scholarly journals SKIP Is Underexpressed in AML Leading to Sphingosine Kinase Hypofunction

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5324-5324
Author(s):  
Essam Ghazaly ◽  
Paul Smith ◽  
Chathunissa Gnanaranjan ◽  
Bryan D Young ◽  
Simon Joel ◽  
...  
Keyword(s):  
Cell Lines ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Annexin V ◽  
Leukemic Cell ◽  
Sphingosine Kinase ◽  
Fold Increase ◽  
Peripheral Blood Mononuclear ◽  
Leukemic Cell Lines ◽  
Intracellular Concentrations

Abstract Background: Sphingosine kinase interacting protein (SKIP) has been shown to be mostly silenced by hypermethylation in AML [1]. SKIP interacts with and regulates the function of sphingosine kinase (SK) enzyme. SK activity results in phosphorylation of sphingosine (SPH) to form sphingosine 1 phosphate (S1P), which promotes cell survival and resistance to apoptosis. On the other hand, S1P precursors ceramide (CER) and SPH mediate antiproliferative and apoptotic responses. SKIP has been reported to negatively regulate SK1 activity in fibroblasts. Therefore, we investigated the consequences of SKIP silencing in primary AML cells. In addition, we studied the effects of SKIP re-expression in leukemic cell lines. Methods: CTS and K562 cells were transfected with SKIP gene using standard techniques. SKIP is normally silenced in both cell lines. Intracellular and extracellular S1P, SPH and CER were measured by UPLC-MS/MS. In addition, intracellular SK activity was determined based on C17 S1P production from C17 SPH substrate. Chemosensitivity to doxorubicin, Imatinib and Ara-C in transfected cells was also studied. Results: In Primary AML cells, intracellular S1P and CER concentrations were reduced compared to G-CSF-mobilized peripheral blood mononuclear cells. In addition, SK activity was found to be downregulated in AML primary cells. When we transfected leukemic cell lines with SKIP gene, S1P and CER showed at least 2 fold increase in intracellular and extracellular basal levels compared to vector alone control (Figure 1). Further studies confirmed a significant increase in intracellular SK activity in SKIP transfected compared to vector alone cells, based on C17 S1P production (8.8 ± 2.6 vs 1.4 ± 0.4 ng/106 cells respectively after 24 hrs, p< 0.05). This increase in S1P and CER was associated with increasing apoptotic signals as evidenced by Annexin V staining and cleaved PARP expression. Moreover, chemosensitivity to Imatinib and AraC was significantly increased in SKIP transfected cell lines. These experiments confirm the regulation of SK1 function by SKIP. Conclusion: These data indicate that SKIP is downregulated in AML leading to reduced SK activity, which ultimately inhibits the apoptosis response. Figure 1. Effect of SKIP transfection on intracellular concentrations of S1P. Figure 1. Effect of SKIP transfection on intracellular concentrations of S1P. 1. Saied, M.H., et al., Genome wide analysis of acute myeloid leukemia reveal leukemia specific methylome and subtype specific hypomethylation of repeats. PLoS One, 2012. 7(3): p. e33213. EAG and PS contributed equally JG and DT contributed equally Disclosures No relevant conflicts of interest to declare.

scholarly journals Preclinical Antileukemia Activity of Tramesan: A Newly Identified Bioactive Fungal Metabolite

2017 ◽  
Vol 2017 ◽  
pp. 1-8 ◽  
Author(s):  
M. R. Ricciardi ◽  
R. Licchetta ◽  
S. Mirabilii ◽  
M. Scarpari ◽  
A. Parroni ◽  
...  
Keyword(s):  
Cell Lines ◽  
Molecular Mechanisms ◽  
Mononuclear Cells ◽  
Leukemic Cell ◽  
Bioactive Compound ◽  
Primary Cells ◽  
Peripheral Blood Mononuclear ◽  
Normal Peripheral Blood ◽  
Leukemic Cell Lines

