Altered Hematopoietic Cell Gene Expression Identifies Patients at Risk for Development of Therapy-Related Leukemia (t-MDS/AML)

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 234-234 ◽  
Author(s):  
Liang Li ◽  
Min Li ◽  
Can-Lan Sun ◽  
Liton Francisco ◽  
Melanie Sabado ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
High Risk ◽  
Cell Cycle Regulation ◽  
Gene Signature ◽  
Training Set ◽  
Test Set ◽  
Gene Sets ◽  
Cd34 Cells ◽  
Cycle Regulation

Abstract Abstract 234 Therapy-related myelodysplasia or acute myeloid leukemia (t-MDS/AML) is a lethal complication of cancer treatment. Study of t-MDS/AML offers a unique opportunity to understand leukemogenesis since known genotoxic exposures can be temporally and causally related to genetic changes associated with development of leukemia. Although development of t-MDS/AML is associated with known genotoxic exposures, its pathogenesis is not well understood, and methods to predict risk of development of t-MDS/AML in individual cancer survivors are not available. To better understand the pathogenetic mechanisms underlying development of t-MDS/AML we performed microarray analysis of gene expression in patients who developed t-MDS/AML after autologous hematopoietic cell transplantation (aHCT) for Hodgkin lymphoma (HL) or non-Hodgkin lymphoma (NHL) and controls that did not develop t-MDS/AML after aHCT. Peripheral blood stem cell (PBSC) samples obtained pre-aHCT from patients who subsequently developed t-MDS post-aHCT (cases) and controls matched for primary diagnosis, age, race/ethnicity, and time since aHCT were studied. In a training set of 18 t-MDS/AML cases and 37 controls, CD34+ cells were selected from PBSC samples using flow cytometry, and gene expression evaluated using Affymetrix HG U133 plus 2.0 Arrays. Differences in gene expression in CD34+ cells from cases and controls were analyzed using conditional logistic model. Significant differences in gene expression were seen in PBSC obtained pre-aHCT from patients who later developed t-MDS/AML compared to controls.(Blood, 2009; 114: 677) PBSC obtained pre-aHCT from patients who subsequently t-MDS/AML after aHCT showed significant downregulation of gene sets related to mitochondria and oxidative phosphorylation, ribosomes, aminoacyl-tRNA biosynthesis, amino acid metabolism, cell cycle regulation, and hematopoietic differentiation. G-protein coupled receptors, hematopoietic regulation, and cell adhesion related genes were upregulated in PBSC from cases. There was reduced expression of genes with binding motifs for the transcription factor NRF2, which regulates oxidative stress and drug detoxification. We then sought to identify a smaller PBSC gene signature that would identify NHL and HL patients at the pre-aHCT timepoint at high risk for developing t- MDS/AML after aHCT (Figure 1). A cross-validated 38-gene classifier was derived from the training set using prediction analysis of microarray (PAM). This gene classifier was applied to an independent test set of PBSC obtained pre-HCT from 16 patients who developed t-MDS/AML after aHCT for NHL or HL, and 20 matched controls that did not develop t-MDS/AML. Application of the 38-gene signature to the test set correctly classified 19 of the 20 subjects (95%) who did not subsequently develop t-MDS/AML, and 14 of the 16 subjects (87.5%) who did develop t-MDS/AML, with significant correlation between predicted and true disease status (P<0.001). These results indicate that the gene expression profile of hematopoietic cells pre-aHCT can identify patients at high risk for t-MDS/AML post-aHCT. GSEA analysis revealed extensive overlap of up and down-regulated gene sets in t-MDS/AML cases in the training and test sets (Figure 2). Gene expression changes related to mitochondria, metabolism, cell cycle regulation and hematopoietic progenitors that were observed in the training set were validated in the test set. These results indicate that genetic programs associated with t-MDS/AML are perturbed long before disease onset, and that PBSC gene signatures can accurately identify patients at high risk of developing this complication. Disclosures: No relevant conflicts of interest to declare.

