Novel insights into the biology and treatment of chronic myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPNs) are clonal disorders of hematopoiesis characterized by a high frequency of genetic alterations, and include chronic myeloid leukemia (CML) and the BCR-ABL1-negative MPNs. Herein we summarize recent advances and controversies in our understanding of the biology and therapy of these disorders, as discussed at the 8th post-American Society of Hematology CML-MPN workshop. The principal areas addressed include the breakthrough discovery of CALR mutations in patients with JAK2/MPL wild type MPN, candidate therapies based on novel genetic findings in leukemic transformation and new therapeutic targets in MPNs, and an appraisal of bone marrow histopathology in MPNs with a focus on the potential new clinical entity of "masked" polycythemia vera.

Prospective blinded study of BRAFV600E mutation detection in cell-free DNA of patients with systemic histiocytic disorders.

Unlabelled: Patients with Langerhans cell histiocytosis (LCH) and Erdheim-Chester disease (ECD) have a high frequency of BRAF(V600E) mutations and respond to RAF inhibitors. However, detection of mutations in tissue biopsies is particularly challenging in histiocytoses due to low tumor content and stromal contamination. We applied a droplet-digital PCR assay for quantitative detection of the BRAF(V600E) mutation in plasma and urine cell-free (cf) DNA and performed a prospective, blinded study in 30 patients with ECD/LCH.

SETBP1 mutations drive leukemic transformation in ASXL1-mutated MDS.

Mutations in ASXL1 are frequent in patients with myelodysplastic syndrome (MDS) and are associated with adverse survival, yet the molecular pathogenesis of ASXL1 mutations (ASXL1-MT) is not fully understood. Recently, it has been found that deletion of Asxl1 or expression of C-terminal-truncating ASXL1-MTs inhibit myeloid differentiation and induce MDS-like disease in mice. Here, we find that SET-binding protein 1 (SETBP1) mutations (SETBP1-MT) are enriched among ASXL1-mutated MDS patients and associated with increased incidence of leukemic transformation, as well as shorter survival, suggesting that SETBP1-MT play a critical role in leukemic transformation of MDS.

The mutational landscape of paroxysmal nocturnal hemoglobinuria revealed: new insights into clonal dominance.

Paroxysmal nocturnal hemoglobinuria (PNH) is a disorder of hematopoietic stem cells that has largely been considered a monogenic disorder due to acquisition of mutations in the gene encoding PIGA, which is required for glycosylphosphatidylinositol (GPI) anchor biosynthesis. In this issue of the JCI, Shen et al. discovered that PNH is in fact a complex genetic disorder orchestrated by many genetic alterations in addition to PIGA mutations.

Recurrent RAS and PIK3CA mutations in Erdheim-Chester disease.

Erdheim-Chester disease (ECD) is a rare histiocytic disorder that is challenging to diagnose and treat. We performed molecular analysis of BRAF in the largest cohort of ECD patients studied to date followed by N/KRAS, PIK3CA, and AKT1 mutational analysis in BRAF wild-type patients. Forty-six of 80 (57.

Somatic alterations and dysregulation of epigenetic modifiers in cancers.

Genomic discovery efforts in patients with cancer have been critical in identifying a recurrent theme of mutations in epigenetic modifiers. A number of novel and exciting basic biological findings have come from this work including the discovery of an enzymatic pathway for DNA cytosine demethylation, a link between cancer metabolism and epigenetics, and the critical importance of post-translational modifications at specific histone residues in malignant transformation. Identification of cancer cell dependency on a number of these mutations has quickly resulted in the development of therapies targeting several of these genetic alterations.

Femoral bone marrow aspiration in live mice.

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed step-by-step technical procedure for an analogous procedure in live mice which allows for serial characterization of cells present in the BM. This procedure facilitates studies aimed to detect the presence of exogenously administered cells within the BM of mice as would be done in xenograft studies for instance.

Frequent ASXL2 mutations in acute myeloid leukemia patients with t(8;21)/RUNX1-RUNX1T1 chromosomal translocations.

Acute myeloid leukemia (AML) with t(8;21) (q22;q22) is considered to have favorable risk; however, nearly half of t(8;21) patients are not cured, and recent studies have highlighted remarkable genetic heterogeneity in this subset of AML. Here we identify somatic mutations in additional sex combs-like 2 (ASXL2) in 22.7% (25/110) of patients with t(8;21), but not in patients with inv(16)/t(16;16) (0/60) or RUNX1-mutated AML (0/26).

Selective activity of the histone deacetylase inhibitor AR-42 against leukemia stem cells: a novel potential strategy in acute myelogenous leukemia.

Most patients with acute myelogenous leukemia (AML) relapse and die of their disease. Increasing evidence indicates that AML relapse is driven by the inability to eradicate leukemia stem cells (LSC). Thus, it is imperative to identify novel therapies that can ablate LSCs.

Hematopoietic stem cell origin of BRAFV600E mutations in hairy cell leukemia.

Hairy cell leukemia (HCL) is a chronic lymphoproliferative disorder characterized by somatic BRAFV600E mutations. The malignant cell in HCL has immunophenotypic features of a mature B cell, but no normal counterpart along the continuum of developing B lymphocytes has been delineated as the cell of origin. We find that the BRAFV600E mutation is present in hematopoietic stem cells (HSCs) in HCL patients, and that these patients exhibit marked alterations in hematopoietic stem/progenitor cell (HSPC) frequencies.

Consensus guidelines for the diagnosis and clinical management of Erdheim-Chester disease.

