Positional Cloning of the Mouse Scat Mutation Reveals a Critical Role for RASA3, a Ras GTPase Activating Protein, in Vertebrate Erythropoiesis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 776-776
Author(s):  
Luanne L. Peters ◽  
Steven L. Ciciotte ◽  
E. Ricky Chan ◽  
Babette Gwynn ◽  
Amy J. Lambert ◽  
...  
Keyword(s):  
Missense Mutation ◽  
Positional Cloning ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Chromosome 8 ◽  
Recessive Mutation ◽  
Severe Thrombocytopenia ◽  
Gtpase Activating Protein ◽  
Form Analysis ◽  
Mild Anemia

Abstract Abstract 776 The spontaneous recessive mutation scat (severe combined anemia and thrombocytopenia) arose on the inbred BALB/cBy (BALB) mouse strain. The phenotype of scat is cyclic. All homozygotes are severely anemic and thrombocytopenic at birth. Leukocytes are also significantly depleted (Table). Approximately 13% die during this first “crisis” episode that lasts, on average, until the 9th postnatal day. Remarkably, a spontaneous remission ensues in those surviving the neonatal crisis period wherein all peripheral blood values revert to normal. A second crisis follows, and 94% of the mice die by 30 days of age. The recessive ENU (N-ethyl-N-nitrosourea)-induced mutation, hlb381, on the C57BL/6J (B6) background, is characterized by severe thrombocytopenia and leukopenia with mild anemia. Unlike scat, hlb381 is not cyclical. The phenotype is present at birth and persists throughout life. Despite the phenotypic differences, scat and hlb381 interact genetically; double heterozygotes show non-cyclical severe thrombocytopenia and leukopenia, and mild anemia (Table). This interaction implies that the scat and hlb381 genetic defects affect the same gene or distinct genes within the same pathway. Both scat and hlb381 mapped to overlapping intervals on mouse chromosome 8. Sequence analysis of genes within the interval identified Rasa3 (GAPIII, GAP1IP4BP) as a strong candidate gene for both scat and hlb381. In scat, Rasa3 carries a missense mutation near the N-terminus (G125V, exon 5) and, in hlb381, a missense mutation near the C-terminus (H794L, exon 23). RASA3 is a GTPase activating protein (GAP) that negatively regulates p21 Ras function by accelerating GTP hydrolysis and converting Ras to the inactive GDP bound form. Analysis in Panther and SIFT predicts that both residues are highly conserved and substitutions are likely to be deleterious. Rasa3 is widely expressed throughout embryonic and fetal development in mice, and is ubiquitously expressed in zebrafish 24 hours post fertilization (hpf). RASA3 protein is detected in erythroid tissues and platelets in the adult mouse. Analysis of scat spleen and bone marrow erythrocyte populations by FACS (dual staining for Ter119 and CD71 followed by forward scatter of the Ter119 high population) reveals a severe block in erythropoiesis during crisis periods. The proerythroblast, EryA (basophilic erythroblasts), and EryB (late basophilic and polychromatophilic erythroblasts) populations are significantly increased in frequency vs. wild type, and the EryC (orthochromatophilic erythroblasts and reticulocytes) population is markedly decreased. Annexin V staining revealed no significant differences in any of these populations. Notably, a similar delay in erythroid maturation, albeit much milder, is also seen in hlb381. In pull-down assays using the Ras-binding domain of Raf1 to affinity purify active GTP-bound Ras followed by detection by western blotting with pan-Ras antibody, active GTP-bound Ras is deficient in scat crisis red cells but recovers during remission. Finally, injection of two independent splice-blocking morpholinos designed to disrupt exon 5 and induce disruption of rasa3 mRNA processing resulted in a major decrease in the number of hemoglobinized cells when stained with o-dianisidine at 48 and 72 hpf in zebrafish. Over 90% of morphants showed no hemoglobinized cells at all, or vastly reduced numbers (<20) of hemoglobinized cells. We conclude that RASA3 plays a critical role in vertebrate erythropoiesis. Differences in the scat and hlb381 phenotypes likely result from allele-specific interactions mediated by the different genetic backgrounds (B6 vs. BALB) or domain-specific functions of the RASA3 protein.Hematological ValuesWBC × 103/μLRBC × 106/μLHb g/dLHct %Reticulocytes %Platelets × 103/μLSpleen weight (% body weight)BALB-scat homozygotes in crisis and remissionBALB-nl5.28.713.241.911.79550.74 scat crisis2.4*2.8*4.2*14.5*51.6*145*2.69*scat remission4.17.912.541.022.95611.22BALB,B6-hlb381 homozygotes and scat/hlb381 double heterozygotesBALB,B6-nl5.19.113.744.08.612190.28 hlb381/hlb3812.1*9.113.343.512.0*24*0.50*hlb381/scat2.4*8.4*12.3*39.9*12.1*14*NAAll mice 18-30 days old. All values X ± SD; nl, normal littermates, NA, no data available.*P < 0.05 vs. normal littermates Disclosures: No relevant conflicts of interest to declare.

