Identification of Sphingosine Kinases As Therapeutic Targets in B-Lineage Acute Lymphoblastic Leukemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1499-1499
Author(s):  
Craig T Wallington-Beddoe ◽  
Kenneth F Bradstock ◽  
Linda J Bendall
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Xenograft Model ◽  
Intracellular Location ◽  
Gene Signatures ◽  
Sphingosine Kinases ◽  
B Lineage

Abstract Abstract 1499 Sphingosine 1-phosphate (S1P) is a bioactive lipid with roles in cell proliferation and survival. S1P is produced by the sphingosine kinases, SphK1 and SphK2. SphK1 is over expressed in a number of malignancies and evidence points overwhelmingly to a pro-survival role. The role of SphK2 is much less well defined and appears to be dependent on its intracellular location with some reports of opposite effects to those of SphK1. Little is known about the interaction of SphKs with intracellular signalling pathways. Here we assess the relevance of SphKs in B-lineage acute lymphoblastic leukemia (B-ALL). Gene expression signatures indicative of activation of SphK1 or SphK2 were not publicly available. Therefore we treated ALL cell lines with the SphK1 or SphK2 specific inhibitors SK1-I and ABC294640 respectively, and analysed gene expression by microarray. Although a signature for SphK1 was not obtained, two independent methods of analysis generated SphK2 gene signatures that segregated control and drug treated cell lines. The signatures consisted of 11 and 35 genes and included reduced expression of the NF-κB inhibitor TRIB3 (NF-κB activity is commonly up regulated in ALL), the C/EBP family transcription factor DDIT3, the protein phosphatase 1 regulatory subunit PPP1R15A and the sphingolipid/cholesterol transporter ABCA1, all validated by quantitative RT-PCR. These gene signatures were used to interrogate a large publicly available gene expression dataset (GSE28497) obtained from pediatric ALL patients at the time of diagnosis. SphK2 activity signatures were more highly expressed in ALL samples (p=0.001 and p=0.027 for the smaller and larger signatures respectively) than normal B-cell progenitors. Although SPHK1 or SPHK2 genes were not over expressed in this dataset, SphK1 protein levels were increased in ALL cell lines and in patient samples. The importance of SphK1 and SphK2 in the development of ALL was examined by transducing B-cell progenitors isolated from WT, SphK1−/− or SphK2−/− mice with the ALL associated p185 form of the oncogenic fusion gene BCR/ABL and injecting transduced cells into sub-lethally irradiated wild type C57BL/6 mice. Twenty-two of 29 mice receiving cells from WT animals developed ALL, with a mean survival of 52.6 days (95% CI 42.3–62.8 days). Mice receiving cells lacking Sphk1 or Sphk2 had a significantly reduced incidence of ALL development with 14 of 30 (mean survival 71.6 days, 95% CI 60.5–82.8 days) and 16 of 29 animals (mean survival of 68.7 days, 95% CI 57.9–79.5 days) respectively (p=0.001). Lymphoblasts with a B-cell progenitor phenotype (B220+, CD19+, IgM−, CD11b−) were present in blood films and livers from leukemic mice and cells recovered from these animals produced a rapidly fatal ALL in secondary recipients. The presence of BCR/ABL and the expected deletion of Sphk1 or Sphk2 were confirmed by PCR in all murine leukemias examined. We have previously reported that inhibitors of SphK1 and/or SphK2 inhibit proliferation and induce cell death in ALL cell lines, and that these agents can synergize with imatinib in Ph+ ALL cell lines. We have furthered these studies to demonstrate that patient derived ALL cells similarly respond to SphK1 and SphK2 inhibition in vitro. The SphK2 inhibitor ABC294640 (100mg/kg) reduced plasma S1P levels in mice consistent with our previous reports of reduced ALL burden following ABC294640 treatment, and using a NOD/SCID γc−/− xenograft model of human ALL we have now shown that ABC294640 extends survival of ALL-bearing mice (p=0.0012 for Ph− xenograft) and further extends the survival when combined with imatinib treatment in mice engrafted with a human Ph+ ALL patient sample (p=0.044). This is the first report to suggest that sphingosine kinase 2 activity is increased in ALL, providing support for targeting SphK2 as a therapeutic strategy. Loss of SphKs reduces the incidence of ALL in a murine model of BCR/ABL-driven disease and ABC294640 reduces disease and extends survival in a human xenograft model of ALL. This compound synergizes with a number of potential therapeutic agents and further extends survival in a xenograft model of Ph+disease when combined with imatinib. This has potential to translate into a useful anti-leukemic strategy. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Precise Modeling of IKZF1 Alterations in Human B-Cell Acute Lymphoblastic Leukemia Cell Lines Reveals Distinct Chemosensitivity, Homing, and Engraftment Properties

