cytogenetic abnormalities (especially +22) and male sex predicted better outcome. Conclusion When the prognostic impact of race, secondary cytogenetic abnormalities, sex, and response to salvage treatment is considered, t(8;21) and inv(16) AMLs seem to be distinct clinical entities and should be stratified and reported separately.

Purpose P-glycoprotein (Pgp) is strongly inhibited by PSC-833. A chemotherapy dose-escalation study was performed with PSC-833 in patients younger than 60 years with untreated acute myeloid leukemia. Clinical rather than pharmacokinetic end points were used to develop two induction therapies containing drugs susceptible to Pgp-mediated efflux and associated with comparable toxicities at the maximum-tolerated doses. Patients and Methods A total of 410 patients were enrolled. Fifteen induction regimens containing variable doses of daunorubicin (DNR) and etoposide (ETOP) and fixed doses of cytarabine were evaluated with (ADEP) or without (ADE) a fixed dose of PSC-833. Results Doses selected for phase III testing were DNR 90 mg/m2 and ETOP 100 mg/m2 in ADE, and DNR and ETOP each 40 mg/m2 in ADEP. Intolerable mucosal toxicity occurred at higher doses of ADEP. Although the design of this study precludes direct comparisons, there was an apparent advantage for receiving ADEP with respect to disease-free and overall survival in patients ≤ 45 years old, despite the significantly lower doses of DNR and ETOP given in ADEP compared with ADE. Conclusion A large clinical data set was used to develop induction regimens containing two drugs susceptible to Pgp-mediated efflux, with and without an inhibitor of Pgp function. The chosen doses have comparable antileukemia activity and toxicity, making them suitable for use in a phase III comparative study of induction chemotherapy for patients with acute myeloid leukemia younger than 60 years. That trial will also clarify whether patients ≤ 45 years old are especially likely to benefit from Pgp inhibition during induction therapy.

Whites have a more favorable prognosis than African Americans for a number of cancers. The relationship between race and outcome is less clear in acute myeloid leukemia (AML). Using data from 7 Cancer and Leukemia Group B studies initiated from 1985 to 1997, we conducted a retrospective cross-sectional analysis of 2570 patients (270 African American and 2300 white) with de novo AML who received induction chemotherapy. African Americans were younger than whites (48 versus 54 years, P

Summary We analysed the nature and prognostic significance of secondary cytogenetic changes in 111 newly diagnosed adults with acute lymphoblastic leukaemia (ALL) and t(9;22)(q34;q11.2) or its variants. Secondary aberrations were seen in 75 (68%) patients. They included, in order of descending frequency: +der(22)t(9;22), +21, abnormalities of 9p, high hyperdiploidy (>50 chromosomes), +8, −7, +X and abnormalities resulting in loss of material from 8p, gain of 8q, gain of 1q and loss of 7p. Eighty patients (72%) had ≥1 normal metaphase in their karyotype. There were four balanced and 12 unbalanced translocations previously unreported in ALL with t(9;22). The t(2;7)(p11;p13) and der(18)t(8;18)(q11.2;p11.2) were seen in two cases each, and have never before been reported in haematological malignancy. All but four patients were treated on front-line Cancer and Leukaemia Group B clinical protocols. The presence of −7 as a sole secondary abnormality was associated with a lower complete remission (CR) rate (P = 0·004), while the presence of ≥3 aberrations was associated with a higher CR rate (P = 0·009) and +der(22)t(9;22) with a higher cumulative incidence of relapse (P = 0·02). It will be of interest to see if newly diagnosed t(9;22)-positive adult ALL patients with these and other secondary aberrations respond differently to treatment regimens that include imatinib mesylate.