Despite improvements that occurred in the last decades in the acute myeloid leukemia (AML) treatment, clinical results are still unsatisfactory. More effective therapies are required, and innovative approaches are ongoing, including the discovery of novel antileukemia natural compounds. Several studies have described the activity of extracts from mushrooms which produce compounds that exhibited immunological and antitumor activities. The latter has been demonstrated to be promoted in vitro by mushroom polysaccharides via induction of apoptosis. However, the antileukemia activity of these compounds on primary cells is still not reported. In the present study, we examined the in vitro effects of Tramesan (TR), a bioactive compound extracted from Trametes versicolor, on leukemic cell lines and primary cells. Our results demonstrated that TR induced a marked growth inhibition of leukemic cell lines and primary cells from AML patients. The antiproliferative effects of TR were associated in primary AML cells with a significant increase of apoptosis. No significant cytotoxic effects were observed in normal peripheral blood mononuclear cells (MNC) from healthy donors. Our data demonstrated a cytotoxic activity of TR on leukemia cells prompting further translational applications. Ongoing studies are elucidating the molecular mechanisms underlying its antileukemic activity.

Regulation of prolactin expression in leukemic cell lines and peripheral blood mononuclear cells

2003 ◽  
Vol 135 (1-2) ◽  
pp. 107-116 ◽  
Author(s):  
Sarah Gerlo ◽  
Wim Vanden Berghe ◽  
Peggy Verdood ◽  
Elizabeth L Hooghe-Peters ◽  
Ron Kooijman
Keyword(s):  
Cell Lines ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Leukemic Cell ◽  
Peripheral Blood Mononuclear ◽  
Leukemic Cell Lines ◽  
Blood Mononuclear Cells

A Novel Peptide Derived from Ginger Induces Apoptosis through the Modulation of p53, BAX, and BCL2 Expression in Leukemic Cell Lines

Planta Medica ◽  
2021 ◽  
Author(s):  
Chawalit Chatupheeraphat ◽  
Sittiruk Roytrakul ◽  
Narumon Phaonakrop ◽  
Kamolchanok Deesrisak ◽  
Sucheewin Krobthong ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Mononuclear Cells ◽  
Raji Cell ◽  
Leukemic Cell ◽  
B Cell Lymphoma ◽  
Leukemic Cells ◽  
Peripheral Blood Mononuclear ◽  
Normal Peripheral Blood ◽  
Peptide Fraction

AbstractDespite the efficacy of chemotherapy, the adverse effects of chemotherapeutic drugs are considered a limitation of leukemia treatment. Therefore, a chemotherapy drug with minimal side effects is currently needed. One interesting molecule for this purpose is a bioactive peptide isolated from plants since it has less toxicity to normal cells. In this study, we extracted protein from the Zingiber officinale rhizome and performed purification to acquire the peptide fraction with the highest cytotoxicity using ultrafiltration, reverse-phase chromatography, and off-gel fractionation to get the peptide fraction that contained the highest cytotoxicity. Finally, a novel antileukemic peptide, P2 (sequence: RALGWSCL), was identified from the highest cytotoxicity fraction. The P2 peptide reduced the cell viability of NB4, MOLT4, and Raji cell lines without an effect on the normal peripheral blood mononuclear cells. The combination of P2 and daunorubicin significantly decreased leukemic cell viability when compared to treatment with either P2 or daunorubicin alone. In addition, leukemic cells treated with P2 demonstrated increased apoptosis and upregulation of caspase 3, 8, and 9 gene expression. Moreover, we also examined the effects of P2 on p53, which is the key regulator of apoptosis. Our results showed that treatment of leukemic cells with P2 led to the upregulation of p53 and Bcl-2-associated X protein, and the downregulation of B-cell lymphoma 2, indicating that p53 is involved in apoptosis induction by P2. The results of this study are anticipated to be useful for the development of P2 as an alternative drug for the treatment of leukemia.

