AF4-MLL Expression Is Necessary and Sufficient for Leukemia Onset in Mice

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 685-685
Author(s):  
Adelheid Bursen ◽  
Karen Schwabe ◽  
Brigitte Rüster ◽  
Anne Wenger ◽  
Martin Ruthardt ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Lymphoblastic Leukemia ◽  
Model System ◽  
Chromosomal Translocations ◽  
Leukemic Cells ◽  
Human Leukemia ◽  
Fusion Genes ◽  
Chromosome 11 ◽  
Mll Gene ◽  
Necessary And Sufficient

Abstract Uniform structural and numerical chromosomal abnormalities are frequently demonstrated in human leukemia and lymphomas, probably as initiating events in cancer formation. Recurrent chromosomal translocations generally result in two derivative chromosomes, both of which are usually present in the leukemic blasts at the time of diagnosis. The human MLL (mixed lineage leukemia) gene on chromosome 11, band q23, participates in a variety of chromosomal translocations, which are assumed to be the initial step of the malignant transformation of haematopoietic cells leading to malignancies of myeloid and/or lymphoid lineage. Translocation t(4;11)(q21;q23) fuses the MLL gene to the AF4 (ALL-1 fused gene on chromosome 4; MLLT2) gene and is one of the most frequent rearrangements of the human MLL gene, being particularly common in infant acute lymphoblastic leukemia (ALL) associated with a poor outcome with treatment. Of note, the fusion of MLL to most other partners results in acute myeloid leukemia (AML). While MLL fusions associated with AML have been successfully established in mice, modeling a t(4;11) associated ALL emerged as more delicate. To generate such a model system in mice and to elucidate a potential association of the resulting fusion genes, MLL-AF4 and AF4-MLL for leukemia phenotype specification, the cDNA constructs of both fusion genes were used in a retroviral transduction/transplantation setup. Therefore murine HSCs (Lin−, Sca-1+) were transduced with either both fusion genes, or with MLL-AF4 or AF4-MLL alone, and subsequently administered by suborbital injection to sublethally irradiated recipient mice. Mice were observed daily and moribund primary AF4-MLL and MLL-AF4/AF4-MLL recipient mice were monitored after a latency of approximately 6 months and with a penetrance of 25% for the AF4-MLL and 40% for the MLL-AF4/AF4-MLL cohort. Diseased mice exhibited the following criteria for classification as a leukemic disorder: All leukemia mice showed enlarged spleen and thymus, and a massive infiltration of lymphoblast-like leukemic cells in the peripheral blood, bone marrow, and other major organs. cDNA cassettes of the fusion genes were transcribed in the analyzed samples, as assessed by RT-PCR. Furthermore, leukemic cells of AF4-MLL and MLL-AF4/AF4-MLL mice could be successfully re-transplanted into secondary recipients with a latency of 3–7 weeks and a penetrance of 90%, phenocopying the primary leukemia. Flow cytometry was used to further characterize the leukemic immunophenotype. Primary AF4-MLL recipients exclusively developed a CD3+ precursor T-cell lymphoblastic leukemia (Pre-T LBL; according to Bethesda proposals for classification of lymphoid neoplasms in mice), and aside from CD3+ Pre-T LBL, one of the MLL-AF4/AF4-MLL leukemia mice displayed a mixed lymphoid/myeloid malignancy. In contrast, expression of the MLL-AF4 fusion protein in muHSCs did not show any detectable effect in recipient mice over an observation period of more than 13 months. Taken together, in this particular model system the expression of the AF4-MLL fusion protein in multi-potent haematopoietic stem cells is necessary and sufficient to cause cancer. Additional expression of the MLL-AF4 fusion protein in murine HSCs indicates an instructive function in lineage determination of the tumor. For further examination of this finding we consider the establishment of a xenograft NOD/SCID mouse model expressing the fusion genes MLL-AF4 and AF4-MLL in human CD34+ cells.

