scholarly journals Differentiation of Acute Myeloid Leukemia from B- and T-Lineage Acute Lymphoid Leukemias by Real-Time Quantitative Reverse Transcription-PCR of Lineage Marker mRNAs

2004 ◽  
Vol 50 (7) ◽  
pp. 1165-1173 ◽  
Author(s):  
Pascale Saussoy ◽  
Jean-Luc Vaerman ◽  
Nicole Straetmans ◽  
Véronique Deneys ◽  
Guy Cornu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Real Time ◽  
Reverse Transcription ◽  
Cell Sorting ◽  
Acute Leukemias ◽  
Ct Value ◽  
Blast Cells ◽  
Lineage Markers ◽  
B Lineage ◽  
Acute Myeloid

Abstract Background: Flow cytometry of lineage markers is useful in the classification of leukemias. Our aim was to assess whether the study of lineage genes at the RNA level would enable differentiation of acute myeloid leukemias (AMLs) from B-and T-lineage acute lymphoid leukemias (ALLs). Methods: We measured mRNA of four lineage markers [CD19, CD79a, CD3e, and myeloperoxidase (MPO)] by reverse transcription followed by real-time quantitative (RTQ)-PCR. We investigated 72 acute leukemias (40 AMLs with 23–93% blast cells plus 27 B-lineage ALLs and 5 T-lineage ALLs) defined by morphologic criteria at diagnosis. RTQ-PCR analysis was performed on bone marrow without cell sorting. The expression of each gene was calculated as the difference in the threshold cycle [ΔCT; CT value of target gene minus CT value of housekeeping gene (Abelson)]. Results: Three patterns of expression were detected. In the first, CD19, CD79a, and MPO mRNAs were less abundant than CD3e. In the second pattern, MPO mRNA was more abundant than the other three mRNAs. In the third, CD19 or CD79a was more highly expressed than CD3e and MPO. The three patterns corresponded to T-ALL, AML, and B-ALL, respectively. The use of cutoffs to establish qualitatively the pattern of coexpression of the four lineage markers provided the same information as the comparison among the four ΔCT values. Prospective use of the scoring system correctly classified each of 13 additional cases (8 AML, 4 B-lineage ALL, and 1 T-lineage ALL). Conclusion: Study of lineage markers at diagnosis by RTQ-PCR allows differentiation of AML from B-ALL or T-ALL without cell sorting, even when the bone marrow contains few blast cells.

scholarly journals Identification of novel B-lineage cells in human fetal bone marrow that coexpress CD7 [see comments]

Blood ◽  
1991 ◽  
Vol 77 (1) ◽  
pp. 64-68 ◽  
Author(s):  
ER Grumayer ◽  
F Griesinger ◽  
DS Hummell ◽  
RD Brunning ◽  
JH Kersey
Keyword(s):  
Bone Marrow ◽  
Cell Sorting ◽  
B Cell Development ◽  
Multiparameter Flow Cytometry ◽  
Fetal Bone ◽  
Lymphoid Development ◽  
Alternate Pathway ◽  
Lineage Markers ◽  
B Lineage

Abstract In the present study we used multiparameter flow cytometry and cell sorting to evaluate fetal bone marrow, a rich source of cells early in lymphoid development. We found CD7 to be expressed on a subset of CD19+ cells, including some that had matured to cytoplasmic mu+ (pre-B) and surface mu+ (B) cells. In addition, a less mature CD7+19+ population was characterized as mu- and CD34+/-. The CD7+19+ population was clearly distinct from the mature T cells. The CD7+19+ cells were negative for nuclear TdT in contrast to CD7–19+ cells, which frequently contained TdT. CD10, which is coexpressed on the cell surface of more than 90% of CD19+ lymphocytes, was detected in a minority of CD7+19+ lymphocytes. The CD7+19+34+ cell population may be B-lineage committed, or may represent uncommitted lymphoid precursors. The biologic role of the expression of CD7 on immature and mature cells, including those of the B lineage, may indicate (1) the presence of CD7+19+ lymphoid precursor cells and/or (2) an alternate pathway of B-cell development, in which cells coexpress CD7 with other B-lineage markers.

