scholarly journals BCL-2 dependence and ABT-737 sensitivity in acute lymphoblastic leukemia

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 2300-2309 ◽  
Author(s):  
Victoria Del Gaizo Moore ◽  
Krysta D. Schlis ◽  
Stephen E. Sallan ◽  
Scott A. Armstrong ◽  
Anthony Letai
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Cancer Cells ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Single Agent ◽  
Primary Cells ◽  
Intrinsic Apoptotic Pathway ◽  
Combination Therapies ◽  
Apoptotic Pathway ◽  
Oncogene Activation

Cancer cells acquire disruptions in normal signal transduction pathways and homeostatic mechanisms that would trigger apoptosis in normal cells. These abnormalities include genomic instability, oncogene activation, and growth factor independent proliferation. Therefore, cancer cells likely require a block in apoptosis in order to survive. Overexpression of the antiapoptotic protein BCL-2 provides a block in apoptosis that is frequently observed in cancer cells. We have developed methods for the detection and analysis of BCL-2 dependence and here apply them to acute lymphoblastic leukemia (ALL). BH3 profiling, a mitochondrial assay that classifies blocks in the intrinsic apoptotic pathway, indicated a dependence on BCL-2 of both ALL cell lines and primary samples. This dependence predicted that BCL-2 would be complexed with select pro-death BH3 family proteins, a prediction confirmed by the isolation of BCL-2 complexes with BIM. Furthermore, the BH3 profiling and protein analysis predicted that ALL cell lines and primary cells would be sensitive to ABT-737 as a single agent. Finally, BH3 profiling and protein studies accurately predicted a relative degree of sensitivity to BCL-2 antagonism in cell lines. The ALL cells studied exhibit BCL-2 dependence, supporting clinical trials of BCL-2 antagonists in ALL as single agents or combination therapies.

Restraining the Proliferation of Acute Lymphoblastic Leukemia Cells by Genistein through Up-regulation of B-cell Translocation Gene-3 at Transcription Level

2019 ◽  
Vol 8 ◽  
Author(s):  
Masoumeh Abedi Nejad ◽  
Mohsen Nikbakht ◽  
Masoomeh Afsa ◽  
Kianoosh Malekzadeh
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cancer Cells ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Suppressor Gene ◽  
Transcription Level ◽  
Hematopoietic Malignancy ◽  
Proliferative Effect ◽  
Cancer Agent

Background: Acute lymphoblastic leukemia (ALL) is a highly prevalent pediatric cancer accounting for approximately 78% of leukemia cases in patients younger than 15 years old. Different studies have demonstrated that B-cell translocation gene 3 (BTG3) plays a suppressive role in the progress of different cancers. Genistein is considered a natural and biocompatible compound and a new anti-cancer agent. In this study, we evaluate the effect of genistein on BTG3 expression and proliferation of ALL cancer cells. Materials and Methods: ALL cell lines (MOLT4, MOLT17, and JURKAT) were cultured in standard conditions. Cytotoxicity of genistein was detected using MTT assay. The cells were treated with different concentrations of genistein (10, 25, 40, and 55μM) for 24, 48, and 72 hours, and then cell viability and growth rate were measured. The quantitative real-time polymerase chain reaction was applied to investigate the effect of genistein on BTG3 expression. Results: The percentage of vital cells treated with genistein significantly decreased compared to the non-treated cells, showed an inverse relationship with an increasing genistein concentration. The present study suggests a dose of 40μM for genistein as a potent anticancer effect. Genistein could elevate BTG3 for 1.7 folds in MOLT4 and JURKAT and 2.7 folds in MOLT17 cell lines at transcription level conveged with 60 to 90% reduction in the proliferation rate of cancer cells. Conclusion: Up-regulation of BTG3 as a tumor suppressor gene can be induced by genistein. It seems that BTG3 reactivation can be introduced as another mechanism of anti-proliferative effect of genistein and could be considered as a retardant agent candidate against hematopoietic malignancy.[GMJ. 2019;inpress:e1229]

The Inhibition of Checkpoint Kinase 1 As a Promising Strategy to Increase the Effectiveness of Different Treatments in Acute Lymphoblastic Leukemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2478-2478
Author(s):  
Andrea Ghelli Luserna Di Rora ◽  
Ilaria Iacobucci ◽  
Enrica Imbrogno ◽  
Enrico Derenzini ◽  
Anna Ferrari ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Acute Lymphoblastic Leukemia ◽  
Cell Viability ◽  
Cytotoxic Activity ◽  
Cell Lines ◽  
Propidium Iodide ◽  
Lymphoblastic Leukemia ◽  
Single Agent ◽  
Dna Damages ◽  
Checkpoint Kinase

