Automated Analysis of Differentiation-Induced Leukemic Cells During All-Trans Retinoic Acid Therapy of Acute Promyelocytic Leukemia

2003 ◽  
Vol 127 (1) ◽  
pp. e4-e10
Author(s):  
Kaoru Tohyama ◽  
Shuichi Shiga ◽  
Hiroyuki Fujimoto ◽  
Yukio Hamaguchi ◽  
Satoshi Ichiyama
Keyword(s):  
Flow Cytometry ◽  
Retinoic Acid ◽  
Fluorescence Intensity ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Differentiation Induction ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Hematology Analyzer ◽  
Automated Hematology Analyzer

Abstract During differentiation-induction therapy of acute promyelocytic leukemia (APL) patients with all-trans retinoic acid (ATRA), a variety of APL-derived bizarre granulocytic cells appear in the peripheral blood. To evaluate the differentiation induction of leukemic cells, we have developed a new scattergram analyzing program with an automated hematology analyzer and compared the data with the flow cytometry measuring the expression of differentiation-associated cell surface antigens, CD11b and CD16. We used the fluorescence intensity and side scatter as parameters of granulocytic maturation in the analysis with the automated hematology analyzer. The analysis of 2 ATRA-treated APL patients and in vitro study using HL-60 cells demonstrated that the levels of fluorescence intensity and side scatter decreased as accompanied with granulocytic maturation, and these changes were parallel with the results of flow cytometry. Our automated scattergram analysis of cell differentiation will contribute to general, objective, and real-time evaluation of differentiation-induction therapy of APL with ATRA.

scholarly journals Resistance to all-trans retinoic acid (ATRA) therapy in relapsing acute promyelocytic leukemia: study of in vitro ATRA sensitivity and cellular retinoic acid binding protein levels in leukemic cells [see comments]

Blood ◽  
1993 ◽  
Vol 82 (7) ◽  
pp. 2175-2181 ◽  
Author(s):  
L Delva ◽  
M Cornic ◽  
N Balitrand ◽  
F Guidez ◽  
JM Miclea ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Acute Promyelocytic Leukemia ◽  
Induction Therapy ◽  
Binding Protein ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Differentiation Induction ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

Abstract All-trans retinoic acid (ATRA) induces leukemic cell differentiation and complete remission (CR) in a high proportion of patients with acute promyelocytic leukemia (AML3 subtype). However, relapses occur when ATRA is prescribed as maintenance therapy, and resistance to a second ATRA-induction therapy is frequently observed. An induced hypercatabolism of ATRA has been suggested as a possible mechanism leading to reduced ATRA sensitivity and resistance. CRABPII, an RA cytoplasmic binding protein linked to RA's metabolization pathway, is induced by ATRA in different cell systems. To investigate whether specific features of the AML3 cells at relapse could explain the in vivo resistance observed, we studied the CRABP levels and in vitro sensitivity to ATRA of AML3 cells before and at relapse from ATRA. Relapse-AML3 cells (n = 12) showed reduced differentiation induction when compared with “virgin”-AML3 cells (n = 31; P < .05). Dose-response studies were performed in 2 cases at relapse and showed decreased sensitivity to low ATRA concentrations. CRABPII levels and in vitro differentiation characteristics of AML3 cells before and at relapse from ATRA therapy were studied concomittantly in 4 patients. High levels of CRABPII (median, 20 fmol/mg of protein) were detected in the cells of the 4 patients at relapse but were not detected before ATRA therapy. Three of these patients showed a decrease in differentiation induction of their leukemic cells, and a failure to achieve CR with a second induction therapy of ATRA 45 mg/m2/day was noted in all patients treated (n = 3). Results from this study provide evidence to support the hypothesis of induced-ATRA metabolism as one of the major mechanisms responsible for ATRA resistance. Monitoring CRABPII levels after ATRA withdrawal may help to determine when to administer ATRA in the maintenance or relapse therapy of AML3 patients.

scholarly journals Resistance to all-trans retinoic acid (ATRA) therapy in relapsing acute promyelocytic leukemia: study of in vitro ATRA sensitivity and cellular retinoic acid binding protein levels in leukemic cells [see comments]

Blood ◽  
1993 ◽  
Vol 82 (7) ◽  
pp. 2175-2181 ◽  
Author(s):  
L Delva ◽  
M Cornic ◽  
N Balitrand ◽  
F Guidez ◽  
JM Miclea ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Acute Promyelocytic Leukemia ◽  
Induction Therapy ◽  
Binding Protein ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Differentiation Induction ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