Collaboration of the Meningioma 1 (MN1) Oncogene with MLL-Fusions in Pediatric Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3786-3786
Author(s):  
Ting Liu ◽  
Dragana Jankovic ◽  
Laurent Brault ◽  
Sabine Ehret ◽  
Vincenzo Rossi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Human Leukemia ◽  
Fusion Genes ◽  
Pediatric Leukemia ◽  
Acute Myeloid

Abstract Expression of meningioma 1 (MN1) has been proposed to be a negative prognostic marker in adult acute myeloid leukemia (AML). In pediatric leukemia, we found overexpression of MN1 in 53 of 88 cases: whereas no MN1 expression was detected in T-cell acute lymphoblastic leukemia (T-ALL), significant amounts of MN1 were found in immature B-cell ALL and most cases of infant leukemia. Interestingly, 17 of 19 cases harboring fusion genes involving the mixed-lineage leukemia (MLL-X) gene showed elevated MN1 expression. Lentiviral siRNA mediated MN1 knock-down resulted in cell cycle arrest and impaired clonogenic growth of 3 MLL-X-positive human leukemia cell lines overexpressing MN1 (THP-1, RS4;11, MOLM-13). In a mouse model of MLL-ENL-induced leukemia we found MN1 to be overexpressed as a consequence of provirus integration. Strikingly co-expression of MN1 with MLL-ENL resulted in significantly reduced latency for induction of an AML phenotype in mice suggesting functional cooperation. Immunophenotyping and secondary transplant experiments suggested that MN1 overexpression seems to expand the L-GMP cell population targeted by the MLL-ENL fusion. Gene expression profiling allowed defining a number of potential MN1 hematopoietic targets. Upregulation of CD34, FLT3, HLF, or DLK1 was validated in bone marrow transiently overexpressing MN1, in MN1-induced mouse acute myeloid leukemia, as well as in pediatric leukemias with elevated MN1 levels. Our work shows that MN1 is overexpressed in a significant fraction of pediatric acute leukemia, is essential for growth of leukemic cells, and that MN1 can act as a cooperating oncogene with MLL-ENL most probably through modification of a distinct gene expression program that leads to expansion of a leukemic progenitor population targeted by MLL-fusion genes.

Interaction of AF4 Wildtype and AF4•MLL Fusion Protein with SIAH Proteins: Indication for T(4;11) Pathobiology?.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4268-4268
Author(s):  
Adelheid Bursen ◽  
Moritz Sven ◽  
Gaussmann Anne ◽  
Dingermann Theo ◽  
Marschalek Rolf
Keyword(s):  
Early Childhood ◽  
Acute Lymphoblastic Leukemia ◽  
High Risk ◽  
Fusion Protein ◽  
Lymphoblastic Leukemia ◽  
Recombination Process ◽  
Fusion Genes ◽  
Terminal Portion ◽  
Mll Gene ◽  
Proteosomal Degradation

Abstract The human AF4 gene (4q21) is recurrently involved in recipro cal translocations to the MLL gene (11q23), correlated with high-risk acute lymphoblastic leukemia (ALL) in infants and early childhood. The t(4;11) translocation is one of the most frequent MLL translocations known today. In general, MLL translocations are the result of an illegitimate recombination process leading to reciprocal fusions of unrelated translocation partner (TP) genes with the MLL gene. Due to the constant presence of the derivative(11) product, the hypothesis was posed that only MLL•TP fusion genes are responsible for the leukemogenic process. This concept has been successfully tested for some known MLL fusions, while some other MLL fusions failed. Here, we demonstrate growth transforming potential of AF4 wildtype and the AF4•MLL fusion protein. The underlying oncogenic mechanism involves the two E3 ubiquitin ligases SIAH1 and SIAH2, the N-terminal portion of AF4 and the protection of the AF4•MLL fusion protein against proteosomal degradation. Supported by grant Ma 1786/4-1 from the DFG.

scholarly journals The Effect of L-Asparaginase on the Nucleic Acid Metabolism and Cell Cycle of Human Leukemia Cells

Blood ◽  
1972 ◽  
Vol 39 (4) ◽  
pp. 575-580 ◽  
Author(s):  
E. Fred Saunders
Keyword(s):  
Cell Cycle ◽  
Nucleic Acid ◽  
Dna Synthesis ◽  
Rna Synthesis ◽  
Lymphoblastic Leukemia ◽  
Buffy Coat ◽  
Leukemic Cells ◽  
Nucleic Acid Metabolism ◽  
Rapid Decline ◽  
Human Leukemia