Abstract Nowadays the effectiveness of the treatments for adult Acute Lymphoblastic Leukemia (ALL) patients is still inadequate and frequently many patients after years of response to treatments develop relapses. Thus there is a need to find novel targets for specific therapies and to maximize the effect of the actual treatments. Recently different Checkpoint Kinase (Chk)1/Chk2 inhibitors has been assessed for the treatment of different type of cancers but only few studies have been performed on hematological diseases. We evaluated the effectiveness of the Chk1 inhibitor, LY2606368, as single agent and in combination with tyrosine kinase inhibitors (imatinib and dasatinib) or with the purine nucleoside antimetabolite clofarabine in B-/T- acute lymphoblastic leukemia (ALL) cell lines and in primary blasts. Human B (BV-173, SUPB-15, NALM-6, NALM-19 and REH) and T (MOLT-4, RPMI-8402 and CEM) ALL cell lines were incubated with increasing concentrations of drug (1-100 nM) for 24 and 48 hours and the reduction of the cell viability was evaluated using WST-1 reagent. LY2606368 deeply reduced the cell viability in a dose and time dependent manner in all the cell lines, with the BV-173 (6.33 nM IC50 24hrs) and RPMI-8402 (8.07 nM IC50 24hrs) being the most sensitive while SUP-B15 (61.4 nM IC50 24hrs) and REH (96.7 nM IC50 24hrs) being the less sensitive cell lines. Moreover the sensitivity to the compound was no correlated with the different sub-type of ALL or with the mutational status of p53, which is a marker of the functionality of the G1/S checkpoint. The cytotoxic activity was confirmed by the significant increment of apoptosis cells (Annexin V/Propidium Iodide), by the increment of gH2AX foci and by the activation of different apoptotic markers (Parp-1 and pro-Caspase3 cleavage). To understand the relationship between the activation of apoptosis and the effect on cell cycle and to identify hypothetical mechanisms of death, different cell cycle analyses were performed (Propidium Iodide staining). The inhibition of Chk1, deeply changed the cell cycle profile. Indeed in all the cell lines the percentage of cells in S phase and in G2/M phase were reduced by the treatment while the numbers of cells in sub-G1 and G1 phase were increased. The hypothetical function of LY2606368 as a chemosensitizer agent was evaluated combining the compound with different drugs normally used in clinical trials. For each drugs the combination strongly reduced the cell viability when compared to the cytotoxic effect of the single drugs. Moreover the combination showed an additive efficacy in term of induction of DNA damages as showed by the increase number of gH2AX foci and the activation of pChk1 (ser 317). The results found on the cell lines were confirmed also using primary leukemic blast isolated from adult Philadelphia-positive ALL patients. Indeed LY2606368 as single agent or in combination with the Tki, imatinib, was able to deeply reduce the cell viability and to induce DNA damages (gH2AX foci). In conclusion LY2606368 showed a strong cytotoxic activity on B-/T-All cell lines and primary blasts as single agent and in combination with other drugs. In our opinion this data are the basis for a future clinical evaluation of this compound in the treatment of leukemia. Supported by ELN, AIL, AIRC, progetto Regione-Università 2010-12 (L. Bolondi), FP7 NGS-PTL project. Disclosures Soverini: Novartis, Briston-Myers Squibb, ARIAD: Consultancy. Cavo:JANSSEN, CELGENE, AMGEN: Consultancy. Martinelli:ROCHE: Consultancy; Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau; Pfizer: Consultancy; Ariad: Consultancy; AMGEN: Consultancy; MSD: Consultancy.

scholarly journals Suppressive Effect of Genistein on Childhood Cell Lineage Acute Lymphoblastic Leukemia by Inducing Dicer and AGO2

2020 ◽  
Author(s):  
Fatemeh Piroozian ◽  
Hoda Bagheri Varkiyani ◽  
Afshin Samiei ◽  
Amirhosein Bradaran Najar ◽  
Masoumeh Afsa ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Cancer Cells ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Mrna Level ◽  
Cell Lineage ◽  
Suppressive Effect ◽  
Inhibitory Effect ◽  
Childhood All ◽  
Proliferative Effect