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 549-549
Author(s):  
Jason H. Rogers ◽  
Rohit Gupta ◽  
Jaime M. Reyes ◽  
Lorenzo Brunetti ◽  
Michael C. Gundry ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cell Lines ◽  
Sanger Sequencing ◽  
Lymphoblastic Leukemia ◽  
Clonal Cell ◽  
Ribonucleoprotein Complexes ◽  
Clonal Cell Lines ◽  
Cell Acute Lymphoblastic Leukemia

Abstract Around 20% of pediatric and the majority of adults with B-cell acute lymphoblastic leukemia (B-ALL) suffer relapse, and prognosis after relapse is very poor. Therefore, identifying those at risk for treatment failure and improving their outcome is imperative. In B-ALL, deletions and mutations of the gene IKAROS family zinc finger 1 (IKZF1) are associated with an increased risk of relapse. IKZF1 encodes the IKAROS protein, which is a master lymphoid regulatory transcription factor and chromatin remodeler. Somatic IKZF1 lesions are thought to be secondarily acquired, arising in lymphoblasts with existing driver genetic lesions, most commonly co-occurring with BCR-ABL1 fusion, activating kinase fusions of Ph-like disease and deregulated DUX4 and ERG. In B-ALL, mono- or bi-allelic deletions of the entire gene, as well as intragenic deletions occur. One of the most common perturbations of IKZF1 in B-ALL is an intragenic deletion of a 50-kilobase (kb) region containing exons 4-7, resulting in the expression of a dominant-negative isoform, IK6. Recently published clinical data show potentially conflicting results over the benefits of therapy intensification in IKZF1-mutant cases (Clappier et al., 2015; Hinze et al., 2017; & Yeoh, et al., 2018). Human cell models of these deletions are needed, as there may be unknown functional differences among mutation types, and the available body of data relies on clinical statistical associations, in vitro RNA interference, viral overexpression of IK6, and mouse models. We used the CRISPR/Cas9 system in the human B-ALL cell lines Nalm-6 and REH by electroporation with sgRNA-Cas9 ribonucleoprotein complexes (RNPs) to generate IKZF1-mutant clones. We identified single cell-derived clonal lines with IKZF1 frameshift mutations in one or both alleles by Sanger sequencing and TIDE decomposition. We confirmed ablation of protein expression by immunoblotting. We treated the IKZF1-mutant clonal cell lines with chemotherapeutic agents commonly used to treat B-ALL and calculated the IC50 by Annexin V/7-AAD double-negative population after 48-72 hour treatment. Compared to IKZF1-wild type Nalm-6 cells, Nalm-6 IKZF1-/- clones exhibited profound resistance to dexamethasone and modest but significant resistance to most other chemotherapeutics tested including vincristine, asparaginase, and daunorubicin. In contrast, these cell lines were more sensitive to the nucleoside analog, cytarabine (Panel A). We next analyzed gene expression profiles by RNA-seq and observed that IKZF1-/- clones are characterized by a stem cell-like gene expression signature and activation of the JAK/STAT pathway (Panel B). Transplantation into immunodeficient NOD scid gamma (NSG) mice demonstrated that IKZF1 deletion leads to enhanced engraftment, significantly increased bone marrow homing, and reduced survival time (Panel D). We also employed a novel CRISPR/Cas9 homology-directed repair (HDR) strategy to generate clonal cell lines expressing IK6 under control of the endogenous promoter, which represents a significant advantage to many previous studies utilizing viral overexpression. We electroporated the cells with sgRNA-Cas9 RNPs along with a 3kb commercially synthesized double-stranded DNA HDR template that knocks-in exon 8 with a GFP tag directly following exon 3. Using this strategy, we were able to isolate heterozygous clones (IKZF1IK6/+) from both Nalm-6 (Panel C) and REH cell lines using flow cytometry sorting for GFP-positive cells. We confirmed precise HDR by Sanger sequencing and immunoblotting. When transplanted into immunodeficient mice, IKZF1IK6/+cells showed delayed engraftment and disease onset, but profound splenic infiltration, consistent with a more indolent, infiltrative disease phenotype (Panels D & E). Ongoing drug treatment assays suggest the chemosensitivity profiles of IKZF1IK6/+ and IKZF1IK6/-clonal cell lines are distinct from their isogenic IKZF1-/-counterparts. Our data support clinical studies reporting that IKZF1-mutated B-ALL is an aggressive, infiltrative, and treatment-resistant disease. Notable differences in drug response and in vivo dynamics in xenografts exist between IKZF1-/-cells and IKZF1IK6/+cells. Detailed delineation of the exact IKZF1 status in ALL patients at diagnosis may be informative in more accurately determining risk stratification and the most effective therapeutic regimen. Disclosures No relevant conflicts of interest to declare.