Abstract The effect of L-asparaginase on the cell cycle and nucleic acid synthesis of leukemic cells was studied in five children with acute lymphoblastic leukemia. Following an intravenous infusion of the drug, serial marrow samples were obtained for buffy coat volume, mitotic index, and autoradiographic assessment of DNA and RNA synthesis using tritiated thymidine and tritiated uridine, respectively. A rapid decline in buffy coat volume indicated a lytic effect on lymphoblasts. There was a greater kill of proliferative (blasts in the cell cycle) than nonproliferative (G0) leukemic cells. Mitotic indices changed little until 24 hr; in contrast, thymidine labeling indices decreased markedly to less than 50% of control by 6 hr. The changes in labeling indices prior to changes in mitotic indices indicated that L-asparaginase blocked the entrance of cells into the DNA synthesis period of the cell cycle. Cells already in DNA synthesis appeared to continue into mitosis. Uridine labeling indices decreased progressively in all patients. Uridine uptake was inhibited equally in both proliferative and nonproliferative blasts. Therefore, inhibition of RNA synthesis by L-asparaginase was independent of the proliferative activity of the marrow.

Expression and Function of the CXCR7 Chemokine Receptor in Leukemias

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2423-2423
Author(s):  
Sergej Konoplev ◽  
Hongbo Lu ◽  
Michael A Fiegl ◽  
Zhihong Zeng ◽  
Wenjing Chen ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cell ◽  
Leukemic Cells ◽  
Human Leukemia ◽  
Rt Pcr ◽  
Hematopoietic Stem ◽  
Leukemic Cell Lines ◽  
And Function

Abstract Background: Bone marrow produced stromal-derived factor-1a (SDF-1a) is a key chemokine involved in chemotaxis, homing, mobilization, and expansion of hematopoietic stem and progenitor cells. While the majority of well-defined functions of SDF-1a are mediated via its receptor CXCR4, recent studies have characterized CXCR7 as an alternative receptor capable of binding SDF-1a. Although the functions of CXCR7 are still incompletely understood, the receptor was reported to promote migration and adhesion in certain cell types and function as a pro-survival factor in breast cancer cells. CXCR7 expression and function in human leukemia cells has not been characterized. In this study, we examined CXCR7 expression in leukemia cell lines and primary samples from patients with acute lymphoblastic leukemia (ALL) and utilized a small molecule inhibitor of CXCR7 to probe CXCR7’s function. Materials and methods: CXCR4 and CXCR7 expression was determined by flow cytometry, real-time PCR (RT-PCR) and immunocytochemistry (ICC) in leukemic cell lines including AML (OCI-AML2, OCI-AML3, HL60, U937 NB4, Molm13), ALL (REH, Raji, RS4; 11, Nalm6, Molt4) and CML (KBM5, K562) cells. In primary ALL patient samples, CD34+CD19+ gating was applied to detect CXCR7 expression on pre-B leukemic cells by flow cytometry. The migration of leukemic cells towards SDF-1a was studied using a transwell system. CXCR4 inhibitor AMD3100 was purchased from Sigma, and CXCR7 inhibitor CCX-733 was provided by ChemoCentryx Inc., Mountain View, CA. Results: CXCR4 was found to be ubiquitously expressed on the cell surface of all leukemic cell lines tested. CXCR7 mRNA and protein expression was detectable only in Burkitt lymphoma Raji cells, as analyzed by flow cytometry (clone 11G8, R&D systems), RT-PCR and ICC. Curiously, CXCR7 expression was significantly induced in MOLM13 cells under hypoxic (6% O2) conditions (p=0.01). Low levels of surface CXCR7 were found in 8 of the 9 primary ALL samples by flow cytometry. To determine the respective roles of CXCR4 and CXCR7 in migration of leukemic cells, we utilized CXCR4 inhibitor AMD3100 and CXCR7 inhibitor CCR733 in Raji (CXCR7 positive) and RS4;11 (CXCR7 negative) cells. AMD3100 at 25μM significantly inhibited SDF-1a induced migration (from 38.5% to 12%); CCR733 at 10μM also inhibited SDF-1a induced migration (from 38.5% to 24%) and the combination of AMD3100 and CCR733 resulted in 81% inhibition of migration (from 38.5% to 7.2%). AMD3100 blocked SDF-1a induced migration of CXCR4+CXCR7− RS4;11 cells (from 36.5% to 15.8%), while CCR733 had no effect (36.5% and 39.2%). In conclusion, these studies demonstrate functional expression of the SDF-1 receptor CXCR-7 on Raji and primary ALL cells and suggest that CXCR7 plays an active role in the migration of leukemic cells. CXCR-7 may serve as an alternative receptor to CXCR4. Studies addressing the role of CXCR7 in adhesion, SDF-1a-mediated signaling and survival of leukemic cells are in progress.

