HOX11L2/TLX3 is transcriptionally activated through T-cell regulatory elements downstream of BCL11B as a result of the t(5;14)(q35;q32).

International audienceThe t(5;14)(q35;q32) chromosomal translocation is specifically observed in up to 20% of childhood T-cell acute lymphoblastic leukemia (T-ALL). It affects the BCL11B/CTIP2 locus on chromosome 14 and the RANBP17-TLX3/HOX11L2 region on chromosome 5. It leads to ectopic activation of TLX3/HOX11L2. To investigate the reasons of the association between t(5;14) and T-ALL, we isolated the translocation breakpoints in 8 t(5;14) patients. Sequence analyses did not involve recombinase activity in the genesis of the translocation. We used DNAse1 hypersensitive experiments to locate transcriptional regulatory elements downstream of BCL11B. By transient transfection experiments, 2 of the 6 regions demonstrated cis-activation properties in T cells and were also effective on the TLX3 promoter. Our data indicate that the basis of the specific association between t(5;14) and T-ALL lies on the juxtaposition of TLX3 to long-range cis-activating regions active during T-cell differentiation

Targeting iron homeostasis induces cellular differentiation and synergizes with differentiating agents in acute myeloid leukemia

Differentiating agents have been proposed to overcome the impaired cellular differentiation in acute myeloid leukemia (AML). However, only the combinations of all-trans retinoic acid or arsenic trioxide with chemotherapy have been successful, and only in treating acute promyelocytic leukemia (also called AML3). We show that iron homeostasis is an effective target in the treatment of AML. Iron chelating therapy induces the differentiation of leukemia blasts and normal bone marrow precursors into monocytes/macrophages in a manner involving modulation of reactive oxygen species expression and the activation of mitogen-activated protein kinases (MAPKs). 30% of the genes most strongly induced by iron deprivation are also targeted by vitamin D3 (VD), a well known differentiating agent. Iron chelating agents induce expression and phosphorylation of the VD receptor (VDR), and iron deprivation and VD act synergistically. VD magnifies activation of MAPK JNK and the induction of VDR target genes. When used to treat one AML patient refractory to chemotherapy, the combination of iron-chelating agents and VD resulted in reversal of pancytopenia and in blast differentiation. We propose that iron availability modulates myeloid cell commitment and that targeting this cellular differentiation pathway together with conventional differentiating agents provides new therapeutic modalities for AML

Epstein-Barr Virus-Induced Gene 3 (EBI3): A Novel Diagnosis Marker in Burkitt Lymphoma and Diffuse Large B-Cell Lymphoma

The distinction between Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL), two types of mature aggressive B-cell lymphomas that require distinct treatments, can be difficult because of forms showing features intermediate between DLBCL and BL (here called BL/DLBCL). They can be discriminated by the presence of c-myc translocations characteristic of BL. However, these are not exclusive of BL and when present in DLBCL are associated with lower survival. In this study, we show that Epstein-Barr virus-induced gene 3 (EBI3) is differentially expressed among BL and DLBCL. Analysis of gene expression data from 502 cases of aggressive mature B-cell lymphomas available on Gene Expression Omnibus and immunohistochemical analysis of 184 cases of BL, BL/DLBCL or DLBCL, showed that EBI3 was not expressed in EBV-positive or -negative BL cases, whereas it was expressed by over 30% of tumoral cells in nearly 80% of DLBCL cases, independently of their subtypes. In addition, we show that c-myc overexpression represses EBI3 expression, and that DLBCL or BL/DLBCL cases with c-myc translocations have lower expression of EBI3. Thus, EBI3 immunohistochemistry could be useful to discriminate BL from DLBCL, and to identify cases of BL/DLBCL or DLBCL with potential c-myc translocations

Formation of Nup98-containing nuclear bodies in HeLa sublines is linked to genomic rearrangements affecting chromosome 11.

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Three Novel AML Cases Harboring the Semi-Cryptic t(7;21)(p22;q22) Translocation Expressing RUNX1-USP42 Fusion Transcripts Associated with Diploidy/Tetraploidy and/or 5q Alterations: a Probably Underestimated Combination