scholarly journals Engineered Bcor Mutations Lead to Acute Lymphoblastic Leukemia of Progenitor B-1 Lymphocyte Origin in a Sensitized Background

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1331-1331
Author(s):  
Mianmian Yin ◽  
Yang Jo Chung ◽  
R. Coleman Lindsley ◽  
Yeulin Zhu ◽  
Robert L. Walker ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Regulatory Elements ◽  
Leukemic Transformation ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Indel Mutation

Abstract Chromosomal translocations resulting in NUP98 fusion genes have been associated with a wide spectrum of hematologic malignancies, including MDS, AML, T-ALL, and B cell precursor (BCP) ALL. Based on gene expression profiles and murine transplantation experiments, it is thought that NUP98 fusions can confer aberrant self-renewal potential to hematopoietic cells. Approximately 90% of mice that express a NUP98-PHF23 (NP23) fusion in the hematopoietic compartment, under the control of Vav1 regulatory elements develop AML and/or T-ALL. However, approximately 10% of NP23 mice develop an aggressive acute lymphoblastic leukemia of B1-lymphocyte progenitor origin (pro B-1 ALL). Whole exome sequencing demonstrated that all NP23 pro-B1-ALL had acquired somatic frameshift mutations of the BCL6 co-repressor (Bcor) gene, and most had acquired mutations in the Jak/Stat pathway. To determine whether experimentally engineered Bcor mutations would lead to pro B-1 ALL, we used CRISPR-Cas9 to introduce Bcor indel mutations into NP23 hematopoietic stem and progenitor cells through the use of Bcor single guide RNAs (Bcor sgRNA). Recipient mice transplanted with NP23 bone marrow (BM) or fetal liver (FL) cells that had been transduced with a Bcor sgRNA developed pro B-1 ALL, characterized by a B-1 progenitor immunophenotype, clonal Igh gene rearrangement, and Bcor indel mutation, whereas control recipients did not. In addition, similar to some human BCP ALL, the Bcor sgRNA/NP23 murine pro B-1 ALL had acquired somatic mutations in Jak kinase genes. A distinct subset of pediatric BCP ALL are characterized by rearrangement and overexpression of the CRLF2 gene (designated CRLF2r); the CRLF2 gene is the receptor for thymic stromal lymphopoietin (TSLP), a cytokine that plays a role in normal progenitor B1 cell development. The NP23 pro-B1 ALL are similar to CRLF2r BCP ALL in terms of a preferential V heavy chain (VH) usage, gene expression profile, and propensity for acquired JAK/STAT pathways mutations. JAK inhibitors (ruxolitinib and tofacitinib) induced apoptosis and inhibited the growth of pro B-1 ALL cell lines established from Bcor sgRNA/NP23 recipients, at clinically achievable concentrations (10-100 nM). Taken together, these findings demonstrate that a CRISPR-induced Bcor frameshift collaborates with an NP23 transgene to predispose B-1 progenitors to leukemic transformation. These two events are unlikely to be sufficient for leukemic transformation, as we detected spontaneous Jak pathway mutations that were required for continued growth of the leukemic cells. This constellation of mutations (NP23 expression leading to increased stem cell self-renewal, Bcor frameshift leading to impaired B cell differentiation, and Jak pathway mutations leading to dysregulated proliferation) is similar to that seen in human BCP ALL patients, and suggests that the NP23/Bcor mutant mice and cell lines will be a useful model for human pro-B1 ALL. Disclosures Aplan: NIH Office of Technolgy Transfer: Employment, Patents & Royalties: NUP98-HOXD13 mice.

