|ognizant Communication Corporation|
Featuring PRECLINICAL AND CLINICAL CANCER THERAPEUTICS
VOLUME 18, NUMBERS 2-3
Oncology Research, Vol. 18, pp. 47-56
0965-0407/09 $90.00 + .00
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Identification of Genes With Differential Expression in Chemoresistant Epithelial Ovarian Cancer Using High-Density Oligonucleotide Microarrays
Woong Ju,1 Byong Chul Yoo,2 Il-Jin Kim,3 Jae Weon Kim,4 Seung Cheol Kim,1 and Hyo Pyo Lee5
1Department of Obstetrics and Gynecology and Medical Research
Institute, College of Medicine, Ewha Womans University, Seoul, South Korea
2Research Institute and Hospital, National Cancer Center, Gyeonggi, South Korea
3Cancer Research Institute, University of California at San Francisco, San Francisco, CA, USA
4Department of Obstetrics and Gynecology, Cancer Research Institute, and Human Genome Research Institute, College of Medicine, Seoul National University, Seoul, South Korea
5Department of Obstetrics and Gynecology, Konkuk University Hospital, Seoul, South Korea
A major obstacle in treatment of epithelial ovarian cancer is chemoresistance. The aim of this study was to determine whether distinct gene expression profiles are associated with chemoresistance in epithelial ovarian carcinoma. We performed global gene expression analysis in 13 primary epithelial ovarian cancer tissues including 5 primary chemosensitive tumors and 8 primary chemoresistant tumors using Affymetrix HGU133A microarray. The gene expression patterns of chemosensitive tumors were compared with those of chemoresistant tumors using fold change. Validity of microarray results was examined by semiquantitative RT-PCR. We identified over 320 genes differentially expressed in chemoresistant epithelial ovarian cancer (>=twofold). Upregulated genes in chemoresistant tumors included cell cycle regulating genes (TOP2A, BCAT1, CDCA8, CCNA2, CENPE), and genes with previously known mechanisms in tumorigenesis (S100A9, APOA1, RNF125, IFI16). Downregulated genes in chemoresistant tumors included genes related to cell adhesion (MUC5B, CITED2), transcription regulating genes (FOXD1, MAD1L1, PAX2), genes involving signal transduction (SOSTDC1, SNX1, SFRP1, FOXA2, PLK2), and stress protein gene (TP53AP1). These data show that gene expression profiling can discriminate primary chemoresistant from primary chemosensitive ovarian cancers. This type of molecular profiling could provide a basis for additional functional studies.
Key words: Ovarian cancer; Gene expression; Chemoresistant tumors; Chemosensitive tumors
Address correspondence to Woong Ju, M.D., Ph.D., Department of Obstetrics and Gynecology and Medical Research Institute, College of Medicine, Ewha Womans University, 911-1 Mok-Dong, Yangcheon-Ku, Seoul, 158-710, South Korea. Tel: +82-2-2650-2779; Fax: +82-2-2647-9860; E-mail: email@example.com
Association Between MTHFR Polymorphism (C677T) With Nonfamilial Colorectal Cancer
Mahdi Montazer Haghighi,1 Ramin Radpour,2 Touraj Mahmoudi,1 Seyed Reza Mohebbi,1 Mohsen Vahedi,1 and Mohammad Reza Zali1
1Research Center for Gastroenterology and Liver Diseases,
Taleghani Hospital, Shaheed Beheshti Medical University, Tehran, Iran
2Laboratory for Prenatal Medicine and Gynecologic Oncology, Women's Hospital/Department of Biomedicine, University of Basel, Basel, Switzerland
The enzyme 5,10-methylene-tetrahydrofolate reductase (MTHFR) is linked to DNA methylation, synthesis, and repair. C677T is one of the most important polymorphisms in the MTHFR gene. The single nucleotide polymorphism C677T has been found to be associated with decreased enzyme activity and plasma folate, and thus may play a crucial role in the etiology of colorectal cancer. This decrease was observable in people with either high or low folate status. We aimed to test the hypothesis that the C677T genotype is involved in colorectal cancer. Using pyrosequencing, we analyzed the MTHFR genotypes in 234 colorectal cancer patients and 257 matched controls. We examined the polymorphisms in MTHFR and folate intake in relation to risk of colon cancer in an Iranian population-based case-control study. Our finding revealed that the CC, CT, and TT genotypes of MTHFR among the colorectal cancer patients were 50%, 29.1%, and 20.9%, respectively. On the other hand, we could find 29.5% of 677CC, 46% of 677CT and 24.5% of 677TT in the controls. A decreased risk of colon cancer for participants with wild-type genotype was observed. Interestingly, this association was stronger at higher levels of folate intake. Our study corroborates previous findings of an inverse association of the MTHFR 677TT genotype with colorectal cancer, in particular at high levels of folate.
