scholarly journals Reductive regulation of BECN1 gene in adult Egyptian patients with do novo AML

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Manal Fawzy Ghozlan ◽  
Botheina Ahmed Thabet Farweez ◽  
Nesma Ahmed Safwat ◽  
Noha Bassiouny Hassan ◽  
Walaa Ali Elsalakawy
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Peripheral Blood ◽  
De Novo ◽  
Quantitative Polymerase Chain Reaction ◽  
Control Group ◽  
Significant Difference ◽  
Egyptian Patients ◽  
Blast Cells ◽  
De Novo Aml

Abstract Background Acute myeloid leukaemia (AML) is a clonal haematopoietic disease characterized by the proliferation of immature blast cells in the bone marrow and peripheral blood. Autophagy is an inherent cellular route by which waste macromolecules are engulfed within autophagosomes prior to their fusion with cytoplasmic lysosomes for degradation. The BECN1 gene encodes the Beclin-1 protein, which regulates autophagy. Few reports have investigated BECN1 gene expression and its value in AML patients. Results This randomized case-control study included 50 newly diagnosed AML patients, in addition to 20 subjects as a control group. BECN1 gene expression was assessed using real-time quantitative polymerase chain reaction (qRT-PCR). The median level of BECN1 gene expression in AML patients was 0.41 (IQR 0.29–1.03) in comparison to 1.12 (IQR 0.93–1.26) in the control group (P = 0.000). Seventy-two percent of AML patients showed reduced BECN1 gene expression, which was highly significantly associated with intermediate and adverse cytogenetic risk. Reduced BECN1 gene expression was associated with older age, higher total leukocyte counts, the presence of peripheral blood blast cells, a higher percentage of bone marrow blast cells, and higher expression of CD34 and CD117. FLT3-ITD mutation was detected in 14 patients (38.9%), all of whom showed reduced BECN1 gene expression (P = 0.006). BECN1 gene expression was also reduced in non-responder AML patients, with a highly statistically significant difference (P = 0.002). Conclusion A reduction in BECN1 gene expression might indicate a poor prognosis in adult Egyptian patients with de novo AML. Decreased BECN1 gene expression is associated with a higher risk of resistance to treatment. Targeting autophagy pathways may help in the treatment of AML patients.

Pro- and Antiapoptotic Proteins Expression on Bone Marrow and Peripheral Blood CD34+Cells in Acute Leukemia (AL) Patients

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 4996-4996
Author(s):  
Elena E. Khodunova ◽  
Elena N Parovichnikova ◽  
Irina V. Galtzeva ◽  
Sergey M. Kulikov ◽  
Valeri G Savchenko
Keyword(s):  
Bone Marrow ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Expression Level ◽  
Leukemia Cells ◽  
Control Group ◽  
Healthy Donors ◽  
Cd34 Cells ◽  
Significant Difference ◽  
Blast Cells