Modulation of Gene Expression Profile and In Vivo Anti-Myeloma Activity Induced by Valproic Acid, a Histone Deacytylase Inhibitor.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4790-4790
Author(s):  
Paola Neri ◽  
Teresa Calimeri ◽  
Mariateresa Di Martino ◽  
Marco Rossi ◽  
Orietta Eramo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Valproic Acid ◽  
Dna Replication ◽  
Gene Expression Profile ◽  
Cell Cycle Regulation ◽  
Cycle Regulation ◽  
Anti Tumor Activity

Abstract Valproic acid (VPA) is a well-tolerated anticonvulsant drug that has been recently recognized as powerful histone deacetylase (HDCA) inhibitor. VPA induces hyperacetylation of histone H3 and H4 and inhibits both class I and II HDCACs. Recently it has been shown that VPA exerts in vitro and in vivo anti-tumor activity against solid cancers and its in vitro anti-Multiple Myeloma (MM) activity has been previously reported. However, the molecular mechanisms are still unclear. Here we have investigated molecular changes induced by VPA as well as its in vivo activity in murine models of MM. We first studied the in vitro activity of VPA against IL-6 independent as well as IL-6 dependent MM cells. A time- and dose-dependent decrease in proliferation and survival of MM cell lines was observed (IC50 in the range of 1–3 mM). Gene expression profile following treatment with VPA at 2 and 5 mM showed down-regulation of genes involved in cell cycle regulation, DNA replication and transcription as well as up-regulation of genes implicated in apoptosis and chemokine pathways. The signaling pathway analysis performed by Ingenuity Systems Software identified the cell growth, cell cycle, cell death as well as DNA replication and repair as the most important networks modulated by VPA treatment. We next evaluated the in vivo activity of VPA using two xenograft models of human MM. A cohort of SCID mice bearing subcutaneous MM1s or OPM1 were treated i.p. daily with VPA (200 mg/kg, and 300 mg/kg, n=5 mice, respectively), or vehicle alone (n=5 mice) for 16 consecutive days. Tumors were measured every 2 days, and survival was calculated using the Kaplan Mayer method. Following VPA treatment, we found a significant (p=0.006) inhibition of tumor growth in mice bearing subcutaneous MM-1s cells treated with VPA at 200 mg/kg compared to control group, which translated into a significant (p= 0.002) survival advantage in the VPA treated animals. Similar results were obtained in animals bearing subcutaneous OPM1 cells. Flow cytometry analysis performed on retrieved tumor tissues from animals showed reduction of G2-M and S phase in tumor specimens following VPA treatment, versus untreated tumors, strongly suggesting in vivo effects of VPA on cell cycle regulation. Taken together, our data demonstrate the in vitro and in vivo anti-tumor activity of VPA, delineate potential molecular targets triggered by this agent and provide a preclinical rationale for its clinical evaluation, both as a single agent or in combination, to improve patient outcome in MM.

In Megakaryocytic Differentiation, the Aryl Hydrocarbon Receptor (AhR) Transcription Factor Regulates Hes1 Expression and Megakaryocytic Polyploidization.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2519-2519
Author(s):  
Stephan Lindsey ◽  
Eleftherios Papoutsakis
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Notch Signaling ◽  
Cell Cycle Regulation ◽  
Ex Vivo ◽  
Megakaryoblastic Leukemia ◽  
Aryl Hydrocarbon ◽  
Human Cd34 ◽  
Cd34 Cells ◽  
Cycle Regulation