BACKGROUND The objective of the current study was to evaluate the efficacy of intensive chemotherapy with and without cranial radiation for central nervous system (CNS) prophylaxis in adults with Burkitt leukemia or lymphoma. METHODS Patients received 18 weeks of therapy. Prophylactic cranial radiation (2400 centigrays) and 12 doses of triple intrathecal chemotherapy were administered to the first cohort of patients. A subsequent cohort received the same therapy, with the exceptions that intrathecal therapy was reduced to six doses and radiotherapy was administered only to high-risk individuals. RESULTS The median follow-up durations were 6.8 years in Cohort 1 and 4.1 years in Cohort 2. Three occurrences of transverse myelitis, 2 severe neuropathies, 3 cases of aphasia, and 1 case of blindness were documented in the first cohort of 52 patients (Cohort 1). In the subsequent cohort of 40 patients (Cohort 2), none of these occurrences were observed, and patients experienced less neurologic toxicity overall (61% vs. 26%; P = 0.001). Responses were similar, and the 3-year event-free survival rate was 0.52 (95% confidence interval, 0.38–0.65) for Cohort 1 and 0.45 (0.29–0.60) for Cohort 2. CONCLUSIONS Intensive, short-duration chemotherapy with less intensive CNS prophylaxis led to control at this sanctuary site with little neurotoxicity and may be curative for adults with Burkitt leukemia or lymphoma. Cancer 2004;100:1438–48. ©2004 by the American Cancer Society.

Cladribine has been reported to have little activity in fludarabine- refractory chronic lymphocytic leukemia (CLL). We sought to determine whether resistance to therapy with cladribine in fludarabine-refractory CLL patients represented primary drug resistance or the inability to tolerate the myelosuppression associated with this therapy. Patients with fludarabine refractory CLL patients without severe thrombocytopenia (platelets/=50 x 10(9)/l) or granulocytopenia (neutrophils1.5 x 10(9)/l) were enrolled. All patients received cladribine (0.14 mg/kg) as a 2-h intravenous infusion daily for 5 days, repeated every 4 weeks. Patients received up to six cycles of therapy. Twenty-eight patients enrolled; 13 had intermediate (Rai stage I or II) and 15 high (Rai stage III and IV) risk stages. No patient had a complete remission, but nine (32%; 95% confidence interval, 15-49%) attained a partial remission when assessed using the modified NCI criteria (1996). The median time to relapse for responders was 12 months, while median progression-free survival for the entire group was 9 months (95% confidence interval, 4-14 months). The median overall survival was 2.2 years (95% confidence interval, 0.8-3.1 years). Response was predicted by pre-treatment Rai status with seven of 13 (54%) intermediate risk vs two of 15 (13%) high-risk patients responding (P = 0.04). Toxicity was myelosuppression and infections (grade 3-5: neutropenia 75%, thrombocytopenia 68%, and infections 43%). Cladribine has modest clinical activity and considerable toxicity in a very selected group of patients with fludarabine-refractory CLL lacking pre-treatment neutropenia and thrombocytopenia.

A World Health Organization (WHO) classification of hematopoietic and lymphoid neoplasms has recently been published. This classification was developed through the collaborative efforts of the Society for Hematopathology, the European Association of Hematopathologists, and more than 100 clinical hematologists and scientists who are internationally recognized for their expertise in hematopoietic neoplasms. For the lymphoid neoplasms, this classification provides a refinement of the entities described in the Revised European-American Lymphoma (REAL) Classification—a system that is now used worldwide. To date, however, there has been no published explanation or rationale given for the WHO classification of the myeloid neoplasms. The purpose of this communication is to outline briefly the WHO classification of malignant myeloid diseases, to draw attention to major differences between it and antecedent classification schemes, and to provide the rationale for those differences.

