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ABSTRACT: In this study, we investigated the effects of ionizing radiation (IR) on human umbilical vein endothelial cells (HUVECs) in the context of senescence. HUVECs at passage number (PN)1, PN2 and PN3 were exposed to irradiation (2 Gy). The growth rate of the HUVECS was measured by proliferation assay and senescence-associated β-galactosidase assay was used to measure the number of senescent cells. Telomerase activity and the expression of telomerase- and angiogenesis-related genes were measured by telomerase assay and real-time PCR, respectively. The number of senescent cells was significantly increased in the irradiated HUVECs at all PNs. Compared to the controls, telomerase activity, the expression of human telomerase reverse transcriptase (hTERT) and c-Myc in the irradiated HUVECs were downregulated during serial passage. The downregulation of vascular endothelial growth factor (VEGF) was observed in the irradiated HUVECs as the PN increased. The data presented in this study may aid in the understanding of the mechanisms behind IR‑induced EC senescence and telomerase- and angiogenesis‑related gene response.
International Journal of Molecular Medicine 03/2013; · 1.98 Impact Factor
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ABSTRACT: In the present study, we investigated the role of matrix metalloproteinase (MMP)-2 and -9 as novel biomarkers in the body fluid of patients with metastatic breast cancer. We measured the expression of MMPs in 37 samples of body fluid (10 peritoneal and 27 pleural fluids) from metastatic breast cancer patients between 2000 and 2009. Zymography and ELISA assays were used to determine the cut-off level and to quantify MMP expression from a positive control, HT-1080 conditioned media. MMP expression in patient samples was measured with ELISA and compared with other clinical parameters. Ascitic carcinoembryonic antigen (CEA) and pleural CEA were measured in patient samples with a chemiluminescent enzyme immunoassay. Body fluid cytology had a positivity of 45% (9/20) for pleural fluid and 28.6% (2/7) for ascites. However, MMP-2 had a positivity of 85.2% (23/27) in 27 pleural fluid samples and 100% (10/10) in ascitic fluid with cut-off levels of 8.6 and 0.14 ng/ml for MMP-2 and -9, respectively. When body fluid CEA and MMP-2 were combined, the positivity improved to 96% in pleural fluid and 100% in ascites. MMP-2 expression in body fluid did not show any significant differences, but MMP-9 expression was lower in ascites than in pleural fluids (p<0.005). Our results suggest that MMP-2 expression in body fluid be used as an additive diagnostic marker for metastatic breast cancer patients.
Oncology letters 03/2012; 3(3):699-703. · 0.11 Impact Factor
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ABSTRACT: Epigenetic regulations play a role in the development and progression of cancer. Therefore, discovering novel epigenetically regulated genes could provide useful information in understanding cancer. Lamin A/C is an intermediate filament protein whose expression is reported to be suppressed in tissues of gastro-intestinal malignancies. We examined expression of lamin A/C in gastric and colorectal cancer cell lines and its association with DNA methylation.
The methylation status of CpG island in 19 gastric, 5 colorectal cancer cells and 1 normal colon cell line were examined with methylation-specific PCR using paired methylated and unmethylated primers. The level of mRNA expression of lamin A/C was detected using RT-PCR.
Eighteen gastric cancer cell lines showed 95% unmethylation of lamin A/C and 1 cell line showed partial methylation. In colorectal cancer, only 1 out of 5 cancer cell lines (20%) was partially methylated and the remaining cell lines, including 1 normal colon cell line was unmethylated. With RT-PCR, all cell lines demonstrated mRNA expression of lamin A/C regardless of methylation status.
We observed that the expression of lamin A/C was not suppressed in gastrointestinal cancer cell lines different from hematologic malignant cells and it is not regulated through DNA methylation.
Hepato-gastroenterology 11/2011; 59(116):1313-8. · 0.66 Impact Factor
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ABSTRACT: Gastric cancer (GC) is a heterogeneous disease that is not well detected by current tumor markers. Identifying molecular markers that can predict the potential for tumor progression is important for appropriate individualized therapy. Using the Cancer Metastasis Research Center microarray database (17K cDNA microarray), we identified genes that were differentially expressed between 96 cancer and 98 normal gastric tissues using significant analysis of microarrays. From these, we selected genes that were overexpressed more than twofold in tumor tissues that encode secreted proteins. The selected genes were validated with ELISA using the sera of 96 GC patients and 48 healthy donors. Our first round of selection included 6510 genes that were differentially expressed between 96 cancer and 98 normal gastric tissues with a minimal false discovery rate of 0.005%. Out of those genes, we picked 386 that encoded secreted proteins based on the SOURCE database. Of these genes, we focused on 55 that were overexpressed more than twofold in GC compared to normal tissues. With Ingenuity Pathway Analysis, we found 34 genes related to cancer. One in particular, chemokine growth-regulated oncogene 1, CXCL1, has been linked to cancer progression in various cancer types, but not yet to GC. Levels of CXCL1 in serum samples of GC patients were significantly higher compared with healthy donors (P < 0.05). Within GC patients, CXCL1 serum levels increased according to tumor stage and lymph node metastasis. The CXCL1 gene appears to be a candidate marker for GC progression.
