scholarly journals Variable transcription of BCR-ABL by Ph+ cells arising from hematopoietic progenitors in chronic myeloid leukemia [see comments]

Blood ◽  
1994 ◽  
Vol 83 (7) ◽  
pp. 1744-1749 ◽  
Author(s):  
A Keating ◽  
XH Wang ◽  
P Laraya
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Internal Control ◽  
Myeloid Leukemia ◽  
Critical Role ◽  
Pcr Amplification ◽  
Chronic Phase ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Polymerase Chain ◽  
Aberrant Gene

Recent studies suggest that the BCR-ABL gene plays a critical role in the pathogenesis of Ph+ chronic myeloid leukemia (CML). We investigated the hematopoietic colonies derived from the marrows of 12 patients with Ph+ CML in chronic phase by reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of BCR-ABL mRNA and by cytogenetics. Colonies were individually harvested and each colony divided into two portions, one for cytogenetics and the other for isolation of total RNA for PCR of BCR-ABL transcripts and for an RNA internal control. We found that 23% +/- 18% (mean +/- SD, range 0% to 60%) of Ph+ colonies did not transcribe the aberrant gene. In each case when BCR-ABL transcription was not detected, normal ABL mRNA was present. The data suggest that hitherto unknown mechanisms may regulate BCR-ABL expression in some Ph+ cells and indicate that caution should be exercised in the interpretation of results using RT-PCR analysis of hematopoietic colonies from clinical specimens and from experiments with antisense oligonucleotides directed at the BCR-ABL gene. These data also raise the notion of a transitional Ph+ precursor cell in which BCR-ABL may become upregulated and lead to a fully expressed phenotype. We conclude that further studies correlating the frequency of Ph+ PCR- progenitors with prognostic clinical variables are warranted.

scholarly journals Variable transcription of BCR-ABL by Ph+ cells arising from hematopoietic progenitors in chronic myeloid leukemia [see comments]

Blood ◽  
1994 ◽  
Vol 83 (7) ◽  
pp. 1744-1749 ◽  
Author(s):  
A Keating ◽  
XH Wang ◽  
P Laraya
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Internal Control ◽  
Myeloid Leukemia ◽  
Critical Role ◽  
Pcr Amplification ◽  
Chronic Phase ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Polymerase Chain ◽  
Aberrant Gene

Abstract Recent studies suggest that the BCR-ABL gene plays a critical role in the pathogenesis of Ph+ chronic myeloid leukemia (CML). We investigated the hematopoietic colonies derived from the marrows of 12 patients with Ph+ CML in chronic phase by reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of BCR-ABL mRNA and by cytogenetics. Colonies were individually harvested and each colony divided into two portions, one for cytogenetics and the other for isolation of total RNA for PCR of BCR-ABL transcripts and for an RNA internal control. We found that 23% +/- 18% (mean +/- SD, range 0% to 60%) of Ph+ colonies did not transcribe the aberrant gene. In each case when BCR-ABL transcription was not detected, normal ABL mRNA was present. The data suggest that hitherto unknown mechanisms may regulate BCR-ABL expression in some Ph+ cells and indicate that caution should be exercised in the interpretation of results using RT-PCR analysis of hematopoietic colonies from clinical specimens and from experiments with antisense oligonucleotides directed at the BCR-ABL gene. These data also raise the notion of a transitional Ph+ precursor cell in which BCR-ABL may become upregulated and lead to a fully expressed phenotype. We conclude that further studies correlating the frequency of Ph+ PCR- progenitors with prognostic clinical variables are warranted.

scholarly journals Concurrent Diagnosis of Chronic Myeloid Leukemia and Follicular Lymphoma: An Unreported Presentation

2018 ◽  
Vol 2018 ◽  
pp. 1-4
Author(s):  
Amy G. Starr ◽  
Sushma R. Jonna ◽  
Joeffrey J. Chahine ◽  
Bhaskar V. Kallakury ◽  
Chaitra S. Ujjani
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Follicular Lymphoma ◽  
Myeloid Leukemia ◽  
Chronic Phase ◽  
Mediastinal Lymphadenopathy ◽  
Pcr Analysis ◽  
Molecular Testing ◽  
Fish Analysis ◽  
Rt Pcr ◽  
Non Hodgkin Lymphoma