The mechanisms governing disease-associated chromatin organization and drug selectivity are poorly understood. Our epigenetic drug screening experiments identify two distinct lineage-specific drug-responsive patterns in myeloid leukemia (MDS/AML) cells. We have established multiple MDS/AML cell lines resistant to nucleic acid analogues with different chemical backbones and have demonstrated a clear relationship between drug-resistance and sugar-phosphate backbone. Based on these results, we used various experimental approaches to elucidate leukemia-associated, drug-responsive chromatin structural changes and their underlying mechanisms. Our data demonstrate that hnRNPK, a conserved factor in heterogeneous nuclear RNA-binding protein (hnRNP) complexes, directly interacts with RNA m5C methyltransferases NSUNs/DMNT2 and RNA-polymerase II (pol-II)/CTD9 as well as erythroid vs myeloid lineage-determining transcription factors (TFs) GATA1 and SPI1/PU.1 to form distinct drug-responsive chromatin structures in MDS/AML cells. hnRNP preferentially binds to methylated polypyrimidine RNA sequences to facilitate transcription elongation. Compared to azacitidine-sensitive MDS/AML cells, there is a marked increase in azacitidine-resistant MDS/AML cells in RNA m5C and NUSNs/ DNMT2 associated with a marked increase in the interaction between hnRNPK and active pol-II/CDK9. By employing a newly-developed nascent RNA capturing technology coupled with super-resolution stimulated-emission depletion confocal microscopy, we demonstrate co-localization of hnRNPK with the TFs and active pol-II at nascent RNA sites in MDS/AML cells, further supporting the existence of hnRNPK-mediated drug-responsive transcription and lineage-specific chromatin structures in MDS/AML cells. Our experiments using clinical specimens demonstrate a positive correlation between MDS/AML progression and increase in expression of hnRNPK and RNA m5C methyltransferases, supporting their importance and clinical relevance. Furthermore, knockdown of hnRNPK and RNA 5mC methyltransferases effectively inhibited the growth of MDS/AML cells. In conclusion, our data demonstrate distinct RNA m5C methyltransferases/hnRNPK-mediated chromatin structures that control the growth and drug sensitivity of MDS/AML cells. Based on our data, we propose a novel working model of RNA m5C methyltransferase/hnRNPK-mediated drug-responsive chromatin structure in MDS/AML cells, in which RNA m5C methyltransferases function as the writers of m5C on nascent RNA, and hnRNPK functions as a reader of the RNA m5C and an operator to regulate transcription elongation and gene activation. Such novel chromatin structure-based drug action models and mechanisms may identify new diagnostic and prognostic biomarkers and therapeutic approaches. Note: This abstract was not presented at the meeting. Citation Format: Jason X. Cheng, Li Cheng, Adam Cloe, Yuan Li, Ming Yue, Michelle M. Le Beau, Richard A. Larson, James W. Vardiman. RNA m5C methyltransferases and hnRNPK mediate disease-associated chromatin structure and drug resistance in leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4470. doi:10.1158/1538-7445.AM2017-4470

The World Health Organization separates acute erythroid leukemia (erythropoiesis in ≥50% of nucleated bone marrow cells; ≥20% myeloblasts of non-erythroid cells) from other entities with increased erythropoiesis - acute myeloid leukemia with myelodysplasia-related changes (≥20% myeloblasts of all nucleated cells) or myelodysplastic syndromes - and subdivides acute erythroid leukemia into erythroleukemia and pure erythroid leukemia subtypes. We aimed to investigate the biological/genetic justification for the different categories of myeloid malignancies with increased erythropoiesis (≥50% of bone marrow cells).We investigated 212 patients (aged 18.5-88.4 years) with acute myeloid leukemia or myelodysplastic syndromes characterized by 50% or more erythropoiesis: 108 had acute myeloid leukemia (77 with acute erythroid leukemia, corresponding to erythroid/myeloid erythroleukemia, 7 with pure erythroid leukemia, 24 with acute myeloid leukemia with myelodysplasia-related changes) and 104 had myelodysplastic syndromes. Morphological and chromosome banding analyses were performed in all cases; subsets of cases were analyzed by polymerase chain reaction and immunophenotyping.Unfavorable karyotypes were more frequent in patients with acute myeloid leukemia than in those with myelodysplastic syndromes (42.6% versus 13.5%; P0.0001), but their frequency did not differ significantly between patients with acute erythroid leukemia (39.0%), pure erythroid leukemia (57.1%), and acute myeloid leukemia with myelodysplasia-related changes (50.0%). The incidence of molecular mutations did not differ significantly between the different categories. The 2-year overall survival rate was better for patients with myelodysplastic syndromes than for those with acute myeloid leukemia (P0.0001), without significant differences across the different acute leukemia subtypes. The 2-year overall survival rate was worse in patients with unfavorable karyotypes than in those with intermediate risk karyotypes (P0.0001). In multivariate analysis, only myelodysplastic syndromes versus acute myeloid leukemia (P=0.021) and cytogenetic risk category (P=0.002) had statistically significant effects on overall survival.The separation of acute myeloid leukemia and myelodysplastic syndromes with 50% or more erythropoietic cells has clinical relevance, but it might be worth discussing whether to replace the subclassifications of different subtypes of acute erythroid leukemia and acute myeloid leukemia with myelodysplasia-related changes by the single entity, acute myeloid leukemia with increased erythropoiesis ≥50%.