Cancer Science 10/2010; 101(10):2200-6. · 3.33 Impact Factor
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ABSTRACT: Here we evaluated the cytotoxic effects of a combination of the histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) and taxanes in human breast cancer cell lines. Combination treatment with taxane and SAHA had a synergistic cytotoxic effect against taxane-resistant breast cancer cells. Oligonucleotide microarray analysis identified 28 genes (MAPK13, ATP2C1, ANKRD57, MT1G, RGL4, C12orf49, EXOC6, RAB4A, TM9SF3, IFNGR1, DMD, HCG9, KIFC3, SYNGR3, NDRG4, NT5E, EOMES, SMC4, LANCL1, SCHIP1, and 8 ESTs) whose expression correlated with the combined effect of paclitaxel and SAHA. Twelve of these genes were down-regulated in cell lines that were paclitaxel-resistant but combination synergistic. SAHA induced NT5E mRNA expression in paclitaxel-resistant YCC-B1 cell. Our results indicate that a combination of taxane and SAHA could be efficacious for the treatment of breast cancer and that genes involved in the synergistic response to paclitaxel and SAHA could serve as biomarkers to predict therapeutic response in breast cancer patients.
Breast Cancer Research and Treatment 03/2010; 125(1):55-63. · 4.43 Impact Factor
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ABSTRACT: We evaluated the cytotoxic effects of combining suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, with taxanes in human gastric cancer cell lines and assessed the pre-treatment difference of gene expression to identify genes that could potentially mediate the cytotoxic response.
Gastric cancer cell lines were treated with SAHA and paclitaxel or docetaxel, and the synergistic interaction between the drugs was evaluated in vitro using the combination index (CI) method. We performed significance analysis of microarray (SAM) to identify chemosensitivity-related genes in gastric cancer cell lines that were concomitantly treated with SAHA and taxane. We generated a correlation matrix between gene expression and CI values to identify genes whose expression correlated with a combined effect of taxanes and SAHA.
Combination treatment with taxane and SAHA had a synergistic cytotoxic effect against taxane-resistant gastric cancer cells. We identified 49 chemosensitivity-related genes via SAM analysis. Among them, nine common genes (SLIT2, REEP2, EFEMP2, CDC42SE1, FSD1, POU1F1, ZNF79, ETNK1, and DOCK5) were extracted from the subsequent correlation matrix analysis.
The combination of taxane and SAHA could be efficacious for the treatment of gastric cancer. The genes that were related to the synergistic response to taxane and SAHA could serve as surrogate biomarkers to predict the therapeutic response in gastric cancer patients.
Journal of Cancer Research and Clinical Oncology 03/2010; 136(12):1901-13. · 2.56 Impact Factor
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ABSTRACT: It has been suggested that vasculogenesis by endothelial progenitor cells (EPC) as well as angiogenesis play an important role in the production of blood vessels in neoplasm. The present study was designed to isolate and characterize the EPC in gastric cancer patients as a tumor specific angiogenesis marker. The cells derived from CD34 positive PBMC presented with a cobblestone appearance at 28 days, revealing differentiation into endothelial cells. They were also positive to the LDL-uptake reaction, showing that they have biological endothelial cell functions. These cells demonstrated tube formation, showing their ability to participate in neovascularization. The cells derived from CD34 positive PBMC expressed CD133 and demonstrated telomerase activity, showing the stem cell character. In xenograft model, EPC derived from CD34 positive PBMC mobilized mainly into tumor area after being injected through tail vein. With isolation, ex vivo amplification and characterization of EPC from gastric cancer patients receiving chemotherapy, endothelial progenitor cells may be used as a candidate prognostic and predictive biomarker for cancer.