Abstract Background Childhood Acute Lymphoblastic Leukemia (ALL) is one of the most prevalent malignancies. Dysregulation of microRNAs in some cancers suggests their role in pathogenesis of the disease. Dicer and AGO2, two factors participate in biogenesis of miRNAs can exert a crucial function in development of the cancers. On the other hand, it has been proved that, genistein has anticancer effects against some cancer cells. Methods In the present study, it was attempted to assess alteration in the mRNA expression of the Dicer and AGO2 genes, and then evaluating inhibitory effect of genistein on Dicer and AGO2 genes. Up-regulation of Dicer and down-regulation of AGO2 were observed in 40 patients with childhood ALL compared to 35 healthy controls. Results Alteration in the expression of these genes directed to a correlation with progression of the disease. Genistein had anti-proliferative effect against ALL cancer cells through increasing mortality rate via induction of apoptosis and decreasing growth rate of malignant cells. The genistein significantly increased the mRNA level of Dicer particularly in two cells (Molt-17 and Nalm-6). Up-regulation of AGO2 that occurred as a result of genistein administration was significant in the 4 cell lines compared to non-treated cells. Conclusions The concordance in alteration of AGO2 and Dicer mRNA expressions in B-ALL cell lines caused by genistein administration suggests existence of another mechanism of this compound as chemotherapeutic agent against ALL cell line.

AMN107, Novel Aminopyrimidine Inhibitor of Bcr-Abl, Is Significantly More Potent Than Imatinib Mesylate Against Philadelphia Chromosome Positive Acute Lymphoblastic Leukemia Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2737-2737
Author(s):  
Mirna Golemovic ◽  
Miloslav Beran ◽  
Francis Giles ◽  
Taghi Manshouri ◽  
Deborah Thomas ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Cell Line ◽  
Imatinib Mesylate ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Single Agent ◽  
Philadelphia Chromosome ◽  
Imatinib Treatment ◽  
Significant Activity ◽  
Mts Assay

Abstract Imatinib mesylate is effective against Philadelphia chromosome (Ph)-positive acute lymphoblastic leukemia (ALL) but, when used as a single agent, responses are transient and most patients relapse within 4–6 months. AMN107 is a novel oral aminopyrimidine ATP-competitive inhibitor of the protein tyrosine kinase activity of Bcr-Abl. Following oral administration to animals, AMN107 is well absorbed, has a good pharmacokinetic profile, and is well tolerated. The activity of AMN107, relative to imatinib, in both Ph-positive (Z-119 and Z-181) and Ph-negative (Z-138) ALL cell lines was studied. Z-119 and Z-181 cells were derived from Ph-positive ALL patients and retained typical B-cell characteristics and phenotypes of the original leukemia, including cytogenetic abnormality t(9;22) and p190 Bcr/Abl kinase. Z-138, a Ph-negative cell line, was derived from a patient with chronic lymphocytic leukemia and supervening ALL. Treatment with AMN107 or imatinib for 3 days (MTS assay) inhibited proliferation of Z-119 cells with the IC50 values of 19.3 nM and 620.0 nM, respectively, revealing AMN107 to be 32 fold more potent than imatinib. Treatment of Z-181 cell line lasted for 4 days (MTS assay) because of lower growth rate of these cells: IC50 for AMN107 and imatinib were 1.6 nM and 63.9 nM, respectively, showing AMN107 to be 40 fold more potent than imatinib. Neither drug showed activity against Ph-negative Z-138 cells. We also compared the activity of AMN107 in Ph-positive ALL cell lines expressing p190 Bcr/Abl protein to that in Ph-positive chronic myeloid leukemia cell lines KBM5 and KBM7 expressing p210 Bcr/Abl protein. The activity was similar with IC50 in KBM5 cells of 11.3 nM and in KBM7 cells of 4.3 nM. In experiments focused on cell cycle analysis we found that at equipotent doses (as determined by MTS assay) both drugs induced cell accumulation in G0/G1 phase in Z-119 but not in Z-181. We demonstrated that increasing equipotent concentrations of AMN107 and imatinib induced activation of caspase-3 that resulted in apoptosis, as assessed by propidium iodide staining, in Z-119 cells, while Z-181 cells showed lack of apoptotic response. Following treatment with a broad range of AMN107 and imatinib doses for 3 hrs, Bcr/Abl expression and phosphorylation were determined in Z-119 cells by immunoprecipitation and Western blotting: Bcr/Abl phosphorylation was inhibited completely with AMN107 at 125.0 nM, and with imatinib at 2500 nM, confirming again the higher potency of AMN107. Finally, similar differential effect of AMN107 and imatinib on Bcr/Abl protein expression and phosphorylation was observed in leukemic cells obtained from blood of Ph-positive ALL patients. We conclude that AMN107 has significant activity against Ph-positive ALL cells and warrants investigation in patients with Ph-positive ALL.