Lymphadenopathy in chronic myeloid leukemia (CML) is usually due to extramedullary involvement with accelerated or blast phases of the disease. The occurrence of non-Hodgkin lymphoma (NHL) as a synchronous malignancy with CML is rare. We report a case of a 73-year-old male who presented with dyspnea and right-sided lower extremity edema in the setting of leukocytosis. Bone marrow evaluation indicated a chronic phase chronic myeloid leukemia (CML), confirmed by molecular testing. Imaging of the chest for persistent dyspnea revealed supraclavicular and mediastinal lymphadenopathy. Biopsy of the cervical node showed expanded lymphoid follicles with atypical germinal centers that were positive for CD10, BCL-2, and BCL-6, consistent with follicular lymphoma (FL). Nodal PCR demonstrated clonal IGH and IGK gene rearrangements, and FISH analysis was positive for IGH-BCL-2 fusion. Together, these tests supported the diagnosis of FL. Additionally, the lymph node showed paracortical expansion by maturing pan-hematopoietic elements, no blastic groups, and positive RT-PCR analysis for BCR-ABL1, indicating concomitant involvement by chronic phase-CML. To our knowledge, this is the first reported case of a patient with a concurrent diagnosis of CML and FL.

scholarly journals BCR-ABL, ABL-BCR, BCR, and ABL genes are all expressed in individual granulocyte-macrophage colony-forming unit colonies derived from blood of patients with chronic myeloid leukemia

Blood ◽  
1995 ◽  
Vol 85 (8) ◽  
pp. 2171-2175 ◽  
Author(s):  
J Diamond ◽  
JM Goldman ◽  
JV Melo
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Peripheral Blood ◽  
Myeloid Leukemia ◽  
Pcr Amplification ◽  
Pcr Screening ◽  
Colony Forming Unit ◽  
Ph Chromosome ◽  
Polymerase Chain ◽  
Macrophage Colony ◽  
Positive Results

It has been suggested that the BCR-ABL gene of chronic myeloid leukemia (CML) is not uniformly expressed in Philadelphia (Ph)-positive cells, and that BCR-ABL gene expression precludes transcription of the normal BCR or ABL genes. Therefore, we have analyzed granulocyte-macrophage colony-forming unit (CFU-GM) colonies derived from peripheral blood of 11 CML patients by cytogenetic and by reverse transcriptase-polymerase chain reaction (PCR) amplification of BCR-ABL, ABL-BCR, BCR, and ABL. All CFU-GM colonies with analyzable metaphases were found to contain a Ph chromosome. In 2 patients, the initial PCR screening failed to detect BCR-ABL transcripts in 2 of 11 and 1 of 7 Ph-positive colonies. However, when amplification for BCR-ABL was repeated in quintuplicate, all but 1 colony from a single patient showed one or more positive results. Amplifications of the four genes in each colony showed that BCR-ABL, ABL-BCR, and the normal BCR and ABL were simultaneously expressed in the majority of CFU-GM colonies. Replicate PCR tests for BCR and for ABL in colonies initially scored as negative also uncovered previously undetected positive amplifications. We conclude that BCR-ABL expression does not suppress transcription from the normal BCR and ABL genes, and that Ph-positive, BCR-ABL-negative colonies derived from peripheral blood CFU-GM are rare or nonexistent.

RT-PCR analysis of Na+/H+ exchanger mRNAs in rat medullary thick ascending limb

1995 ◽  
Vol 268 (6) ◽  
pp. F1224-F1228 ◽  
Author(s):  
P. Borensztein ◽  
M. Froissart ◽  
K. Laghmani ◽  
M. Bichara ◽  
M. Paillard
Keyword(s):  
Rat Kidney ◽  
Pcr Amplification ◽  
Restriction Enzymes ◽  
Pcr Analysis ◽  
Thick Ascending Limb ◽  
Rt Pcr ◽  
Medullary Thick Ascending Limb ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Nhe Isoforms