All-trans retinoic acid (ATRA) induces leukemic cell differentiation and complete remission (CR) in a high proportion of patients with acute promyelocytic leukemia (AML3 subtype). However, relapses occur when ATRA is prescribed as maintenance therapy, and resistance to a second ATRA-induction therapy is frequently observed. An induced hypercatabolism of ATRA has been suggested as a possible mechanism leading to reduced ATRA sensitivity and resistance. CRABPII, an RA cytoplasmic binding protein linked to RA's metabolization pathway, is induced by ATRA in different cell systems. To investigate whether specific features of the AML3 cells at relapse could explain the in vivo resistance observed, we studied the CRABP levels and in vitro sensitivity to ATRA of AML3 cells before and at relapse from ATRA. Relapse-AML3 cells (n = 12) showed reduced differentiation induction when compared with “virgin”-AML3 cells (n = 31; P < .05). Dose-response studies were performed in 2 cases at relapse and showed decreased sensitivity to low ATRA concentrations. CRABPII levels and in vitro differentiation characteristics of AML3 cells before and at relapse from ATRA therapy were studied concomittantly in 4 patients. High levels of CRABPII (median, 20 fmol/mg of protein) were detected in the cells of the 4 patients at relapse but were not detected before ATRA therapy. Three of these patients showed a decrease in differentiation induction of their leukemic cells, and a failure to achieve CR with a second induction therapy of ATRA 45 mg/m2/day was noted in all patients treated (n = 3). Results from this study provide evidence to support the hypothesis of induced-ATRA metabolism as one of the major mechanisms responsible for ATRA resistance. Monitoring CRABPII levels after ATRA withdrawal may help to determine when to administer ATRA in the maintenance or relapse therapy of AML3 patients.

Chemokine induction by all-trans retinoic acid and arsenic trioxide in acute promyelocytic leukemia: triggering the differentiation syndrome

Blood ◽  
2009 ◽  
Vol 114 (27) ◽  
pp. 5512-5521 ◽  
Author(s):  
Maaike Luesink ◽  
Jeroen L. A. Pennings ◽  
Willemijn M. Wissink ◽  
Peter C. M. Linssen ◽  
Petra Muus ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Arsenic Trioxide ◽  
Acute Promyelocytic Leukemia ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Differentiation Therapy ◽  
Chemokine Production ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Pulmonary Infiltration

Abstract In acute promyelocytic leukemia (APL), differentiation therapy with all-trans retinoic acid (ATRA) and/or arsenic trioxide can induce a differentiation syndrome (DS) with massive pulmonary infiltration of differentiating leukemic cells. Because chemokines are implicated in migration and extravasation of leukemic cells, chemokines might play a role in DS. ATRA stimulation of the APL cell line NB4 induced expression of multiple CC-chemokines (CCLs) and their receptors (> 19-fold), resulting in increased chemokine levels and chemotaxis. Induction of CCL2 and CCL24 was directly mediated by ligand-activated retinoic acid receptors. In primary leukemia cells derived from APL patients at diagnosis, ATRA induced chemokine production as well. Furthermore, in plasma of an APL patient with DS, we observed chemokine induction, suggesting that chemokines might be important in DS. Dexamethasone, which efficiently reduces pulmonary chemokine production, did not inhibit chemokine induction in APL cells. Finally, chemokine production was also induced by arsenic trioxide as single agent or in combination with ATRA. We propose that differentiation therapy may induce chemokine production in the lung and in APL cells, which both trigger migration of leukemic cells. Because dexamethasone does not efficiently reduce leukemic chemokine production, pulmonary infiltration of leukemic cells may induce an uncontrollable hyperinflammatory reaction in the lung.

scholarly journals Do High-mobility Group Box 1 Gene Polymorphisms Affect the Incidence of Differentiation Syndrome in Acute Promyelocytic Leukemia?

2021 ◽  
Author(s):  
Ghazaleh Hoseinzadeh ◽  
Zahra Mohammadzadeh ◽  
Bahram Chahardouli ◽  
Kamran Ali Moghaddam ◽  
Seyed Asadollah Mousavi ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Arsenic Trioxide ◽  
Acute Promyelocytic Leukemia ◽  
High Mobility ◽  
Promyelocytic Leukemia ◽  
High Mobility Group ◽  
Leukemic Cells ◽  
Genotype Distribution ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