Targeting HMG-CoA Reductase in Acute Lymphoblastic Leukemia (ALL): Statins Inhibit Proliferation and Induce Apoptosis in ALL Cells

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2927-2927
Author(s):  
David Teachey ◽  
Cecilia Sheen ◽  
Alix Eden Seif ◽  
Valerie I. Brown ◽  
Stephan A. Grupp
Keyword(s):  
Signal Transduction ◽  
Acute Lymphoblastic Leukemia ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Cholesterol Biosynthesis ◽  
Large Population ◽  
Single Agent ◽  
Cytotoxic Agents ◽  
Hmg Coa Reductase ◽  
Coa Reductase

Abstract HMG-CoA reductase inhibitors (statins) are commonly used, FDA-approved agents for the treatment of hypercholesterolemia. In addition to reducing serum cholesterol, statins have been shown to have anti-inflammatory properties and anti-proliferative effects on a number of cell types, including lymphocytes. Many of these effects occur because inhibition of HMG-CoA reductase results in depletion of a number of important cellular intermediates required for signal transduction through the AKT, Ras, and Erk pathways. Statins have been demonstrated to have anti-proliferative effects in a number of malignancies, including acute myeloid leukemia using preclinical models; however, there are no published reports evaluating the efficacy of these agents against acute lymphoblastic leukemia. Large population studies have also suggested that statins may reduce the incidence of a number of cancers, furthering interest in the use of these agents in malignancies, especially since the dose-limiting toxicities (hepatitis and rhabdomyolysis) do not overlap with the majority of commonly used cytotoxic agents. We hypothesized that ALL cells are dependent on the function of HMG-CoA reductase for survival. We tested this hypothesis by studying the efficacy of statins against ALL cells. We studied the effects of 5 different compounds (mevastatin, pravastatin, fluvastatin, lovastatin, and simvastatin) against 7 ALL cell lines (4 human and 3 murine). We found that the more potent 2nd generation statins (fluvastatin, lovastatin, and simvastatin) had marked effects on ALL cells, whereas the less potent first generation statins (mevastatin and pravastatin) had less significant effects. We found that fluvastatin, lovastatin, and simvastatin inhibited proliferation of all 7 ALL cell lines using MTT assay (p <0.05). We also found these three statins induced apoptosis, leading to profound cell death in all 7 cell lines (p<0.05) as assessed by flow cytometry for Annenix-V staining and 7-AAD. IC50 dosing for the three agents ranged between 500nM and 5uM depending on the cell line, levels easily obtainable in humans. The most likely explanation for this remarkable, single-agent effect on ALL is that ALL cells are indeed dependent on cholesterol biosynthesis. Other potential explanations include the possibility that ALL cells are dependant on one of the signal transduction pathways affected by targeting HMG-CoA reductase, or the formal possibility of an off-target effect of statins separate from the inhibition of cholesterol biosynthesis. To determine if the effects of the statins were due to a direct effect on HMG-CoA reductase, we treated cells with melavonolactone, the product of conversion of HMG-CoA by HMGCoA reductase. We found that the addition of melavonolactone completely reversed the effects of all statins even at high doses (>10uM) in all cell lines, making the explanation of an off-target, non-HMG-CoA reductase-based mechanism unlikely. Ongoing work includes the testing of statins in NOD/SCID xenograft models of primary human ALL models, assessing the effects of statins on AKT, Ras, and Erk in ALL cells, and assessing the combination of statins with cytotoxic agents in ALL. In conclusion, we found that ALL cells are dependent of HMG-CoA reductase for survival. Since statins are safe and well-tolerated, this class of agents should be further explored in patients with ALL.

scholarly journals Mine the Stability of the G2/M Checkpoint to Break Down Acute Lymphoblastic Leukemia Defenses Against Antineoplastic Drugs

