Identification of gene expression determinants of radiosensitivity in bladder cancer (BCa) cell lines.

470 Background: Trimodality therapy with TURBT followed by concurrent chemoradiation is an alternative to cystectomy in selected patients with muscle invasive BCa. Identifying genetic determinants of radiation response may help select patient for organ-sparing treatments with curative intent. Methods: Molecular characterization of 20 cell lines was previously performed, including mutation analysis, CNA (high density SNP arrays) and gene expression (RNAseq). Cell line identity was authenticated by genomic fingerprinting. We performed colony forming assays (CFA) and cumulative survival was quantified using the area under the cell survival curve (AUC, range 0-8Gy) to create a radiosensitivity index. Pre and post-radiation proliferative capacity was determined by cell titer glo (CTG) assay. Gene (R v3.5.1) and network (Ingenuity Pathyway Analysis) level expression differences were determined as a function of AUC to identify biologically relevant associations with radiation response. Results: Of 20 cell lines (7 basal, 13 luminal subtype), both CFA and CTG data was successfully obtained for 13 cell lines (6 basal, 7 luminal) treated with high dose rate (HDR) radiation. AUCs for the colony forming assay (CFA) survival curves ranged 1.79-3.27. RNAseq identified 18,634 unique transcripts mapping to distinct loci and 196 genes were identified with AUC correlation p values <0.01 (FDR <0.5, mean FPKM>0.5). These genes were strongly enriched for membership in the peroxisome proliferator (PPAR) pathway (IPA, p = 9.40E-03) and STAT3 pathway (IPA, p = 1.56E-3). Validation studies confirmed PPARγ, ICAM2, JAK3, IL1B, OAS1 and OAS2 genes to be associated with differential response to radiation, with expression upregulated in radio-resistant cells. A basal subtype was strongly associated with a radiosensitive phenotype (Chi-Squared χ2 p=0.0083). Conclusions: BCa radio-sensitivity was associated with baseline gene expression differences in vitro. The basal subtype and repression of either the PPAR or STAT3 pathways predicted increased radiosensitivity. This study nominates candidate biomarkers for mechanistic studies and clinical validation in BCa.

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Anti-Apoptotic Proteins, HSP90 and Activated STAT3 Contribute to Busulfan Resistance of Myeloid Leukemia Cells.

Blood ◽

10.1182/blood.v110.11.3472.3472 ◽

2007 ◽

Vol 110 (11) ◽

pp. 3472-3472

Author(s):

Borje S. Andersson ◽

Ben C. Valdez ◽

David Murray ◽

Latha Ramdas ◽

Marcos de Lima ◽

...

Keyword(s):

Gene Expression ◽

Cell Lines ◽

Leukemia Cell ◽

Myelogenous Leukemia ◽

Leukemia Cells ◽

Treatment Modalities ◽

High Dose ◽

Hematopoietic Stem ◽

Conditioning Treatment ◽

Apoptotic Genes

Abstract Busulfan(Bu)-based chemotherapy is a conditioning treatment prior to hematopoietic stem cell transplantation (HSCT) of patients with acute and chronic myelogenous leukemia (AML, CML). A major obstacle to successful HSCT is Bu resistance, which might be attributed to individual differences in drug pharmacokinetics and metabolism, or inherent resistance of cancer cells. We hypothesize that gene expression profiling of leukemia cells can be used to dissect the factors that contribute to their Bu resistance. Two Bu-resistant leukemia cell lines were established, characterized and analyzed by microarray and real-time RT-PCR techniques to identify differentially expressed genes. The CML B5/Bu2506 cells are 4.5-fold more resistant to Bu than their parental B5 cells. The AML KBM3/Bu2506 cells are 4.0-fold more Bu-resistant than KBM3 parental cells. Both resistant sublines evade Bu-mediated G2-arrest and apoptosis with constitutively up-regulated anti-apoptotic genes (BCL-XL, BCL2, BCL2L10, BAG3 and IAP2/BIRC3) and down-regulated pro-apoptotic genes (BIK, BNIP3, and LTBR).). Bu-induced apoptosis is partly mediated by activation of caspases; use of the inhibitor Z-VAD-FMK completely abrogated PARP1 cleavage and reduced apoptosis by ∼ 50%. Furthermore, Bu resistance in these cells may be attributed in part to up-regulation of HSP90 protein and activation of STAT3. Inhibition of HSP90 with geldanamycin attenuated phosphorylated STAT3 and made B5/Bu2506 and KBM3/Bu2506 more Bu-sensitive. Analysis of cells derived from patients classified as either clinically resistant or sensitive to high-dose Bu-based chemotherapy had alterations in gene expression that were analogous to those observed in the in-vitro model cell lines, confirming the potential clinical relevance of this model for Bu resistance. Our results suggest the important roles of apoptotic signaling mechanism, HSP90 and STAT3 and should be considered in the classification of AML patients who will likely benefit from busulfan-based pretransplant conditioning therapy and those who should be offered alternative treatment modalities.