Abstract Abstract 2519 Poster Board II-496 Understanding the mechanisms underlying megakaryocytic (Mk) differentiation and maturation is vital to the discovery of novel approaches to treating Mk and platelet disorders such as thrombocytopenia, megakaryoblastic leukemia, and thrombocythemia. The number of platelets released is proportional to the amount of DNA present in a given Mk, so insights into the molecular basis of Mk polyploidization could inspire improved ex vivo culturing methods to promote Mk commitment, expansion, and differentiation, leading to improved autologous transfusion protocols to offset thrombocytopenia associated with HSC transplants following high-dose chemotherapy or MDS progression. Microarray analyses on ex vivo Mk-differentiated primary human CD34+ cells showed that mRNA levels of the Aryl Hydrocarbon Receptor (AhR) increased during Mk differentiation and was elevated 4–6 fold in Mks compared to isogenic granulocytic cultures. These data were further confirmed by quantitative(Q)-RT-PCR analysis of differentiating Mks derived from primary human CD34+ cells as well as from CHRF cells (human megakaryoblastic leukemia). We have shown that CHRF cells are a valid model of human Mk differentiation (Fuhrken PG et al. Exp Hematol, 2007; 35:476–489). Thus, we hypothesized that AhR may act as a novel Mk transcription factor, possibly by influencing or regulating Mk polyploidization. Known as a “toxin sensor”, AhR is involved in the mechanism of action of environmental toxins, likely by altering cell cycle regulation. Epidemiological studies of toxic waste spills and Vietnam veterans suggest that exposure to known AhR ligands may result in increased platelet counts proportional to dioxin exposure level (Webb K et al. Am J Ind Med, 1987;11:685–691, Michalek JE Arch Environ Health, 2001; 56:396–405). These studies offer the intriguing possibility that AhR activation modulates megakaryocyte differentiation and/or platelet production. Interestingly, AhR influences the differentiation of other myeloid lineages including monocytes (Hayashi S et al. Carcinogenesis, 1995; 16:1403–1409) and is upregulated after leukocyte activation (Crawford RB et al. Mol Pharmacol, 1997; 52:921–927). Western blot analyses determined that although initially expressed in both the cytoplasm and nucleus, AhR became solely nuclear in differentiating CHRF cells. EMSA analysis using CHRF nuclear extracts demonstrated that AhR binding to a consensus binding sequence increased as megakaryopoiesis progressed (n=3). Increased AhR-DNA binding during CHRF Mk differentiation correlated with 4.6-fold increased mRNA expression of the AhR transcriptional target Hes1 (n=3, p<0.005), a known cell-cycle regulator and mediator of notch signaling. In order to examine the functional role of AhR in megakaryopoiesis, we generated 3 independent AhR knockdown (KD) CHRF cell lines. Depending on the day of culture, AhR-KD CHRF cell lines differentiated into Mk cells expressed 2-3 fold less AhR mRNA (n=3; p<0.02), 40–60% less AhR protein (n=3), 2.7 times less Hes1 mRNA (n=3; p=0.018), displayed Mk-ploidy distributions shifted towards lower ploidy classes, and were incapable of reaching higher ploidy classes (i.e., ≥32n) seen in control cells. Ploidy levels on day 7 (maximal ploidy in control cells) were 3-fold lower in AhR-KD CHRF cells (n=3; p=0.012 or p=0.005 depending on KD cell line). AhR KD resulted in increased DNA synthesis of low ploidy (<8n; n=3; p<0.05) without influencing apoptosis (n=3, p=0.37). These data suggest that AhR may regulate the cell cycle differently in Mks compared to other cell types, where loss of AhR results in cell cycle blockage and increased apoptosis. As such, AhR deregulation provides a mechanistic explanation for chemical-induced thrombocytopenia, including chemotherapy, and suggests that AhR agonists may provide novel therapies for megakaryoblastic leukemia. AhR-mediated expression of Hes1, an established regulator of the Notch signaling pathway, provides a novel molecular model of endomitotic entry and Mk polyploidization; in drosophila, Notch cell-cycle regulation controls the initial switch toward endomitosis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals A gene expression network model of type 2 diabetes links cell cycle regulation in islets with diabetes susceptibility

2008 ◽  
Vol 18 (5) ◽  
pp. 706-716 ◽  
Author(s):  
M. P. Keller ◽  
Y. Choi ◽  
P. Wang ◽  
D. Belt Davis ◽  
M. E. Rabaglia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Type 2 Diabetes ◽  
Cell Cycle ◽  
Network Model ◽  
Cell Cycle Regulation ◽  
Diabetes Susceptibility ◽  
Gene Expression Network ◽  
Cycle Regulation