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2808-2808
Author(s):  
Andrea Ghelli Luserna di Rorà ◽  
Ilaria Iacobucci ◽  
Neil Beeharry ◽  
Maria Vittoria Falzacappa ◽  
Chiara Ronchini ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Single Agent ◽  
Leukemic Cell ◽  
Leukemic Cells ◽  
Antineoplastic Drugs ◽  
Primary Cells ◽  
Dna Damages ◽  
Leukemic Cell Lines

Abstract Although impressive developments have been made in the treatment of Acute Lymphoblastic Leukemia (ALL) patients, the overall survival is still very poor. With the exception of novel therapeutic strategies based on monoclonal antibodies (Bi-specific T-cell engagers, BiTEs) or immunogenic cells (CART cells), the therapeutic approaches for adult ALL patients are still base on non-selective chemotherapy or on tyrosine kinase inhibitors (TKIs) for the patients harboring the BCR-ABL1 fusion transcript. In addition a large percentage of initial successfully treated patients frequently develop relapses. Thus there is a need to improve the efficacy of conventional therapies, in particular those related to TKIs and to DNA damaging agents, in order to reduce the off-target toxicity and avoid relapses. In the present study we evaluated the in vitro, ex vivo and in vivo efficacy of MK-1775, a specific Wee1 inhibitor, in single agent and in combination with different therapeutic agents normally used for the treatment of B-/T-ALL. We firstly started by evaluated the efficacy of the compound in single agent on a panel of human B and T ALL cell lines (n=8) and on primary cells isolated from the bone marrow of adult B-ALL patients (n=8). The inhibition of Wee1 deeply reduced the cell viability and the proliferation rate, induced the apoptosis and increased the DNA damages of both leukemic cell lines and primary cells. Further cell-cycle analysis showed that in leukemic cell lines the treatment increased the number of cell in late S and G2/M phase. Light microscopy analyses, looking for nuclei morphology, confirmed that MK-1775 increased the number of mitotic cells but it interfered with normal mitotic division (induction of aberrant mitosis as showed by the increment of DNA bridges and micro-nuclei). The effects of the compound on the cell cycle profile and on the G2/M checkpoint were confirmed also in immunoblotting analyses, by the increment of phospho-HH3(ser10) and of Myt1 (mitotic isoform), and by gene expression analysis looking to specific genes involved in the G2/M checkpoints (PrimePcr DNA damage assay, Biorad). In particular genes like GADD45A and CCNB1/CCNB2 were significantly up-regulated between treated and untreated samples. Finally using a T-ALL mouse model we evaluated the effect of MK-1775 in single agent. Although no significative differences were seen between treated and un-treated samples, due to a very aggressive phenotype of the disease (all animal died after only 18 days from the engraftment), molecular analyses confirmed that the treatment induced DNA damages (increase of H2A.X and p-Chk1 ser317) and inhibited Wee1 functionality (reduction of pCDC2) on leukemic blasts isolated from both spleens and bone marrows. To evaluate if the inhibition of the G2/M checkpoint could sensitize leukemic cells to the toxicity of antineoplastic drugs, Philadelphia-negative ALL cell lines and primary leukemic cells (n=9) where treated with increasing concentration of MK-1775 and increasing concentration of the nucleotide analogue, clofarabine. Statistical analyses (Combination index value) confirmed the synergy of the combination in the reduction of the cell viability, in the inhibition of the proliferation and in the induction of the apoptosis. Similar results were seen on Philadelphia-positive ALL cell lines and primary cells (n=3) combining the MK-1775 with the TKI, bosutinib. The simultaneously inhibition of the Wee1 and the BCR-ABL downstream pathway resulted in a synergic inhibition of the cell viability, reduction of the proliferation and induction of apoptosis. In our opinion the pre-clinical results of this study are the basis for a future clinical evaluation of MK-1775 for the treatment of ALL patients. Acknowledgments: ELN, AIL, AIRC, progetto Regione-Università 2010-12 (L. Bolondi), Fondazione del Monte di Bologna e Ravenna, FP7 NGS-PTL project. Disclosures Martinelli: Novartis: Speakers Bureau; BMS: Speakers Bureau; Amgen: Consultancy, Speakers Bureau; Roche: Consultancy, Speakers Bureau; MSD: Consultancy; Pfizer: Consultancy, Speakers Bureau; Ariad: Consultancy, Speakers Bureau; Genentech: Consultancy; Celgene: Consultancy, Speakers Bureau.