Abstract Abstract 2708 RUNX1 is implicated in numerous chromosomal abnormalities acquired in acute myeloid leukemia (AML). The most frequent one, the t(8;21) is associated with a particular morphology together with a favorable prognosis. This is not the case for other 21q abnormalities, that are much less frequent and for which the prognosis is quite different. Moreover, beside point mutations, conventional cytogenetics failed to detect some of chromosomal alterations involving RUNX1. Recently 3 cases of the rare and semi-cryptic t(7;21)(p22;q22) translocation expressing the RUNX1-USP42 fusion transcripts have been reported, demonstrating the recurrence of this abnormality in AML. We describe here 3 additional cases with the same translocation and fusion transcripts, associated to 5q alterations leading to EGR1 and CSF1R heterozygous losses. In all our patients, the t(7;21)(p22.1;q22.3) was initially detected by the systematic FISH evaluation of the blastic populations using ETO-AML1 Dual Fusion probe. Patient#1 bone marrow karyotype was characterized by a tetraploid clone (89,XXYY) with loss of chromosomes 15, 17 and 18 in addition to the t(7;21), and a unbalanced translocation der(5)t(5;13)(q23;q?) between long arms of chromosomes 5 and 13, resulting in a heterozygous loss of EGR1 and CSF1R. Patient #2 blood and bone marrow karyotypes revealed a diploid clone with a del(5)(q31q33) associated with the t(7;21). The FISH analysis confirmed EGR1 and CSF1R deletions. In patient #3, the bone marrow karyotype showed diploid/tetraploïd clones, both harboring the t(7;21)(p22;q22), confirmed by FISH experiments (WCP7, AML1 probes). In addition, a der(5)t(1;5)(q3?2;q21-23) was identified within the tetraploïd clone, resulting in the loss of EGR1 and CSF1R, confirmed by FISH. In all three cases a RUNX1-USP42 fusion transcript was detected using RT-PCR, as well as the reciprocal transcript. Sequence analysis of RT-PCR products showed that the breakpoints occurred exactly in the same introns of USP42 and RUNX1 as in the previously described cases. For patient #1 and #3 a chimeric transcript was found formed of the RUNX1 exon 7 fused to the USP42 exon 3. In patient #2, a shorter chimeric transcript arised from the fusion of the RUNX1 exon 5 to the exon 3 of USP42. As already noticed in the previous reports, an alternative splicing of the RUNX1 exon 6 has been detected in these three cases. The description of these 3 novel t(7;21) confirm the recurrence of this balanced translocation in AML, and shows that this chromosomal abnormality is often associated with diploid/tetraploid clones and/or 5q alterations. Special attention should be paid in karyotype analysis of AML with diploid or tetraploid clones harboring 5q alterations. In such cases RUNX1 rearrangements should be explored using FISH analysis, and RUNX1-USP42 fusion transcript should be searched by RT-PCR in positive cases. Prospective and retrospective studies of AML have now to be settled in order to assess the incidence and clinical relevance of this cryptic translocation. Disclosures: No relevant conflicts of interest to declare. </jats:sec

Abnormalities of the Hematopoietic Stem Cell Compartment in Children After in Utero Exposure to AZT

Abstract Abstract 1123 Mother to child HIV-1 transmission prophylaxy with antiretroviral (ARV) during pregnancy is remarkably effective. Although now based on various ARV-combinations, azydothymidine (AZT) remains the cornerstone drug in all guidelines. AZT hematologic toxicity after in utero exposure consisted in a reversible anemia but also in a persistant slight decrease on platelet, neutrophils and lymphocytes count up to 2 years of age. This long lasting biological effect evokes an alteration of the hematopoietic stem cell compartment. The very long term impact is unknown. Twenty six neonates from HIV-1 infected mothers treated with an AZT based combination during pregnancy in Paris area were included in the study and compared to 20 neonates from healthy mothers. Umbilical cord blood was sampled after obtaining written informed consent from the mother. The experimental scheme included phenotypical and functional analysis of the hematopoietic compartment as well as transcriptome analysis of hematopoietic progenitors. Phenotypic analysis revealed a tendancy to higher numbers of CD34+ cells in the AZT group as compared to the control group. A similar finding was observed for hematopoietic stem cells defined on the basis of a Lin-CD34+ CD38loCD90+CD45RA- phenotype. Multipotent progenitors Lin-CD34+ CD38loCD90-CD45RA- were also increased at a statistically significant level. Affymetrix analysis of gene expression profiles was performed on CD34+ cells sorted from 8 AZT and 6 controls. Ingenuity analysis revealed significant differences in genes involved in hematopoiesis and cell cycle. Among hematopoietic genes for which differences of expression were noted between the two groups, 14 have a known role in the maintenance and differentiation of hematopoietic stem cells. Quantitative RT-PCR are actually ongoing to further characterize the gene expression profile of AZT CD34+ cells. These results support the hypothesis of an alteration of the hematopoietic stem cell compartment in children after in utero exposure to AZT. Potential hematotoxicity of other ARV drugs must be evaluated to delineate the safest as possible mother to child HIV-1 transmission prophylaxy. Disclosures: No relevant conflicts of interest to declare. </jats:sec