Choladda V. Curry

DNMT3A Loss Drives Enhancer Hypomethylation in FLT3-ITD-Associated Leukemias

DNMT3A, the gene encoding the de novo DNA methyltransferase 3A, is among the most frequently mutated genes in hematologic malignancies. However, the mechanisms through which DNMT3A normally suppresses malignancy development are unknown. Here, we show that DNMT3A loss synergizes with the FLT3 internal tandem duplication in a dose-influenced fashion to generate rapid lethal lymphoid or myeloid leukemias similar to their human counterparts. Loss of DNMT3A leads to reduced DNA methylation, predominantly at hematopoietic enhancer regions in both mouse and human samples. Myeloid and lymphoid diseases arise from transformed murine hematopoietic stem cells. Broadly, our findings support a role for DNMT3A as a guardian of the epigenetic state at enhancer regions, critical for inhibition of leukemic transformation.

Erratum: DNMT3A Loss Drives Enhancer Hypomethylation in FLT3-ITD-Associated Leukemias (Cancer Cell (2016) 29(6) (922–934) (S1535610816302082) (10.1016/j.ccell.2016.05.003))

1 Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas 77030, USA 2 Dan L. Duncan Cancer Center and Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas 77030, USA 3 Department of Bioinformatics, School of Life sciences and Technology, Tongji University, Shanghai 20092, China. 4 Stem Cells and Regenerative Medicine Center, Baylor College of Medicine, Houston, Texas 77030, USA 5 Department of Pathology and Immunology, Texas Children’s Hospital, Baylor College of Medicine, Houston, Texas 77030, USA 6 Department of Systems Biology, Division of Cancer Medicine, University of Texas MD Anderson Cancer Center, Houston, Texas, USA. 7 Department of Pediatrics, Baylor College of Medicine, Houston, Texas 77030, USA 8 Division of Oncology, Washington University School of Medicine, St. Louis, Missouri 63110, USA 9 Wellcome Trust/MRC Stem Cell Institute, Cambridge, UK 10 Greehey Childrens Cancer Research Institute and Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, USA 11 Rice University, Houston, Texas 77030

Clear cell sarcoma of soft tissue: diagnostic utility of fluorescence in situ hybridization and reverse transcriptase polymerase chain reaction.

A 7‐year‐old girl presented with pain and progressive swelling on the left plantar surface. Biopsy of a 2.5 cm mass showed nests of large round to oval neoplastic cells with abundant amphophilic to clear cytoplasm, prominent nucleoli and high mitotic activity. Occasional cells showed spindled morphology. Infrequent melanin pigment was present. Melanocytic markers (HMB45, S‐100) were diffusely positive. A diagnosis of clear cell sarcoma of soft tissue (CCSS) was made, and the mass was re‐excised with negative margins. 28 months later, a 1.0 cm pulmonary nodule was identified and wedge excision showed metastatic CCSS. Cytogenetics showed a complex karyotype (unbalanced translocation der(12;14)(q10;q10), additional chromosome 22 material of unknown origin). Although the CCSS translocation t(12;22)(q13;q12) was not identified, EWSR1 gene rearrangement was detected by fluorescence in situ hybridization (FISH). Reverse transcription polymerase chain reaction (RT‐PCR) showed an EWS‐ATF1 fusion transcript, confirmed by direct sequencing. CCSS requires differentiation from malignant melanoma, because of overlapping clinical presentations, sites of involvement, histomorphology, immunocytochemical profiles and ultrastructure. In many circumstances, definitive diagnosis is only possible with confirmation of the CCSS‐defining translocation.