scholarly journals Identification of novel B-lineage cells in human fetal bone marrow that coexpress CD7 [see comments]

Blood ◽  
1991 ◽  
Vol 77 (1) ◽  
pp. 64-68 ◽  
Author(s):  
ER Grumayer ◽  
F Griesinger ◽  
DS Hummell ◽  
RD Brunning ◽  
JH Kersey
Keyword(s):  
Bone Marrow ◽  
Cell Sorting ◽  
B Cell Development ◽  
Multiparameter Flow Cytometry ◽  
Fetal Bone ◽  
Lymphoid Development ◽  
Alternate Pathway ◽  
Lineage Markers ◽  
B Lineage

In the present study we used multiparameter flow cytometry and cell sorting to evaluate fetal bone marrow, a rich source of cells early in lymphoid development. We found CD7 to be expressed on a subset of CD19+ cells, including some that had matured to cytoplasmic mu+ (pre-B) and surface mu+ (B) cells. In addition, a less mature CD7+19+ population was characterized as mu- and CD34+/-. The CD7+19+ population was clearly distinct from the mature T cells. The CD7+19+ cells were negative for nuclear TdT in contrast to CD7–19+ cells, which frequently contained TdT. CD10, which is coexpressed on the cell surface of more than 90% of CD19+ lymphocytes, was detected in a minority of CD7+19+ lymphocytes. The CD7+19+34+ cell population may be B-lineage committed, or may represent uncommitted lymphoid precursors. The biologic role of the expression of CD7 on immature and mature cells, including those of the B lineage, may indicate (1) the presence of CD7+19+ lymphoid precursor cells and/or (2) an alternate pathway of B-cell development, in which cells coexpress CD7 with other B-lineage markers.

scholarly journals Expression of the hematopoietic growth factor receptor FLT3 (STK- 1/Flk2) in human leukemias

Blood ◽  
1996 ◽  
Vol 87 (3) ◽  
pp. 1089-1096 ◽  
Author(s):  
CE Carow ◽  
M Levenstein ◽  
SH Kaufmann ◽  
J Chen ◽  
S Amin ◽  
...  
Keyword(s):  
Growth Factor ◽  
Progenitor Cells ◽  
Western Blotting ◽  
Growth Factor Receptor ◽  
Hematopoietic Growth Factor ◽  
Acute Leukemias ◽  
Rnase Protection ◽  
Flt3 Expression ◽  
B Lineage ◽  
Acute Myeloid

Normal expression of the hematopoietic growth factor receptor FLT3 (STK- 1@Flk2) is limited to CD34+ stem/progenitor cells. We have evaluated the expression of FLT3 by RNase protection assay and Western blotting in 161 primary bone marrow (BM) samples from patients with leukemia. FLT3 RNA was found to be expressed at a higher level than in normal BM controls in 33 of 33 B-lineage acute leukemias, 11 of 12 acute myeloid leukemias (AMLs), and 3 of 11 T-cell acute leukemias (T-ALLs). Expression of FLT3 RNA was also observed in some cases of blast crisis CML. The FLT3 signal resulted from expression on the leukemic blasts, and was not caused by increased FLT3 expression on normal CD34+ stem/progenitor cells in the leukemic samples. To determine if FLT3 protein was also overexpressed, proteins were extracted from leukemic BM samples and screened by Western blotting with anti-FLT3 antisera. FLT3 protein was not detected in normal BM controls, but was found in 14 of 14 B-lineage ALLs, 36 of 41 AMLs, and 1 of 4 T-ALLs. Stimulation of patient samples with FLT3 ligand resulted in autophosphorylation of the FLT3 receptor, suggesting the receptor is functional in these cells. These data show that FLT3 RNA and protein are aberrantly expressed by AML and ALL cells in that CD34 expression and FLT3 expression are no longer synchronous, and suggest the possibility that overexpression of FLT3 could play a role in the survival and/or proliferation of malignant clones in acute myeloid and lymphoid leukemias.