PURPOSE: To prospectively compare cytogenetics and reverse transcriptase–polymerase chain reaction (RT-PCR) for detection of t(8;21)(q22;q22) and inv(16)(p13q22)/t(16;16)(p13;q22), aberrations characteristic of core-binding factor (CBF) acute myeloid leukemia (AML), in 284 adults newly diagnosed with primary AML. PATIENTS AND METHODS: Cytogenetic analyses were performed at local laboratories, with results reviewed centrally. RT-PCR for AML1/ETO and CBFβ/MYH11 was performed centrally. RESULTS: CBF AML was ultimately identified in 48 patients: 21 had t(8;21) or its variant and AML1/ETO, and 27 had inv(16)/t(16;16), CBFβ/MYH11, or both. Initial cytogenetic and RT-PCR analyses correctly classified 95.7% and 96.1% of patients, respectively (P = .83). Initial cytogenetic results were considered to be false-negative in three AML1/ETO-positive patients with unique variants of t(8;21), and in three CBFβ/MYH11-positive patients with, respectively, an isolated +22; del(16)(q22),+22; and a normal karyotype. The latter three patients were later confirmed to have inv(16)/t(16;16) cytogenetically. Only one of 124 patients reported initially as cytogenetically normal was ultimately RT-PCR–positive. There was no false-positive cytogenetic result. Initial RT-PCR was falsely negative in two patients with inv(16) and falsely positive for AML1/ETO in two and for CBFβ/MYH11 in another two patients. Two patients with del(16)(q22) were found to be CBFβ/MYH11-negative. M4Eo marrow morphology was a good predictor of the presence of inv(16)/t(16;16). CONCLUSION: Patients with t(8;21) or inv(16) can be successfully identified in prospective multi-institutional clinical trials. Both cytogenetics and RT-PCR detect most such patients, although each method has limitations. RT-PCR is required when the cytogenetic study fails; it is also required to determine whether patients with suspected variants of t(8;21), del(16)(q22), or +22 represent CBF AML. RT-PCR should not replace cytogenetics and should not be used as the only diagnostic test for detection of CBF AML because of the possibility of obtaining false-positive or false-negative results.