scholarly journals Restraining the Proliferation of Acute Lymphoblastic Leukemia Cells by Genistein through Up-regulation of B-cell Translocation Gene-3 at Transcription Level

2019 ◽  
Vol 8 ◽  
pp. e1229
Author(s):  
Masoumeh Abedi Nejad ◽  
Mohsen Nikbakht ◽  
Masoomeh Afsa ◽  
Kianoosh Malekzadeh
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cancer Cells ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Suppressor Gene ◽  
Transcription Level ◽  
Hematopoietic Malignancy ◽  
Proliferative Effect ◽  
Cancer Agent

Background: Acute lymphoblastic leukemia (ALL) is a highly prevalent pediatric cancer accounting for approximately 78% of leukemia cases in patients younger than 15 years old. Different studies have demonstrated that B-cell translocation gene 3 (BTG3) plays a suppressive role in the progress of different cancers. Genistein is considered a natural and biocompatible compound and a new anti-cancer agent. In this study, we evaluate the effect of genistein on BTG3 expression and proliferation of ALL cancer cells. Materials and Methods: ALL cell lines (MOLT4, MOLT17, and JURKAT) were cultured in standard conditions. Cytotoxicity of genistein was detected using MTT assay. The cells were treated with different concentrations of genistein (10, 25, 40, and 55μM) for 24, 48, and 72 hours, and then cell viability and growth rate were measured. The quantitative real-time polymerase chain reaction was applied to investigate the effect of genistein on BTG3 expression. Results: The percentage of vital cells treated with genistein significantly decreased compared to the non-treated cells, showed an inverse relationship with an increasing genistein concentration. The present study suggests a dose of 40μM for genistein as a potent anticancer effect. Genistein could elevate BTG3 for 1.7 folds in MOLT4 and JURKAT and 2.7 folds in MOLT17 cell lines at transcription level conveged with 60 to 90% reduction in the proliferation rate of cancer cells. Conclusion: Up-regulation of BTG3 as a tumor suppressor gene can be induced by genistein. It seems that BTG3 reactivation can be introduced as another mechanism of anti-proliferative effect of genistein and could be considered as a retardant agent candidate against hematopoietic malignancy.[GMJ.2019;8:e1229]

scholarly journals Bovine Dialyzable Leukocyte Extract IMMUNEPOTENT-CRP Induces Selective ROS-Dependent Apoptosis in T-Acute Lymphoblastic Leukemia Cell Lines

2020 ◽  
Vol 2020 ◽  
pp. 1-17 ◽  
Author(s):  
Helen Yarimet Lorenzo-Anota ◽  
Ana Carolina Martínez-Torres ◽  
Daniel Scott-Algara ◽  
Reyes S. Tamez-Guerra ◽  
Cristina Rodríguez-Padilla
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Immune System ◽  
Acute Lymphoblastic Leukemia ◽  
Cancer Cells ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cells ◽  
Cell Death Mechanism ◽  
Nuclear Alterations

Immunotherapies strengthen the immune system to fight multiple diseases such as infections, immunodeficiencies, and autoimmune diseases, and recently, they are being used as an adjuvant in cancer treatment. IMMUNEPOTENT-CRP (I-CRP) is an immunotherapy made of bovine dialyzable leukocyte extract (bDLE) that has chemoprotective and immunomodulatory effects in different cellular populations of the immune system and antitumor activity in different cancer cell lines. Our recent results suggest that the antineoplastic effect of I-CRP is due to the characteristics of cancer cells. To confirm, we evaluated whether the selectivity is due to cell lineage or characteristics of cancer cells, testing cytotoxicity in T-acute lymphoblastic leukemia cells and their cell death mechanism. Here, we assessed the effect of I-CRP on cell viability and cell death. To determine the mechanism of cell death, we tested cell cycle, mitochondrial and nuclear alterations, and caspases and reactive oxygen species (ROS) and their role in cell death mechanism. Our results show that I-CRP does not affect cell viability in noncancer cells and induces selective cytotoxicity in a dose-dependent manner in leukemic cell lines. I-CRP also induces mitochondrial damage through proapoptotic and antiapoptotic protein modulation (Bax and Bcl-2) and ROS production, nuclear alterations including DNA damage (γ-H2Ax), overexpression of p53, cell cycle arrest, and DNA degradation. I-CRP induced ROS-dependent apoptosis in leukemic cells. Overall, here, we show that I-CRP cytotoxicity is selective to leukemic cells, inducing ROS-dependent apoptosis. This research opens the door to further exploration of their role in the immune system and the cell death mechanism that could potentially work in conjunction with other therapies including hematological malignances.