Abstract Differentiation syndrome (DS) is an inflammatory complication seen in some patients with acute promyelocytic leukemia (APL) undergoing differentiation therapy with all-trans retinoic acid (ATRA) and/or arsenic trioxide (ATO). It is unknown how DS occurs, but it is believed that it is caused by inflammatory cytokines release from differentiating leukemic cells. High mobility group box-1 (HMGB1) is a DNA-binding protein that acts as a cytokine outside of cells and may play a role in inflammation. This study was conducted to determine whether HMGB1 polymorphisms (rs1360485, rs2249825 and rs1060348) are associated with the incidence of differentiation syndrome in acute promyelocytic leukemia patients treated with all-trans retinoic acid and arsenic trioxide. One hundred and thirty APL patients and 100 healthy controls were included. Seventeen patients with differentiation syndrome were selected according to the PETHEMA criteria. Tetra-primer ARMS polymerase chain reaction (tetra-ARMS PCR) was used to determine the genotype distribution of polymorphisms. DNA sequencing was done to validate the results. In both healthy and APL patients, AA was the most frequent genotype in rs1360485 followed by AG and GG. CC, CG, and GG were the most frequent genotypes in rs2249825 polymorphism in the order mentioned. CC was more frequent than CT, and CT was more frequent than TT in rs1060348. There was no correlation between HMGB1 polymorphisms and the incidence of differentiation syndrome based on genetic models (p-value > 0.05). As a result, HMGB1 polymorphisms are not probably associated with DS development in APL patients treated with ATRA and ATO.

scholarly journals Complete Remission of t(11;17) Positive Acute Promyelocytic Leukemia Induced by All-trans Retinoic Acid and Granulocyte Colony-Stimulating Factor

Blood ◽  
1999 ◽  
Vol 94 (1) ◽  
pp. 39-45 ◽  
Author(s):  
J.H. Jansen ◽  
M.C. de Ridder ◽  
W.M.C. Geertsma ◽  
C.A.J. Erpelinck ◽  
K. van Lom ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Acute Promyelocytic Leukemia ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Leukemia Cells ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Stimulating Factor

The combined use of retinoic acid and chemotherapy has led to an important improvement of cure rates in acute promyelocytic leukemia. Retinoic acid forces terminal maturation of the malignant cells and this application represents the first generally accepted differentiation-based therapy in leukemia. Unfortunately, similar approaches have failed in other types of hematological malignancies suggesting that the applicability is limited to this specific subgroup of patients. This has been endorsed by the notorious lack of response in acute promyelocytic leukemia bearing the variant t(11;17) translocation. Based on the reported synergistic effects of retinoic acid and the hematopoietic growth factor granulocyte colony-stimulating factor (G-CSF), we studied maturation of t(11;17) positive leukemia cells using several combinations of retinoic acid and growth factors. In cultures with retinoic acid or G-CSF the leukemic cells did not differentiate into mature granulocytes, but striking granulocytic differentiation occurred with the combination of both agents. At relapse, the patient was treated with retinoic acid and G-CSF before reinduction chemotherapy. With retinoic acid and G-CSF treatment alone, complete granulocytic maturation of the leukemic cells occurred in vivo, followed by a complete cytogenetical and hematological remission. Bone marrow and blood became negative in fluorescense in situ hybridization analysis and semi-quantitative polymerase chain reaction showed a profound reduction of promyelocytic leukemia zinc finger–retinoic acid receptor- fusion transcripts. This shows that t(11;17) positive leukemia cells are not intrinsically resistant to retinoic acid, provided that the proper costimulus is administered. These observations may encourage the investigation of combinations of all-trans retinoic acid and hematopoietic growth factors in other types of leukemia.

scholarly journals CCAAT/enhancer binding proteins alpha and epsilon cooperate with all-trans retinoic acid in therapy but differ in their antileukemic activities

Blood ◽  
2006 ◽  
Vol 108 (7) ◽  
pp. 2416-2419 ◽  
Author(s):  
Young-Jin Lee ◽  
Letetia C. Jones ◽  
Nikolai A. Timchenko ◽  
Danilo Perrotti ◽  
Daniel G. Tenen ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Binding Proteins ◽  
Retinoic Acid Receptor ◽  
Chromosomal Translocation ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Enhancer Binding Proteins ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Retinoic Acid Receptor Α

Abstract CCAAT/enhancer binding proteins (C/EBPs) play critical roles in myelopoiesis. Dysregulation of these proteins likely contributes to the pathogenesis of myeloid disorders characterized by a block in granulopoiesis. In one such disease, acute promyelocytic leukemia (APL), a promyelocytic leukemia–retinoic acid receptor α (PML-RARα) fusion protein is expressed as a result of a t(15;17) chromosomal translocation. Treatment of PML-RARα leukemic cells with all-trans retinoic acid (ATRA) causes them to differentiate into mature neutrophils, an effect thought to be mediated by C/EBPs. In this study, we assess the potential for cooperativity between increased C/EBP activity and ATRA therapy. We demonstrate that although both C/EBPα and C/EBPϵ can significantly prolong survival in a mouse model of APL, they are not functionally equivalent in this capacity. We also show that forced expression of C/EBPα or C/EBPϵ in combination with ATRA treatment has a synergistic effect on survival of leukemic mice compared with either therapy alone.