Pioglitazone Inhibits the Growth of Human Leukemic Cell Lines and Primary Leukemic Cells in Vitro.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4493-4493 ◽  
Author(s):  
Yoshihiro Hatta ◽  
Minoru Saiki ◽  
Yuko Enomoto ◽  
Shin Aizawa ◽  
Umihiko Sawada ◽  
...  
Keyword(s):  
Cell Lines ◽  
Colony Formation ◽  
Mononuclear Cells ◽  
Hematopoietic Cells ◽  
Leukemic Cell ◽  
Leukemic Cells ◽  
Peroxisome Proliferator ◽  
Leukemic Cell Lines ◽  
Ppar Γ

Abstract Troglitazone and pioglitazone are one of thiazolidinediones that are high affinity ligand for the nuclear receptor called peroxisome proliferator-activated receptor gamma (PPAR-γ). Troglitazone is a potent inhibitor of clonogenic growth of acute myeloid leukemia cells when combined with a retinoid. However, the effect of pioglitazone to neoplastic cells and normal hematopoietic cells has not been studied yet. Adult T-cell leukemia (ATL), prevalent in western Japan, is a highly aggressive malignancy of mature T lymphocyte. Therefore, we studied antitumor effect of pioglitazone against leukemic cells including ATL as well as normal hematopoietic cells. With 300 μM of pioglitazone, colony formation of ATL cell lines (MT1, MT2, F6T, OKM3T, and Su9T01) was completely inhibited. Colony formation of HUT102, another ATL cell line, was 12 % compared to untreated control. Clonogenic cells of other leukemic cell lines (K562, HL60, U937, HEL, CEM, and NALM1) was also inhibited to 0–30% of control. Colony formation of primary leukemic cells from 5 AML patients was decreased to 15 %. However, normal hematopoietic cells were weakly inhibited with 300 μM pioglitazone; 77 % of CFU-GM, 70 % of CFU-E, and 33 % of BFU-E survived. Cell cycle analysis showed that pioglitazone decreased the ratio of G2/M phase in HL60 cells, suggesting the inhibition of cell division. By Western blotting, PPAR-γ protein level was similar in all leukemic cells and normal bone marrow mononuclear cells. Taken together, pioglitazone effectively eliminate leukemic cells and could be used as an antitumor agent in vivo.

Genome-Wide Identification of Human Micrornas Located in Leukemia-Associated Genomic Alterations.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1287-1287
Author(s):  
Daniel T. Starczynowski ◽  
Ryan Morin ◽  
Andrew McPherson ◽  
Martin Hirst ◽  
Marco A. Marra ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Cell Lines ◽  
Small Rna ◽  
Conflicts Of Interest ◽  
Leukemic Cell ◽  
Fragile Sites ◽  
Small Rna Sequencing ◽  
Genomic Alterations ◽  
Aberrant Expression ◽  
Leukemic Cell Lines

Abstract Abstract 1287 Poster Board I-309 Cytogenetic alterations, such as amplifications, deletions, or translocations, contribute to myeloid malignancies. MicroRNAs (miRNAs) have emerged as critical regulators of hematopoietic processes and their aberrant expression has been associated with various leukemias. Genomic regions containing sequence alterations and fragile sites in human and mouse cancers are enriched with miRNA genes, however the potentially relevant miRNAs within these regions of genomic instability have not been evaluated on a global basis. Here we investigated miRNAs relevant to acute myeloid leukemia (AML) by: 1) mapping miRNAs within leukemia-associated genomic alterations in models of human AML by high-resolution genome arrays, and 2) evaluated absolute expression of these miRNAs by deep small RNA sequencing. We determined ∼75% (542/706) of miRNAs mapped to leukemia-associated copy-number alterations (CNA) in the cell lines, however, only 20% (99/542) of these miRNAs are expressed at levels above background. Small RNA sequencing allowed us to also identify 28 putative novel miRNAs, 18 of which map to leukemia-associated CNA in the cell lines. Our detailed genomic and small RNA analysis analysis of human leukemic cell lines has identified a subset of leukemia-associated miRNAs warranting further validation. Disclosures No relevant conflicts of interest to declare.