Cancer letters 08/2009; 288(1):124-32. · 4.86 Impact Factor
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ABSTRACT: We investigated the biological role of thymidine phosphorylase (TP), an angiogenic factor, in gastric cancer cell migration and invasion and explored a therapeutic approach for high TP-expressing tumors using TP enzymatic inhibitor (TPI) and rapamycin. We established TP cDNA overexpressing gastric cancer cell lines (MKN-45/TP and YCC-3/TP) and did invasion and adhesion assays with Matrigel-coated transwell membranes. The related signal pathway using recombinant human TP (rhTP), deoxy-d-ribose (D-dRib), and signal pathway inhibitors (wortmannin, LY294002, and rapamycin) was investigated. First, AGS and MKN-1 gastric cancer cell lines showed dose-dependent up-regulation of invasiveness through Matrigel following treatment with rhTP or D-dRib. TP-overexpressing cancer cell lines displayed increased migration and invasion activity, which doubled with rhTP and D-dRib treatment. This activity depended on the enzymatic activity of TP, and TP stimulated the adhesion of cancer cells onto Matrigel and induced actin filament remodeling. Finally, we showed that this activity is related to increased phosphatidylinositol 3-kinase activity in TP-overexpressing cells and that combination treatment with rapamycin and TP enzymatic inhibitor produces an additive effect to abrogate TP-induced invasion. Taken together, TP increases the migration and invasion of gastric cancer cells, especially in TP-expressing cells. Therapies targeting TP might diminish the propensity for invasion and metastasis in gastric cancer.
Molecular Cancer Research 11/2008; 6(10):1554-66. · 4.29 Impact Factor
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ABSTRACT: Gastric cancer is one of the most common cancers worldwide, and there are clinical caveats in predicting tumor response to chemotherapy. This study describes the construction of an in vitro pharmacogenomic database, and the selection of genes associated with chemosensitivity in gastric cancer cell lines. Gene expression and chemosensitivity databases were integrated using the Pearson correlation coefficient to give the GC-matrix. The 85 genes were selected that were commonly associated with chemosensitivity of the major anticancer drugs. We then focused on the genes that were highly correlated with each specific drug. Classification of cell lines based on the set of genes associated with each drug was consistent with the division into resistant or sensitive groups according to the chemosensitivity results. The GC-matrix of the gastric cancer cell line database was used to identify different sets of chemosensitivity-related genes for specific drugs or multiple drugs.
Genomics 10/2008; 93(1):52-61. · 3.02 Impact Factor
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ABSTRACT: The telomeric G-rich 3' overhang is important for the maintenance of chromosomal integrity by stabilizing T-loop structure in which the 3' overhang invades the double-stranded telomeric DNA. However, the 3' overhang length has not been examined in different human cell lines, and its regulatory mechanism has not been revealed. In this study, we examined overhang length in 56 human cancerous cell lines and five normal cell lines, originated from various tissues. In cancer cells, relative overhang length existed in a wide range from 23% to 308% and showed no significant association with tissue types although short overhang was noted in brain, cervix, and colorectal cells. Normal cells exhibited overhangs in the range from 92% to 202%, which were relatively longer than those seen in cancer cells (p = 0.002). The overhang length was positively correlated with telomere length (p < 0.001), and showed no correlation with mRNA levels of hTERT, a catalytic protein of telomerase, POT1, an overhang binding protein and TPP1, a POT1 interacting protein. This study demonstrates a broad distribution of overhang length in human cells, suggesting a dynamic regulation of 3' overhang length. The overhang length seems to be closely associated with telomere length and might be regulated by multiple mechanisms.
Cancer Letters 07/2008; 264(1):107-18. · 4.24 Impact Factor
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ABSTRACT: In order to identify genes which could predict chemosensitivity in colorectal cancer, gene expression and chemosensitivity were examined in colorectal cancer cell lines. Gene expression profiling of 5 colorectal cancer and 3 normal cell lines was performed using a 22K spotted oligonucleotide microarray. The IC50s of 17 anticancer drugs were determined using the MTT assay for chemosensitivity. The SOURCE database, KEGG Pathway database, and Molecular Diagnosis Score (MDS) were used for data analysis. Two representative colorectal cancer cell lines were identified which were resistant or sensitive to drugs commonly used for colon cancer treatment (5-FU, irinotecan and topotecan). Six hundred and eighty-three genes that were up- or down-regulated by >4-fold between the two cell lines were selected. Pathway analysis was performed with 147 of the 683 genes using the KEGG Pathway database. This analysis revealed 27 genes in the apoptosis, MAPK signaling, and focal adhesion pathways, which could explain the mechanism of chemosensitivity in colorectal cancer cell lines. In addition, the chemosensitivity of other colorectal cancer and normal cell lines was predictable with the selected 27 genes. These genes may act as putative predictive markers for chemosensitivity in chemo-naive colorectal cancer patients following functional analysis and clinical validation.