The NOD/SCID Xenograft Model Provides Clinically-Relevant Insights Into Glucocorticoid-Induced Gene Expression In Childhood B-Cell Precursor Acute Lymphoblastic Leukemia (ALL)

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2493-2493
Author(s):  
Vivek A Bhadri ◽  
Mark J Cowley ◽  
Warren Kaplan ◽  
Richard B Lock
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
Mouse Model ◽  
B Cell ◽  
Differential Expression ◽  
Differentially Expressed Genes ◽  
Lymphoblastic Leukemia ◽  
Xenograft Model ◽  
Differentially Expressed ◽  
Cell Precursor

Abstract Abstract 2493 Introduction. Glucocorticoids (GC) such as prednisolone (Pred) and dexamethasone (Dex) are critical drugs in multi-agent chemotherapy protocols used to treat acute lymphoblastic leukemia (ALL). The NOD/SCID ALL xenograft mouse model is a clinically relevant model in which the mice develop a systemic leukemia which retains the fundamental biological characteristics of the original disease. Here we report the results of a study evaluating the NOD/SCID xenograft model to investigate GC-induced gene expression. Methods. Cells from a GC-sensitive xenograft derived from a child with B-cell precursor ALL were inoculated into NOD/SCID mice. Engraftment, defined as the proportion of human vs mouse CD45+ cells in the peripheral blood, was monitored by serial weekly tail-vein sampling. When engraftment levels reached >50%, the mice were randomised and treated with either dexamethasone 15 mg/kg or vehicle control by intraperitoneal injection, and harvested at 0, 8, 24 or 48 h thereafter. The 48 hour groups received a second dose of vehicle or Dex at 24 hours. At the defined timepoints, the mice were euthanized and lymphoblasts harvested from the spleen. RNA was extracted, amplified and hybridised onto Illumina WG-6 V3 chips. The data was pre-processed using variance-stabilisation transformation, and quantile normalisation. Differential expression was determined using limma by comparing all treated groups to time 0, with the positive False Discovery Rate correction for multiple testing. Hierarchical clustering was used to compare groups to each other. The stability of results when reducing the number of replicates was assessed using the Recovery Score method. Functional analysis was performed using gene set enrichment analysis (GSEA) and comparison to publicly available microarray data using parametric GSEA. Results. The 8 hour Dex-treated timepoint had the most number of significantly differentially expressed genes (see Table), with fewer observed at the 24 and 48 hour Dex-treated timepoints. There was minimal significant differential gene expression across the time-matched controls. At the 8 hour timepoint, ZBTB16, a known GC-induced gene, was the most significantly upregulated gene. Other significantly differentially expressed genes included TSC22D3 and SOCS1, both downstream targets of the glucocorticoid receptor (upregulated), and BCL-2 and C-MYC (downregulated). GSEA at 8 hours revealed a significant upregulation of catabolic pathways and downregulation of pathways associated with cell proliferation, particularly C-MYC. GSEA at 24 hours revealed enrichment of pathways associated with NF-kB. Replicate analysis revealed that at the 8 hour Dex treated timepoint, a dataset with high signal and differential expression, using data from 3 replicates instead of 4 resulted in excellent recovery scores of >0.9. However at other timepoints with less signal very poor recovery scores were obtained using 3 replicates. We compared our data to publicly available datasets of GC-induced genes in ALL (Schmidt et al, Blood 2006; Rainer et al, Leukemia 2009) using parametric GSEA, which revealed that the 8 hour gene expression data obtained from the NOD/SCID xenograft model clustered with data from primary patient samples (Schmidt) rather than the cell line data (Rainer). The 24 and 48 hour datasets clustered separately from all other datasets by this method, reflecting fewer and predominantly downregulated gene expression at these timepoints. Conclusions: The NOD/SCID xenograft mouse model provides a reproducible experimental model system in which to investigate clinically-relevant mechanisms of GC-induced gene regulation in ALL; the 8 hour timepoint provides the highest number of significantly differentially expressed genes; time-matched controls are redundant and excellent recovery scores can be obtained with 3 replicates. Disclosures: No relevant conflicts of interest to declare.