Background: BETs are evolutionarily conserved proteins that specifically bind to acetyl-lysines on histones and other proteins to recruit transcriptional machinery and influence gene expression (Shi and Vakoc. 2014 Mol Cell 54, 728-736). Small molecules that target BETs, such as JQ1 and I-BET, can effectively inhibit the growth of midline carcinoma cells that have a BRD4-NUT gene fusionas well as leukemia cells, such as AML, which have a diverse genetic background and do not have BRD4 rearrangements. DNA methyltransferase (DNMT) inhibitors, such as azacitidine (5-AZA) and decitabine, have been proven effective in some patients with hematologic malignancies including AML and MDS. However, the detailed mechanisms underlying the selectivity and resistance of these epigenetic-modifying drugs remain elusive. Our previous genome-wide epigenetic profiling data have demonstrated epigenetic inactivation of the SPI1/PU.1 pathway in cytogenetically normal MDS with erythroid predominance (Cheng et al. 2013 Leukemia 27, 1291-1300). This study aims to elucidate the role of chromatin structure in the regulation of response to epigenetic drugs in MDS and AML. Methods: Co-immunoprecipitation (co-IP) coupled with RNase/DNase digestion, super-resolution two-photon stimulated-emission depletion (STED) confocal microscopy, high-throughput sequencing and chromatin conformation capture (3C) were used. Results: Our data demonstrate that GATA1 and SPI1/PU.1, erythroid- vs. myeloid-determining transcription factors (TFs), selectively interact with specific sets of DNA/histone modifiers, such as TET2, DNMTs and EZH2, to form distinct lineage-specific, drug-responsive chromatin structures at specific gene loci. The interactions between the TFs, DNA/histone modifiers and chromatin structures are highly sensitive to RNase and DNase digestions, i.e., dependent on native chromatin environment. Our experiments show that hnRNPK, a conserved factor in the heterogeneous nuclear RNA-binding protein (hnRNP) complexes, directly interact with the lineage-determining TFs and DNA/histone modifiers as well as RNAs/DNAs to form distinct chromatin-structures in MDS/AML cells. Epigenetic drugs, such as 5-AZA, efficiently disrupt the hnRNPK-mediated chromatin structures in 5-AZA-sensitive MDS/AML cells, but not in 5-AZA-resistant MDS/AML cells. Our data demonstrate a marked (~4-6 fold) increase in BRD4, BRD2 and DNMT1 in the 5-AZA-resistant MDS/AML cell lines compared to the 5-AZA-sensitive MDS/AML cell lines. Strikingly, a massive amount of BRD4-associated RNA polymerase II (pol-II) is observed in the 5-AZA-resistant MDS/AML cells, but not in the 5-AZA-senstive MDS/AML, and there is approximately 70-80 fold increase in BRD4-associated pol-II, but not in BRD4-associated GATA1, in the 5-AZA-resistant cells compared to the 5-AZA-sensitive cells. Such BRD4-associated pol-II is almost exclusively in a processive pol-II complex that is associated with the phosphorylated C-terminal domain (S2p/S5p-CTD) of pol-II, and sensitive to JQ1. Furthermore, JQ1 significantly decreases the interactions between BRD4 and lineage-determining TFs GATA1 and SPI1/PU.1, in the 5-AZA-sensitive MDS/AML cells, but not in the 5-AZA-resistant MDS/AML cells. A very low concentration (0.01 uM) of JQ1 can re-sensitize the 5-AZA-resistant MDS/AML cell lines and completely reverse the drug resistance to 5-AZA, suggesting a BET-mediated 5-AZA-resistance in those cells. Genome-wide chromatin immunoprecipitation coupled with high-throughput sequencing has been carried out to map the BRD4-associated RNA pol-II at the genome-wide level in the 5-AZA resistant and sensitive MDS/AML cells. Our experiments using clinical specimens demonstrate a positive correlation between MDS/AML progression and increase in expression of hnRNPK, BRD2 and BRD4, confirming their importance and clinical relevance. Conclusion: Our study has revealed a novel drug resistance mechanism that operates through BET-mediated recruitment of an active pol-II complex and induction of drug-responsive chromatin structural changes in MDS/AML cells (Fig 1). Such BET-mediated changes in pol-II complexes and chromatin structure could become useful biomarkers to predict drug responses, leading to rational designs and actionable targets to reprogram therapeutic drug susceptibility for more effective therapy in MDS and AML. Disclosures No relevant conflicts of interest to declare.