77 Nobiletin supplementation affects gene expression profiles of the Akt pathway in bovine embryos invitro

2020 ◽  
Vol 32 (2) ◽  
pp. 164
Author(s):  
Y. N. Cajas ◽  
K. Cañón-Beltrán ◽  
C. L. V. Leal ◽  
M. E. González ◽  
A. Gutierrez-Adán ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Cell Cycle ◽  
Embryonic Development ◽  
Cell Cycle Regulation ◽  
Akt Pathway ◽  
Cleavage Rate ◽  
Cell Stage ◽  
Cycle Regulation ◽  
And Oxidative Stress

Embryonic genome activation (EGA) is a critical event in early embryonic development and occurs in 8-16-cell stage embryos in bovine. Invitro embryo production increases reactive oxygen species (ROS), leading to low yield and cell death. Nobiletin is an antioxidant that inhibits ROS production and affects cell cycle regulation. The aim of this study was to evaluate the effect of nobiletin supplementation in two key periods of early embryo development on blastocyst yield and expression of candidate genes of the Akt pathway. Invitro-produced zygotes were cultured in synthetic oviductal fluid supplemented with 5% FCS (control; C); C with 5 or 10 µM nobiletin (MedChemExpress; N5, N10) or C with 0.03% dimethyl sulfoxide (CD vehicle for nobiletin dilution) during the minor (2-8-cell stage; MNEGA) or major (8-16-cell stage; MJEGA) phase of EGA, considered as two separate experiments. For all groups, the speed of development was considered, and normally developing embryos that reach ≥8 cells at 54h post-insemination and ≥16 cells at 96h post-insemination were selected and cultured in control medium until Day 8, respectively. Embryos at ≥8-cell stage (N5/N10 MNEGA), 16-cell stage (N5/N10MJEGA), and Day 7 blastocysts of both periods were snap-frozen in LN2 for gene expression analysis. Cleavage rate and blastocyst yield (Day 7-8) were evaluated. The mRNA abundance of candidate genes related to the Akt pathway (CDK2, PGC1A, PPARG, RPS6KB1) and oxidative stress (GPX1) was measured by quantitative PCR. The H2AFZ and ACTB genes were used as housekeeping genes. Statistical analysis was assessed by one-way ANOVA. Nobiletin supplementation during MNEGA showed no differences in cleavage rate, whereas the blastocyst yield at Day 8 was higher (P&lt;0.001) for N5 (42.9±1.4%) and N10 (45.3±2.1%) compared with C (32.9±1.1%) and CD (32.6±1.4%) groups. When nobiletin was supplemented during MJEGA, no differences were found in cleavage rate; however, Day 8 blastocyst yield was higher (P&lt;0.001) for N10 (61.8±0.7%) compared with C (45.2±1.7%), CD (43.6±1.4%), and N5 (52.1±2.1%) groups, whereas N5 was higher (P&lt;0.05) compared with both control groups. The mRNA abundance of CDK2 significantly increases in 8-cell stage embryos from N5 and N10 groups during MNEGA, whereas 16-cell stage embryos from N10 group during MJEGA showed a significant increase compared with both controls (P&lt;0.05). The expression of PGC1A was significantly higher in blastocysts from N5, N10 during MNEGA, and N10 during MJEGA groups compared with both controls (P&lt;0.05). No differences were observed for PPARG and RPS6KB1 in any group from both phases. GPX1, an oxidative indicator gene, was up-regulated in all nobiletin-supplemented groups from both phases compared with controls (P&lt;0.05). In conclusion, supplementation of embryo culture during MNEGA or MJEGA with nobiletin improves embryo development and induces changes in the transcriptional genes related to cell cycle regulation and oxidative stress. This suggests that nobiletin acts through the Akt pathway during the first stages of embryonic development. Funding was provided by MINECO-Spain AGL2015-70140-R&amp;RTI2018-093548-B-I00; Y. N. Cajas, SENESCYT-Ecuador; C. L. V. Leal, FAPESP-Brasil 2017/20339-3.

Sign in / Sign up

Export Citation Format

Share Document