MLL genomic DNA Breakpoints In Infant Acute Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1350-1350
Author(s):  
Grigory Tsaur ◽  
Claus Meyer ◽  
Alexander Popov ◽  
Olga Plekhanova ◽  
Anatoly Kustanovich ◽  
...  
Keyword(s):  
Acute Leukemia ◽  
Genomic Dna ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Prognostic Significance ◽  
Chromosome 11 ◽  
Long Distance ◽  
Mll Gene ◽  
Stable Pattern ◽  
Breakpoint Location

Abstract Background Infant acute leukemia is characterized by high incidence of MLL gene rearrangements. Purpose To evaluate the distribution of MLL genomic DNA breakpoints and their relation to several diagnostic parameters among infant acute leukemia. Methods 72 infants with MLL-rearranged acute lymphoblastic leukemia (ALL) (n=52), acute myeloid leukemia (AML) (n=19) and mixed phenotype acute leukemia (n=1) were included in this study based on the availability of DNA material at diagnosis. In the observed group there were 28 boys (39%) and 44 girls (61%) with median age of 4.9 mo (range 0.03-11.9). Genomic DNA breakpoint detection in MLL gene and translocation partner genes (TPG) was performed by long-distance inverse PCR (LDI-PCR). Exon-intron numbering of MLL gene was done according to I. Nilson et al, 1996. Results Majority of ALL cases (n=28; 54%) was characterized by presence of MLL-AF4 fusion gene (FG), less frequently MLL-MLLT1 (n=12; 23%), MLL-MLLT3 (n=7; 13%) and others were found (Table 1). The most common breakpoint location within MLL gene in ALL patients was intron 11, detected in 25 cases (48%). The highest variability of MLL breakpoints was found in MLL-AF4-positive patients: only 11 of 28 (39%) had breakpoints in intron 11. The most stable pattern of MLL genomic DNA breakpoints was observed in MLL-MLLT1-positive patients: 8 of 12 (67%) had breakpoints in intron 11. In AML patients two the most prevalent FGs were MLL-MLLT3 (n=7, 37%) and MLL-MLLT10 (n=5, 26%). The remaining ones are listed in Table 1. The most frequent breakpoints location was intron 8 (8 out of 19, 42%). The most stable pattern was revealed for MLL-MLLT10 FG: MLL breakpoints in 4 of 5 (80%) cases were found in intron 9 (Table 1). ALL patients who had breakpoints in intron 11 were significantly younger (median 3.0 mo, range 0.03-11.6) than all others (median 5.6 mo, range 0.7-11.9) (p=0.025) and than patients with MLL breakpoints in intron 9 (median 6.6 mo, range 3.1-11.9) (p=0.017). For AML cases we did not find any relation between age and breakpoints locations. Distribution of MLL DNA breakpoints was similar in boys and girls and did not depend on type of TPG. Genetic recombinations involving MLL gene predominantly resulted in reciprocal chromosomal translocations (n=62; 86%). Beside them, 6 (11%) insertions were identified in all MLL-MLLT10-positive cases and MLL-SEPT6-positive one. In 11 (15%) patients we found breakpoints within the regions located from 0.7 Kb to 25.4 Kb 3' of the first exon of TPGs (MLLT1 n=9; EPS15 n=1; MYO1F n=1), however fusion transcripts at cDNA level were identified and sequenced in all these cases, indicating a spliced fusion mechanism. 3-way translocations were found in 5 patients and in 1 case we found combination of insertion with interstitial deletion of chromosome 11. The list of reciprocal genes involved in these 6 cases was as follows: CEP164, DNAH6, DCPA1, MCL1 as well as non-coding regions of 2q21.2 and 2p21. We also analyzed breakpoints in TPGs. Except above mentioned spliced fusion cases, the remaining 3 breakpoints in MLLT1 as well as 3 of 4 breakpoints in EPS15 and all breakpoints in MLLT11 were within intron 1 of corresponding genes. In AF4 the major breakpoint region included intron 3 (n=19), intron 4 (n=6) and intron 5 (n=2). We also revealed 2 rare breakpoints in intron 6 and 10. In MLLT3 the most frequent breakpoint location was intron 5 (n=12), additionally 2 cases in intron 5 were identified. In MLLT10 two separate breakpoint locations were found: intron 3 (n=1) and intron 8 (n=3) in combination with intron 9 (n=1). We estimated prognostic significance of MLL breakpoint locations in 31 cases of infant ALL treated by MLL-Baby protocol. 3-year cumulative incidence of relapse was remarkably higher in patients with breakpoints in intron 11 (n=18) in comparison to patients with breakpoint localized from intron 7 to exon 11, inclusively (n=13) (0.85±0.01 and 0.57±0.02, respectively), although difference between these two groups did not achieve statistical significance (p=0.261). Median follow-up time in the observed group was 30 months (range 6–42). Conclusion In the current study we estimated clinical and prognostic significance of MLL and TPG genomic DNA breakpoints in infant acute leukemia. Our data provide additional information of molecular genetic features of MLL-rearranged infant acute leukemia. Disclosures: No relevant conflicts of interest to declare.