Abstract Abstract 4996 It was shown that drug resistance, poor-risk cytogenetics and poor prognosis in AL is associated with high level of Bcl-2 expression and low Bax/Bcl-2 ratio (<0,3). Fas-antigen (CD95) as a protein triggering the extrinsic apoptotic pathway is differently expressed on hematopoietic precursors. More immature CD34+/CD38- AML blast cells have lower expression of Fas/Fas-L and lower Fas-induced apoptosis than CD34+/CD38+cells. CD34+/CD38− leukemia precursors also have a reduced sensitivity to daunorubicin in vitro and increased expression of multidrug resistance genes (mrp/lrp). CD34+ leukemia cells have not yet been properly characterized regarding the expression of angiotensin converting enzyme (ACE) which regulatory influence on hematopoiesis is now beeing extensively investigated. ACE expression on blast cells is high, but it's still unknown how CD34+ACE+ leukemia cells behave after chemotherapy. Recent publications indicate that CD34+ACE+ hematopoietic precursors transplanted into NOD/SCID mice contribute 10-fold higher numbers of multilineage blood cells than their CD34+ACE- counterparts. We have studied the dynamics of Bcl-2, Bax, CD95 and ACE expression on CD34+ cells in peripheral blood (PB) and bone marrow (BM) in AL pts during treatment. PB and BM samples were collected before and on +36 day after chemotherapy. The antigens were detected by flow cytometry using monoclonal antibodies. We calculated 10 000 cells in each sample. 19 pts were included in the study: 10 - AML and 9 - ALL. The control group comprised 8 healthy donors. At time of diagnosis there were 40±5,7% of CD34+ cells in BM and 26±4,9% - in PB. There was no significant difference between AML and ALL. CD34+ cells in BM and PB of healthy donors constituted 1,6% and 0,27%, respectively. After induction therapy (+36 day) CD34+ cells decreased in BM to 6,1%±3,3 (p=0,0001), in PB to 3,7%± 2,7 (p=0,0008) in all pts. The data on antigens expression on CD34+ cells of BM and PB are presented in table 1 CD34+/Bcl-2+ CD34+/Bax+ CD34+/CD95+ CD34+/ACE+ BM PB BM PB BM PB BM PB AML pts n=10 0 day 38±11,6* 41±14 24,4±7,9 29,2±7,6* 16,4±8,5 23,2±7,8 21,7±9,5 20,8±8,7* 36 day 13,5±3,4** 23,7±5** 46,2±11,5 50,3±11 19,9±5,5 36,4±10 34±6,6 35±9,2** ALL pts n=9 0 day 36±11 33,7±12 46,2±9,4 37,4±3,7* 3,4±1,1* 7,1±2,5* 41±10,9 33,2±9,7* 36 day 18,4±5,8 26±8,9 38±11,8 40,5±10 26,2±9,1** 40,9±9,2** 34±10 62,8±10** Donors n=8 11,7±1,6 26,1±5,9 22,8±4 67,8±6,7 13,4±3,2 47,7±11,6 28±5,3 68,2±10,2 * − p<0.05 compare with donors ** − p<0.05 compare with day 0 CD34/Bcl-2 expression in BM in AML pts is significantly higher (p=0,04) at the diagnosis comparing with donors. CD34/Bcl-2 expression in PB in AML pts and in BM and PB in ALL pts is higher too, but not significantly. This expression level decreased substantially in BM and PB in AML pts on +36 day comparing with day 0 (p<0,05). We did not found significant changes in ALL pts. CD34/Bax expression in PB is significantly lower (p=0,003) both in AML and ALL pts in comparison with donors. In AML, not in ALL, chemotherapy caused augmentation of Bax expression in CD34+ BM and PB cells on +36 day. BM and PB CD34+ cells in donors had different expression characteristics of Bcl-2 and Bax, demonstrating much higher level of pro- and antiapoptotic markers in PB cells. On the contrast CD34+ leukemia cells in BM and PB had similar characteristics regarding CD34/Bcl-2 and CD34/Bax expression. This fact demonstrates the heterogeneity of donor CD34+cells in BM and PB and points that leukemia CD34+cells in BM and PB are rather similar. CD95 expression on CD34+ BM and PB before treatment is significantly lower (p=0,01, p=0,008) in ALL pts in comparison with donors, and this expression level increased after chemotherapy (p<0,05). CD34/CD95 expression in AML pts is similar with donors, and we didn't find changes after treatment. CD34/ACE coexpression in BM cells of leukemia pts and donors did not differ much at any time of evaluation. But CD34/ACE expression in PB cells of AML and ALL pts was much lower (p<0,05) than in donors and substantially increased on the day 36. So, our data demonstrate that Bcl-2, Bax, CD95 and ACE expression on CD34+ cells in AL pts and donors significantly differs. The chemotherapy provokes critical changes in CD34/CD95 expression in BM and PB in ALL pts, CD34/Bcl-2 expression in AML pts and ÑÂ34/ACE expression in PB in all AL pts. Disclosures: No relevant conflicts of interest to declare.

Expression of BECN1 gene in Acute Myeloid Leukemia: Association with Hematological and Clinical Parameters

QJM ◽  
2021 ◽  
Vol 114 (Supplement_1) ◽  
Author(s):  
Mohamed Moustafa Ahmed ◽  
Manal Fawzy Ghozlan ◽  
Walaa Ali Mohamed ◽  
Nesma Ahmed Safwat ◽  
Noha Bassiouny Hassan
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Peripheral Blood ◽  
Myeloid Leukemia ◽  
Control Group ◽  
Fish Analysis ◽  
Newly Diagnosed ◽  
Significant Difference ◽  
Acute Myeloid