Failure of Erythropoiesis and Megakaryocytopoiesis in RASA3 Mutant Scat Mice

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 680-680
Author(s):  
Lionel Blanc ◽  
Babette Gwynn ◽  
Steven L. Ciciotte ◽  
Luanne L. Peters
Keyword(s):  
Bone Marrow ◽  
Positional Cloning ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Inbred Mouse Strain ◽  
Recessive Mutation ◽  
Initial Increase ◽  
Severe Anemia ◽  
Wild Type ◽  
Hematopoietic Stem

Abstract Abstract 680 Scat (severe combined anemia and thrombocytopenia) is a spontaneous, autosomal recessive mutation coisogenic with the BALB/cBy inbred mouse strain. Homozygous scat mice present a cyclic phenotype with alternating episodes of crisis and remission. As its name implies, crisis episodes are characterized by severe anemia and thrombocytopenia, but significant lymphocyte depletion occurs as well. The first crisis episode begins in utero, lasts until postnatal day (P) 9 on average, and is associated with 10–15% mortality. Remarkably, in homozygotes that survive the first crisis, a remission phase occurs wherein the disease phenotype reverts to normal. This remission is transient, however, and is followed by a second crisis episode during which 94% of scat/scat mice die by P30. Previously we showed that the scat phenotype is transferrable via the hematopoietic stem cells and is also recapitulated in scat/scat, Hox11−/− double homozygotes in which a spleen does not develop, indicating that the splenic micro-environment plays little or no role in disease appearance or progression. Positional cloning of scat revealed a missense mutation in Rasa3 encoding a GTPase activating protein (GAP) that negatively regulates Ras function by accelerating GTP hydrolysis and converting Ras to the inactive GDP bound form. We further showed that Rasa3 is a conserved gene critical to vertebrate erythropoiesis via morpholino knockdowns in zebrafish which resulted in profound anemia. Here we report data that shed further light on RASA3 function during hematopoiesis. Overall, the data indicate that defects in RASA3 profoundly and negatively impact erythropoiesis and megakaryocytopoieis through, at least in part, a Ras-mediated mechanism. FACS analyses of scat spleen and bone marrow erythroid populations reveal a severe block in erythropoiesis during crisis periods. In the spleen, despite an initial increase in size due to expansion of Ter-119+ cells, there is ultimately a loss of compensatory erythropoiesis resulting in a return to normal cellularity and a striking loss of hemoglobinized cells as the crisis phenotype deepens. In addition, the bone marrow shows loss of Ter-119+ cells and overall cell depletion during crisis. Megakaryocyte numbers are increased in scat crisis BM and spleen. By transmission electron microscopy, scat crisis megakaryocytes display features characteristic of a significant developmental delay: a disorganized demarcation membrane system with no platelet forming areas and few granules with hypersegmented nuclei and excess rough endoplasmic reticulum. In addition to the severe anemia and thrombocytopenia, a significant lymphopenia occurs in scat crisis mice. However, the scat phenotype is not lymphocyte mediated, as the scat phenotype is completely recapitulated in mice doubly homozygous for scat and the immunodeficient mutations, scid and Rag1tm1Mom, in which B- and T-lymphocytes are completely depleted. Together these results suggest that lymphopenia is a secondary phenomenon in scat, and the severe anemia and thrombocytopenia aspect of the phenotype neither follows from nor is dependent upon loss of lymphocytes. Despite the delay observed in erythroid differentiation, some mature red cells are produced although ∼50% of these are reticulocytes. By confocal microscopy, we show that RASA3 protein localizes to the plasma membrane as well as internal membrane compartments in wild type reticulocytes, where it partially colocalizes with CD71. Western blot analyses of reticulocytes after Percoll gradient purification show that RASA3 is lost during the maturation step, both in vivo and in vitro. Interestingly, in scat, RASA3 is present in reticulocytes, but appears to be mislocalized, the protein being found in the cytosol. Preparation of ghosts from wild type and scat reticulocytes confirms that RASA3 is not attached to the membrane in scat animals during crisis. In pull-down assays active GTP-bound Ras is increased in scat crisis reticulocytes when compared to wild type, suggesting that scat is a RASA3 loss of function mutation due to its mislocalization and demonstrating a critical role for the RASA3-Ras axis during mammalian erythropoiesis. Disclosures: No relevant conflicts of interest to declare.