Differentiating Between Burkitt Lymphoma and CD10+ Diffuse Large B-Cell Lymphoma: The Role of Commonly Used Flow Cytometry Cell Markers and the Application of a Multiparameter Scoring System

The goal of this study was to evaluate routine flow cytometric (FC) immunophenotypic markers in differentiating between Burkitt lymphoma (BL) and CD10+ diffuse large B-cell lymphoma (DLBCL). We performed retrospective analysis of FC data from 55 patients. We evaluated 9 FC parameters: forward and side scatter (FSC and SSC); mean fluorescent intensity (MFI) for CD20, CD10, CD38, CD79b, CD43, and CD71; and the percentage of neoplastic cells positive for CD71 (%CD71). The FSC; MFIs of CD10, CD43, CD79b, and CD71; and %CD71 cells were significantly different between BL and CD10+ DLBCL (P < .05; Student t test). A 5-point scoring system (FSC, %CD71, and MFIs of CD43, CD79b, and CD71) was devised, and 6 (60%) of 10 BLs scored 3 or greater and 1 (10%) of 10 CD10+ DLBCLs scored 3 (P = .04; χ(2)). Our findings indicate that routine FC parameters can aid in differentiating BL from CD10+ DLBCL.

The transcription factors Ik-1 and MZF1 downregulate IGF-IR expression in NPM-ALK+ T-cell lymphoma

BackgroundThe type I insulin-like growth factor receptor (IGF-IR) tyrosine kinase promotes the survival of an aggressive subtype of T-cell lymphoma by interacting with nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) oncogenic protein. NPM-ALK+ T-cell lymphoma exhibits much higher levels of IGF-IR than normal human T lymphocytes. The mechanisms underlying increased expression of IGF-IR in this lymphoma are not known. We hypothesized that upregulation of IGF-IR could be attributed to previously unrecognized defects that inherently exist in the transcriptional machinery in NPM-ALK+ T-cell lymphoma.Methods and resultsScreening studies showed substantially lower levels of the transcription factors Ikaros isoform 1 (Ik-1) and myeloid zinc finger 1 (MZF1) in NPM-ALK+ T-cell lymphoma cell lines and primary tumor tissues from patients than in human T lymphocytes. A luciferase assay supported that Ik-1 and MZF1 suppress IGF-IR gene promoter. Furthermore, ChIP assay showed that these transcription factors bind specific sites located within the IGF-IR gene promoter. Forced expression of Ik-1 or MZF1 in the lymphoma cells decreased IGF-IR mRNA and protein. This decrease was associated with downregulation of pIGF-IR, and the phosphorylation of its interacting proteins IRS-1, AKT, and NPM-ALK. In addition, overexpression of Ik-1 and MZF1 decreased the viability, proliferation, migration, and anchorage-independent colony formation of the lymphoma cells.ConclusionsOur results provide novel evidence that the aberrant decreases in Ik-1 and MZF1 contribute significantly to the pathogenesis of NPM-ALK+ T-cell lymphoma through the upregulation of IGF-IR expression. These findings could be exploited to devise new strategies to eradicate this lymphoma.

Pediatric subcutaneous panniculitis-like T-cell lymphoma with features of hemophagocytic syndrome.