The Reliability and Specificity of c-kit for the Diagnosis of Acute Myeloid Leukemias and Undifferentiated Leukemias

Blood ◽  
1998 ◽  
Vol 92 (2) ◽  
pp. 596-599 ◽  
Author(s):  
M.C. Bene ◽  
M. Bernier ◽  
R.O. Casasnovas ◽  
G. Castoldi ◽  
W. Knapp ◽  
...  
Keyword(s):  
Acute Leukemia ◽  
De Novo ◽  
Lymphoblastic Leukemia ◽  
Cell Lineage ◽  
Specific Marker ◽  
Acute Leukemias ◽  
Routine Basis ◽  
Immunological Classification ◽  
B Lineage ◽  
Acute Myeloid

Abstract We document findings on c-kit (CD117) expression in 1,937 pediatric and adult de novo acute leukemia cases, diagnosed in five single European centers. All cases were well characterized as to the morphologic, cytochemical, and immunologic features, according to the European Group for the Immunological Classification of Leukemias (EGIL). The cases included 1,103 acute myeloid leukemia (AML), 819 acute lymphoblastic leukemia (ALL), 11 biphenotypic acute leukemia (BAL), and 4 undifferentiated (AUL). c-kit was expressed in 741 (67%) AML cases, regardless of the French-American-British (FAB) subtype, one third of BAL, all four AUL, but only in 34 (4%) of ALL cases. The minority of c-kit+ ALL cases were classified as: T-cell lineage (two thirds), mainly pro-T–ALL or T-I, and B lineage (one third); cells from 62% of these ALL cases coexpressed other myeloid markers (CD13, CD33, or both). There were no differences in the frequency of c-kit+ AML or ALL cases according to age being similar in the adult and pediatric groups. Our findings demonstrate that c-kit is a reliable and specific marker to detect leukemia cells committed to the myeloid lineage, and therefore should be included in a routine basis for the diagnosis of acute leukemias to demonstrate myeloid commitment of the blasts. c-kit expression should score higher, at least one point, in the system currently applied to the diagnosis of BAL, as its myeloid specificity is greater than CD13 and CD33. Findings in ALL and AUL suggest that c-kit identifies a subgroup of cases, which may correspond to leukemias either arising from early prothymocytes and/or early hematopoietic cells, both able to differentiate to the lymphoid and myeloid pathways.

Cellular pharmacokinetics of daunorubicin: relationships with the response to treatment in patients with acute myeloid leukemia.

1988 ◽  
Vol 6 (5) ◽  
pp. 802-812 ◽  
Author(s):  
E Kokenberg ◽  
P Sonneveld ◽  
W Sizoo ◽  
A Hagenbeek ◽  
B Löwenberg
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Response To Treatment ◽  
Remission Induction ◽  
Pharmacokinetic Parameters ◽  
Induction Treatment ◽  
Blast Cells ◽  
Acute Myeloid

In an attempt to identify pharmacokinetic factors that determine the response of acute myeloid leukemia (AML) patients to induction chemotherapy, we determined the concentrations of daunorubicin (DNR) and the main metabolite daunorubicinol (DOL) in vivo and particularly evaluated the concentrations in blood and bone marrow nucleated cells. Cell measurements were obtained in 37 evaluable patients during their first remission induction treatment with DNR and cytarabine (ara-C) and directly compared with the plasma distribution kinetics of DNR. We show that (1) plasma DNR concentrations do not correlate with DNR concentrations in bone marrow nucleated cells; but (2) plasma area under the curve (AUC) values of DNR correlate inversely (P less than .01) with AUC values of DNR in WBCs; (3) concentrations of DNR in WBCs correlate positively (P less than .01) with DNR concentrations in bone marrow nucleated cells; and (4) the concentrations of DNR in WBCs show a negative correlation (P less than .01) with the numbers of peripheral blast cells at diagnosis. We then tested whether the pharmacokinetic parameters had predictive value for the clinical outcome of therapy, but none of the plasma levels or WBC and bone marrow concentrations of DNR predicted treatment outcome. The inverse correlation between the concentrations of DNR in WBC and the numbers of peripheral blast cells suggests that the effective DNR concentrations achieved intracellularly are mainly a function of the tumor load so that lesser amounts of DNR accumulate intracellularly when the AML cell numbers in blood are higher.