Coexistance of Multiple Myeloma and Myelodysplastic Syndrome or Lymphoid Diseases At Diagnosis: Evidence for Two Clonal and Independent Chromose Abnormalities

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 5063-5063
Author(s):  
Hossein Mossafa ◽  
Sabine Defasque ◽  
Christine Fourcade ◽  
JeanPierre Hurst ◽  
Bertrand Joly
Keyword(s):  
Multiple Myeloma ◽  
Plasma Cells ◽  
Chromosomal Abnormalities ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Chromosome 8 ◽  
Trisomy 8 ◽  
Hematopoietic Stem ◽  
Monosomy 7 ◽  
Trisomy 12

Abstract Abstract 5063 Introduction, We describe the simultaneous presentation of multiple myeloma (MM) and yeloproliferative disorders (MPD) or lymphoid diseases (LD) at diagnosis. Therapy-related myelodysplasia (tMDS) occurring during the course of MM is generally believed as a result from hematopoietic stem cell-toxic therapies, such as ionizing radiation and alkylating agent-based chemotherapies (melphalan, nitrosoureas).Patients and methods, We study a total of 342 patients (151 F, 191 M; median age 68.1 years; range 42 to 93 Years), diagnosed with MM based on the International Staging System. The basis for inclusion of patients in this study was with previous untreated MM ones. The study was performed in accordance with the declaration of Helsinki. To determine whether chemotherapies for MM factors play the critical role in the development of secondary disease, simultaneously two different cultures were processed, an unstimulated 96 hours culture (U96HC) on whole BM(WBM), a short-time 24 hours culture (ST24HC) after CD138+ plasma cells (PCs) depleted on negative fraction (CD138- cells) of BM and the FISH was investigated on purified CD138+.All samples were enriched in PCs by the Automated Magnetic Cell Sorter (Miltenyi technology)proceeded with anti-CD138 specific antibodies applied. The CD138+ PCs and the CD138- cells were collected in different tubes. The CD138− cells were used for a ST24HC. FISH was performed on the purified CD138+, PCs with a recommended FISH panel (MM International Working Group). Screening was performed systematically for the following unbalanced alterations and reciprocal rearrangements: del(13)(q14)(D13S25), del(17)(p13)(TP53),+3(D3Z), +9(D9Z1), +15(D15Z14), t(4;14)(p16;q32)/IGH-FGFR3, t(11;14)(q13;q32)/IGH-CCND1 (Abbott).After observing the results of U96HC on whole BM (CD138+ and CD138− cells), ST24HC (CD138− cells) and FISH for each patient, two clone cytogenetically were distinct and unrelated chromosomal abnormalities were found in 40 (11.7%) of the 342 MM patients (6 F, 34 M; median age 74 years; range 42 to 87 Years) 34 had a MPD and 6 had a LD. A second immunophenotyping analysis confirmed the presence of those LD/MM simultaneous haematological malignancy. In the cases of the patients with MM/ MPD, the frequency of cytogenetic abnormality unrelated to the myeloma clone was respectively; the 20q deletion, detected for 13 the 34 patients, the 20q- is a sole abnormality for 12 cases and associated with a complex caryotype in 1 case. The trisomy of chromosome +8 was observed in 7 cases, the del(7q) or monosomy 7 in 5 cases, loss of gonosome Y in 4 cases, del(11) for 2 cases, translocation t(9;22) in one case, 5q abnormality in one case and trisomy 9 with JAK2 V617F mutation in one case. For the patients with MM/LD, 5 patients had a trisomy +12 and or trisomy +18 like sole abnormality or associated with others cytogenetics abnormalities and one patient had 6q deletion. Discussion, Whereas in the literature the most common cytogenetic abnormalities typifying MPD after alkylator-based therapy include partial or complete deletions of chromosomes 5, 7, and 20 as well as trisomy 8. In our study we observed those abnormalities with the same frequency for the patients had simultaneous MPD associated in untreated MM at diagnosis. Six patients had simultaneous LD and MM. The marginal zone lymphoma was confirmed for 3 patients. The CC observed a trisomy +12 for those three patients associated with +18 and +19 for 2 cases and del(13) and trisomy 3 for one among them. We demonstrated in untreated MM patients the coexistence of MM and MPD or LD at diagnosis with MPD-type or LD-type chromosome abnormalities within MM signature karyotype. We hence recommend that CC studies, 96 hours WBM, 24 hours on negative fraction CD138− cells and FISH on purified CD138+ PCs, the three should be an integral part of the evaluation of patients with MM at diagnosis into clinical trials using HDT is warranted to determine whether patients who are predisposed to developing tMDS/sAML, they can be identified prospectively. Disclosures: No relevant conflicts of interest to declare.