Since 1995, the European Association of Pathologists and the Society for Hematopathology have been developing a new World Health Organization (WHO) classification of hematologic malignancies. The classification includes lymphoid, myeloid, histiocytic, and mast cell neoplasms. The WHO project involves 10 committees of pathologists, who have developed lists and definitions of disease entities. A Clinical Advisory Committee of international hematologists and oncologists was formed to ensure that the classification will be useful to clinicians. A meeting was held in November 1997 to discuss clinical issues related to the classification. The WHO has adopted the Revised European-American Classification of Lymphoid Neoplasms, published in 1994 by the International Lymphoma Study Group, as the classification of lymphoid neoplasms. This approach to classification is based on the principle that a classification is a list of "real" disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features. The relative importance of each of these features varies among diseases, and there is no one "gold standard." The WHO classification has applied the principles of the Revised European-American Classification of Lymphoid Neoplasms to myeloid and histiocytic neoplasms. The classification of myeloid neoplasms recognizes distinct entities defined by a combination of morphology and cytogenetic abnormalities. The Clinical Advisory Committee meeting, which was organized around a series of clinical questions, was able to reach a consensus on most of the questions posed. The questions and the consensus are discussed in detail in this article. Among other things, the Clinical Advisory Committee concluded that clinical grouping of lymphoid neoplasms was neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the international prognostic index. The experience of developing the WHO classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world. This should facilitate progress in the understanding and treatment of hematologic malignancies.

Introduction Since 1995, the European Association of Pathologists (EAHP) and the Society for Hematopathology (SH) have been developing a new World Health Organization (WHO) classification of haematological malignancies. The classification includes lymphoid, myeloid, histiocytic and mast cell neoplasms. Design The WHO project involves 10 committees of pathologists, who have developed lists and definitions of disease entities. A Clinical Advisory Committee (CAC) of international haematologists and oncologists was formed to ensure that the classification will be useful to clinicians. A meeting was held in November 1997 to discuss clinical issues related to the classification. Results The WHO has adopted the ‘Revised European–American Classification of Lymphoid Neoplasms’ (REAL), published in 1994 by the International Lymphoma Study Group (ILSG), as the classification of lymphoid neoplasms. This approach to classification is based on the principle that a classification is a list of ‘real’ disease entities, which are defined by a combination of morphology, immunophenotype, genetic features and clinical features. The relative importance of each of these features varies among diseases, and there is no one ‘gold standard’. The WHO classification has applied the principles of the REAL classification to myeloid and histiocytic neoplasms. The classification of myeloid neoplasms recognizes distinct entities defined by a combination of morphology and cytogenetic abnormalities. The CAC meeting, which was organized around a series of clinical questions, was able to reach a consensus on most of the questions posed. The questions and the consensus are discussed in detail below. Among other things, the CAC concluded that clinical groupings of lymphoid neoplasms was neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumour type, if applicable, and clinical prognostic factors such as the international prognostic index (IPI). Conclusion The experience of developing the WHO classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world, which should facilitate progress in the understanding and treatment of haematological malignancies.

Background: Controversy in lymphoma classification dates back to the first attempts to formulate such classifications. Over the years, much of this controversy arose from the assumption that there had to be a single guiding principle a gold standard' - for classification, and from the existence of multiple different classifications. Design: The International Lymphoma Study Group (I.L.S.G.) developed a consensus list of lymphoid neoplasms, which was published in 1994 as the Revised European -American Classification of Lymphoid Neoplasms' (R.E.A.L.). The classification is based on the principle that a classification is a list of real' disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features, The relative importance of each of these features varies among diseases, and there is no one gold standard'. In some tumors morphology is paramount, in others it is immunophenotype, a specific genetic abnormality, or clinical features. An international study of 1300 patients, supported by the San Salvatore Foundation, was conducted to determine whether the R.E.A.L. Classification could be used by expert pathologists and had clinical relevance. Since 1995, the European Association of Pathologists (EAHP) and the Society for Hematopathology (SH) have been developing a new World Health Organization (WHO) Classification of hematologic malignancies, using an updated R.E.A.L. Classification for lymphomas and applying the principles of the R.E.A.L. Classification to myeloid and histiocytic neoplasms. A Clinical Advisory Committee (CAC) was formed to ensure that the WHO Classification will be useful to clinicians. Results: The International Lymphoma Study showed that the R.E.A.L. Classification could be used by pathologists, with inter-observer reproducibility better than for other classifications ( > 85%). Immunophenotyping was helpful in some diagnoses, but not required for many others. New entities not specifically recognized in the Working Formulation accounted for 27% of the cases. Diseases that would have been lumped together as 'low grade' or intermediate/high grade' in the Working Formulation showed marked differences in survival, confirming that they need to be treated as distinct entities. Clinical features such as the International Prognostic Index were also important in determining patient outcome. The WHO Clinical Advisory Committee concluded that clinical groupings of lymphoid neoplasms was neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the International Prognostic Index (IPI). Conclusions: The experience of developing the WHO Classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world, which should facilitate progress in the understanding and treatment of hematologic malignancies.