Identification of novel markers for monitoring minimal residual disease in acute lymphoblastic leukemia

Blood ◽  
2001 ◽  
Vol 97 (7) ◽  
pp. 2115-2120 ◽  
Author(s):  
Jiann-Shiuh Chen ◽  
Elaine Coustan-Smith ◽  
Toshio Suzuki ◽  
Geoffrey A. Neale ◽  
Keichiro Mihara ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Minimal Residual Disease ◽  
Residual Disease ◽  
Lymphoblastic Leukemia ◽  
Cdna Array ◽  
Leukemic Cells ◽  
B Lineage ◽  
Minimal Residual

Abstract To identify new markers of minimal residual disease (MRD) in B-lineage acute lymphoblastic leukemia (ALL), gene expression of leukemic cells obtained from 4 patients with newly diagnosed ALL was compared with that of normal CD19+CD10+ B-cell progenitors obtained from 2 healthy donors. By cDNA array analysis, 334 of 4132 genes studied were expressed 1.5- to 5.8-fold higher in leukemic cells relative to both normal samples; 238 of these genes were also overexpressed in the leukemic cell line RS4;11. Nine genes were selected among the 274 overexpressed in at least 2 leukemic samples, and expression of the encoded proteins was measured by flow cytometry. Two proteins (caldesmon and myeloid nuclear differentiation antigen) were only weakly expressed in leukemic cells despite strong hybridization signals in the array. By contrast, 7 proteins (CD58, creatine kinase B, ninjurin1, Ref1, calpastatin, HDJ-2, and annexin VI) were expressed in B-lineage ALL cells at higher levels than in normal CD19+CD10+ B-cell progenitors (P < .05 in all comparisons). CD58 was chosen for further analysis because of its abundant and prevalent overexpression. An anti-CD58 antibody identified residual leukemic cells (0.01% to 1.13%; median, 0.03%) in 9 of 104 bone marrow samples from children with ALL in clinical remission. MRD estimates by CD58 staining correlated well with those of polymerase chain reaction amplification of immunoglobulin genes. These results indicate that studies of gene expression with cDNA arrays can aid the discovery of leukemia markers.

scholarly journals Biomarker Profile for Prediction of Patient Response to Smac Mimetic Monotherapy in Pediatric Precursor B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2082-2082
Author(s):  
Julia Zinngrebe ◽  
Ferdinand Schlichtig ◽  
Malcolm Meyer ◽  
Johann M. Kraus ◽  
Elena Boldrin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Clinical Trials ◽  
Cell Death ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Expression Pattern ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Survival Rates ◽  
Patient Response