Abstract Background In acute myeloid leukemia (AML), there is copy number loss in autophagic genes such as BECN1. Accordingly, decreased autophagy and the development of AML are related. BECN1 is a critical mediator that influences the onset and progress of autophagy. Objective To investigate the expression status of BECN1 gene in newly diagnosed adult AML patients and its association with various hematological parameters and clinical outcomes. Methods Case control study to study BECN1 gene expression variability between 50 newly diagnosed adult AML patients and 20 healthy age and sex matched controls, with follow up of the patients to detect its effect on induction therapy. All AML patients underwent full history taking, through clinical examination, laboratory investigations such as complete blood count (CBC) with examination of peripheral blood and bone marrow Leishman stained films, immunophenotyping, cytogenetic analysis (karyotyping/FISH analysis) and BECN1 gene expression analysis using real-time quantitative polymerase chain reaction (qRT-PCR). Results In our study, a highly significant difference was found as regards reduced expression of BECN1 gene in patients group compared to control group. We also found reduced BECN1 gene expression in both intermediate and adverse risk groups compared to favorable risk group. Reduced expression of BECN1 gene was associated with increasing age and total leukocytic count (TLC), peripheral blood (PB) and bone marrow (BM) blasts, the presence of FLT3-ITD mutation, CD34 and CD117 and in non-responders group. No statistically significant difference was found as regards haemoglobin (Hb) level, platelet (PLT) count and FAB subtypes. Conclusion Autophagy plays an important role in the pathogenesis of AML. Furthermore; the reductive regulation of the BECN1 gene may carry a poor prognosis and is associated with many well established bad prognostic factors especially FLT3-ITD mutation. Targeting autophagy pathways especially its major regulator (BECN1 gene) may become an effective and promising new line of therapy for AML patients.

scholarly journals Clinical observations of bone marrow transfusion for promoting bone marrow reconstruction after chemotherapy for AIDS-related lymphoma

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Yixuan Liu ◽  
Suhong Xie ◽  
Lei Li ◽  
Yanhui Si ◽  
Weiwei Zhang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Immune System ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Autologous Bone Marrow ◽  
Control Group ◽  
Peripheral Vein ◽  
Significant Difference ◽  
Autologous Bone ◽  
Aids Related Lymphoma

Abstract Background This study investigates the effect of autologous bone marrow transfusion (BMT) on the reconstruction of both bone marrow and the immune system in patients with AIDS-related lymphoma (ARL). Methods A total of 32 patients with ARL participated in this study. Among them, 16 participants were treated with conventional surgery and chemotherapy (control group) and the remaining 16 patients were treated with chemotherapy followed by autologous bone marrow transfusion via a mesenteric vein (8 patients, ABM-MVI group) or a peripheral vein (8 patients, ABM-PI group). Subsequently, peripheral blood and lymphocyte data subsets were detected and documented in all patients. Results Before chemotherapy, no significant difference in indicators was observed between three groups of ARL patients. Unexpectedly, 2 weeks after the end of 6 courses of chemotherapy, the ABM-MVI group, and the ABM-PI group yielded an increased level of CD8+T lymphocytes, white blood cells (WBC), and platelet (PLT) in peripheral blood in comparison to the control group. Notably, the number of CD4+T lymphocytes in the ABM-PI group was significantly higher than that in the other two groups. Additionally, no significant difference in haemoglobin levels was observed before and after chemotherapy in both the ABM-MVI and ABM-PI groups, while haemoglobin levels in the control group decreased significantly following chemotherapy. Conclusions Autologous bone marrow transfusion after chemotherapy can promote the reconstruction of both bone marrow and the immune system. There was no significant difference in bone marrow recovery and reconstruction between the mesenteric vein transfusion group and the peripheral vein transfusion group.

Prolonged Survival of a Patient with De Novo Acute Myeloid Leukemia (AML) and Trisomy 13 with Intensive Chemotherapy and Autologous Peripheral Blood Transplantation (PBSCT): A Case Report.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4519-4519
Author(s):  
Nitin D. Joshi ◽  
Alpesh Amin ◽  
Rajneesh Nath
Keyword(s):  
Bone Marrow ◽  
Complete Remission ◽  
Bone Marrow Biopsy ◽  
Induction Chemotherapy ◽  
Peripheral Blood ◽  
De Novo ◽  
Trisomy 13 ◽  
Autologous Pbsct ◽  
De Novo Aml ◽  
First Complete Remission