Acute Myeloid Leukemia In Patient Over 90 Years Old: Where Do We Go From Here? Case Report

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5034-5034 ◽  
Author(s):  
Marcelo Bellesso ◽  
Daniela Ferreira Dias ◽  
Renato Centrone ◽  
Rodrigo Santucci ◽  
Izabel Pernambuco Nicodemo
Keyword(s):  
Quality Of Life ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Bone Marrow Aspirate ◽  
Chronological Age ◽  
Acceptable Quality ◽  
Blast Cells ◽  
Acute Myeloid

Abstract Introduction Acute Myeloid Leukemia (AML) widely affects elderly patients, with disappointing survival rates with increasing age. Although chronological age is an independent prognostic risk, it is really important to understand that this group is heterogeneous, so a geriatric assessment may be helpful before making decisions regarding therapy. We describe a 91 year-old patient with AML treated with decitabine, achieving a complete response and good quality of life for 10 months. Case Report In May 2012, a 91 year-old woman with myelodysplastic syndrome was admitted at the emergency department presenting asthenia, pallor, pain and edema of the left inferior limb. Her blood count showed: hemoglobin 10.7g/dL, leukocytes 81.800/mm³ with 95% blast cells, and platelets 45.000/mm³. In addition, a Doppler ultrasound evidenced deep venous thrombosis in her left leg. A bone marrow aspirate confirmed AML with myelodysplasia-related alterations, with 95% blast cells, with positive expression of CD45, CD33, MPO, and CD117 (CD45+; CD33+; MPO+; CD117+). Her karyotype was 46,XX [20 cells analyzed]. She was first treated with hydroxyurea. Afterwards, as her performance status improved and in spite of her age, we decided to treat her with Decitabine 20mg/m²/day for 5 days, because she had no severe comorbidity or any severe impairment of every-day-life instrumental activities. After a first cycle with grade 4 neutropenia and thrombocytopenia, she was discharged. After 40 days, complete hematologic recovery was observed. She received seven treatment cycles, and the most important symptom of toxicity was febrile grade 3 neutropenia in the 3rd cycle, with the only inpatient treatment. After this intercurrence, the doses were reduced by 25%. The patient achieved complete remission, spent eleven months without needing blood transfusions and with an acceptable quality of life, but then she relapsed, presenting persistent neutropenia and 44% of blasts in a bone marrow aspirate (05/27/2013). She died on 07/02/2013 due to the progression of the disease. Conclusion This case shows that, despite her advanced chronological age, our patient, affected by a fatal disease, lived approximately 13 months with optimal response to treatment (with Decitabine) and enjoying an acceptable quality of life for ten months. Considering this patient profile, could we conclude that this outcome is the best our treatment can aim to achieve? Disclosures: No relevant conflicts of interest to declare.

scholarly journals Megakaryoblastic leukemia presenting as acute myelofibrosis -- a study of four cases with the platelet-peroxidase reaction

Blood ◽  
1981 ◽  
Vol 58 (2) ◽  
pp. 206-213 ◽  
Author(s):  
BJ Bain ◽  
D Catovsky ◽  
M O'Brien ◽  
HG Prentice ◽  
E Lawlor ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Myeloid Leukemia ◽  
Ultrastructural Level ◽  
Bone Marrow Fibrosis ◽  
Megakaryoblastic Leukemia ◽  
Peroxidase Reaction ◽  
Blast Cells ◽  
Immature Myeloid Cells ◽  
Marrow Fibrosis ◽  
Acute Myeloid

