p53 in chronic myelogenous leukemia. Study of mechanisms of differential expression.

The p53 is a nuclear protein that is associated with normal cellular proliferation and can cooperate with Ha-ras in causing cellular transformation in vitro. Lineage association is known to exist between p53 expression and normal lymphopoiesis, but not myelopoiesis. We studied the expression of p53 using chronic myelogenous leukemia (CML) cell lines, somatic hybrids of these cells, and leukemic cells from CML patients. Lymphoid CML lines expressed both p53 mRNA and protein. We also analyzed p53 synthesis by two B-lymphoid lines from the same CML patient; cells of one line were derived from the neoplastic clone, cells of the other were derived from the normal clone. Both synthesized equal amounts of a phosphorylated p53 protein. None of the myeloid CML lines expressed detectable p53 protein and two of four expressed negligible p53 mRNA. Two other myeloid CML lines and myeloid cells from three of four patients expressed p53 mRNA. These findings suggest that expression of the gene is not regulated normally in CML. Several approaches were pursued to explore the differential expression of p53. Southern blot analyses showed no gross alterations in the p53 gene from cells of either the expressing or the nonexpressing lines. No difference in the pattern of demethylated CpG sites was noted in the region of the p53 gene in cells from K562 (myeloid p53 nonexpressor) and in BV173 (lymphoid p53 expressor). The sites of demethylation clustered in and around the p53 promoter in both cell lines. Somatic hybrids formed between a p53 mRNA nonexpressor myeloid line (K562) and the parental p53 expressor lymphoid lines (Daudi, PUT) produced p53 mRNA and protein, suggesting that p53 is a dominantly expressed protein and that lack of expression in myeloid cells is not mediated by a trans-acting negative regulatory protein. DNA transfection experiments performed using the indicator gene chloramphenicol acetyltransferase attached to promoter sequences of p53 showed that these constructs were equally activated in BV173 (p53 expressor) and K562 (p53 mRNA nonexpressor). The mechanism of p53 regulation in CML remains unclear.

Download Full-text

Inhibition of polyamine synthesis induces p53 gene expression but not apoptosis

AJP Cell Physiology ◽

10.1152/ajpcell.1999.276.4.c946 ◽

1999 ◽

Vol 276 (4) ◽

pp. C946-C954 ◽

Cited By ~ 46

Author(s):

Li Li ◽

Ji Li ◽

Jaladanki N. Rao ◽

Minglin Li ◽

Barbara L. Bass ◽

...

Keyword(s):

Gene Expression ◽

Growth Inhibition ◽

P53 Protein ◽

Specific Inhibitor ◽

P53 Gene ◽

Mrna Levels ◽

Intestinal Epithelial ◽

Polyamine Biosynthesis ◽

Fetal Bovine ◽

P53 Mrna

The nuclear phosphoprotein p53 acts as a transcription factor and is involved in growth inhibition and apoptosis. The present study was designed to examine the effect of decreasing cellular polyamines on p53 gene expression and apoptosis in small intestinal epithelial (IEC-6) cells. Cells were grown in DMEM containing 5% dialyzed fetal bovine serum in the presence or absence of α-difluoromethylornithine (DFMO), a specific inhibitor of polyamine biosynthesis, for 4, 6, and 12 days. The cellular polyamines putrescine, spermidine, and spermine in DFMO-treated cells decreased dramatically at 4 days and remained depleted thereafter. Polyamine depletion by DFMO was accompanied by a significant increase in expression of the p53 gene. The p53 mRNA levels increased 4 days after exposure to DFMO, and the maximum increases occurred at 6 and 12 days after exposure. Increased levels of p53 mRNA in DFMO-treated cells were paralleled by increases in p53 protein. Polyamines given together with DFMO completely prevented increased expression of the p53 gene. Increased expression of the p53 gene in DFMO-treated cells was associated with a significant increase in G1 phase growth arrest. In contrast, no features of programmmed cell death were identified after polyamine depletion: no internucleosomal DNA fragmentation was observed, and no morphological features of apoptosis were evident in cells exposed to DFMO for 4, 6, and 12 days. These results indicate that 1) decreasing cellular polyamines increases expression of the p53 gene and 2) activation of p53 gene expression after polyamine depletion does not induce apoptosis in intestinal crypt cells. These findings suggest that increased expression of the p53 gene may play an important role in growth inhibition caused by polyamine depletion.