Erdheim-Chester disease (ECD) is a rare, non-Langerhans histiocytosis. Recent findings suggest that ECD is a clonal disorder, marked by recurrent BRAFV600E mutations in >50% of patients, in which chronic uncontrolled inflammation is an important mediator of disease pathogenesis. Although ∼500 to 550 cases have been described in the literature to date, increased physician awareness has driven a dramatic increase in ECD diagnoses over the last decade.

Integrated genomic analysis illustrates the central role of JAK-STAT pathway activation in myeloproliferative neoplasm pathogenesis.

Genomic studies have identified somatic alterations in the majority of myeloproliferative neoplasms (MPN) patients, including JAK2 mutations in the majority of MPN patients and CALR mutations in JAK2-negative MPN patients. However, the role of JAK-STAT pathway activation in different MPNs, and in patients without JAK2 mutations, has not been definitively delineated. We used expression profiling, single nucleotide polymorphism arrays, and mutational profiling to investigate a well-characterized cohort of MPN patients.

Recent discoveries in molecular characterization of acute myeloid leukemia.

Acute myeloid leukemia (AML) is a clinically heterogeneous disease, yet it is one of the most molecularly well-characterized cancers. Risk stratification of patients currently involves determination of the presence of cytogenetic abnormalities in combination with molecular genetic testing in a few genes. Several new recurrent genetic molecular abnormalities have recently been identified, including TET2, ASXL1, IDH1, IDH2, DNMT3A, and PHF6.

Efficacy of intermittent combined RAF and MEK inhibition in a patient with concurrent BRAF- and NRAS-mutant malignancies.

Vemurafenib, a RAF inhibitor, extends survival in patients with BRAF(V600)-mutant melanoma but activates extracellular signal-regulated kinase (ERK) signaling in RAS-mutant cells. In a patient with a BRAF(V600K)-mutant melanoma responding to vemurafenib, we observed accelerated progression of a previously unrecognized NRAS-mutant leukemia. We hypothesized that combining vemurafenib with a MAP-ERK kinase (MEK) inhibitor would inhibit ERK activation in the melanoma and prevent ERK activation by vemurafenib in the leukemia, and thus suppress both malignancies.

D-2-hydroxyglutarate produced by mutant IDH2 causes cardiomyopathy and neurodegeneration in mice.

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) have been discovered in several cancer types and cause the neurometabolic syndrome D2-hydroxyglutaric aciduria (D2HGA). The mutant enzymes exhibit neomorphic activity resulting in production of D2-hydroxyglutaric acid (D-2HG). To study the pathophysiological consequences of the accumulation of D-2HG, we generated transgenic mice with conditionally activated IDH2(R140Q) and IDH2(R172K) alleles.

Focus on the epigenome in the myeloproliferative neoplasms.

The discovery of mutations activating JAK-STAT signaling in the majority of patients with myeloproliferative neoplasms (MPNs) led to identification of tyrosine kinase activation as a predominant mechanism driving MPN pathogenesis. Despite this, the existence of additional genetic events that modify the MPN phenotype, predate JAK2 mutations, and/or contribute to leukemic transformation of MPNs has been suggested. Recently, mutations in several epigenetic modifiers have been described in patients with MPNs, including mutations in ASXL1, DNMT3A, EZH2, IDH1, IDH2, and TET2.

The importance of subclonal genetic events in MDS.

In this issue of Blood, Papaemmanuil et al present results from the largest single targeted sequencing effort in myelodysplastic syndrome (MDS) to date both in terms of number of patients studied (738 patients, of which 603 had MDS) and genes sequenced (111 genes).

Deletion of Asxl1 results in myelodysplasia and severe developmental defects in vivo.

Somatic Addition of Sex Combs Like 1 (ASXL1) mutations occur in 10-30% of patients with myeloid malignancies, most commonly in myelodysplastic syndromes (MDSs), and are associated with adverse outcome. Germline ASXL1 mutations occur in patients with Bohring-Opitz syndrome. Here, we show that constitutive loss of Asxl1 results in developmental abnormalities, including anophthalmia, microcephaly, cleft palates, and mandibular malformations.

Myelodysplastic syndromes are induced by histone methylation–altering ASXL1 mutations.

Recurrent mutations in the gene encoding additional sex combs-like 1 (ASXL1) are found in various hematologic malignancies and associated with poor prognosis. In particular, ASXL1 mutations are common in patients with hematologic malignancies associated with myelodysplasia, including myelodysplastic syndromes (MDSs), and chronic myelomonocytic leukemia. Although loss-of-function ASXL1 mutations promote myeloid transformation, a large subset of ASXL1 mutations is thought to result in stable truncation of ASXL1.

Emerging therapeutic paradigms to target the dysregulated Janus kinase/signal transducer and activator of transcription pathway in hematological malignancies.

Over the past decade, there has been increasing biochemical evidence that the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway is aberrantly activated in malignant cells from patients with a wide spectrum of cancers of the blood and immune systems. The emerging availability of small molecule inhibitors of JAK and other signaling molecules in the JAK/STAT pathway has allowed preclinical studies validating an important role of this pathway in the pathogenesis of many hematologic malignancies, and provided motivation for new strategies for treatment of these diseases. Here, a round-table panel of experts review the current preclinical and clinical landscape of the JAK/STAT pathway in acute lymphoid and myeloid leukemias, lymphomas and myeloma, and chronic myeloid neoplasms.

Antagonistic activities of the immunomodulator and PP2A-activating drug FTY720 (Fingolimod, Gilenya) in Jak2-driven hematologic malignancies.

FTY720 (Fingolimod, Gilenya) is a sphingosine analog used as an immunosuppressant in multiple sclerosis patients. FTY720 is also a potent protein phosphatase 2A (PP2A)-activating drug (PAD). PP2A is a tumor suppressor found inactivated in different types of cancer.