Abstract Trisomies are uncommon cytogenetic abnormalities in patient with de novo AML. Survival of patients with trisomy 13 ranges from 0.5 to 14.7 months. We present the treatment outcome of a 71-year-old man with de novo AML and trisomy 13 who had PBSCT in first complete remission. A 71-year Puerto Rican male was diagnosed with AML in April 2003. His CBC showed WBC count 177 K/mm3, hemoglobin 10.3 gm/dl, platelets 43 K/mm3 and blast cells 75%. Flow cytometry revealed that the leukemic blasts were CD33, CD13, CD11c and CD56 positive but negative for CD34. Cytogenetics failed to yield any metaphases. Peripheral blood FISH studies revealed trisomy 13 positivity in 300 of 325 cells analyzed. Patient received induction chemotherapy with high dose Ara-c (HiDAC) 3g/m2 QD x 5 doses and mitoxantrone 80mg/m2 on day # 2. Bone marrow done day 28 post induction chemotherapy revealed residual leukemic blasts. Cytogenetics showed that one out twenty metaphases had trisomy 13 along with translocation t (9:18) (q34; q10). 11.9% of cells had trisomy 13 by FISH analysis. The patient then received a second cycle of chemotherapy with HiDAC at 2 g/m2 Q12 x 12 doses. Bone marrow biopsy on day 35 following reinduction chemotherapy revealed normocellular-regenerating marrow in remission and FISH was negative for trisomy 13. On the third cycle of chemotherapy, patient received Etoposide 11 mg/kg. Neupogen was started on day #3 and 10.3 x 106 CD34 positive cells/kg were collected. The patient then underwent autologous PBSCT using Melphalan 160 mg/m2 as the preparative regimen. On Day +87 and Day +182 post transplant, bone marrow biopsy showed complete remission with FISH negative for trisomy 13. The patient is still alive 27 months after initial treatment and 22 months post PBSCT. Autologous PBSCT in first complete remission for AML with trisomy 13 may provide a superior survival than chemotherapy alone.

Study of Sequential Treatment of Newly Diagnosed De Novo AML Patients with DA and CAG Regimens as Remission Induction Therapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4630-4630
Author(s):  
Hui ping Sun ◽  
Wei Hong Liu ◽  
Jun min Li ◽  
Qiu sheng Chen ◽  
Yu Chen
Keyword(s):  
Bone Marrow ◽  
Induction Therapy ◽  
Peripheral Blood ◽  
De Novo ◽  
Classification Criteria ◽  
Sequential Treatment ◽  
Remission Induction ◽  
Newly Diagnosed ◽  
Fab Classification ◽  
De Novo Aml

Abstract Objectives To evaluate the efficacy and safety of sequential treatment of newly diagnosed de novo AML patients with DA and CAG regimens as induction therapy. Methods Those who were newly diagnosed as de novo AML (FAB classification criteria) were enrolled and DA regimen chemotherapy were administered. Bone marrow aspirates were performed and BM smears were examined at 48 hours since the end of chemotherapy. If severe hypocellularities were not achieved, the percentage of blasts in BM was between 20%–60% and peripheral WBC was in the range of (0.5–10) x109/L, the patients would receive CAG regimen therapy since 72 hours. Patients’ general status and the important parameters, such as peripheral blood count, liver function, renal function, thrombosis and hemostasis parameters were monitored throughout the course of the treatment and thereafter. When the clinical symptoms were relieved and peripheral blood counts returned to normal, or it was the end of the second or third week since the end of the CAG regimen, Bone marrow were examined again to evaluate the efficacy of the sequential therapy. Results 14 patients consisted of 9 male and 5 female patients were enrolled. Out of them, 2 were M1, 5 M2, 4 M4 and 3 M5 according to FAB classification criteria. Median of blasts in BM were 38.5%(20%–60%) before CAG regimen. Of the 14 patients, 10 reached CR, 2 PR and 2 NR. CR rate was 71.4% (10/14) and total response rate was 85.7%(12/14). Time to achieve CR was on 15th(14th–29th)day medianly since the end of the treatment. During the CAG therapy courses, the nadir of peripheral blood cell counts and the time when it occurred were as follows: WBC 1.0(0.2–3.5)(x109/L),10(1–23)(d); Hb 57.5(44–69) (g/L), 10(1–27)(d)and PLT 11.5(10–65)(x109/L), 12(3–23)(d), respectively. Neutropenia (WBC<1.0x109/L) and thrombocytopenia (PLT<20.0x109/L) were lasted for 0(0–24) and 11(0–21)days, respectively. Median units of transfusions of platelets and red blood cells required by each patient were 3(0–10)(u) and 4(0–12)(u), respectively. The most commonly observed side effect of the regimen was bone marrow proliferation inhibition. Infections, usually respiratoy tract infections, were the second. However, sepsis was rare, which appeared in 1 out of 14 patients. Conclusions DA and CAG regimens sequential treatment as remission induction chemotherapy in patients with newly diagnose de novo AML was highly effective and well tolerated. It would be beneficial for those who might not be sensitive enough to DA regimen chemotherapy only.