Key words: Nonfamilial colorectal cancer; Methylenetetrahydrofolate reductase (MTHFR); Pyrosequencing; Polymorphism; Folate
Address correspondence to Dr. Mahdi Montazer Haghighi, Research Center for Gastroenterology and Liver Diseases, Taleghani Hospital, Shaheed Beheshti Medical University, 1985711151 Tehran, Iran. Tel: +98 21 22432515; Fax: +98 21 22432517; E-mail: firstname.lastname@example.org
Polymorphism of FGFR4 Gly388Arg Does Not Confer an Increased Risk to Breast Cancer Development
R. Naidu,1 Y. C. Har,2,3 and N. A. Taib2,3
1School of Medicine and Health Sciences, Monash University
Sunway Campus, Selangor Darul Ehsan, Malaysia
2Department of Surgery, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia
3University Malaya Medical Center, University of Malaya, Kuala Lumpur, Malaysia
The genotype analysis of the Gly and Arg allele at codon 388 of fibroblast growth factor receptor-4 (FGFR4) gene was evaluated using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in a hospital-based Malaysian population. Peripheral blood samples were collected from 387 breast cancer patients and 252 normal and healthy women who had no history of any malignancy. The aim of the present study was to evaluate the association between the FGFR4 Gly388Arg polymorphism and breast cancer risk as well as clinicopathological parameters of the patients. The Gly/Gly, Gly/Arg, Arg/Arg, and Arg allele genotypes were detected in 46.3%, 44.4%, 9.3%, and 53.7% of breast cancer cases, respectively. The distribution of genotype (p = 0.204) and allele (p = 0.086) frequencies of FGFR4 polymorphism were not significantly different between the breast cancer cases and normal individuals. Women who were Arg/ Arg homozygotes (OR = 1.714, 95% CI 0.896-3.278), Gly/Arg heterozygotes (OR = 1.205, 95% CI 0.863-1.683), carriers of Arg allele genotype (OR = 1.269, 95% CI 0.921-1.750), or Arg allele (OR = 1.246, 95% CI 0.970-1.602) were not associated with breast cancer risk. The Arg allele genotype was significantly associated with lymph node metastases (i = 0.001) but not with other clinicopathological parameters. Our findings suggest that the polymorphic variant at codon 388 of FGFR4 gene does not confer increased risk to breast cancer development but it may be a potential genetic marker for tumor prognosis.
Key words: Fibroblast growth factor receptor-4 (FGFR4); Polymorphism; Breast cancer
Address correspondence to Dr. Rakesh Naidu, School of Medicine and Health Sciences, Monash University Sunway Campus, Jalan Lagoon Selatan, 46150 Bandar Sunway, Selangor Darul Ehsan, Malaysia. Tel: +603 55146345; E-mail: email@example.com
Combined Effect of Rapamycin and Cisplatin on Survival of Hep-2 Cells In Vitro
Wenbin Lei, Tao Jia, Zhenzhong Su, Weiping Wen, and Xiaolin Zhu
National Key Disciplines of Otorhinolaryngology, Otorhinolaryngology Hospital, Otorhinolaryngology Institute, The First Affiliated Hospital, Sun Yat-sen University, Guangdong, P.R. China
The cytotoxic effects and mechanism of action of cisplatin and the mTOR inhibitor rapamycin on Hep-2 laryngeal cancer cells were investigated. Hep-2 cells were cultured in the presence of different concentrations of rapamycin, cisplatin, or the two combined. Inhibition of cell growth, apoptosis, and AKT, mTOR, S6K, and ERCC1 protein levels were assessed. All combinations of rapamycin and cisplatin resulted in synergistic inhibition of cell growth (as indicated by q values determined using Jin's formula >1.15). Rapamycin inhibited Hep-2 cell growth, induced G1 arrest, and when combined with cisplatin, enhanced apoptosis. p-mTOR and S6K expressions were significantly downregulated by rapamycin. ERCC1 expression was significantly upregulated with cisplatin treatment. Combined cisplatin and rapamycin treatment resulted in significant downregulated p-mTOR and S6K expression, but no change in ERCC1 expression. Rapamycin and cisplatin act in a synergistic manner, increasing the cytotoxic effect on Hep-2 cells. Rapamycin may facilitate increased Hep-2 cell apoptosis with cisplatin via inhibiting downstream expression of proteins in the AKTmTOR signaling pathway.