KAUTILYA ON THE SCOPE AND METHODOLOGY OF ACCOUNTING, ORGANIZATIONAL DESIGN AND THE ROLE OF ETHICS IN ANCIENT INDIA

2004 ◽  
Vol 31 (2) ◽  
pp. 125-148 ◽  
Author(s):  
Balbir S. Sihag
Keyword(s):  
Economic Development ◽  
Economic Performance ◽  
Organizational Design ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Ethical Values ◽  
Allocation Of Resources ◽  
Economic Data ◽  
Ancient India

Kautilya, a 4th century B.C.E. economist, recognized the importance of accounting methods in economic enterprises. He realized that a proper measurement of economic performance was absolutely essential for efficient allocation of resources, which was considered an important source of economic development. He viewed philosophy and political science as separate disciplines but considered accounting an integral part of economics. He specified a very broad scope for accounting and considered explanation and prediction as its proper objectives. Kautilya developed bookkeeping rules to record and classify economic data, emphasized the critical role of independent periodic audits and proposed the establishment of two important but separate offices - the Treasurer and Comptroller-Auditor, to increase accountability, specialization, and above all to reduce the scope for conflicts of interest. He also linked the successful enforcement of rules and regulations to their clarity, consistency and completeness. Kautilya believed that such measures were necessary but not sufficient to eliminate fraudulent accounting. He also emphasized the role of ethics, considering ethical values as the glue which binds society and promotes economic development.

scholarly journals My D88-Dependent Interplay Between Myeloid and Endothelial Cells in the Initiation and Progression of Obesity-Associated Inflammatory Diseases

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. SCI-44-SCI-44
Author(s):  
Xiaoxia Li
Keyword(s):  
Insulin Resistance ◽  
Endothelial Cells ◽  
Adipose Tissue ◽  
Systemic Inflammation ◽  
Conflicts Of Interest ◽  
Ex Vivo ◽  
Inflammatory Diseases ◽  
Critical Role ◽  
Low Grade

Abstract Low-grade systemic inflammation is often associated with metabolic syndrome, which plays a critical role in the development of the obesity-associated inflammatory diseases, including insulin resistance and atherosclerosis. Here, we investigate how Toll-like receptor-MyD88 signaling in myeloid and endothelial cells coordinately participates in the initiation and progression of high fat diet-induced systemic inflammation and metabolic inflammatory diseases. MyD88 deficiency in myeloid cells inhibits macrophage recruitment to adipose tissue and their switch to an M1-like phenotype. This is accompanied by substantially reduced diet-induced systemic inflammation, insulin resistance, and atherosclerosis. MyD88 deficiency in endothelial cells results in a moderate reduction in diet-induced adipose macrophage infiltration and M1 polarization, selective insulin sensitivity in adipose tissue, and amelioration of spontaneous atherosclerosis. Both in vivo and ex vivo studies suggest that MyD88-dependent GM-CSF production from the endothelial cells might play a critical role in the initiation of obesity-associated inflammation and development of atherosclerosis by priming the monocytes in the adipose and arterial tissues to differentiate into M1-like inflammatory macrophages. Collectively, these results implicate a critical MyD88-dependent interplay between myeloid and endothelial cells in the initiation and progression of obesity-associated inflammatory diseases. Disclosures No relevant conflicts of interest to declare.