scholarly journals SIRT1 Expression Is Higher in Human Follicular Lymphoma and Correlates with Ki67 and Bcl-6 Overexpression

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1649-1649
Author(s):  
Raffaele Frazzi ◽  
Marco Tigano ◽  
Riccardo Valli ◽  
Ione Tamagnini ◽  
Valentina Fragliasso ◽  
...  
Keyword(s):  
B Lymphocytes ◽  
Histone Deacetylases ◽  
Mononuclear Cells ◽  
Conflicts Of Interest ◽  
Tissue Bank ◽  
Fold Increase ◽  
Proliferation Index ◽  
Gene Promoters ◽  
Peripheral Blood Mononuclear ◽  
Real Time Quantitative Pcr

Abstract Introduction. SIRT1 is a histone/protein deacetylase belonging to the family of Sirtuins and is the human homologue of the yeast Sir2 enzyme1. Histone deacetylases (HDACs) play a key role in several cancer-related mechanisms and the evidences of the efficacy of HDACs inhibitors in the treatment of lymphomas is rapidly growing2. SIRT1 binds and deacetylates the master transcriptional regulator Bcl-6 with implications in the pathogenesis of diffuse large B-cell lymphoma3. Ki67 is the proliferation index and is a diagnostic and useful marker currently used in the pathological evaluation of lymphoma samples4. Our aim is the characterization of SIRT1 in the B lymphocytes derived from follicular lymphomas (FL) or follicular hyperplasias and their comparison to Ki67, Bcl-6 and HIC-1. Methods. Immunohistochemical stainings (IHC) have been performed on a total of 61 formalin fixed paraffin embedded (FFPE) tissue sections including 36 FL and 25 follicular hyperplasias. All these FFPE samples were stained also for the proliferation index Ki67/MIB1. HIC-1 was also evaluated on 36 FL and 17 follicular hyperplasias. IHCs have been evaluated independently by two pathologists. B lymphocytes were immunomagnetically sorted starting from frozen biopsies collected in a Tissue Bank. Flow cytometry was used to determine the phenotype of the purified B populations. Genomic DNA and total RNA were extracted from purified B lymphocytes. Real-time, quantitative PCR (qPCR) was performed with Evagreen. qPCR was also performed on 4 samples of peripheral blood mononuclear cells (PBMCs) used as a reference. Data analysis and statistics were performed with GraphPad Prism 5. Results. IHCs show that SIRT1 localizes preferentially in the centroblasts of the follicles. In the 36 FL analyzed, SIRT1 is expressed in 33/36 cases (91.7%) and is stronger in the centroblasts of 23/36 cases (63.9%). Interestingly, the Ki67 staining correlates positively with SIRT1 in the FL where SIRT1 localizes in the centroblasts. However, SIRT1 positivity does not correlate with FL grade in our cohort. HIC1 reactivity in the same cohorts of samples resulted only in a weak staining of the stroma but the lymphocytes were negative. In order to investigate the expression levels of the two transcriptional variants of SIRT1 (SIRT1_001 and SIRT1_002) and Bcl-6 in the tumoral lymphocytes, we sorted and purified B lymphocytes from a total of 24 biopsies of FL and 10 follicular hyperplasias. qPCR revealed that FL have a statistically significant higher SIRT1_001 expression when compared to follicular hyperplasias and PBMCs. Furthermore, the FL having a Bcl-6 fold increase higher than 10 have also SIRT1_001 significantly higher than the FL with a Bcl-6 fold increase less than 10. Conclusions. Collectively, our data show that Ki67 positivity correlates with SIRT1 localization in the nuclei of the centroblasts. Interestingly, the transcriptional isoform SIRT1_001 is higher in the B lymphocytes of FL than in follicular hyperplasias and seems also to be positively correlated to Bcl-6 overexpression in FL samples. Our investigations are now aimed at defining the significance of these data in terms of epigenetic regulation of key gene promoters in the tumoral B lymphocytes. 1 Heltweg, B. et al.Cancer research66, 4368-4377, doi:10.1158/0008-5472.CAN-05-3617 (2006). 2 Murawski, N. & Pfreundschuh, M. The lancet oncology11, 1074-1085, doi:10.1016/S1470-2045(10)70210-2 (2010). 3 Liu, T., Liu, P. Y. & Marshall, G. M. Cancer research69, 1702-1705, doi:10.1158/0008-5472.CAN-08-3365 (2009). 4 Yamamoto, E. et al.Cancer science104, 1670-1674, doi:10.1111/cas.12288 (2013). Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Study On the Expression of 14-3-3ζ in Acute Myeloid Leukemic Cell Lines