Acute myelofibrosis (AM) or malignant myelosclerosis is a myeloprofilerative syndrome in which bone marrow fibrosis is associated with a proliferation of immature myeloid cells. In four patients with typical AM, investigated by the platelet-peroxidase reaction at ultrastructural level, the blast cells were found to be megakaryoblasts. One patient, treated with the drug combination DAT, achieved a complete remission of 5 mo duration. This study supports the view that megakaryoblastic leukemia is the most frequent underlying cause of AM and proposes that it should be classified as a form of acute myeloid leukemia.

scholarly journals Microvesicles microRNAs Reflect and Affect Progression of Acute Myeloid Leukemia and Could Serve As a Biomarker of Disease Dynamics

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1664-1664
Author(s):  
Inna Tzoran ◽  
Annie Rebibo-Sabbah ◽  
Benjamin Brenner ◽  
Anat Aharon
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Healthy Controls ◽  
Disease Dynamics ◽  
Rt Pcr ◽  
Blast Cells ◽  
Acute Myeloid

Abstract Background: Acute myeloid leukemia (AML) is characterized by rapid growth of abnormal blast cells that accumulate in the bone marrow and interfere with the production of normal blood cells. Microvesicles (MVs) are shedding from various cells and express antigens reflecting their cellular origin. Our previous study has demonstrated a correlation between the level of MVs originating from blast cells (CD117+ MVs) in the peripheral blood and the amount of CD117 positive blast cells in the bone marrow (BM) at diagnosis and in remission (r=0.49, p=0.0025; r=0.6, p=0.01; respectively). Additional assessment of CD117 expression on MVs obtained at diagnosis in patients who died despite achieving a remission revealed significantly higher values compared to those found in patients who were alive at 3 years of follow-up (1.9 vs. 0.6; p=0.01). A similar trend was found in CD34 positive MVs (1.2 vs. 0.13; p=0.01). It has, therefore, been concluded that circulating MVs of AML patients might serve as a biomarker of leukemia progression. MVs also contain cytokines and micro-RNA (miRNA) that are critical for cell development, proliferation and apoptosis. MVs are the major transport vehicle for miRNAs, and serve as a unique mode of genetic exchange between cells. To this end, several studies have shown that cancer stem cells regulate tumor environment through MVs. The current study aimed to explore the potential role of MV miRNAs as a biomarker of disease progression in AML and to study MV effects on the bone marrow leukemic niche. Methods: Blood and bone marrow samples were collected from healthy controls and patients with AML at diagnosis and upon achievement of first remission. MV effects on the BM mesenchymal stem cells (BM-MSC) and endothelial cells were studied using confocal microscopy, migration and proliferation assays and the RT-PCR method. miRNA expression was screened by NanoString technology and validated by RT-PCR. Results: The study was approved by the Institutional Review Board of the Rambam Health Care Campus (Approval #0351-10). Blood and bone marrow samples were collected from 43 AML patients and 4 random healthy volunteers after obtaining written informed consent. Sixty seven percent of patients remained alive and achieved remission following induction chemotherapy about one month after the diagnosis. Co-incubation of fluorescent-labeled BM-MVs (CD33+/CD117+) of AML patients with unlabeled BM-MSC resulted in incorporation of >80% of MVs to the cells. Patient BM-MVs of obtained at diagnosis induced a higher migration rate of BM-MSC compared to MVs obtained from healthy controls (p<0.01). Patient BM-MVs also induced a significantly higher proliferation rate of BM-MSC (p<0.05). Screening of patient BM-MVs demonstrated that the expression of some miRNAs was high at diagnosis but decreased in remission, while other miRNAs exhibited an opposite trend. Notably, these alterations were observed in miRNA-181a, that is known to play a role in normal and malignant hematopoiesis. Specifically, miRNA-181a levels in BM-MVs of AML patients were at least 10 times higher at diagnosis than in remission or in healthy controls (p<0.05). Similar results were found in miRNA-181a of circulating MVs of these patients, particularly in those who were alive at 1 year of follow-up. However, in patients that died within the first year, miRNA-181a expression at diagnosis was lower compared to healthy controls (2 times less, p=0.035) or to patients who were alive at the follow-up evaluation (7 times less, p= 0.0097). Co-incubation of BM-MSC with patient BM-MVs, obtained at diagnosis, resulted in a 4-time higher expression of miRNA-181a in these cells compared to untreated ones (p<0.05). Conclusion: MV-miRNAs of AML patients are involved in the regulation of tumor BM microenvironment, affecting BM-MSC migration, proliferation and gene expression. MV-miRNAs reflect and affect AML progression and may serve as a biomarker of disease dynamics. Disclosures No relevant conflicts of interest to declare.