Download Full-text

Liposomal delivery of methylphosphonate antisense oligodeoxynucleotides in chronic myelogenous leukemia [see comments]

Blood ◽

10.1182/blood.v84.2.601.601 ◽

1994 ◽

Vol 84 (2) ◽

pp. 601-607 ◽

Cited By ~ 35

Author(s):

AM Tari ◽

SD Tucker ◽

A Deisseroth ◽

G Lopez-Berestein

Keyword(s):

Growth Inhibition ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Fusion Gene ◽

Hematologic Malignancy ◽

Control Cell ◽

Selective Inhibition ◽

Inhibitory Effect ◽

Myelogenous Leukemia ◽

Ph Chromosome

Abstract Chronic myelogenous leukemia (CML) is a hematologic malignancy characterized by the presence of the Philadelphia (Ph) chromosome. Bcr- abl, the fusion gene associated with the Ph chromosome, expresses a p210bcr-abl protein that promotes a selective expansion of mature myeloid progenitor cells. Methylphosphonate (MP) oligodeoxynucleotides complementary to specific regions of the bcr-abl mRNA were incorporated in liposomes. We studied the effects of liposomal MP (L-MP) on the growth inhibition of CML-like cell lines. L-MP targeted to the breakpoint junctions of the bcr-abl mRNA inhibited the growth of CML cells. Fifty percent inhibition was achieved at approximately 1 mumol/L of L-MP oligonucleotide concentrations. The inhibitory effect was selective because growth inhibition was observed only with CML but not with control cell lines. Moreover, CML cell growth inhibition was dependent on the sequence of the MP oligodeoxynucleotides incorporated in the liposomes. The growth inhibition of CML cells by L-MP resulted from selective inhibition of the expression of the p210bcr-abl protein.

Download Full-text

Terminal deoxynucleotidyl transferase activities in human blood leukocytes and lymphoblast cell lines: high levels in lymphoblast cell lines and in blast cells of some patients with chronic myelogenous leukemia in acute phase

Blood ◽

10.1182/blood.v47.1.11.11 ◽

1976 ◽

Vol 47 (1) ◽

pp. 11-20 ◽

Cited By ~ 126

Author(s):

PS Sarin ◽

PN Anderson ◽

RC Gallo

Keyword(s):

T Cell ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Human Blood ◽

Lymphocytic Leukemia ◽

Myelogenous Leukemia ◽

Thymus Gland ◽

Terminal Deoxynucleotidyl Transferase ◽

Blast Phase ◽

Blast Cells

Abstract Terminal deoxynucleotidyl transferase, an enzyme which catalyzes the polymerization of deoxyribonucleoside triphosphates, elongating oligo- or polydeoxynucleotide chains, but without direction from a nucleic acid template, is thought to be specific for thymus gland and thymus- derived cells. We have confirmed the observations that high levels are characteristic of thymus gland with both human and calf tissue and that elevated levels may be found in some cases of acute lymphocytic leukemia. High levels were also found in human lymphoblast cell lines with T-cell characteristics, and insignificant activity was observed in leukocytes of patients with chronic myelogenous leukemia not in acute blast phase of the disease, chronic lymphocytic leukemia, human B- cells, and normal human blood lymphocytes even after stimulation with phytohemagglutinin. However, high levels (approximately 200 nmoles/hr/10(9) cells) equivalent to those in thymus tissue and lymphoblast cell lines with T-cell characteristics were found in the peripheral blood blast cells of four patients with chronic myelogenous leukemia in an acute blast phase of their disease. One hypothesis that may explain the present results is that in chronic myelogenous leukemia in acute blast phase of the disease the proliferative blast response may not always be myeloblasts but in some cases it may be lymphoblasts.

Download Full-text

eIF3b regulates the cell proliferation and apoptosis processes in chronic myelogenous leukemia cell lines via regulating the expression of C3G

Biotechnology Letters ◽

10.1007/s10529-020-02878-1 ◽

2020 ◽

Vol 42 (7) ◽

pp. 1275-1286

Author(s):

Laiquan Huang ◽

Zhongling Wei ◽

Xiangxiang Chang ◽

Xinyuan Zheng ◽

Jiawei Yan ◽

...