Dnmt3b Is Dispensable for Hematopoietic Stem Cell Differentiation, but Acts Synergistically with Dnmt3a to Control the Balance Between Self-Renewal and Differentiation

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 848-848
Author(s):  
Allison Mayle ◽  
Grant Anthony Challen ◽  
Deqiang Sun ◽  
Mira Jeong ◽  
Min Luo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Bone Marrow ◽  
Peripheral Blood ◽  
De Novo ◽  
Annexin V ◽  
Dna Methyltransferases ◽  
Self Renewal ◽  
Minimal Impact ◽  
Hsc Transplantation

Abstract Abstract 848 DNA methylation is an epigenetic modification in vertebrate genomes critical for regulation of gene expression. DNA methylation is catalyzed by a family of DNA methyltransferase enzymes, Dnmt1, Dnmt3a, and Dnmt3b. Dnmt1 is primarily a maintenance methyltransferase, targeting hemimethylated DNA to reestablish methylation marks after DNA replication. Dnmt3a and Dnmt3b are de novo methyltransferases that are essential for normal embryonic development. In humans, somatic mutations in DNTM3A have been identified in ∼20% of human acute myeloid leukemia (AML) and ∼10% of myelodysplastic syndrome (MDS) patients, but the mechanisms through which these mutations contribute to pathogenesis is not well understood. Congenital mutations in DNMT3B can cause ICF (immunodeficiency, centromeric instability, and facial anomalies) syndrome. These patients exhibit chromosomal instability due to heterochromatin decondensation and demethylation of satellite DNA. Our group has recently reported that Dnmt3a is essential for HSC differentiation (Challen Nature Genetics, 2011). Conditional knockout of Dnmt3a (Dnmt3a-KO) resulted in HSCs that could not sustain peripheral blood generation after serial transplantation, but phenotypically defined HSCs accumulated in the bone marrow. Dnmt3b is also highly expressed in HSCs, but its contribution to gene regulation in hematopoiesis is unclear. Here, we examine the role of Dnmt3b, alone and in combination with Dnmt3a KO, in the regulation of hematopoiesis. We performed conditional ablation of Dnmt3b, as well as Dnmt3a and Dnmt3b simultaneously using the Mx1-cre system. Unlike the Dnmt3a-KO HSCs, loss of Dnmt3b had a minimal impact on blood production. Even after several rounds of transplantation, 3b-KO HSCs performed similarly to WT controls. However, the Dnmt3ab-dKO (double knock-out) peripheral blood contribution was quickly and severely diminished, accompanied by a dramatic accumulation of Dnmt3ab-dKO HSCs in the bone marrow (Figure 1). The dKO phenotype paralleled that of the 3a-KO HSC, but was more extreme. To examine the impact of loss of Dnmt3a and -3b on DNA methylation in HSCs, we performed Whole Genome Bisulfite Sequencing (WGBS) on Dnmt3a-KO, Dnmt3ab- dKO and control HSCs. As we previously found with more limited DNA methylation analysis, loss of Dnmt3a led to both increases and decreases of DNA methylation at distinct genomic regions (Challen, Nature Genetics, 2011). However, loss of both Dnmt3a and -3b primarily resulted in loss of DNA methylation that was much more extensive than that seen in the 3a-KO. In addition, RNAseq of the mutant HSCs revealed increased expression of repetitive elements, inappropriate splicing, and truncation of 3ÕUTRs. To gain insight into the accumulation of Dnmt3ab-dKO HSCs in the bone marrow, we performed a time course analysis of the proliferation and apoptosis status of the HSCs. Every four weeks after transplantation of HSCs, we sacrificed a cohort of 3 control and 3 dKO mice, counted donor derived HSCs in the bone marrow, and analyzed their Ki67 and Annexin V expression. Up to 12 weeks post-transplant, no significant differences are seen in the expression of Ki67 or Annexin V. These data show that while Dnmt3b alone has minimal impact on DNA methylation in HSCs, Dnmt3a and -3b act synergistically to effect gene expression changes that permit HSC differentiation. In the absence of both of these de novo DNA methyltransferases, there is an immediate and extreme shift toward self-renewal of dKO HSCs. The Ki67 and Annexin V expression patterns suggest that a lack of de novo DNA methylation does not affect the proliferation or apoptosis of HSCs, but instead that the accumulation of HSCs and lack of peripheral blood contribution is primarily due to an imbalance between self-renewal and differentiation. By understanding the mechanisms through which Dnmt3a and -3b exert these effects, we should identify genes that are critical for normal hematopoietic differentiation. These genes may serve as targets for therapeutic intervention in malignancies caused by defective DNA methyltransferases. Figure 1: HSC composition of the bone marrow after secondary transplantation of control (left) and double Dnmt3a/3b KO (right) HSCs. After control HSC transplantation, HSCs comprise ∼0.01% of whole bone marrow. After transplantation of dKO HSCs, phenotypically-defined HSCs (KLS CD34–Flk2–) comprise ∼0.48% of bone marrow. Figure 1:. HSC composition of the bone marrow after secondary transplantation of control (left) and double Dnmt3a/3b KO (right) HSCs. After control HSC transplantation, HSCs comprise ∼0.01% of whole bone marrow. After transplantation of dKO HSCs, phenotypically-defined HSCs (KLS CD34–Flk2–) comprise ∼0.48% of bone marrow. Disclosures: No relevant conflicts of interest to declare.