Introduction Since 1995, the European Association of Pathologists and the Society for Hematopathology have been developing a new World Health Organization (WHO) classification of hematologic malignancies. The classification includes lymphoid, myeloid, histiocytic, and mast cell neoplasms. Materials and methods The WHO project involves ten committees of pathologists, who have developed lists and definitions of disease entities. A Clinical Advisory Committee (CAC) of international hematologists and oncologists was formed to ensure that the classification will be useful to clinicians. A meeting was held in November 1997 to discuss clinical issues related to the classification. Results WHO has adopted the 'Revised European-American Classification of Lymphoid Neoplasms' (REAL), published in 1994 by the International Lymphoma Study Group (ILSG), as the classification of lymphoid neoplasms. This approach to classification is based on the principle that a classification is a list of 'real' disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features. The relative importance of each of these features varies among diseases, and there is no one 'gold standard'. The WHO classification has applied the principles of the REAL classification to myeloid and histiocytic neoplasms. The classification of myeloid neoplasms recognizes distinct entities defined by a combination of morphology and cytogenetic abnormalities. The CAC meeting, which was organized around a series of clinical questions, was able to reach a consensus on most of the questions posed. The questions and the consensus are discussed in detail below. Among other things, the CAC concluded that clinical groupings of lymphoid neoplasms were neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the international prognostic index (IPI). Conclusion The experience of developing the WHO classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world, which should facilitate progress in the understanding and treatment of hematologic malignancies.

Summary Introduction Since 1995, the European Association of Pathologists (EAHP) and the Society for Hematopathology (SH) have been developing a new World Health Organization (WHO) Classification of hematologic malignancies. The classification includes lymphoid, myeloid, histiocytic, and mast cell neoplasms. Design The WHO project involves 10 committees of pathologists, who have developed lists and definitions of disease entities. A Clinical Advisory Committee (CAC) ) of international hematologists and oncologists was formed to ensure that the classification will be useful to clinicians. A meeting was held in November, 1997, to discuss clinical issues related to the classification. Results The WHO has adopted the ‘Revised European–American Classification of Lymphoid Neoplasms’ (R.E.A.L.), published in 1994 by the International Lymphoma Study Group (ILSG), as the classification of lymphoid neoplasms. This approach to classification is based on the principle that a classification is a list of ‘real’ disease entities, which are defined by a combination of morphology, immunophenotype, genetic features, and clinical features. The relative importance of each of these features varies among diseases, and there is no one ‘gold standard’. The WHO Classification has applied the principles of the R.E.A.L. Classification to myeloid and histiocytic neoplasms. The classification of myeloid neoplasms recognizes distinct entities defined by a combination of morphology and cytogenetic abnormalities. The CAC meeting, which was organized around a series of clinical questions, was able to reach a consensus on most of the questions posed. The questions and the consensus are discussed in detail below. Among other things, the CAC concluded that clinical groupings of lymphoid neoplasms were neither necessary nor desirable. Patient treatment is determined by the specific type of lymphoma, with the addition of grade within the tumor type, if applicable, and clinical prognostic factors such as the international prognostic index (IPI). Conclusion The experience of developing the WHO Classification has produced a new and exciting degree of cooperation and communication between oncologists and pathologists from around the world, which should facilitate progress in the understanding and treatment of hematologic malignancies.