scholarly journals Impact of Platelet Count on Bleeding in the Setting of Anti-Platelet Therapy

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 18-18
Author(s):  
Robert Hugh Lee ◽  
Wolfgang Bergmeier
Keyword(s):  
Real Time ◽  
Conflicts Of Interest ◽  
Thrombotic Event ◽  
Platelet Inhibition ◽  
Intravital Imaging ◽  
Severe Thrombocytopenia ◽  
Platelet Counts ◽  
Anti Platelet Therapy ◽  
The Impact

Anti-platelet therapy (APT) is used for secondary prevention of thrombosis. The most commonly prescribed anti-platelet drugs are aspirin and P2Y12 inhibitors, including clopidogrel, prasugrel and ticagrelor. Dual anti-platelet therapy (DAPT) consisting of aspirin and a P2Y12 inhibitor is often used in the first 1-12 months after an initial thrombotic event and has a greater anti-thrombotic effect than single agents, but is also associated with a higher risk of bleeding. Due to this risk of hemorrhage, the appropriate use of DAPT in patients requiring percutaneous coronary intervention (PCI) with baseline or periprocedural thrombocytopenia remains unclear. To study the impact of thrombocytopenia on bleeding with APT, we used intravital imaging in a murine hemostasis model and adoptive platelet transfer to generate mice with specific platelet counts with or without platelet inhibition. To generate experimental mice, we used transgenic mice in which platelets express a chimeric GPIb receptor with the extracellular domain replaced with a domain of the human IL-4R (hIL-4R/GPIb-Tg). Endogenous platelets were depleted by injection of anti-hIL-4R antibody, and the recipient mice were then transfused with wild-type (WT) platelets from donor mice treated, or not, with single or dual APT (aspirin 20 mg/kg; clopidogrel 25 mg/kg) to achieve specific platelet counts ranging from 50,000 to 400,000 platelets/μL. We also compared these mice with WT mice (with normal platelet counts, ~1,200,000 platelets/μL) treated with APT. Platelet inhibition was confirmed prior to performing in vivo experiments. Hemostasis was determined by intravital imaging in our saphenous vein laser injury model, in which a 50 μm injury was induced by laser ablation. Real-time top-down epifluorescence imaging was used to determine time to initial hemostasis, rebleeding events, and platelet and fibrin accumulation. In each mouse, 3-5 injuries were induced at different sites and each injury was visualized for 10 minutes. Following real-time imaging, spinning disk confocal Z-stacks of platelet plugs were obtained for 3D reconstruction to compare platelet plug volume. In untreated WT mice, hemostasis was achieved in ~20 seconds. In WT mice treated with DAPT, initial hemostasis was often rapidly achieved but this was followed by significant rebleeding events. Paradoxically, platelet accumulation was increased in WT + DAPT mice due to extravascular accumulation of platelets which occurred during bleeding. However, in plugs that stabilized, plug volume was reduced in WT + DAPT mice. In hIL-4R/GPIb-Tg mice with reduced platelet counts, untreated platelets were able to form a stable hemostatic plug even at 50,000/μL, although time to hemostasis was slightly prolonged. However, as platelet counts decreased in mice with DAPT-treated platelets, initial hemostasis became more prolonged and many injuries never achieved initial hemostasis. These results suggest that DAPT may not be safe in the setting of severe thrombocytopenia. Disclosures No relevant conflicts of interest to declare.

Values, Other-Interest, and Ethical Behavior: The Critical Role of Moral Emotions

2020 ◽  
Author(s):  
Jason Kautz ◽  
M. Audrey Korsgaard ◽  
Sophia So Young Jeong
Keyword(s):  
Conflicts Of Interest ◽  
Critical Role ◽  
Moral Values ◽  
Moral Emotions ◽  
The Self ◽  
Collective Rationality ◽  
Ethical Challenges ◽  
Self Interest ◽  
Organizational Stakeholders ◽  
Rational Reasoning