scholarly journals AML1-ETO Collaborates with the Homeobox Gene Meis1 in Inducing Acute Leukemia in the Mouse Bone Marrow Transplantation Model.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 928-928 ◽  
Author(s):  
Vegi M. Naidu ◽  
Vijay P.S. Rawat ◽  
Christina Schessl ◽  
Konstantin Petropoulus ◽  
Monica Cusan ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Leukemia ◽  
Insertional Mutagenesis ◽  
Fusion Gene ◽  
Genetic Alterations ◽  
Control Group ◽  
Mouse Bone Marrow ◽  
Acute Leukemias ◽  
Hox Gene ◽  
Blast Cells

Abstract AML1-ETO is the most frequent fusion gene in human AML. Previously, we and others have demonstrated that the fusion is not able to cause leukaemia on its own in experimental murine models, but that it needs collaborative partners. However, although mutations such as the FLT3-length mutation and C-KIT mutations were defined as important collaborative genetic events in AML1-ETO positive AML, most human AML1-ETO cases do not carry these mutations, indicating the presence of unkown collaborative partners in these patients. On the other hand Meis1, a HOX gene co-factor, belonging to the TALE family of homeodomain proteins, has a well established function as a protooncogene with a strong collaborative potential in Hox gene associated AML in mice. First we confirmed expression of MEIS1 in some patients with AML1-ETO positive AML by real-time PCR. Based on this we sought to determine if AML1-ETO can collaborate with Meis1 in inducing acute leukemias: single constructs or both genes were co-transfected in 5-FU treated primary murine bone marrow cells by retroviral gene transfer, using MSCV retroviral constructs with an IRES–GFP or YFP cassette. Mice were transplanted with BM cells expressing Meis1 alone (n=10), with BM cells solely expressing the fusion gene (n=10) or EGFP (n=7, control) or with BM expressing both genetic alterations (n=14). None of the mice in the Meis1 and AML1-ETO as well as in the control group developed disease. In contrast, 14 mice transplanted with BM co-expressing AML1-ETO and Meis1 developed lethal disease after a median latency of 102 days. Three mice succumbed to a myeloproliferative syndrome and nine mice died by acute leukemia (6 mice developed AML, 3 mice ALL), which was serially transplantable into secondary recipients (median = 57 days). Immunohistochemistry of various organs of leukemic mice showed massive infiltration with blast cells. In MPS and AML 85 ± 9.3 % of the blast cells co-expressed Gr-1+ and Mac1+. In ALL cases 40 ± 19.9 % of the malignant cells co-expressed Mac1 and the lymphoid-associated B220 antigen. Analysis of retroviral integration did not reveal recurrent integration sites as an indication for insertional mutagenesis. In summary, our data demonstrate for the first time that AML1-ETO can collaborate with Meis1 and identify a novel collaborative partner in t(8;21) positive AML. Furthermore, our analyses demonstrate that Meis1 can collaborate with non-homeobox genes in inducing acute leukemia.

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