Key words: Rapamycin; AKT/mTOR; Cisplatin; Hep-2 cells; Apoptosis
Address correspondence to Zhenzhong Su, Professor of Otorhinolaryngology, The First Affiliated Hospital, Sun Yat-sen University, Zhongshan 2nd Road 58, Guangzhou 510080, P.R. China. Tel: +0086-20-87333733; Fax: 0086-20-87333733; E-mail: firstname.lastname@example.org
v-3 and v-6 Polyunsaturated Fatty Acids Enhance Arsenic Trioxide Efficacy in Arsenic Trioxide-Resistant Leukemic and Solid Tumor Cells
Melanie Wirtitsch, Erich Roth, Thomas Bachleitner-Hofmann, Barbara Wessner, and Sanda Sturlan
Surgical Research Laboratories, Department of Surgery, Medical University of Vienna, Vienna, Austria
Recently we showed that the polyunsaturated fatty acid (PUFA) docosahexaenoic acid (DHA) sensitizes arsenic trioxide (As2O3)-resistant tumor cells to a clinically achievable concentration (1 mM) of As2O3 via a reactive oxygen species (ROS)-dependent mechanism. The aim of the present study was to evaluate, whether this combined effect of As2O3 and DHA is also applicable to other PUFAs [i.e., eicospentaenoic acid (EPA), arachidonic acid (AA), and gamma-linolenic acid (GLA)]. Fourteen tumor cell lines were incubated with As2O3 (1 mM), PUFA (25-100 mM), or the combination thereof (±vitamin E). Cell viability (colorimetric), apoptosis (bivariate annexin V/propidium iodide staining, detection of hypodiploid DNA), and thiobarbituric acid reactive substances (TBARS) were evaluated. Twelve of 14 As2O3-resistant cell lines tested were resistant to PUFA monotherapy. However, combined treatment with As2O3 and either PUFA significantly reduced cell viability in a dose-dependent manner with AA being the most potent As2O3 enhancer. The combined cytotoxic effect of As2O3/AA treatment was due to induction of apoptosis, preceded by increased intracellular TBARS and was abolished by the antioxidant vitamin E. Importantly, the combined effect of As2O3 and AA was selectively toxic for malignant cells because no cytotoxic effect was observed in normal skin fibroblasts and human microvascular endothelial cells. In conclusion, our study shows that also other PUFAs than DHA-and in particular the v-6-PUFA AA-can be used as effective modulators of tumor cell chemosensitivity to clinically achievable concentrations of As2O3. Enhanced lipid peroxidation most likely constitutes the key mechanism for the combined effect.
Key words: Polyunsaturated fatty acids; Arsenic trioxide; Apoptosis; Lipid peroxidation; Tumor cells
Address correspondence to Erich Roth, Surgical Research Laboratories, Department of Surgery, Medical University of Vienna, Wa¨hringer Gu¨rtel 18-20, A-1090 Vienna, Austria. Tel: +43-1-40400-5621; Fax: +43-1-40400-6782; E-mail: email@example.com
The Inhibitor of Growth 1 (ING1) Proteins Suppress Angiogenesis and Differentially Regulate Angiopoietin Expression in Glioblastoma Cells
Gesche Tallen,1* Sonja Farhangi,2* Mona Tamannai,1 Nikola Holtkamp,2 Dorothea Mangoldt,3 Sitar Shah,5 Keiko Suzuki,5 Matthias Truss,4 Guenter Henze,1 Karl Riabowol,5 and Andreas von Deimling6
1Department of Pediatric Oncology/Haematology, Charité-Universitätsmedizin
Berlin, 13353 Berlin, Germany
2Department of Neuropathology, Charité-Universitätsmedizin Berlin, Berlin, Germany
3Department of Dermatology, Allergology, Venerology, Charité-Universitätsmedizin Berlin, Berlin, Germany
4Department of Pediatrics, Laboratory for Molecular Biology, Charité-Universitätsmedizin Berlin, Berlin, Germany
5Departments of Biochemistry and Molecular Biology and Oncology, Faculty of Medicine, The University of Calgary, Calgary, Alberta, Canada
6Department of Neuropathology, University of Heidelberg, Heidelberg, Germany