Organizations and their agents regularly face ethical challenges as the interests of various constituents compete and conflict. The theory of other-orientation provides a useful framework for understanding how other concerns and modes of reasoning combined to produce different mindsets for approaching ethical challenges. To optimize outcomes across parties, individuals can engage in complex rational reasoning that addresses the interests of the self as well as others, a mindset referred to as collective rationality. But collective rationality is as difficult to sustain as it is cognitively taxing. Thus, individuals are apt to simplify their approach to complex conflicts of interest. One simplifying strategy is to reduce the relevant outcome set by focusing on self-interests to the neglect of other-interest. This approach, referred to as a rational self-interest mindset, is self-serving and can lead to actions that are deemed unethical. At the other extreme, individuals can abandon rational judgment in favor of choices based on heuristics, such as moral values that specify a given mode of prosocial behavior. Because this mindset, referred to as other-oriented, obviates consideration of outcome for the self and other, it can result in choices that harm the self as well as other possible organizational stakeholders. This raises the question: how does one maintain an other-interested focus while engaging in rational reasoning? The resolution of this question rests in the arousal of moral emotions. Moral emotions signal to the individual the opportunity to express, or the need to uphold, moral values. Given that moral values direct behavior that benefits others or society, they offset the tendency to focus on self-interest. At extreme levels of arousal, however, moral emotions may overwhelm cognitive resources and thus influence individuals to engage in heuristic rather than rational reasoning. The effect of moral emotions is bounded by attendant emotions, as individuals are likely to experience multiple hedonic and moral emotions in the same situation. Deontic justice predicts that the arousal of moral emotions will lead individuals to retaliate in response to injustice, regardless of whether they experience personal benefit. However, evidence suggests that individuals may instead engage in self-protecting behavior, such as withdrawal, or self-serving behaviors, such as the contagion of unjust behavior. These alternative responses may be due to strong hedonic emotions, such as fear or schadenfreude, the pleasure derived from others’ misfortunes, overpowering one’s moral emotions. Future research regarding the arousal levels of moral emotions and the complex interplay of emotions in the decision-making process may provide beneficial insight into managing the competing interests of organizational stakeholders.

Acute Lymphoblastic Leukemia (ALL) with t(8;14)(q11.2;q32): B-Lineage Disease with High Proportion of Down Syndrome. A Children’s Oncology Group (COG) Study.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1477-1477
Author(s):  
Yoav H. Messinger ◽  
Rodney Higgins ◽  
Meenakshi Devidas ◽  
Stephen P. Hunger ◽  
Andrew J. Carroll ◽  
...  
Keyword(s):  
Down Syndrome ◽  
Trisomy 21 ◽  
Lymphoblastic Leukemia ◽  
Critical Role ◽  
Prognostic Significance ◽  
Philadelphia Chromosome ◽  
Chromosome 8 ◽  
Excellent Outcome ◽  
Children With Down Syndrome ◽  
B Lineage

Abstract Recurrent chromosome translocations play a critical role in the pathogenesis of ALL and many translocations have important prognostic significance. The t(8;14)(q11.2;q32) is a recurrent translocation that fuses the chromosome 8 CEBPD (CCAAT enhancer binding protein delta) gene to the IgH (immunoglobulin heavy chain) gene, leading most likely to deregulated CEPD expression. We previously reported the clinical data of 10 such patients (Leukemia, 2000;14:238–240). Using the COG ALL cytogenetics database we expand the report to include 22 patients (13 females and 9 males) with the t(8;14)(q11.2;q32). All patients with available immunophenotyping data (n=20) had B-lineage ALL. The median age at diagnosis was 9.2 years (range: 1.6 months – 17.1 years). Median diagnostic white blood cell count (WBC) was 9,950 (range: 700–172,000). Of the 22 patients, 7 were NCI standard risk and 15 were high risk. The most common additional cytogenetic abnormality was trisomy 21. Of the ten cases with trisomy 21, seven were constitutional (Down Syndrome), thus confirming previous reports that a significant fraction of t(8;14) patients (7/22; 31.8%) have Down Syndrome, which is much higher than the 3% (80/2811) overall rate of children with Down Syndrome in a recent COG ALL trial (p&lt;0.0001). In addition to the Down Syndrome cases, one case with phenotypic Turner syndrome had a mosaic constitutional r(Y)(p11q11.2); thus, eight cases (36%) had constitutional abnormalities. Secondary abnormalities included additional X (n=5); additional 5 (n=3); Philadelphia chromosome (n=1). Two cases had a second der(14)t(8;14) and two had loss of the der(8) t(8;14), consistent with the der(14)t(8;14) as the significant abnormality. Four cases (18%) had 9p deletions compared with 11% in the overall ALL population, and two cases (9%) had abnormalities of 13q compared with 2% in the overall population. Numerically, the cases were pseudodiploid (n=12) and hyperdiploid (n=10). Children with Down Syndrome had superior estimated 5-year event-free survival of 100%, compared to non-Down patients with 50.1±17.7% (p=0.04). Overall survival was also different but did not reach statistical significance: 100% vs. 60.9±17.0% (p=0.088). In summary, the rare t(8;14)(q11.2;q32) is associated with B-lineage phenotype and occurs with greatly increased frequency in children with Down Syndrome, who have an excellent outcome with standard COG therapy as compared to non-Down Syndrome patients with this translocation.