Keyword(s):

Cell Proliferation ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Leukemia Cell ◽

Myelogenous Leukemia ◽

Chronic Myelogenous Leukemia Cell ◽

Leukemia Cell Lines ◽

Proliferation And Apoptosis

Download Full-text

Constitutive expression of SOCS3 confers resistance to IFN-α in chronic myelogenous leukemia cells

Blood ◽

10.1182/blood-2002-01-0073 ◽

2002 ◽

Vol 100 (8) ◽

pp. 2926-2931 ◽

Cited By ~ 67

Author(s):

Ikuya Sakai ◽

Kazuto Takeuchi ◽

Hayato Yamauchi ◽

Hirosi Narumi ◽

Shigeru Fujita

Keyword(s):

Cell Line ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Blast Crisis ◽

Constitutive Expression ◽

Myelogenous Leukemia ◽

Cytokine Signaling ◽

Interferon Α ◽

Socs3 Expression ◽

Blast Cells

Because suppressor of cytokine signaling (SOCS) proteins are negative regulators of cytokine-induced signaling, it has been hypothesized that aberrant SOCS expression confers resistance against cytokine therapy. This study reports on the constitutive expression of SOCS3 in most chronic myelogenous leukemia (CML) cell lines, which are resistant to treatment with interferon α (IFN-α). In contrast, the KT-1/A3 cell line, in which constitutive expression of SOCS3 is barely detectable, is sensitive to IFN-α treatment. Forced expression of SOCS3 in the KT-1/A3 cell line confers resistance to IFN-α treatment. Furthermore, most of the blast cells from patients in CML blast crisis, which are usually resistant to IFN-α therapy, showed constitutive expression of SOCS3. These findings indicate that constitutive SOCS3 expression affects the IFN-α sensitivity of CML cell lines and blast cells from patients with CML blast crisis.

Download Full-text

Mutation of the p53 gene in human acute myelogenous leukemia

Blood ◽

10.1182/blood.v77.7.1500.1500 ◽

1991 ◽

Vol 77 (7) ◽

pp. 1500-1507 ◽

Cited By ~ 75

Author(s):

JM Slingerland ◽

MD Minden ◽

S Benchimol

Keyword(s):

Acute Myelogenous Leukemia ◽

P53 Protein ◽

Point Mutations ◽

P53 Gene ◽

Myelogenous Leukemia ◽

Wild Type ◽

P53 Expression ◽

Coding Sequence ◽

Acute Myelogenous ◽

Blast Cells

Abstract Heterogeneity of p53 protein expression is seen in blast cells of patients with acute myelogenous leukemia (AML). p53 protein is detected in the blasts of certain AML patients but not in others. We have identified p53 protein variants with abnormal mobility on gel electrophoresis and/or prolonged half-life (t 1/2). We have sequenced the p53 coding sequence from primary blast cells of five AML patients and from the AML cell line (OCIM2). In OCIM2, a point mutation in codon 274 was identified that changes a valine residue to aspartic acid. A wild type p53 allele was not detected in these cells. Two point mutations (codon 135, cysteine to serine; codon 246, methionine to valine) were identified in cDNA from blasts of one AML patient. Both mutations were present in blast colonies grown from single blast progenitor cells, indicating that individual leukemia cells had sustained mutation of both p53 alleles. The cDNAs sequenced from blast samples of four other patients, including one with prolonged p53 protein t 1/2 and one with no detectable p53 protein, were fully wild type. Thus, the heterogeneity of p53 expression cannot be explained in all cases by genetic change in the p53 coding sequence. The prolonged t 1/2 of p53 protein seen in some AML blasts may therefore reflect changes not inherent to p53. A model is proposed in which mutational inactivation of p53, although not required for the evolution of neoplasia, would confer a selective advantage, favoring clonal outgrowth during disease progression.

Download Full-text

Chronic myelogenous leukemia blast cell proliferation is inhibited by peptides that disrupt Grb2-SoS complexes

Blood ◽

10.1182/blood.v98.6.1773 ◽

2001 ◽

Vol 98 (6) ◽

pp. 1773-1781 ◽

Cited By ~ 38

Author(s):

Christian Kardinal ◽

Birgit Konkol ◽

Hui Lin ◽

Manfred Eulitz ◽

Enrico K. Schmidt ◽

...