The inhibitor of growth 1 (ING1) homologue ING4 has previously been implicated as a negative regulator of angiogenesis in a murine glioma and a multiple myeloma model. An association between ING1 and angiogenesis has not been reported yet. Our previous studies using tumor samples from patients have shown that ING1 levels are downregulated in glioblastoma multiforme (GBM), one of the most highly vascularized malignancies. Based on this background, the goal of this study was to test the effects of the major ING1 splicing isoforms, p47ING1a and p33ING1b, on pathological angiogenesis induced by human GBM cells. We used a chorioallantoic membrane (CAM) assay to examine whether LN229 human GBM cells can induce angiogenesis and whether alterations in ING1 expression, such as ING1 knockdown by siRNA or ectopic ING1 overexpression using ING1a and ING1b expression constructs, can affect this process. Increased ING1 protein expression significantly suppressed LN229 cell-induced angiogenesis in the CAM assay. While no effects on the proangiogenic factors VEGF or IL-8 were noted, the expression of angiopoietins (Ang) 1 and 4 were increased by the p47ING1a, but not by the p33ING1b isoform. Levels of Ang-2 were not sensitive to altered ING1 levels. Our data are the first to suggest that ING1 proteins suppress neoangiogenesis in GBM. Moreover, our results may support the idea that ING1 proteins regulate the expression of proteins that are critical for angiogenesis in GBM such as the angiopoietins.
Key words: Inhibitor of growth 1 (ING1); Glioblastoma multiforme; Angiogenesis; Angiopoietins
Address correspondence to Gesche Tallen, Department of Pediatric Oncology/Haematology, Charité-Universitätsmedizin Berlin, Campus Virchow-Klinikum, Augustenburger Platz 1, 13353, Berlin, Germany. Tel: 49-30-450-566032; Fax: 49-30-450-566906; E-mail: firstname.lastname@example.org
*Authors contributed equally to this work.
Kiss-1 Suppresses MMP-9 Expression by Activating p38 MAP Kinase in Human Stomach Cancer
Kyung Hee Lee1 and Jae-Ryong Kim2,3
1Department of Hematology-Oncology, College of Medicine,
Yeungnam University, Daegu, Korea
2Department of Biochemistry and Molecular Biology, College of Medicine, Yeungnam University, Daegu, Korea
3Aging-Associated Vascular Disease Research Center, College of Medicine, Yeungnam University, Daegu, Korea
Kiss-1 has been identified as a putative metastasis suppressor gene in various human malignancies. However, there is little information about its possible role in gastric carcinoma. In this study, we determined whether the Kiss-1 gene negatively regulates MMP-9 expression. cDNA microarray technology was used to identify the genes associated with metastasis by hepatocyte growth factor (HGF) in the gastric cancer cell lines, NUGC-3 and MKN-28. The levels of Kiss-1 RNA and protein were confirmed to be upregulated in HGF-treated gastric cancer cells. HGF induced Kiss-1 and MMP-9 production in a dose-dependent manner. In order to investigate roles of HGF signaling in tumor progression and metastasis, we measured effects of a specific MEK1 inhibitor (PD 098059) and a p38 kinase inhibitor (SB 203580) on HGF-mediated cell proliferation and MMP-9. Pretreatment with PD 098059 reduced MMP-9 and HGF-mediated cell proliferation, but increased Kiss-1 expression. In contrast, SB 203580 pretreatment enhanced MMP-9 and cell proliferation, but decreased Kiss-1 expression. Cotreatment of PD098059 and SB203580 increased the p38 phosphorylation stimulated by HGF. These results suggest that the HGF-mediated Kiss-1 overexpression is regulated mainly by the p38 activation and, furthermore, the activation of ERK might affect HGF-mediated Kiss-1 expression indirectly by the regulation of p38 kinase. Consistent with this result, p38 phosphorylation was strongly repressed by the knock-down of Kiss-1. Downregulation of Kiss-1 using Kiss-1 shRNA also increased in vitro cell invasion. In conclusion, Kiss-1 suppresses MMP-9 expression by activating the p38 MAP kinase signaling pathway.