P15 and P16 Promoters Methylation Status in APL Patients Treated by Arsenic Trioxide.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4682-4682
Author(s):  
Omid Yeganeh ◽  
Kamran Alimoghaddam ◽  
Seyed Hamidolah Ghaffari ◽  
Shahrbano Rostami ◽  
Mahdi Jalili ◽  
...  
Keyword(s):  
Arsenic Trioxide ◽  
Tumor Suppressor Genes ◽  
Promoter Region ◽  
Conflicts Of Interest ◽  
Critical Role ◽  
Methylation Status ◽  
Early Mortality ◽  
Newly Diagnosed ◽  
Sodium Bisulphite ◽  
Disease Free

Abstract Abstract 4682 Introduction Inactivation of tumor suppressor genes P15 & P16 due to Hypermethylation has a critical role in progression of cancer. We analyzed the meltylation status of promoter region of these two genes and their correlation with patients survival in APL patients treated with ATO without chemotherapy and /or ATRA. Methods Patients treated by Arsenic Trioxide without ATRA or chemotherapy and 45 newly diagnosed and 5 relapsed were enrolled in our study. Whole blood DNA was extracted and modified by sodium bisulphite reaction. Then methylation status was studied by MS-PCR and gel electrophoresis. Also for newly diagnosed patients, early mortality, disease-free and overall survival compared between methylated or non-methylated groups. Results Of 45 new cases, 31patients(69%) had methylated P15 but no patient had methylated P16. Eighty percent of relapsed patients had methylated P15 too, and no relapsed patient had methylated P16. DFS and OS correlated significantly with methylation status of P15 ( P=0.05 and 0.04) for patients who achieve to CR after treatment but it has no correlation with early motility during treatment course. Actually methyaltion of P15 improve chance of DFS and OS. Conclusion P15 but not P16 is frequently methylated in APL. Although in some studies, methyaltion of P15 negatively affected prognosis of APL patients treated by ATRA but it has no adverse effect when Arsenic Trioxide used in induction and cosolidation. Disclosures: No relevant conflicts of interest to declare.

Minimal Disseminated Disease Is An Independent Poor Prognosis Marker Among High Risk Burkitt's Lymphoma Patients .

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2944-2944
Author(s):  
Lara Mussolin ◽  
Marta Pillon ◽  
Gloria Tridello ◽  
Maria Paola Boaro ◽  
Emanuele S d'Amore ◽  
...  
Keyword(s):  
High Risk ◽  
Poor Prognosis ◽  
Conflicts Of Interest ◽  
Burkitt’S Lymphoma ◽  
Chromosome 8 ◽  
Burkitt's Lymphoma ◽  
Long Distance ◽  
Igh Locus ◽  
Minimal Disseminated Disease ◽  
Disseminated Disease