Oncology Reports 09/2007; 18(3):593-9. · 1.84 Impact Factor
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ABSTRACT: Significant variability in the efficacy and toxicity of an anticancer drug is observed in cancer patients. Currently, there are no standard tools for prediction of a patient's tumor response or his risk of adverse events to chemotherapy.
We investigate an association between polymorphisms of gemcitabine metabolism-related genes and its chemosensitivity in vitro using 62 human cancer cell lines of various origins. Polymorphisms of gemcitabine metabolism-related genes of deoxycytidine monophosphate deaminase (DCTD), deoxycytidine kinase (DCK) and ribonucleotide reductase M1 (RRM1) were evaluated using the CEQ8000 Genetic analysis system and GeneDoc software. Chemosensitivity of gemcitabine was expressed as an IC50 using MTT assay.
The frequency of the polymorphisms was 21% in DCTD 315T>C, 45.2% in RRM1 1082C>A, 59.7% in RRM1 2455A>G, and 79% in RRM1 2464G>A. When examining the association between these polymorphisms and IC50, only the RRM1 2464G>A showed the tendency to be more chemosensitive to gemcitabine (P=0.011), and haplotypes containing 2464G>A polymorphism also showed the association with chemosensitivity when compared to wild-type RRM1 (G2464G). We could not see the significant differences of mRNA expression level with real-time PCR between cell lines according to G2464A polymorphism. In oligonucleotide microarray 73 GenBank Accession Number (69 genes) were selected which expressed differently between RRM1 wild-type and the G2464A polymorphism.
RRM1 2464G>A polymorphism demonstrated an association with gemcitabine sensitivity, which needs functional studies with co-expressing genes and prospective clinical studies for the clinical application as a predictive bio-marker.
Pharmacogenetics and Genomics 07/2006; 16(6):429-38. · 3.48 Impact Factor
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Chan Hee Park,
Ha Jin Jeong,
Yeon Ho Choi,
Sang Cheol Kim,
Hei Chul Jeong,
Kyu Hyun Park,
Gui Yeon Lee, Tae Soo Kim,
Sang Wha Yang,
Sung Whan Ahn,
Yang Seok Kim,
Sun Young Rha,
Hyun Cheol Chung
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ABSTRACT: cDNA microarray-based CGH (Microarray-CGH) is a useful technique for detecting genomic aberrations with a high resolution. However, the criteria for determining a genomic alteration have not been determined. We evaluated the genome-wide measurement of copy number of each gene in normal gastric and placenta tissues with both sex-matched, direct and sex-mismatched, indirect designs using 17K cDNA microarray. The results revealed the range of genomic copy number of normal tissues to be +/-0.3 of the log(2) ratio (gain >0.3, loss <-0.3) in the autosomal genes with direct and indirect designs. The copy number at a gene level from the X chromosomal genes using the direct and indirect sex-mismatched designs was +/-0.68 of the log(2) ratio (amplification >0.68, deletion <-0.68). In summary, the suggested method can be used as a guideline for analysis of genomic aberration using a Microarray-CGH in both direct and indirect designs.
International Journal of Molecular Medicine 02/2006; 17(2):261-7. · 1.98 Impact Factor
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Woo Young Shim,
Kyu Hyun Park,
Hei-Chul Jeung,
Yong Tae Kim, Tae Soo Kim,
Woo Jin Hyung,
Sung Hwan An,
Sang Hwa Yang,
Sung Hoon Noh,
Hyun Cheol Chung,
Sun Young Rha
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ABSTRACT: Real-time TRAP assay was developed to improve the sensitivity and quantitative detection of telomerase activity in the body fluids of cancer patients. The sensitivity and clinical significance of the real-time TRAP assay was investigated. Real-time PCR protocol was modified by using ACX primer and SYBR green mixture from the process of TS primer extension. Real-time TRAP showed high correlation (r2=0.843, p=0.001) and sensitivity (25 times higher) compared to conventional TRAP. Of 164 samples, there were 8 positives in cytology (4.9%), 7 (4.3%) using the conventional TRAP, and 41 (25%) using real-time TRAP. In cytology positive samples, real-time TRAP showed a higher positivity than conventional TRAP (75% vs 63%) suggesting a higher sensitivity in the body fluids. There was a tendency towards a longer progression-free duration in telomerase negative patients than in positive patients, as determined by conventional and real-time TRAP. The diagnostic interval between the first positivity documentation and clinical progression was short in the order of real-time TRAP, conventional TRAP and cytology. In conclusion, real-time TRAP assay can detect telomerase activity at an earlier phase of cancer progression and can replace conventional TRAP assay for detecting the telomerase activity in body fluids.