Keyword(s):

Map Kinase ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Protein Complexes ◽

Therapeutic Option ◽

Selective Inhibition ◽

Myelogenous Leukemia ◽

Blast Cells ◽

Signal Transduction Proteins ◽

Cell Fractions

Abstract Chronic myelogenous leukemia (CML) is commonly characterized by the presence of the p210Bcr-Abl oncoprotein. Many downstream effectors of Bcr-Abl have been described, including activation of the Grb2-SoS-Ras–MAP kinase (Erk) pathway. The precise contributions of these signal-transduction proteins in CML blast cells in human patients are not yet well defined. To gain further insight into the importance of Grb2 for CML, peptides that disrupt Grb2-SoS complexes were tested. These high-affinityGrb2-binding peptides (HAGBPs) can autonomously shuttle into cells and function by binding to the N-terminal SH3 domain of Grb2. The HAGBPs were analyzed for their effects on Bcr-Abl–expressing cell lines and freshly isolated CML blast cells from patients. They induced a dramatic decrease in the proliferation of CML cell lines. This was not observed with point-mutated control peptides with abolished Grb2SH3(N) binding. As expected, Grb2-SoS complexes were greatly diminished in the HAGBP-treated cells, and MAP kinase activity was significantly reduced as determined by an activation-specific phospho-MAPK antibody. Furthermore, cell fractions that are enriched for blast cells from CML patients with active disease were also incubated with the Grb2 blocker peptides. The HAGBPs led to a significant proliferation reduction of these cells in the majority of the isolates, but not in all patients' cells. These results show that, in addition to the direct targeting of Bcr-Abl, selective inhibition of Grb2 protein complexes may be a therapeutic option for a significant number of CML patients.

Download Full-text

A 41-kilodalton protein is a potential substrate for the p210bcr-abl protein-tyrosine kinase in chronic myelogenous leukemia cells.

Molecular and Cellular Biology ◽

10.1128/mcb.12.3.1312 ◽

1992 ◽

Vol 12 (3) ◽

pp. 1312-1323 ◽

Cited By ~ 6

Author(s):

E Freed ◽

T Hunter

Keyword(s):

Tyrosine Kinase ◽

Cell Lines ◽

Chronic Myelogenous Leukemia ◽

Protein Tyrosine Kinase ◽

Leukemia Virus ◽

Lymphoid Cells ◽

Myelogenous Leukemia ◽

Two Dimensional ◽

Kinase Gene ◽

Protein Tyrosine

Chronic myelogenous leukemia (CML) is characterized by a translocation involving the c-abl protein-tyrosine kinase gene. A chimeric mRNA is formed containing sequences from a chromosome 22 gene (bcr) at its 5' end and all but the variable exon 1 of c-abl sequence. The product of this mRNA, p210bcr-abl, has constitutively high protein-tyrosine kinase activity. We examined K562 cells and other lines established from CML patients for the presence of phosphotyrosine (P-Tyr)-containing proteins which might be p210bcr-abl substrates. Two-dimensional gel separation of 32P-labeled proteins followed by phosphoamino acid analysis of 25 phosphoproteins, which comprised the major alkali-stable phosphoproteins, indicated that three related proteins of 41 kDa are the most prominent P-Tyr-containing proteins detected by this method. The 41-kDa phosphoproteins are found in two other CML lines that we examined but not in lines of similar lineage isolated from patients with distinct leukemic disease. A protein that comigrates with the major form of pp41 (pp41A) and contains P-Tyr is also found in murine fibroblasts and B-lymphoid cells transformed by Abelson murine leukemia virus, which encodes the v-abl protein, and in platelet-derived growth factor-treated fibroblasts, in which it has been described previously. We analyzed three pairs of Epstein-Barr virus-immortalized B-cell lines from individual CML patients and found that only the lines in which active p210bcr-abl was present contained detectable pp41. We also performed immunoblotting with anti-P-Tyr antibodies on the same CML cell lines and detected at least four other putative substrates of p210bcr-abl, which were undetected with use of the two-dimensional gel technique.

Download Full-text