The thick ascending limb (TAL) of rat kidney absorbs bicarbonate secondary to proton secretion, but displays both basolateral and luminal Na+/H+ exchange (NHE) activity. Several NHE genes, including NHE-1, NHE-2, NHE-3, and NHE-4, are expressed in the kidney. To identify the NHE isoforms expressed in the rat medullary TAL (MTAL), we used the reverse transcription-polymerase chain reaction (RT-PCR) to detect the mRNAs for NHE in microdissected MTAL. RT-PCR amplification from total RNA was performed between two specific primers for each NHE isoform. In rat kidney homogenate, the four NHE isoform mRNAs were detected, and the identity of the PCR products was demonstrated by the sizes of the fragments, digestion with restriction enzymes, and Southern blot analysis. In microdissected rat MTAL, NHE-3 was strongly expressed and NHE-1 mRNA was also detected, whereas NHE-2 and NHE-4 mRNAs were not detected. Therefore, NHE-3 could be the apical Na+/H+ exchanger, and NHE-1 could be the basolateral isoform in the MTAL.

scholarly journals A new method of "in-cell reverse transcriptase-polymerase chain reaction" for the detection of BCR/ABL transcript in chronic myeloid leukemia patients

Blood ◽  
1996 ◽  
Vol 87 (9) ◽  
pp. 3822-3827 ◽  
Author(s):  
N Testoni ◽  
G Martinelli ◽  
P Farabegoli ◽  
A Zaccaria ◽  
M Amabile ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chronic Myeloid Leukemia ◽  
Messenger Rna ◽  
Myeloid Leukemia ◽  
Residual Disease ◽  
Single Cells ◽  
Fluorescent Microscopy ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

Methods of detecting minimal residual disease (MRD) in chronic myeloid leukemia (CML) include chromosome analysis, Southern blotting, polymerase chain reaction (PCR) and fluorescent in situ hybridization (FISH) techniques. We report a novel method to detect intracellular messenger RNA (mRNA) by combining the techniques of reverse transcription (RT) and PCR performed directly inside the cells, without extraction of the nucleic acid. We applied this method, which we call “in-cell RT-PCR”, to detect hybrid BCR/ABL transcript within single cells. After cellular permeabilization and fixation of single cells in suspension, the neoplastic mRNA was reverse transcribed into cDNA, and the cDNA was amplified by PCR with fluorescent primers, specific for bcr/abl. Flow cytometry was used to detect cells positive for the amplified DNA within the cell cytoplasm. After transferring the amplified cells onto slides by cytospin, the positive cells for BCR/ABL cDNA were observed by fluorescent microscopy. The technique was capable of detecting low abundancy signals and distinguishing different levels of gene expression. The amplification products were found in the cells and supernatants. The distribution was critically affected by the protease digestion condition. The specificity of amplification was confirmed by a nested RT-PCR of BCR/ABL performed on extracted mRNA from the same sample, and by reamplification of supernatants. We have used the technique to study 10 Ph+ CML patients and three normal subjects as controls. Four patients were 100% Ph+ at diagnosis time and RT-PCR+ at cytogenetic and molecular analysis, respectively. In-cell RT- PCR showed that the residual non-neoplastic cells could be observed in all cases. In two patients undergoing interferon-alpha (IFN-alpha) therapy and in four bone-marrow transplanted patients, the in-cell RT- PCR was used to compare the level of Ph+ positivity detected by cytogenetic analysis with the number of cells expressing BCR/ABL transcript. In this manner, we could estimate the MRD. Our preliminary application of the technique suggests that it is capable of accurately identifying cells transcribing bcr/abl, and that it may have significant clinical applications in the detection of MRD.

scholarly journals Philadelphia-negative (Ph-) chronic myeloid leukemia (CML): comparison with Ph+ CML and chronic myelomonocytic leukemia. The Groupe Francais de Cytogenetique Hematologique

Blood ◽  
1991 ◽  
Vol 78 (1) ◽  
pp. 205-211 ◽  
Author(s):  
P Martiat ◽  
JL Michaux ◽  
J Rodhain
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Chronic Phase ◽  
Chronic Myelomonocytic Leukemia ◽  
Ras Mutation ◽  
Ph Chromosome ◽  
Polymerase Chain ◽  
Atypical Cml ◽  
Myelomonocytic Leukemia ◽  
Monocytic Lineage