PRMT1 Activates Leukemic Stem Cell Program in MLL-Rearranged Leukemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 3493-3493
Author(s):  
Wing Chi Lui ◽  
Yuen Fan Chan ◽  
Ray Ng
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Fusion Protein ◽  
Fusion Proteins ◽  
Leukemic Cells ◽  
Leukemic Stem Cell ◽  
Self Renewal ◽  
Mll Gene ◽  
Acute Myeloid

Abstract In MLL-rearranged leukemia, the Mixed Lineage Leukemia (MLL) gene undergoes chromosomal translocation that results in the loss of C-terminal histone methyltransferase SET domain, whereas the N-terminal of MLL gene fuses in-frame with one of the 60 identified partner genes. The resultant MLL fusion proteins lead to a characteristic aberrant gene expression pattern in human acute myeloid and lymphoblastic leukemia. Epigenetic dysregulation mediated by MLL fusion proteins has been suggested to be a key event in MLL-rearranged leukemia. It has been demonstrated that MLL-EEN/PRMT1 oncogenic complex induces transformation of primary myeloid progenitors via introduction of aberrant H4R3me2 at target Hoxloci. PRMT1 is the predominant protein arginine methyltransferase in mammals and is responsible for over 85% of arginine methylation activity in mammalian cells. Dysregulation of PRMT1 has been implicated in different cancers such as leukemia, suggesting the expression of PRMT1 is positively correlated with cancer progression and clinical parameters. Nevertheless, the leukemogenic role of PRMT1 in the establishment of leukemic stem cell (LSC) remains unclear. Previously we have demonstrated that a MLL fusion protein, MLL-EEN, can strongly enhance the self-renewal ability of murine primary hematopoietic cells through multiple rounds of replating assays. We have created a conditional Mll-Een invertor mouse model (MllEen/+) in which the expression of fusion protein is restricted to hematopoietic progenitors. Immunophenotypic analysis demonstrated a significant increase in the immature myeloid cell population (c-kit+Mac-1+) in bone marrow of MllEen/+ mice, suggesting that the expression of Mll-Een induces the development of acute myeloid leukemia. We have also established an Mll-Een expressing cell line from the bone marrow of MllEen/+ mouse. These leukemic cells can persistently form colonies and they also demonstrated deregulation of Hox genes, which is frequently observed in human leukemia cases. The leukemogenicity of Mll-Een is closely associated with Prmt1, which was demonstrated through knockdown of Prmt1. Strikingly, we discovered a subpopulation of CD41+Mll-Een expressing cells, which showed enhanced self-renewal ability in the serial colony forming assays. The percentage of CD41+ leukemic cells is reduced once Prmt1 was knocked down, suggesting that Prmt1 is crucial in the maintenance of this subpopulation of cells. In addition, the CD41+ cells showed enhanced expression of genes associated with hematopoietic stem cell (HSC) activities (Bmi-1, Runx1, Tal-1 and Lmo2), implying that part of the HSC transcriptional program has been re-activated in these cells. We therefore speculate that the CD41+ cells may represent a group of MLL leukemic cells that harbors strong stem cell features, and presumably functions as LSCs. The CD41+ leukemic cells will be further characterized by their LSC functions and CD41 can potentially serve as a novel LSC marker in MLL-rearranged leukemia. Taken together, studies on the role of PRMT1 can provide novel insights on the establishment of LSC and the development of effective clinical treatment for MLL-rearranged leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals Rearrangement of the chromosome 11 bcl-1 locus in centrocytic lymphoma: analysis with multiple breakpoint probes