Bcl-2 and mTOR as Effective Targets for Molecular Therapy of Acute Lymphoblastic Leukemia

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3228-3228
Author(s):  
Stefano Iacovelli ◽  
Maria Rosaria Ricciardi ◽  
Andrea Miele ◽  
Paola Bergamo ◽  
Roberto Licchetta ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Growth Inhibition ◽  
Cell Line ◽  
Cell Lines ◽  
Synergistic Effects ◽  
Lymphoblastic Leukemia ◽  
Mtor Inhibitors ◽  
Primary Cells ◽  
Apoptosis Induction ◽  
Combined Use

Abstract Abstract 3228 Although they frequently achieve complete remission (CR), adult patients with acute lymphoblastic leukemia (ALL) subsequently experience leukemia relapse, which represents an unresolved therapeutic problem. Based on the observation that ALL cells are frequently characterized by the deregulation of the apoptotic machinery, we and others have evaluated pre-clinically the activity of ABT-737 (kindly provided by Abbott Laboratories), a BH3-mimetic Bcl-2/Bcl-XL inhibitor, displaying a potent growth-inhibitory activity in ALL cell lines and primary cells. However ABT-737 binds to the anti-apoptotic protein Mcl-1 with low affinity and Mcl-1 expression may mediate resistance to ABT-737. Since ALL is also characterized by the aberrant activation of the mTOR and related signalling pathways, in the present study we further evaluated the combined Bcl-2/Bcl-XL (by ABT-737) and mTOR (by CCI-779) inhibition focusing, in particular, on the activity of combined molecularly targeted therapies on resistant cells. In MOLT-4 cells, ABT-737 induced dose and time-dependent growth inhibition (IC-50= 198nM) followed, at higher concentrations (250-500nM), by apoptosis induction. In contrast, the CEM-S, CEM-R, JURKAT, DAUDI and RAJI cells proved resistant (IC-50 >5 μM). When we explored the effects of CCI-779 on the aforementioned cell lines, only minor cytostatic effects were observed (IC-50 0.5–28.2μM). MOLT-4 cells, for example, showed a flat dose-response curve (35-55% growth inhibition) at concentrations ranging between 1 and 5000 nM (IC50=9,87μM) and apoptosis induction was not seen until 5000 nM. We next investigated the effects of the combined use of ABT-737 and CCI-779 (each at 1000nM) in the ABT-resistant JURKAT cells. A significant (p= 0.04) induction of apoptosis was observed with the combination, as compared with single agents, after 24 h (47.7% ±5.9 of cells with sub-G1 DNA content with ABT-737 + CCI-779, compared to 17.4% ±1.5 and 4.2% ±1.5 with ABT-737 and CCI-779 as single agents, respectively). Similarly, when we exposed CEM-R cells to the drug combination (ABT-737 1000nM and CCI-779 5000nM) for 24 h, a strikingly stronger apoptosis induction (sub-G1 peak= 75.3% ±16.8) was observed, compared to single agents (15.8% ±7.2 and 4.2% ±1.9 with ABT-737 and CCI-779 alone, respectively) (p=0.0003). These effects were confirmed by measuring Annexin V binding. WB analysis showed decreased Mcl-1 levels, following exposure to CCI-779 and further downregulation in response to combined ABT-737+CCI-779 in the CEM-R cell line. These effects, however, were not seen in the parental CEM-S cell line. Primary cells, obtained from 10 ALL patients, showed an increase of the sub-G1 peak in 7/10 and in 4/10 samples, after exposure to ABT-737 (50nM) and CCI-779 (5000 nM), while synergistic effects on apoptosis induction were observed in 4/10 samples after exposure to the combination. In summary, we observed that the combined use of Bcl-2/Bcl-XL and mTOR inhibitors may exert synergistic cytotoxic effects in some resistant ALL models and this effect is associated with CCI-779-induced Mcl-1 down-regulation. Synergistic effects between these inhibitors were also found in a proportion of primary ALL samples, thus supporting further studies of combined Bcl-2/Bcl-XL and mTOR inhibitors, to overcome ALL resistance. Disclosures: Petrucci: Celgene: Honoraria; Janssen Cilag: Honoraria. Tafuri:Sigma-Tau: Research Funding.

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