To the Editor: We read with interest the recently published article by Huppmann et al. [1] describing a cohort of 16 pediatric patients with subcutaneous panniculitis-like T-cell lymphoma (SPLTCL). In their series, they reported only one case with possible hemophagocytic syndrome (HPS). We have encountered another pediatric case of SPLTCL with features of HPS. The patient was a 13-year-old female with a 1-month history of fevers, chills, and progressive 1–3 cm painful subcutaneous plaques and nodules on her face, chest, back, and extremities. Full-body PET scan revealed innumerable lesions involving the trunk and limbs (Fig. 1A). Laboratory evaluation showed leukopenia, anemia, negative antinuclear antibodies and rheumatoid factor, and elevated erythrocyte sedimentation rate, liver transaminases, lactate dehydrogenase, ferritin, and soluble interleukin-2 receptor level. Fig. 1 Full body PET image, pre-therapy. There are innumerable FDG-avid nodules and subcutaneous plaques involving the trunk and limbs from below the base of the neck. There are also enlarged FDG-avid bilateral axillary and external iliac lymph nodes (A). Full-body ... A skin punch biopsy from the left arm revealed dense infiltrates predominantly confined to the lobules of subcutaneous tissue (Fig. 2A), which displayed a lobular panniculitis-like process consisting of predominantly atypical lymphoid cells rimming around individual fat cells (Fig. 2B). The atypical lymphoid cells were of intermediate size with irregular nuclear contours, hyperchromasia, occasional lobated nuclei, and inconspicuous nucleoli. Increased histiocytes with prominent hemophagocytosis were seen engulfing nucleated cells with associated apoptosis and karyorrhectic debris (Fig. 2C). Scattered plasma cells were also present throughout the infiltrate (Fig. 2D). The atypical lymphoid cells were positive for CD2, CD3, CD7, CD8, and βF1, and were negative for CD4, CD5, CD15, CD30, CD56, TCRγ, and EBV EBER in situ hybridization stain (Fig. 2E, F). Monoclonal T-cell receptor beta chain and gamma chain gene rearrangements were detected by polymerase chain reaction. A bone marrow biopsy revealed increased histiocytes with focal hemophagocytosis. Fig. 2 Histologic findings of the skin punch biopsy from the left arm. Dense, lobular subcutaneous infiltrates are seen with sparing of the epidermis and dermis (A). The infiltrates show a lobular panniculitis-like process with atypical lymphoid cells rimming ... Our case showed characteristics of SPLTCL which is rare in children and is usually described as having an indolent course. However, an association with HPS has been shown to have a more aggressive course and poorer outcome [2]. In the largest series of SPLTCL published so far, which included both pediatric (n = 12) and adult (n = 51) cases, 17% of total patients had HPS [3]; we could not extrapolate, however, if any pediatric cases had HPS. In children, associated HPS appears to be extremely rare, as there was only one possible case with HPS in the series of 16 children by Huppmann et al. [1], and only two other children with HPS have been reported in the Asian population [2]. Of note, the presence of plasma cells, seen in the majority of pediatric SPLTCLs as recently reported [1], was also present in our case, making the differential diagnosis with lupus panniculitis challenging. Our patient was treated with prednisone followed by hydroxychloroquine and then bexarotene plus prednisone. The response was mixed; some nodules and plaques resolved, while others persisted and new nodules developed with recurrent episodes of fever, chills, night sweats, and persistent leukopenia. A systemic chemotherapy regimen of SMILE (steroids, methotrexate, ifosfamide, L-asparaginase, and etoposide) was then initiated. Follow up full-body PET scans at 6 months and 10 months after initial PET scan revealed resolution of the lesions (Fig. 1B). The patient was alive with no evidence of disease at 24 months after diagnosis.

Childhood Nodal Marginal Zone Lymphoma with Unusual Clinicopathologic and Cytogenetic Features for the Pediatric Variant: A Case Report

Nodal marginal zone lymphoma (NMZL) is a B-cell lymphoma that shares morphologic and immunophenotypic features with extranodal and splenic marginal zone lymphomas but lacks extranodal or splenic involvement at presentation. NMZL occurs mostly in adults with no sex predilection, at advanced stage (III or IV), with frequent relapses and a high incidence of tumoral genetic abnormalities including trisomies 3 and 18 and gain of 7q. Pediatric NMZL, however, is a rare but distinct variant of NMZL with characteristic features including male predominance, asymptomatic and localized (stage I) disease, low relapse rates with excellent outcomes, and a lower incidence of essentially similar genetic aberrations compared to adult NMZL. Here we describe a unique case of childhood NMZL with unusual clinicopathologic features for the pediatric variant including generalized lymphadenopathy, high-stage disease with persistence after therapy, unusual immunophenotype (CD5, CD23, and BCL6 positive), and unique chromosomal abnormalities including monosomy 20 and add(10)(p11.2).

MonoMAC versus idiopathic CD4+ lymphocytopenia. Comment to Haematologica. 2011;96(8):1221-5.