Blood ◽  
1991 ◽  
Vol 78 (2) ◽  
pp. 493-498 ◽  
Author(s):  
ME Williams ◽  
TC Meeker ◽  
SH Swerdlow
Keyword(s):  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Chromosomal Translocation ◽  
Large Cell ◽  
Chromosomal Translocations ◽  
Cell Type ◽  
Chromosome 11 ◽  
Myc Oncogene ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Ig Heavy Chain

Centrocytic lymphoma is a B-cell non-Hodgkin's lymphoma (NHL) composed of lymphocytes resembling cleaved follicular center cells (centrocytes). Previous studies have suggested an association between t(11;14) chromosomal translocations and bcl-1 rearrangement in centrocytic and related intermediate lymphocytic lymphomas. To further characterize the association between bcl-1 and centrocytic lymphoma, Southern blot analysis was performed on samples from 23 patients using four separate bcl-1 breakpoint probes spanning 63 kb of the chromosome 11 bcl-1 locus. Rearrangements were identified in six patients with the major translocation cluster (MTC) probe and in another six with probe p94PS, located about 24 kb 5′ of MTC. Eleven of these 12 cases showed comigration of rearranged bcl-1 and Ig heavy chain-joining genes, consistent with the t(11;14) chromosomal translocation. No rearrangements were observed with the bcl-1 locus probes p210 or p11EH located 5′ of p94PS, nor with bcl-2 or c-myc oncogene probes. No bcl-1 rearrangements were identified in B-cell follicular NHL (15), small noncleaved cell (Burkitt's and non-Burkitt's) NHL (8), T-cell NHL (4), multiple myeloma (14), and pre-B-cell acute lymphoblastic leukemia (9). One of 23 B-cell NHL of large cell type and one of 19 chronic lymphocytic leukemias or small lymphocytic NHL had MTC rearrangement. Thus, bcl-1 rearrangement occurred at MTC or p94PS in 12 of 23 centrocytic lymphomas (52%), confirming a nonrandom association and suggesting a pathogenetic role for the bcl-1 locus in this immunohistologic subtype of NHL.

scholarly journals Childhood acute lymphoblastic leukemia with chromosomal breakpoints at 11q23

Blood ◽  
1989 ◽  
Vol 73 (6) ◽  
pp. 1627-1634 ◽  
Author(s):  
SC Raimondi ◽  
SC Peiper ◽  
GR Kitchingman ◽  
FG Behm ◽  
DL Williams ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cells ◽  
Chromosome 11 ◽  
Myeloid Antigens ◽  
Hla Dr ◽  
Chromosomal Breakpoints ◽  
Leukocyte Counts ◽  
Structural Chromosomal Abnormality ◽  
B Lineage