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Restoration of sterol-regulatory-element-binding protein-1c gene expression in HepG2 cells by peroxisome-proliferator-activated receptor-γ co-activator-1α

Biochemical Journal ◽

10.1042/bj20040173 ◽

2004 ◽

Vol 381 (2) ◽

pp. 357-363 ◽

Cited By ~ 19

Author(s):

Hannes OBERKOFLER ◽

Elisabeth SCHRAML ◽

Franz KREMPLER ◽

Wolfgang PATSCH

Keyword(s):

Gene Expression ◽

Cell Lines ◽

Hepg2 Cells ◽

Binding Protein ◽

Regulatory Element ◽

Brown Adipocyte ◽

Peroxisome Proliferator ◽

Peroxisome Proliferator Activated Receptor ◽

Element Binding Protein ◽

Sterol Regulatory Element

The expression of SREBP-1 (sterol-regulatory-element-binding protein-1) isoforms differs between tissues and cultured cell lines in that SREBP-1a is the major isoform in established cell lines, whereas SREBP-1c predominates in liver and most other human tissues. SREBP-1c is transcriptionally less active than SREBP-1a, but is a main mediator of hepatic insulin action and is selectively up-regulated by LXR (liver X receptor) agonists. LXR-mediated transactivation is co-activated by PGC-1α (peroxisome-proliferator-activated receptor-γ co-activator-1α), which displays deficient expression in skeletal-muscle-derived cell lines. In the present paper, we show that PGC-1α expression is also deficient in HepG2 cells and in a human brown adipocyte cell line (PAZ6). In transient transfection studies, PGC-1α selectively amplified the LXR-mediated transcription from the human SREBP-1c promoter in HepG2 and PAZ6 cells via two LXR-response elements with extensive similarity to the respective murine sequence. Mutational analysis showed that the human LXR-response element-1 (hLXRE-1) was essential for co-activation of LXR-mediated SREBP-1c gene transcription by PGC-1α. Ectopic overexpression of PGC-1α in HepG2 cells enhanced basal SREBP-1c and, to a lesser extent, -1a mRNA expression, but only SREBP-1c expression was augmented further in an LXR/RXR (retinoic X receptor)-dependent fashion, thereby inducing mRNA abundance levels of SREBP-1c target genes, fatty acid synthase and acetyl-CoA carboxylase. These results indicate that PGC-1α contributes to the regulation of SREBP-1 gene expression, and can restore the SREBP-1 isoform expression pattern of HepG2 cells to that of human liver.

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Stable reporter cell lines for peroxisome proliferator-activated receptor γ (PPARγ)-mediated modulation of gene expression

Analytical Biochemistry ◽

10.1016/j.ab.2011.02.032 ◽

2011 ◽

Vol 414 (1) ◽

pp. 77-83 ◽

Cited By ~ 33

Author(s):

Linda Gijsbers ◽

Hai-Yen Man ◽

Samantha K. Kloet ◽

Laura H.J. de Haan ◽

Jaap Keijer ◽

...

Keyword(s):

Gene Expression ◽

Cell Lines ◽

Peroxisome Proliferator ◽

Reporter Cell ◽

Peroxisome Proliferator Activated Receptor ◽

Modulation Of Gene Expression

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Troglitazone and pioglitazone interactions via PPAR-γ-independent and -dependent pathways in regulating physiological responses in renal tubule-derived cell lines

AJP Cell Physiology ◽

10.1152/ajpcell.00396.2006 ◽

2007 ◽

Vol 292 (3) ◽

pp. C1137-C1146 ◽

Cited By ~ 27

Author(s):

Francesco Turturro ◽

Robert Oliver ◽

Ellen Friday ◽

Itzhak Nissim ◽

Tomas Welbourne

Keyword(s):

Gene Expression ◽

Cell Lines ◽

Biological Effects ◽

Cellular Growth ◽

Glutamine Metabolism ◽

Peroxisome Proliferator ◽

Renal Tubules ◽

Erk Activation ◽

Ppar Γ

Troglitazone (Tro) and pioglitazone (Pio) activation of peroxisome proliferator-activated receptor (PPAR)-γ and PPAR-γ-independent pathways was studied in cell lines derived from porcine renal tubules. PPAR-γ-dependent activation of PPAR response element-driven luciferase gene expression was observed with Pio at 1 μM but not Tro at 1 μM. On the other hand, PPAR-γ-independent P-ERK activation was observed with 5 μM Tro but not with Pio (5–20 μM). In addition, Pio (1–10 μM) increased metabolic acid production and activated AMP-activated protein kinase (AMPK) associated with decreased mitochondrial membrane potential, whereas Tro (1–20 μM) did not. These results are consistent with three pathways through which glitazones may act in effecting metabolic processes (ammoniagenesis and gluconeogenesis) as well as cellular growth: 1) PPAR-γ-dependent and PPAR-γ-independent pathways, 2) P-ERK activation, and 3) mitochondrial AMPK activation. The pathways influence cellular acidosis and glucose and glutamine metabolism in a manner favoring reduced plasma glucose in vivo. In addition, significant interactions can be demonstrated that enhance some physiological processes (ammoniagenesis) and suppress others (ligand-mediated PPAR-γ gene expression). Our findings provide a model both for understanding seemingly opposite biological effects and for enhancing therapeutic potency of these agents.