Differential Expression Of TREM-1 In Myelogenous Leukemia Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4951-4951 ◽  
Author(s):  
Huiyu Li ◽  
Wenying Li ◽  
Xiaoling Yi ◽  
Shiang Huang ◽  
Wei Liu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Complete Remission ◽  
Patient Group ◽  
Peripheral Blood ◽  
Myelogenous Leukemia ◽  
Leukemia Cells ◽  
Control Group ◽  
Healthy Controls ◽  
Significant Difference ◽  
Patient Groups

Abstract Objectives Triggering receptor expressed on myeloid cells (TREM) -1 is a receptor as a member of the immunoglobulin superfamily expressed on the cell-surface of neutrophils, monocytes and macrophages. This receptor amplifies the inflammatory response, activating the signaling pathway. TREM-1 expression is associated with mature myeloid cell development. TREM-1 is shed from the membrane of activated macrophages without the transmembrane and intracellular domains, and can be found as soluble TREM (sTREM)-1. Soluble TREM-1 is thought to negatively regulate TREM receptor signaling. Some studies currently reported that TREM-1 regulates the malignant behavior of cancer cells in lung cancer and HCC. However, no related studies about the role of TREM-1 in leukemia have been carried out. The aims of this study was investigated the TREM-1 expression in myelogenous leukemia cells. Methods Thirty-five patients with AML, twenty-five patients with CML and a control group of eleven healthy people were subjected to the study. TREM-1 expressions on the surfaces of leukemia cells were measured by flow cytometry. Plasma sTREM-1 levels were measured by ELISA. Results In this study, our results provide the first evidence that TREM-1 was differentially expressed in myelogenous leukemia cells. The TREM-1 mean ratio of median fluorescence intensity (mean ratio of MFI) was 3.13±0.88 and 2.52±0.40 in CML and AML patients, respectively. The TREM-1 mean ratio of MFI was 3.03±1.40 in myelogenous leukemia cell lines (K562, HL60, THP-1). The TREM-1 mean ratio of MFI was 5.37±0.88 in healthy controls. Compared to healthy controls, myelogenous leukemia cells had decreased TREM-1 expressions (P<0.001). The TREM-1 mean ratio of MFI was 4.89±0.60 in patients who are in complete remission after Novartis's Gleevec therapy. Compared with CML patient groups, patients who are in complete remission after Gleevec therapy had rising TREM-1 expressions (P<0.01). TREM-1 expressions of patients who are in complete remission after Gleevec therapy were slightly lower than the healthy controls, but this did not reach significance. No significant difference in TREM-1 expressions was seen between AML and CML patient groups, male and female patient groups, and cells derived from peripheral blood and bone marrow of the same leukemia patients (p>0.1). In addition, the plasma sTREM-1 levels were measured by ELISA. sTREM-1 levels was 48.54±57.63pg/mL for AML group and 43.72±23.93pg/mL for CML group. Results indicated that plasma sTREM-1 levels significantly higher in AML and CML patients than that in healthy controls (P<0.01). However, there was no significant difference in plasma sTREM-1 levels observed in AML patient group compared with CML patient group, male patients group compared with female patients group, and plasma from peripheral blood compared with plasma from bone marrow of the same leukemia patients (p>0.1). An ongoing project focuses on the relationship between the function of TREM-1 and occurrence, progression and prognosis of myelogenous leukemia, advances will be reported in time. Conclusion TREM-1 expression on leukemia cells was significantly lower in patients with AML and CML than those in healthy controls and patients in complete remission had increased TREM-1 expression. Patients with AML and CML had increased plasma soluble TREM-1. The TREM-1 expression on leukemia cells had an inverse correlation with plasma sTREM-1 level in AML and CML patients. Disclosures: No relevant conflicts of interest to declare.