Twenty-one (5.7%) of 368 cases of acute lymphoblastic leukemia (ALL), studied fully for karyotype and immunophenotype, had breakpoints in the q23 region of chromosome 11. This abnormality resulted from reciprocal translocation in 17 cases [with chromosomes 4 (n = 5), 10 (n = 2), and variable chromosomes (n = 10)], from deletions in three cases, and from a duplication in one case. The 17 children with 11q23 translocations had higher leukocyte counts (P less than .01) and were more likely to be black (P less than .01) and younger (P = .08) as compared with each of the following non-11q23 translocation groups: t(1;19), t(9;22), random translocations, and cases without translocations. Event-free survival at 3 years for the 11q23 translocation group did not differ significantly from that of the t(1;19), t(9;22), or random translocation groups. Leukemic cells from ten of the 21 patients with an 11q23 structural chromosomal abnormality had an immunophenotype indicative of B-lineage ALL (HLA-DR+, CD19+, CD2-, CD3-); this was confirmed by the presence of rearranged immunoglobulin heavy-chain genes in seven cases. In eight of these ten B-lineage cases, the blasts were negative for expression of the CD10 antigen, indicating a primitive stage of B-cell development. Four cases were classified as T- cell ALL, and seven others were characterized by blasts that failed to react with our panel of lineage-associated monoclonal antibodies (MoAbs). Myeloid antigens were expressed by leukemic cells in three of the cases that were tested. The initial clinical features associated with translocations involving the 11q23 chromosomal region may define a distinct subtype of ALL. Whether the constellation of findings relates to a breakpoint at 11q23 per se or to the specific translocation will require further study.

scholarly journals Targeted Therapy for ETV6-RUNX1-Driven B-Cell Precursor Acute Lymphoblastic Leukemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 62-62
Author(s):  
Roel Polak ◽  
Marc B. Bierings ◽  
Cindy S. van der Leije ◽  
Rosanna E.S. van den Dungen ◽  
Mathijs A. Sanders ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Fusion Protein ◽  
Cell Viability ◽  
B Cell ◽  
Cell Survival ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Growth Arrest ◽  
Leukemic Cells ◽  
Cell Precursor

Abstract Background: Translocation t(12;21), resulting in the ETV6-RUNX1 fusion protein, is present in 25% of pediatric patients with B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Despite the favorable prognostic parameters of this B-ALL subgroup, relapse and resistance to chemotherapeutics occur and treatment-induced side effects are considerable. The molecular mechanisms underlying ETV6-RUNX1-driven leukemia are largely unknown. Increased knowledge of these mechanisms is essential to develop novel therapeutic strategies to selectively target ETV6-RUNX1-positive leukemia. Objectives: This study aims to identify and target the molecular drivers behind ETV6-RUNX1-positive BCP-ALL. Results: Gene expression profiling of leukemic blasts of 654 ALL patients revealed that the class III PI3-kinase Vps34, an important regulator of autophagy, was exclusively up-regulated in ETV6-RUNX1-positive compared to ETV6-RUNX1-negative BCP-ALL patients (2.7-fold; p ≤ 10-30). In addition, ectopic expression of ETV6-RUNX1 in cord blood-derived hematopoietic progenitor cells (CB-HPCs) significantly induced expression of Vps34 1.3-fold already 40 hours after transduction (p ≤ 0.05). This suggests that the Vps34-autophagy pathway is activated by ETV6-RUNX1, which may mechanistically explain the leukemogenic and pro-survival properties ascribed to ETV6-RUNX1. In correspondence, Ingenuity Pathway Analysis (IPA) predicted a pro-survival and pro-proliferative phenotype in ETV6-RUNX1 transduced CB-HPCs and highlighted a network of up-regulated transcription factors, including HEY1, EGR1, GATA1 and GATA2 (2 – 25-fold up-regulation; p ≤ 0.05). Luciferase reporter assays revealed that not only the ETV6-RUNX1 fusion protein, but also the ETV6-RUNX1-induced target genes HEY1, EGR1 and GATA1 positively regulate Vps34 promoter activity (5 – 13-fold up-regulation; p ≤ 0.01).Lentiviral knockdown experiments were performed to elucidate the importance of Vps34 expression in ETV6-RUNX1-positive BCP-ALL cells. Knockdown of all Vps34 transcript variants, with two independent constructs, led to complete growth arrest of the ETV6-RUNX1-positive cell lines REH and AT2, while this only led to a decrease in proliferation of the ETV6-RUNX1-negative cell line NALM6. This growth arrest was caused by a significant induction of apoptosis (more than 4-fold 7 days after transduction; p ≤ 0.001) and a significantly reduced percentage of cycling cells (1.3-fold 7 days after transduction; p ≤ 0.05). Analysis of p62 protein expression by western blot and reverse phase protein arrays revealed that the levels of autophagy were significantly higher in ETV6-RUNX1-positive compared to ETV6-RUNX1-negative BCP-ALL patients (p ≤ 0.001). In addition, knockdown of ETV6-RUNX1 and Vps34 significantly reduced autophagy, quantified with confocal microscopy, in ETV6-RUNX1-positive cells with 50% and 84%, respectively (p ≤ 0.01). Furthermore, pharmacological inhibition of autophagy with hydroxychloroquine (HCQ) significantly reduced cell viability of BCP-ALL cell lines and primary patient-derived BCP-ALL cells (p ≤ 0.001). Treatment of the ETV6-RUNX1-positive BCP-ALL cell lines REH and AT2 with 20 µg/mL HCQ resulted in a 82% and 95% reduced cell viability, while the viability of ETV6-RUNX1-negative BCP-ALL cell lines and T-ALL cell lines were reduced to a lesser extent (NALM6: 43%; TOM-1: 50%; Loucy: 40%; Jurkat: 0%). Importantly, HCQ selectively sensitized ETV6-RUNX1-positive leukemic cells to L-asparaginase treatment in clinically relevant concentrations. Treatment of primary ETV6-RUNX1-positive patient cells with 10 µg/mL HCQ resulted in a 70% reduction in cell survival during L-asparaginase exposure (p ≤ 0.01). This sensitization was not observed in ETV6-RUNX1-negative BCP-ALL cells. Conclusion: The ETV6-RUNX1 fusion protein activates autophagy via Vps34, which is essential for survival and proliferation of ETV6-RUNX1-positive cells. Inhibition of autophagy in primary ETV6-RUNX1-positive leukemic cells inhibited cell survival and sensitized these cells to L-asparaginase treatment. These results indicate that autophagy inhibition may provide a novel means to sensitize L-asparaginase-resistant ETV6-RUNX1-positive BCP-ALL patients. Disclosures No relevant conflicts of interest to declare.