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Antagonistic Property of G2013 (α-L-Guluronic Acid) on Gene Expression of MyD88, Tollip, and NF-κB in HEK293 TLR2 and HEK293 TLR4

Endocrine Metabolic & Immune Disorders - Drug Targets ◽

10.2174/1871530319666181126153752 ◽

2019 ◽

Vol 19 (2) ◽

pp. 144-149 ◽

Cited By ~ 1

Author(s):

Laleh Sharifi ◽

Asghar Aghamohammadi ◽

Somaye Aletaha ◽

Razieh Bigdeli ◽

Vahid Asgary ◽

...

Keyword(s):

Oxidative Stress ◽

Gene Expression ◽

Cell Lines ◽

Low Dose ◽

Signaling Cascade ◽

High Dose ◽

Quantitative Real Time Pcr ◽

Gene Expressions ◽

Guluronic Acid ◽

Immunosuppressive Property

Introduction: Inhibition of Toll-like receptors (TLRs) signaling plays a crucial role in suppressing the inflammation and available data presenting G2013 as an immunomodulatory agent, therefore, we designed this study to answer whether G2013 can affect the signaling pathway of TLR2 and TLR4. Methods: Cytotoxicity study of G2013 was performed by MTT assay. HEK293 TLR2 and HEK293 TLR4 cell lines were cultured and treated with low dose (5µg/ml) and high dose (25µg/ml) of G2013 for 24 hours. Gene expressions of MyD88, Tollip, and NF-κB were defined by quantitative real-time PCR. Results: The cytotoxicity assay showed that the concentrations lesser than 125μg/ml of G3012 had no apparent cytotoxicity, however, the concentrations of 5µg/ml and 25µg/ml could suppress the mRNA expression of MyD88, Tollip and NF-κB in HEK293 TLR2 and HEK293 TLR4 cell lines. Conclusion: in our study, we verified the linkage between the immunosuppressive property of G2013 and TLR2, TLR4 signaling cascade; but so far, the specific target of G2013 and its molecular mechanism has not been detected yet. We recommend further studies on other Patten Recognition Receptors (PRRs)and other mechanisms of inflammation like oxidative stress to be conducted in the future.

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Multidrug Resistance Protein-4 Influences Aspirin Toxicity in Human Cell Line

Mediators of Inflammation ◽

10.1155/2015/607957 ◽

2015 ◽

Vol 2015 ◽

pp. 1-9 ◽

Cited By ~ 10

Author(s):

Isabella Massimi ◽

Ambra Ciuffetta ◽

Flavia Temperilli ◽

Francesca Ferrandino ◽

Alessandra Zicari ◽

...

Keyword(s):

Gene Expression ◽

Multidrug Resistance ◽

High Dose ◽

Peroxisome Proliferator ◽

Multidrug Resistance Protein ◽

Cell Toxicity ◽

Hek 293 ◽

Dose Aspirin ◽

Resistance Protein ◽

High Dose Aspirin

Overexpression of efflux transporters, in human cells, is a mechanism of resistance to drug and also to chemotherapy. We found that multidrug resistance protein-4 (MRP4) overexpression has a role in reducing aspirin action in patients after bypass surgery and, very recently, we found that aspirin enhances platelet MRP4 levels through peroxisome proliferator activated receptor-α(PPARα). In the present paper, we verified whether exposure of human embryonic kidney-293 cells (Hek-293) to aspirin modifies MRP4 gene expression and its correlation with drug elimination and cell toxicity. We first investigated the effect of high-dose aspirin in Hek-293 and we showed that aspirin is able to increase cell toxicity dose-dependently. Furthermore, aspirin effects, induced at low dose, already enhance MRP4 gene expression. Based on these findings, we compared cell viability in Hek-293, after high-dose aspirin treatment, in MRP4 overexpressing cells, either after aspirin pretreatment or in MRP4 transfected cells; in both cases, a decrease of selective aspirin cell growth inhibition was observed, in comparison with the control cultures. Altogether, these data suggest that exposing cells to low nontoxic aspirin dosages can induce gene expression alterations that may lead to the efflux transporter protein overexpression, thus increasing cellular detoxification of aspirin.

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