To better understand the Philadelphia-negative (Ph-) chronic myeloid leukemia (CML) and its relationships with Philadelphia-positive (Ph+) CML and chronic myelomonocytic leukemia (CMML), a study was undertaken by the Groupe Francais de Cytogenetique Hematologique. Thirty-five Ph- CML patients were investigated and compared with 55 chronic phase Ph+ CML and 100 CMML patients. There were 12 M-BCR positive (M-BCR+) and 23 M-BCR negative (MBCR+) patients. No clinical or biologic differences were found between Ph+ and Ph-, M-BCR+ patients. In the Ph- group, M- BCR+ and M-BCR- patients differed significantly in age (47.7 +/- 6.6 v 67.0 +/- 6.1 years, respectively; P = .001), leukocytosis (153.4 +/- 135.1 v 58.5 +/- 37.7 10(9)/L, P = .002), relative monocytosis (1.8% +/- 1.2% v 5.6% +/- 1.4%, P = .048), absolute basophilia (8.5 +/- 9.7 v 0.9 +/- 1.5 10(9)/L, P = .001), percentage of immature myeloid precursors (IMP) in peripheral blood (29.0% +/- 9.5% v 15.3% +/- 8.1%, P = .001), and percentage of erythroblasts in bone marrow (BM) (6.5% +/- 3.5% v 14.6% +/- 3.6%, P = .001). Karyotypic abnormalities other than the Ph chromosome occurred in 0 of 12 M-BCR- at diagnosis and 7 of 23 M-BCR- Ph- CML (P = .033). None of the 13 investigated BCR- patients had detectable BCR/ABL transcripts using polymerase chain reaction (PCR) and none had an N-RAS mutation. Cytologic findings showed a marked morphologic difference between M-BCR+ and M-BCR- patients, especially in the monocytic lineage. Dysmyelopoietic features in CMML and M-BCR- patients were very similar, and the differences were of quantitative order only. Using four criteria (monocytosis, percentage of IMP, basophilia, and percentage of erythroblasts in BM), patients could be divided into typical and atypical CML and this classification correlated well with molecular findings. We conclude that, while Ph-, M- BCR+, and Ph+ CML are identical diseases, Ph-, M-BCR- CML, and CMML have many similarities and might be only different aspects of a same entity.

scholarly journals Early detection of BCR-ABL transcripts by quantitative reverse transcriptase–polymerase chain reaction predicts outcome after allogeneic stem cell transplantation for chronic myeloid leukemia

Blood ◽  
2001 ◽  
Vol 97 (6) ◽  
pp. 1560-1565 ◽  
Author(s):  
Eduardo Olavarria ◽  
Edward Kanfer ◽  
Richard Szydlo ◽  
Jaspal Kaeda ◽  
Katayoun Rezvani ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Stem Cell Transplantation ◽  
Reverse Transcriptase ◽  
Allogeneic Stem Cell Transplantation ◽  
Myeloid Leukemia ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

The reverse transcriptase–polymerase chain reaction (RT-PCR) has become widely used for monitoring minimal residual disease after allogeneic stem cell transplantation (SCT) for chronic myeloid leukemia (CML). However, most of these studies were performed using qualitative RT-PCR, and the interpretation of the results obtained has been conflicting. The correlation of a quantitative RT-PCR test performed early after SCT (at 3 to 5 months) and long-term outcome of CML patients surviving for more than 6 months was studied. Between January 1991 and June 1999, data from 138 CML patients who received allografts were evaluated. Early RT-PCR results were classified as (1) negative if there were no BCR-ABLtranscripts detected (n = 61), (2) positive at low level if the total number of BCR-ABL transcripts was less than 100 per μg RNA and/or the BCR-ABL/ABL ratio was less than 0.02% (n = 14), or (3) positive at high level if transcript levels exceeded the thresholds defined above (n = 63). Three years after SCT the cumulative incidence of relapse was 16.7%, 42.9%, and 86.4%, respectively (P = .0001). The relationship betweenBCR-ABL transcript level and probability of relapse was apparent whether patients had received sibling or unrelated donor SCT and also whether or not the transplantation was T cell depleted. The results suggest that quantitative RT-PCR performed early after SCT is useful for predicting outcome and may help to define the need for further treatment.