We read with interest a recent article in Haematologica describing a novel defined disorder known as MonoMAC (monocytopenia with Mycobacterium avium complex).[1][1] It is not infrequent for patients with MonoMAC to present with a low CD4 count, leading to a consideration of idiopathic CD4-positive (

TrkA is a binding partner of NPM‐ALK that promotes the survival of ALK+ T‐cell lymphoma

Nucleophosmin‐anaplastic lymphoma kinase‐expressing (NPM‐ALK+) T‐cell lymphoma is an aggressive neoplasm that is more commonly seen in children and young adults. The pathogenesis of NPM‐ALK+ T‐cell lymphoma is not completely understood. Wild‐type ALK is a receptor tyrosine kinase that is physiologically expressed in neural tissues during early stages of human development, which suggests that ALK may interact with neurotrophic factors. The aberrant expression of NPM‐ALK results from a translocation between the ALK gene on chromosome 2p23 and the NPM gene on chromosome 5q35. The nerve growth factor (NGF) is the first neurotrophic factor attributed to non‐neural functions including cancer cell survival, proliferation, and metastasis. These functions are primarily mediated through the tropomyosin receptor kinase A (TrkA). The expression and role of NGF/TrkA in NPM‐ALK+ T‐cell lymphoma are not known. In this study, we tested the hypothesis that TrkA signaling is upregulated and sustains the survival of this lymphoma. Our data illustrate that TrkA and NGF are expressed in five NPM‐ALK+ T‐cell lymphoma cell lines and TrkA is expressed in 11 of 13 primary lymphoma tumors from patients. In addition, we found evidence to support that NPM‐ALK and TrkA functionally interact. A selective TrkA inhibitor induced apoptosis and decreased cell viability, proliferation, and colony formation of NPM‐ALK+ T‐cell lymphoma cell lines. These effects were associated with downregulation of cell survival regulatory proteins. Similar results were also observed using specific knockdown of TrkA in NPM‐ALK+ T‐cell lymphoma cells by siRNA. Importantly, the inhibition of TrkA signaling was associated with antitumor effects in vivo, because tumor xenografts in mice regressed and the mice exhibited improved survival. In conclusion, TrkA plays an important role in the pathogenesis of NPM‐ALK+ T‐cell lymphoma, and therefore, targeting TrkA signaling may represent a novel approach to eradicate this aggressive neoplasm.

Are micromegakaryocytes specific for refractory cytopenia of childhood (RCC)? A study of 38 pediatric patients with thrombocytopenia unrelated to RCC☆

BACKGROUND Micromegakaryocytes (microMKs) are considered the most reliable dysplastic feature for myelodysplastic syndrome (MDS), particularly refractory cytopenia of childhood (RCC); there is no minimal threshold for the diagnosis of RCC. Since most RCC patients present with thrombocytopenia, the presence of microMKs should raise concern for MDS/RCC. This study attempted to investigate the prevalence of microMKs and associated marrow fibrosis in patients with thrombocytopenia unrelated to MDS/RCC and the need for establishing a threshold for microMKs for the diagnosis of MDS/RCC. DESIGN Bone marrow biopsies of pediatric patients with thrombocytopenia unrelated to RCC were examined for microMKs and fibrosis by CD61 immunohistochemical and reticulin stains respectively. RESULT Thirty eight patients (1-18 years old) were included: 33 immune thrombocytopenia (ITP), 3 chronic thrombocytopenia, and 2 inherited macrothrombocytopenia. Fourteen cases (37%) had microMKs; four cases showed increased marrow fibrosis associated with microMKs (two had ITP and two had macrothrombocytopenia). All patients are alive and none developed MDS (follow up: 3months to 4 years). CONCLUSION MicroMKs can be seen in pediatric patients with thrombocytopenia unrelated to RCC. Hence the mere presence of microMKs is insufficient for the diagnosis of RCC in the pediatric population, and a quantitative threshold needs to be established.