Utility of Telomere in Diagnosis of Bone Marrow Failure Syndromes

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4793-4793
Author(s):  
Hasan Ahmed Abdel-ghaffar ◽  
Hosam Zaghloul ◽  
Ahmed El-Waseef ◽  
Mohamed El-Naggar ◽  
Mohamed Mabed ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Telomere Length ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Quantitative Polymerase Chain Reaction ◽  
Control Group ◽  
Hematopoietic Stem ◽  
Relative Telomere Length ◽  
Bone Marrow Failure Syndromes ◽  
Significant Difference

Abstract Background and aim of the work: Bone marrow failure syndromes (BMFS) includes inherited and acquired conditions. Inherited bone marrow failure includes a number of syndromes; with Fanconi anemia (FA) being the most common one of them. Telomeres are eroded with cell division, but in hematopoietic stem cell, maintenance of their length is mediated by telomerase. Short telomeres can result in instability of cell function where diseases occur. Bone Marrow Failure might be developed due to low telomerase activity or short telomeres. Our study is aiming to evaluate the utility of Real Time Quantitative-Polymerase Chain Reaction (RT-qPCR) in measuring the relative telomere length and its significance in diagnosis and prognosis of patients with BMFS. Materials and methods: The study includes 3 groups: A group of congenital BMF (29 patients), a group of acquired BMF (10 patients) and a third control group (15 cases). The relative telomere length is evaluated for them using RT-qPCR. Results: We have found that there is a significant difference in relative telomere length between congenital group and controls (p=0.001), also a significant difference between acquired group and controls (p= 0.029). However, there is no significant difference between congenital and acquired groups (p= 0.479). There is no significant correlation between the telomere length and the overall survival or prognosis of the patients of BMFS. Conclusion: We conclude that the telomere length is significantly altered in patients with BMFS whether being congenital or acquired compared to the control group. Disclosures No relevant conflicts of interest to declare.

Influence of Maternal Folic Acid Supplementation on Hematopoietic Factors in Offspring.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4276-4276
Author(s):  
Kimberly J. Johnson ◽  
David A. Largaespada ◽  
Tucker W. LeBien ◽  
Raha Allaei ◽  
Julie A. Ross
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Folic Acid ◽  
Lymphoblastic Leukemia ◽  
Surface Marker ◽  
Control Group ◽  
Offspring Survival ◽  
Childhood All ◽  
Significant Difference ◽  
B Lineage

Abstract Observational studies of childhood acute lymphoblastic leukemia (ALL) have generally supported a protective role for maternal intake of foods or vitamins rich in folic acid during pregnancy. Using C57BL/6J mice, we evaluated the effect of dietary folic acid intake in the perigestational period on surface marker and gene expression in B-lineage cells as a potential model to further elucidate the role of maternal diet in the etiology of childhood ALL. Thirty-six female mice were randomized to one of three folic-acid varied diets (low=0.3, control=2.0, high=8.0 mg/kg) one month prior to mating and maintained on the diet until pup weaning at three weeks. Reproductive characteristics and offspring red blood cell (RBC) folate were recorded, and bone marrow lymphocytes were isolated for surface marker characterization by flow cytometry. For gene expression analyses, CD19 positive B-lineage bone marrow cells were purified from three pups per dietary group using anti-CD19 microbeads. Gene expression levels were interrogated using the Agilent mouse whole genome microarray. No statistically significant differences were found among dietary groups with respect to mating success, litter size, estimated 21 day offspring weights, or the percentage of male offspring. However, there were significant differences among the 3 diets in offspring survival to the end of the experiment (39%, 84%, and 30% for the 0.3, 2.0, and 8.0 mg/kg groups, respectively). Pup RBC folate values were positively correlated with dietary folic acid dose with mean values of 428, 615, and 724 ng/ml, respectively. Mice were euthanized at 3 weeks. There was no significant difference in the expression of CD19, IgM, or kappa light chains on bone marrow lymphocytes in the offspring among dietary groups. ANOVA analysis, using Genedata Expressionist Pro software, showed differential expression of 267 genes using a Benjamini and Hochburg false discovery rate cutoff of 10%. Three main patterns of gene expression between pups of different dietary groups were identified through unsupervised hierarchical clustering analysis; 152 and 86 genes were up- and down-regulated, respectively, in both the high and the low folic acid groups relative to the control group. Twenty-seven genes showed increasing expression with increasing dietary folic acid dose. In conclusion, dietary folic acid given to dams prior to conception through weaning had a significant effect on offspring survival and gene expression. Additional analyses are underway to determine the relevance of genes found to be differentially expressed with respect to risk of B-cell lineage leukemias followed by validation of relevant gene candidates through quantitative RT-PCR. These data will be used to help inform future etiologic studies of childhood ALL.