Discontinuation of tyrosine kinase inhibitors in chronic myeloid leukemia: when is this a safe option to consider?

Hematology ◽  
2013 ◽  
Vol 2013 (1) ◽  
pp. 184-188 ◽  
Author(s):  
Kendra Sweet ◽  
Vivian Oehler
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Imatinib Mesylate ◽  
Myeloid Leukemia ◽  
Kinase Inhibitors ◽  
Deep Level ◽  
Chronic Phase ◽  
Cytogenetic Response ◽  
Response To Therapy ◽  
Molecular Response ◽  
Rt Pcr

Abstract Mrs G is a 54-year-old woman with a diagnosis of chronic-phase chronic myeloid leukemia dating back 8 years. She had a low-risk Sokal score at diagnosis and was started on imatinib mesylate at 400 mg orally daily within one month of her diagnosis. Her 3-month evaluation revealed a molecular response measured by quantitative RT-PCR of 1.2% by the International Scale. Within 6 months of therapy, she achieved a complete cytogenetic response, and by 18 months, her BCR-ABL1 transcript levels were undetectable using a quantitative RT-PCR assay with a sensitivity of ≥ 4.5 logs. She has maintained this deep level of response for the past 6.5 years. Despite her excellent response to therapy, she continues to complain of fatigue, intermittent nausea, and weight gain. She is asking to discontinue imatinib mesylate and is not interested in second-line therapy. Is this a safe and reasonable option for this patient?

The t(8;22) in chronic myeloid leukemia fuses BCR to FGFR1: transforming activity and specific inhibition of FGFR1 fusion proteins

Blood ◽  
2001 ◽  
Vol 98 (13) ◽  
pp. 3778-3783 ◽  
Author(s):  
Asuman Demiroglu ◽  
E. Joanna Steer ◽  
Carol Heath ◽  
Kerry Taylor ◽  
Mark Bentley ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Messenger Rna ◽  
Myeloid Leukemia ◽  
Kinase Inhibitor ◽  
Growth Factor Receptor ◽  
Chronic Phase ◽  
Mek Inhibitor ◽  
Farnesyltransferase Inhibitors ◽  
Rt Pcr ◽  
Comparable Rate

Abstract This report describes 2 patients with a clinical and hematologic diagnosis of chronic myeloid leukemia (CML) in chronic phase who had an acquired t(8;22)(p11;q11). Analysis by fluorescence in situ hybridization (FISH) and reverse transcription-polymerase chain reaction (RT-PCR) indicated that both patients were negative for the BCR-ABL fusion, but suggested that the BCR gene was disrupted. Further FISH indicated a breakpoint within fibroblast growth factor receptor 1 (FGFR1), the receptor tyrosine kinase that is known to be disrupted in a distinctive myeloproliferative disorder, most commonly by fusion to ZNF198. RT-PCR confirmed the presence in both cases of an in-frame messenger RNA fusion between BCR exon 4 and FGFR1 exon 9. Expression of BCR-FGFR1 in the factor-dependent cell line Ba/F3 resulted in interleukin 3-independent clones that grew at a comparable rate to cells transformed with ZNF198-FGFR1. The growth of transformed cells was inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002, the farnesyltransferase inhibitors L744832 and manumycin A, the p38 inhibitors SB202190 and SB203580 but not by the MEK inhibitor PD98059. The growth of BaF3/BCR-FGFR1 and BaF3/ZNF198-FGFR1 was not significantly inhibited by treatment with STI571, but was inhibited by SU5402, a compound with inhibitory activity against FGFR1. Inhibition with this compound was associated with decreased phosphorylation of ERK1/2 and BCR-FGFR1 or ZNF198-FGFR1, and was dose dependent with an inhibitory concentration of 50% of approximately 5 μM. As expected, growth of BaF3/BCR-ABL was inhibited by STI571 but not by SU5402. The study demonstrates that the BCR-FGFR1 fusion may occur in patients with apparently typical CML. Patients with constitutively active FGFR1 fusion genes may be amenable to treatment with specific FGFR1 inhibitors.

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