Abstract Abstract 2944 Poster Board II-920 Introduction: The chromosomal translocation t(8;14)(q24;q32) represents a specific tumor marker in Burkitt's lymphoma (BL). This chromosomal aberration involves the MYC oncogene on chromosome 8 and the immunoglobulin heavy-chain (IgH) locus on chromosome 14. We have previously demonstrated that this genetic abnormality can be used as a marker of Minimal Disseminated Disease (MDD) in BL (Mussolin et al, JCO, 2007). The aim of the study was to assess of the prevalence of MDD in BL at diagnosis in children enrolled in the AIEOP LNH-97 clinical protocol and the evaluation of its impact on prognosis. Patients and Methods: We established a simplified long-distance PCR (LD-PCR) assay which can amplify up to 15-20 Kb DNA sequence making it possible to detect the t(8;14) at the genomic level with the sensitivity of 10-4. The assay was based on 4 separate PCR reactions in which one primer complementary to the first exon of the MYC gene is used with one of four primers for the IgH locus (1 for the JH region and 1 for each of the 3 constant regions). Results: LD-PCR was applied to prospectively study 124 BL biopsies and detected a specific PCR product in 88 of them (71%). Of the 88 positive BL patients we studied both the tumor and the bone marrow (BM) at diagnosis in 76: BM was positive by LD-PCR in 25 patients (33%), whereas only 10 (13%) were positive at the standard morphological and/or immunophenotypical analyses. Most of the MDD positive patients (88%) belonged to the R4 Risk Group according to BFM definition (stage III or stage IV according to St. Jude staging classification and LDH≥1000 U/l). The 3-year progression-free survival (PFS) was 68% (SE 10%) in MDD positive R4 patients compared with 96% (SE 4%) in MDD negative R4 patients (p= 0.02), whereas there was no difference in PFS between children with morphological involvement of BM at diagnosis versus those who had negative BM (PFS=62.5% (SE 17%) vs. PFS= 87% (SE 6%), respectively, p= 0.09). By multivariate analysis (including MDD, gender, LDH, CNS involvement) MDD was predictive of higher risk of failure (Hazard Ratio: 8.4 , p= 0.04). Conclusions: We demonstrated that MDD identifies a poor prognosis subgroup among high risk Burkitt's lymphoma patients. We suggest that a more effective risk-adapted therapy, possibly including anti-CD20 monoclonal antibody, should be considered in these patients. Disclosures: No relevant conflicts of interest to declare.

Loss of the Gata1 Gene IE Exon Leads to Variant Transcript Expression and the Production of GATA1 Protein Lacking the N-Terminal Domain.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3648-3648
Author(s):  
Eri Kobayashi ◽  
Ritsuko Shimizu ◽  
Yuko Kikuchi ◽  
Satoru Takahashi ◽  
Masayuki Yamamoto
Keyword(s):  
Gene Expression ◽  
Translation Initiation ◽  
Knockout Mice ◽  
Conflicts Of Interest ◽  
Hematopoietic Cells ◽  
Full Length ◽  
Initiation Codon ◽  
Severe Thrombocytopenia ◽  
Exon 2 ◽  
Translation Initiation Codon

Abstract Abstract 3648 Poster Board III-584 GATA1 is a transcription factor essential for the differentiation of erythroid cells and megakaryocytes. Since GATA1 regulates genes related to the survival, proliferation and differentiation of hematopoietic cells, regulation of the Gata1 gene expression is critically important for the understanding of hematopoiesis. The Gata1 locus contains multiple untranslated first exons plus five common coding exons. Of these first exons, erythroid first exon (IE exon) is important for the Gata1 gene expression in the hematopoietic lineages. However, due to the embryonic lethality of this IE exon knockdown mice, less is understood about the contribution of the IE exon to adult hematopoiesis. Here, we achieved specific deletion of the IE exon in adulthood by crossing the IE-floxed mice with the interferon-inducible Mx1-Cre transgenic mice. This conditional IE-deletion mouse (ΔIE mouse) showed severe thrombocytopenia with increased premature megakaryocytes similarly to the phenotypes reported in the conditional Gata1 knockout mice in which the entire Gata1 gene was deleted in adulthood. In addition, the ΔIE mice showed severe anemia with skewed erythroid maturation, and importantly this erythroid phenotypes substantially differed from those observed in the conditional Gata1 knockout mice. Further analyses revealed that the Gata1 mRNA level in the megakaryocytic lineage was significantly downregulated. By contrast, in the erythroid lineage, Gata1 mRNA was retained at a comparable level to that in control mice utilizing two alternative first exons; one was the IEb/c, which was previously reported as a first exon rarely used in hematopoietic cells, and the other was newly identified IEd exon located within the second intron. Surprisingly, in the ΔIE mice these transcripts failed to produce full-length GATA1 protein, but instead inefficiently yielded GATA1 lacking the N-terminal 83 amino acids. This form of GATA1 is often observed in Down syndrome-associated transient myeloproliferative disorder and acute megakaryoblastic leukemia. Of note, the transcript derived from exon IEb/c preserved the first translation initiation codon in exon 2 but lost the potential to select the first translation initiation codon or failed to produce full-length GATA1. The present study demonstrates that the IE exon is instrumental to adult erythropoiesis by regulating the proper level of transcription and by selecting the correct translation start site for production of adequate full-length GATA1 protein. Disclosures: No relevant conflicts of interest to declare.

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