Tissue transglutaminase contributes to the all-trans-retinoic acid–induced differentiation syndrome phenotype in the NB4 model of acute promyelocytic leukemia

Blood ◽  
2010 ◽  
Vol 116 (19) ◽  
pp. 3933-3943 ◽  
Author(s):  
Krisztián Csomós ◽  
István Német ◽  
László Fésüs ◽  
Zoltán Balajthy
Keyword(s):  
Retinoic Acid ◽  
Acute Promyelocytic Leukemia ◽  
Tissue Transglutaminase ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Neutrophil Granulocytes ◽  
Phagocytic Capacity ◽  
Nb4 Cells ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

Abstract Treatment of acute promyelocytic leukemia (APL) with all-trans-retinoic acid (ATRA) results in terminal differentiation of leukemic cells toward neutrophil granulocytes. Administration of ATRA leads to massive changes in gene expression, including down-regulation of cell proliferation–related genes and induction of genes involved in immune function. One of the most induced genes in APL NB4 cells is transglutaminase 2 (TG2). RNA interference–mediated stable silencing of TG2 in NB4 cells (TG2-KD NB4) coupled with whole genome microarray analysis revealed that TG2 is involved in the expression of a large number of ATRA-regulated genes. The affected genes participate in granulocyte functions, and their silencing lead to reduced adhesive, migratory, and phagocytic capacity of neutrophils and less superoxide production. The expression of genes related to cell-cycle control also changed, suggesting that TG2 regulates myeloid cell differentiation. CC chemokines CCL2, CCL3, CCL22, CCL24, and cytokines IL1B and IL8 involved in the development of differentiation syndrome are expressed at significantly lower level in TG2-KD NB4 than in wild-type NB4 cells upon ATRA treatment. Based on our results, we propose that reduced expression of TG2 in differentiating APL cells may suppress effector functions of neutrophil granulocytes and attenuate the ATRA-induced inflammatory phenotype of differentiation syndrome.

All-trans retinoic acid pharmacokinetics and bioavailability in acute promyelocytic leukemia: intracellular concentrations and biologic response relationship.

1995 ◽  
Vol 13 (10) ◽  
pp. 2517-2523 ◽  
Author(s):  
A Agadir ◽  
M Cornic ◽  
P Lefebvre ◽  
B Gourmel ◽  
M Jérôme ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Acute Promyelocytic Leukemia ◽  
Promyelocytic Leukemia ◽  
Leukemic Cells ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Group A ◽  
In Vitro Response

PURPOSE This study investigated the in vitro pharmacologic behavior and disposition kinetics of all-trans retinoic acid (ATRA) in acute myeloid leukemic (AML) cells, their sensitivity to its differentiating effect, and the in vivo response of acute promyelocytic leukemia (APL) patients after therapy. PATIENTS AND METHODS Fresh leukemic cells from 14 AML patients (nine APL and five non-APL), were incubated in suspension culture in the absence or presence of 10(-6) mol/L ATRA. Intracellular ATRA concentration and ATRA metabolism was determined by high-performance liquid chromatography (HPLC). RESULTS Immediate uptake is observed with maximal intracellular levels (Cmax) achieved after 24 hours of incubation. At this time, ATRA levels were variable, ranging from 20 to 230 pmol/10(6) cells (median, 100 pmol/10(6) cells). Comparison of ATRA intracellular levels with the in vitro response of patients' cell samples as measured by the percentage of nitro blue tetrazolium (NBT)-positive cells after a 3-day incubation period allowed us to discriminate a group of APL patients (n = 6) with high Cmax (group A; median, 200 pmol/10(6) cells) and maximal differentiation at day 3 (median, 80%), and a group of patients (n = 8, three APL and five non-APL) with low Cmax (group B; median, 35 pmol/10(6) cells) and poor in vitro response (median, 40%; APL cases only). Interestingly, all APL patients, except one included in group A (rapid in vitro ATRA uptakers), achieved a complete remission. CONCLUSION These findings suggest that intracellular ATRA concentrations are determinant for ATRA response and should be taken into account when monitoring the efficacy of ATRA differentiation therapeutic trials in malignant disorders.

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