International Journal of Molecular Medicine 12/2005; 16(5):857-63. · 1.98 Impact Factor
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ABSTRACT: We evaluated the genome-wide gene expression profiles of various cancer cell lines to identify the gastrointestinal tract cancer cell-related genes.
Gene expression profilings of 27 cancer cell lines and 9 tissues using 7.5K human cDNA microarrays in indirect design with Yonsei reference RNA composed of 11 cancer cell line RNAs were done. The significant genes were selected using significant analysis of microarray in various sets of data. The selected genes were validated using real-time PCR analysis.
After intensity-dependent, within-print-tip normalization by loess method, we observed that expression patterns of cell lines and tissues were substantially different, divided in two discrete clusters. Next, we selected 115 genes that discriminate gastrointestinal cancer cell lines from others using significant analysis of microarray. Among the expression profiles of five gastric cancer cell lines, 66 genes were identified as differentially expressed genes related to metastatic phenotype. YCC-16, which was established from the peripheral blood of one advanced gastric cancer patient, produced a unique gene expression pattern resembling the profiles of lymphoid cell lines. Quantitative real-time reverse transcription-PCR results of selected genes, including PXN, KRT8, and ITGB5, were correlated to microarray data and successfully discriminate the gastrointestinal tract cancer cell lines from hematologic malignant cell lines.
A gene expression database could serve as a useful source for the further investigation of cancer biology using the cell lines.
Clinical Cancer Research 02/2005; 11(1):79-86. · 7.74 Impact Factor
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ABSTRACT: DNA microarray technology has become an essential part of biological research. It enables the genome-scale analysis of gene expression in various types of model systems. Manufacturing high quality cDNA microarrays of microdeposition type depends on some key factors including a printing device, spotting pins, glass slides, spotting solution, and humidity during spotting. UsingEthe Microgrid II TAS model printing device, this study defined the optimal conditions for producing high density, high quality cDNA microarrays with the least amount of cDNA product. It was observed that aminosilane-modified slides were superior to other types of surface modified-slides. A humidity of 30+/-3% in a closed environment and the overnight drying of the spotted slides gave the best conditions for arraying. In addition, the cDNA dissolved in 30% DMSO gave the optimal conditions for spotting compared to the 1X ArrayIt, 3X SSC and 50% DMSO. Lastly, cDNA in the concentration range of 100-300 ng/ micro l was determined to be best for arraying and post-processing. Currently, the printing system in this study yields reproducible 9000 spots with a spot size 150 mm diameter, and a 200 nm spot spacing.
International Journal of Molecular Medicine 06/2004; 13(5):675-9. · 1.98 Impact Factor
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ABSTRACT: We evaluated matrix metalloproteinase (MMP)-2 and -9 as novel biomarkers in the body fluid of advanced gastric cancer with peritoneal and pulmonary metastasis.
MMPs activity from zymography was quantified with ELISA to determine the cut-off expression levels of MMPs. The expression of MMPs in patient samples were evaluated with ELISA and compared with clinical parameters. Ascitic CEA (aCEA) and pleural CEA (pCEA) were measured by chemiluminescent enzyme immunoassay.
MMP-2 and -9 cut-off levels were 8.6ng/mL and 0.14ng/mL, respectively. Ascitic fluid cytology of 93 patients revealed a positivity of 55.9% while for MMP-2 it was 93.3%, for MMP-9 35.2% and for aCEA 86.7%. Combining biomarkers, the positivity increased to 99.1% in patients with MMP-2 or aCEA expression. We found a negative correlation between MMP-2 expression and survival when a new prognostic cut-off of 22.6ng/mL was used. Patients with high MMP-2 expression (≥22.6ng/mL) had a median survival of 45 days and those with low MMP-2 expression (<22.6ng/mL) had a median survival of 69 days (p<0.01).
These results suggest that MMPs could be used as diagnostic markers in body fluid and MMP-2 might be a prognostic marker in ascites of advanced gastric patients with disseminated metastasis.
Hepato-gastroenterology 58(112):2015-9. · 0.66 Impact Factor