scholarly journals Cloning of several species of MLL/MEN chimeric cDNAs in myeloid leukemia with t(11;19)(q23;p13.1) translocation

Blood ◽  
1995 ◽  
Vol 85 (8) ◽  
pp. 2017-2024 ◽  
Author(s):  
K Mitani ◽  
Y Kanda ◽  
S Ogawa ◽  
T Tanaka ◽  
J Inazawa ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Fusion Protein ◽  
Leukemic Cells ◽  
Cdna Libraries ◽  
New Class ◽  
Mll Gene ◽  
Unknown Gene ◽  
Alternatively Spliced ◽  
11Q23 Abnormalities ◽  
Chimeric Transcription Factor

The t(11;19)(q23;p13.1) translocation is thought to play an important role in pathogenesis of myeloid leukemias in older patients. The MLL gene involved in other 11q23 abnormalities was also rearranged by this translocation. Screening of cDNA libraries of the t(11;19)(q23;p13.1)-carrying leukemic cells resulted in the isolation of several species of fusion cDNAs between the MLL gene and an unknown gene on 19p13.1, named MEN (myeloid eleven-nineteen translocation), which is ubiquitously expressed. Although the MLL gene was alternatively spliced, the fusion protein should contain an N-terminal half of the MLL, including AT hook motifs, that is fused to the MEN protein with a lysine-rich sequence, suggesting that the MLL/MEN fusion protein could be a chimeric transcription factor. The MLL/MEN fusion transcripts of 8.0 kb were detected in leukemic cells of two cases with the translocation. The MLL/MEN fusion was consistent in all three cases of the t(11;19)(q23;p13.1)-carrying leukemia examined by RNA-based polymerase chain reaction. These findings strongly suggest that the t(11;19)(q23;p13.1) results in the fusion formation encoding a new class of potential chimeric transcription factor that contributes to leukemogenesis of myeloid lineage.

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