Key words: Kiss-1; Hepatocyte growth factor (HGF); Matrix metalloproteinase-9 (MMP-9); Metastasis
Address correspondence to Jae-Ryong Kim, M.D., Ph.D., Department of Biochemistry and Molecular Biology, College of Medicine, Yeungnam University, 317-1 Daemyung-Dong, Daegu 705-717, Republic of Korea. Tel: 82-53-620-4342; Fax: 82-53-654-6651; E-mail: email@example.com
VEGF165 Promotes the Osteolytic Bone Destruction of Ewing's Sarcoma Tumors by Upregulating RANKL
Hui Guan, Zhichao Zhou, Ying Cao, Xiaoping Duan, and Eugenie S. Kleinerman
Division of Pediatrics, The University of Texas M. D. Anderson Cancer Center, Houston, TX, USA
The purpose of this study was to determine whether vascular endothelial growth factor-165 (VEGF165) contributed to the osteolytic process in Ewing's sarcoma. VEGF165 induced osteoclast formation from murine bone marrow cells. Tartrate-resistant acid phosphatase (TRAP) staining demonstrated significantly fewer osteoclasts in VEGF-inhibited TC/siVEGF7-1 tumors compared to TC71 parental or TC/si-control tumors. Receptor activator NF-kB (RANKL), a critical osteoclastogenic factor, was decreased in TC/siVEGF7-1 cells. Incubation of these cells with recombinant VEGF165 upregulated RANKL in a dose- and time-dependent manner. The induction of (RANKL) by VEGF165 was also demonstrated in MC3T3-E1 mouse osteoblast cells and bone marrow stromal cells. This upregulation was transcriptionally mediated by an effect on the RANKL promoter. Both VEGF and EWS/FLI-1 increased RANKL promoter activity. Taken together, these data suggest that modulation of RANKL by VEGF165 may be one of the mechanisms responsible for the osteolytic process induced by Ewing's sarcoma cells. VEGF165 may, therefore, play an important role in modulating RANKL gene expression in the bone marrow microenvironment during the metastatic process, thereby contribution to tumor induced bone lysis.
Key words: VEGF<sb>165; Ewing's sarcoma; RANKL; Osteolysis
Address correspondence to Eugenie S. Kleinerman, M.D., Division of Pediatrics, The University of Texas M. D. Anderson Cancer Center, 1515 Holcombe Boulevard, Unit 87, Houston, TX 77030, USA. Tel: 713-792-8110; Fax: 713-794-5042; E-mail: firstname.lastname@example.org
Is Cyclin D2 a Marker of B-CLL Cell Activation?
Agata Kosmaczewska,1* Lidia Ciszak,1* Aleksandra Szteblich,1 Anna Laba,1 Marcin Wojtowicz,2 Dariusz Wolowiec,3 and Irena Frydecka1,3
1Department of Experimental Therapy, Institute of Immunology
and Experimental Therapy, Polish Academy of Sciences, Wroclaw, Poland
2Department of Hematology, Regional Hospital, Opole, Poland
3Department of Hematology, Neoplastic Diseases, and Bone Marrow Transplantation, Wroclaw Medical University, Wroclaw, Poland
The most recent studies emphasize a link between B-cell proliferation in vivo and clinical outcome of B-cell chronic lymphocytic leukemia (B-CLL). The expression of cyclin D2 in B-CLL cells isolated from the peripheral blood of 27 untreated patients in relation to the apoptosis ratio both before and 72 h after culture in the absence of growth factors was analyzed by immunocytochemistry. The significant associations between cell death in culture and both cyclin D2 expression in freshly isolated cells and the rate of its decrement in culture found in this study confirm the special role of cyclin D2 in enhancing the longevity of these cells in vivo. As cyclin D2 is inducible in the early G1 phase, its increased expression might also reflect the activation of cells attempting to replicate in vivo. Furthermore, the finding that B-CLL progression positively correlates with the gradual increase in the proportion of apoptotic B lymphocytes in culture seems to support the notion of cells striving to undergo division in the absence of growth factors. All together, these results indicate the possibility that cyclin D2+ cells represent a pool of leukemic cells with the potential to enter the dividing compartment.
Key words: B-cell chronic lymphocytic leukemia; Cyclin D2; Apoptosis; Proliferation
Address correspondence to Agata Kosmaczewska, M.D., Ph.D., Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, R. Weigla 12, 53-114 Wroclaw, Poland. Tel: +48 71-3371172; Fax: +48 71-3371382; E-mail: email@example.com
*These authors contributed equally to this work.