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4820-4820
Author(s):  
Rong Liang ◽  
Hua Xiong ◽  
Zhe Wang ◽  
Qun xie Chen ◽  
Xun guang Gao ◽  
...  
Keyword(s):  
Cell Lines ◽  
Laser Scanning ◽  
Conflicts Of Interest ◽  
Leukemic Cell ◽  
Subcellular Location ◽  
Multidrug Resistant ◽  
Immunofluorescence Assay ◽  
Leukemic Cell Lines ◽  
Mdr1 Mrna ◽  
Acute Myeloid

Abstract Abstract 4820 Objectives: It is known that one of main reasons of refractory and relapsed acute myeloid leukemia (AML) is multidrug resistant (MDR). Despite it was showed that 14-3-3ζ was over- expressed in HL-60/VCR MDR cells than in HL-60 sensitive AML cells by examining the difference of gene expression profile with Affymetrix GeneChip Human Genome U133 set A. Yet, the understanding on role of 14-3-3ζ in the survival of AML cells remains poor. Methods: Semi-quantitative RT-PCR method was used to examine the expression of mdr1 mRNA in AML cell lines to validate the results of microarray. Western blot was performed to investigate Pgp#x2610;A14-3-3ζ#x2610;ABCL-2#x2610;AMCL-1 proteins expression. Immunofluorescence assay was used to detect the subcellular location of 14-3-3ζ protein in HL-60 and HL-60/VCR cells by laser scanning confocal microscope. 14-3-3ζ knockdown cells were obstained by transduction with lentivirus-mediated shRNA to silence 14-3-3ζ in AML cell lines. MTT and cell count method were used to analyze the changes of growth of AML cells. Results: mdr1 mRNA and Pgp was not expressed in HL-60 cells, but significantly overexpressed in HL-60/VCR cells(P<0.01). Except 14-3-3σ, the expression of other subtypes of 14-3-3 was higher in HL-60/VCR cells than in HL-60 cells, especially 14-3-3ζ(P<0.01). The higher increased expression of 14-3-3ζ, BCL-2, MCL-1 protein was observed in HL-60/VCR cells than in HL-60 cells. These results were same as gene chip. It was also noticed that14-3-3ζ was located in the cytoplasma and nuclear of AML cell lines, especially over-expressed in HL-60/VCR cells(P<0.05). Furthermore, suppression of14-3-3ζ by RNA interference resulted in inhibition of the proliferation of AML cells with BCL-2 and MCL-1 decreased protein expression, especially in HL-60/VCR cells(P<0.01). Conclusion: It was implied that14-3-3ζ played an important role in AML MDR and was associated with BCL-2 and MCL-1 expression. These results suggested that development of therapy targeting 14-3-3ζ may provide novel, effective strategies for refractory and relapsed AML. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Melittin—A Natural Peptide from Bee Venom Which Induces Apoptosis in Human Leukaemia Cells