Acute lymphoblastic leukemia (ALL) is the most common malignancy in childhood. While improved multi-agent chemotherapy regimens with individualized risk stratification have led to increased survival rates of approximately 80 percent, 20 percent of patients respond poorly to therapy or relapse. Therefore, novel therapeutic avenues are urgently needed to improve treatment outcome, overcome resistance and reduce side effects. Failure to undergo cell death represents a key survival mechanism of cancer cells and results in drug resistance and clonal escape. Since inhibitor of apoptosis proteins (IAPs) are often overexpressed in malignant cells and their overexpression correlates with inferior survival rates, they provide an attractive molecular target for therapeutic intervention. Small molecule inhibitors have been developed that act as SMAC mimetics (SMs) to counteract the cell death inhibitory function of IAPs. SMs can activate and/or modulate cell death pathways, and are currently being evaluated in clinical trials. Their successful therapeutic implementation requires identification of patients who could benefit from a SM-based treatment regimen ideally before start of therapy. Here, we analyzed the intrinsic activity of two monovalent (AT406 and LCL161) and two bivalent (Birinapant or BV6) SMs on 29 unselected patient-derived pediatric precursor B-cell (BCP)-ALL samples and identified a subset of BCP-ALL primografts to be sensitive to SM treatment (n=8). When we compared gene expression of SM-sensitive (n=8) and SM-insensitive (n=6) patient-derived BCP-ALL samples, we identified a characteristic gene expression signature with 127 differentially regulated genes, amongst them upregulation of TNFRSF1A (TNFR1) in the SM-sensitive subset. In line with previous reports, we confirmed a critical role of the TNF/TNFR1-axis for SM-induced cell death in BCP-ALL by functional analysis. Expression of TNFRSF1A alone, however, did not correlate with sensitivity to SM-induced cell death indicating that TNFR1 is not the only factor regulating cell fate decisions in response to SM treatment. To identify potential biomarker genes for prediction of patient response to SM monotherapy in BCP-ALL, we compared differentially regulated genes of SM responders and non-responders from our cohort with data from a published cohort. Interestingly, we found 4 genes to overlap between these two cohorts. Of these 4 genes TSPAN7, FAM69C, and TNFRSF1A were upregulated whereas MTX2 was downregulated in SM-sensitive samples. The signature identified may reflect a particular TNF network. Analysis of expression levels of these 4 genes in BCP-ALL cell lines (Nalm6, Reh, UoCB6 and RS4;11) revealed that Reh cells, sensitive to SM-induced cell death, exhibited the biomarker profile of primograft sensitivity, i.e. upregulation of TSPAN7, FAM69C, TNFRSF1A and downregulation of MTX2. Nalm6 cells resembled the expression pattern of SM-insensitive samples with a downregulation of TSPAN7, FAM69C, TNFRSF1A and an upregulation of MTX2 and were resistant to SM-induced cell death. RS4;11 and UoCB6 cells showed no pattern. Based on these findings we hypothesized that the respective expression patterns of TSPAN7, FAM69C, TNFRSF1A and MTX2 could predict sensitivity to SMs. An extended screen of additional primary BCP-ALL samples for their expression levels of TSPAN7, FAM69C, TNFRSF1A and MTX2 and response to SMs substantiated this hypothesis. In summary, the subset of primary BCP-ALL samples with sensitivity to SMs is characterized by a gene signature with MTX2 low and TSPAN7, FAM69C and TNFRSF1A high. By using this expression profile, sensitivity to SMs in BCP-ALL could be identified in cell lines and additional primografts. Based on these results, we suggest the identified gene expression pattern as a biomarker for selecting patients to be treated by SM monotherapy in clinical trials. Disclosures No relevant conflicts of interest to declare.

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]

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