Deep Digital Sequencing Identifies an AML Subclone with Enhanced in Vitro and in Vivo Growth Properties Associated with Disease Relapse

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 407-407
Author(s):  
Jeffery M. Klco ◽  
David H Spencer ◽  
Chris Miller ◽  
Tamara Lamprecht ◽  
Robert S. Fulton ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Peripheral Blood ◽  
De Novo ◽  
Hematopoietic Cells ◽  
Bone Marrow Sample ◽  
Clonal Architecture ◽  
Growth Properties ◽  
De Novo Aml

Abstract Abstract 407 Acute myeloid leukemia (AML) is a biologically heterogenous malignancy of hematopoietic cells. All AML samples are comprised of a founding clone and usually one or more subclones that are derived from the founding clone; subclones can gain or lose mutations as they evolve from the founding clone, and often become dominant at relapse1. The clonal architecture of an AML sample can be identified using single nucleotide variants (SNVs) that cluster according to discrete variant allele frequencies (VAFs). To accurately identify clusters with common VAFs, deep digital sequencing must be performed using all of the SNVs present in each genome (hundreds of events). In this report, we studied the subclonal architecture of AML samples from UPN 452198, collected from a 55-year old woman with normal karyotype acute monocytic leukemia (FAB M5) with a high peripheral WBC count (72,700/mm3) at presentation. This bone marrow sample contained a founding clone and 3 subclones at presentation. The SNVs of the founding clone had a mean VAF of 46.4% (i.e. heterozygous mutations found in 92.8% of the cells in the bone marrow sample), including DNMT3A R882H and NPM1 W288 frameshift mutations. The mean VAFs of Subclones 1, 2 and 3 were 31.2%, 12.0% and 2.4%, respectively, and they contained all of the variants in the founding clone, along with additional variants, most notably FLT3 D835H and IDH1 R132H mutations in Subclone 1. The tumor at relapse consisted entirely of Subclone 3, which also contained 42 relapse-specific variants. We designed an oligonucleotide capture reagent to track all 118 de novo and relapse-specific variants, and obtained deep read counts (mean coverage per site: 412 reads/SNV) on the de novo AML sample under different experimental and biological conditions, as follows: 1) We showed that peripheral blood and bone marrow leukemia samples obtained at the same time had nearly identical clonal architectures. We verified this correlation using 4 additional AML samples, suggesting that clonal architecture is preserved in the peripheral blood for many AML samples. 2) We flow-sorted the leukemic peripheral blood sample into blasts, monocytes, and lymphocytes based on side-scatter characteristics and expression of CD45 and CD33. The founding clone and all three subclones were detected in the monocyte population, which was the predominant leukemic cell population in the peripheral blood. By flow cytometry, blasts comprised only 3.3% of the cells, but were strongly enriched for variants in Subclone 3 (mean VAF in sorted blasts 33.9% versus 3.0% in unsorted peripheral blood, p<0.001). Purified lymphocytes, in contrast, contained no leukemia-specific variants, implying that the founding clone for this sample did not contribute to lymphopoiesis. 3) We tested the growth properties of subclones in the de novo sample in vitro and in vivo. We injected 1 million cells from the de novo AML sample into 3 immunodeficient NSG mice, and harvested human AML blasts (co-expression of human CD45 and CD33) 14 weeks later. Although two mice engrafted with the founding clone and Subclone 1 (which comprised the vast majority of the cells in the sample), the 3rd mouse had a tumor composed entirely of the relapse-specific Subclone 3 (which accounted for only 2.4% of the variants in the de novo sample), suggesting that this subclone had a significant growth or engraftment advantage in vivo. In support of this observation, de novo AML cells were strongly enriched for Subclone 3 when grown in the presence of hematopoietic cytokines (SCF, IL3, IL6, TPO and FLT3L) for 7 days on HS27 stroma (VAF at day 7–19.7%; p<0.001) or MS5 stroma (VAF at day 7–22.8%; p<0.001), implying that this clone also had a strong in vitro growth advantage. In summary, a small subclone of AML cells at presentation — that was known to eventually contribute to relapse — had unique growth properties that were revealed using deep digital sequencing of all variants. This approach has allowed us to dissect the evolutionary history of AML clones, and to define their relationship to other hematopoietic cells in a given sample. Similar studies on additional AML samples should allow us to define the mutational profiles of subclones that are destined to contribute to relapse. This data will be essential for improving therapeutic approaches for AML patients. Disclosures: Ley: Washington University: Patents & Royalties.

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