Biomolecules ◽  
2020 ◽  
Vol 10 (2) ◽  
pp. 247 ◽  
Author(s):  
Michal Ceremuga ◽  
Maksymilian Stela ◽  
Edyta Janik ◽  
Leslaw Gorniak ◽  
Ewelina Synowiec ◽  
...  
Keyword(s):  
Cell Lines ◽  
Mononuclear Cells ◽  
Complex Mixture ◽  
Annexin V ◽  
Dry Mass ◽  
Bee Venom ◽  
Leukaemia Cell ◽  
Peripheral Blood Mononuclear ◽  
Dna Damages ◽  
Vast Number

Bee venom is a very complex mixture produced and secreted by the honeybee (Apis mellifera). Melittin is a major component of bee venom that accounts for about 52% of its dry mass. A vast number of studies have been dedicated to the effects of melittin’s regulation of apoptosis and to the factors that induce apoptosis in various types of cancer such as breast, ovarian, prostate, lung. The latest evidence indicates its potential as a therapeutic agent in the treatment of leukaemia. The aim of our present study is to evaluate melittin’s ability to induce apoptosis in leukaemia cell lines of different origin acute lymphoblastic leukaemia (CCRF-CEM) and chronic myelogenous leukaemia (K-562). We demonstrated that melittin strongly reduced cell viability in both leukaemia cell lines but not in physiological peripheral blood mononuclear cells (PMBCs). Subsequent estimated parameters (mitochondrial membrane potential, Annexin V binding and Caspases 3/7 activity) clearly demonstrated that melittin induced apoptosis in leukaemia cells. This is a very important step for research into the development of new potential anti-leukaemia as well as anticancer therapies. Further analyses on the molecular level have been also planned (analysis of proapoptotic genes expression and DNA damages) for our next research project, which will also focus on melittin.

A Novel and Highly Selective Dual PI3K/mTOR Kinase Inhibitor VS-5584, Shows Promising Therapeutic Potential For The Treatment Of Multiple Myeloma

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4433-4433
Author(s):  
Nurulhuda Mustafa ◽  
Stefan Hart ◽  
Wee-Joo Chng
Keyword(s):  
Multiple Myeloma ◽  
Cell Lines ◽  
Mononuclear Cells ◽  
Kinase Inhibitor ◽  
Conflicts Of Interest ◽  
Therapeutic Potential ◽  
Synergistic Activity ◽  
Peripheral Blood Mononuclear ◽  
Wide Range

Currently, most multiple myeloma (MM) patients experience relapse and develop resistance to standard treatments. A recent study showed that patients who relapsed had poor outcomes, with an overall survival of only 6 months and an event-free survival of 1 month. The PI3K/mTOR/AKT pathway represents a critical target in MM because it stimulates proliferation, survival, and drug resistance of MM cells. VS-5584 is a novel agent, with specific and equipotent activity against mTOR and all 4 Class I PI3K isoforms, without relevant activity on 400 other lipid and protein kinases. Here we report that VS-5584 is highly efficacious against a wide panel of MM cells including Velcade- and Doxorubucin- resistant cell lines. This efficacy is maintained even in the presence of additional MM growth factors, IL-6 and IGF-1, and seems independent of PTEN status in the cell lines. Importantly, VS-5584 shows similar efficacy in patient myeloma cells and preferential tumor cell targeting compared to healthy peripheral blood mononuclear cells. Further testing in a myeloma xenograft mouse model further confirmed the potency of this compound in vivo. We have also observed synergistic activity in combination with both MM clinical therapeutic Dexamethasone, and novel anti-MM candidate Panabinostat. Comparing the basal expression profile of hypersensitive (H929) vs less sensitive (OPM2, U266) cell lines have identified the interferon alpha/beta pathway as a marker for association with sensitivity. Just recently, VS-5584 has been reported to evidence very favourable pharmaceutical and pharmacological properties in a wide range of solid tumors resistant to standard care therapies. Taken together with our data, this offers a compelling rationale for its clinical development as a single or combination therapy in multiple myeloma. Disclosures: No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document