CANCER GENOMICS & PROTEOMICS
Volume 5, Number
2, March-April 2008
CONTENTS
PAGE
Microarray-based Expression of DNA Repair Genes Does not Correlate with Growth Inhibition of Cancer Cells by Natural Products Derived from Traditional Chinese Medicine. V.S.B. KONKIMALLA, G. WANG, B. KAINA,
T. EFFERTH (Heidelberg; Mainz, Germany; Detroit, MI, USA)
79
Proteomic Based Identification of Manganese Superoxide Dismutase 2 (SOD2) as a Metastasis Marker for Oral Squamous Cell Carcinoma. H.YE, A. WANG, B.S. LEE, T.YU, S. SHENG, T. PENG, S. HU, D.L. CROWE, X. ZHOU (Chicago, IL;Atlanta, GA, Los Angeles, CA, USA; Guangzhou, China)
85
Feasibility and Relevance of Global Expression Profiling of Gene Transcripts in Serum from Breast Cancer Patients Using Whole Genome Microarrays and Quantitative RTPCR. L. O’DRISCOLL, E. KENNY, J.P. MEHTA, P. DOOLAN, H. JOYCE, P. GAMMELL, A. HILL, B. O’DALY, D. O’GORMAN, M. CLYNES (Dublin, Ireland)
95
Highaccuracy Prediction of Carcinogenicity by Global Quantitative Analysis of Posttranslational Modifications in a 28-Day In Vivo Rat Study.
H. YAMANAKA, Y. YAKABE, K. SAITO, M. SEKIJIMA, H. SATO, T. SHIRAI (Saitama; Osaka; Ibaraki; Tokyo; Nagoya, Japan)
105
Galectin3 Expression in Human Papillary Thyroid Carcinoma. P. SAWANGAREETRAKUL, C. SRISOMSAP, D. CHOKCHAICHAMNANKIT, J. SVASTI (Bangkok, Thailand)
117
Protein Phosphatase and TRAIL Receptor Genes as New Candidate Tumor Genes on Chromosome 8p in Prostate Cancer. M. HORNSTEIN, M.J. HOFFMANN, A. ALEXA, M. YAMANAKA, M. MULLER, V. JUNG, J. RAHNENFUHRER, W.A. SCHULZ (Dusseldorf; Saarbrucken; Homburg; Dortmund, Germany)
123
CANCER GENOMICS & PROTEOMICS 5: 79-84 (2008)
Microarray-based Expression of DNA Repair Genes Does not Correlate with Growth Inhibition of Cancer Cells by Natural Products Derived from Traditional Chinese Medicine
V.S. BADIREENATH KONKIMALLA1, GAN WANG2, BERND KAINA3 THOMAS EFFERTH1
1Pharmaceutical Biology (C015), German Cancer Research Center, Heidelberg, Germany; 2Institute of Environmental Health Sciences, Wayne State University, Detroit, MI, U.S.A.; 3Institute of Toxicology, University of Mainz, Mainz, Germany
Abstract: Drug resistance represents a major obstacle in cancer chemotherapy. As chemically characterized compounds derived from plants used in traditional Chinese medicine (TCM) may have molecular targets different from those of standard antitumor drugs, they might be attractive candidates for novel therapeutics with improved pharmacological features. DNA repair is frequently involved in the development of resistance to established anticancer drugs, e.g. alkylating agents. Using a database of 531 chemically characterized TCM compounds from medicinal plants recently established by us, the IC50 values of 60 N.C.I. tumor cell lines for these 531 natural products were tested for correlation with the microarray-based mRNA expression of six genes involved in nucleotide excision repair (ERCC1, XPA, XPC, DDB2, ERCC4, ERCC5). No compound correlated with the expression of these genes, indicating that mRNA expression of these genes is not associated with resistance of the cell lines to these TCM compounds. The same is true for another six genes of the base excision repair pathway (MPG, APEX1, OGG1, XRCC1, LIG3, POLB). Microarray-based COMPARE analyses were performed to identify other candidate genes that are able to predict responsiveness of tumor cells to TCM-derived natural products. As an example, diallyl disulfide from garlic (Allium sativum L., Chinese name: dashuan) was chosen. Eighteen genes were identified whose mRNA expression predicted sensitivity or resistance to diallyl disulfide in hierarchical cluster analyses. Apart from some genes with still unknown function, genes were identified from different functional groups, e.g. signal transducers, regulators of GTPase activity, those associated with cytoskeleton formation and regulation, constituents of the ribosome. Remarkably, none of these genes have been described to be involved in DNA repair. In conclusion, our data indicate that TCM-derived natural products are worth being further investigated as novel compounds to eradicate tumors which reveal resistance to established anti-cancer drugs.
Proteomic Based Identification of Manganese Superoxide Dismutase 2 (SOD2) as a Metastasis Marker for Oral Squamous Cell Carcinoma
HUI YE1, ANXUN WANG2, BAO-SHIANG LEE3, TIANWEI YU4, SHIHU SHENG2, TINGSHENG PENG5, SHEN HU6, DAVID L. CROWE1,7, XIAOFENG ZHOU1,7,8
1Center for Molecular Biology of Oral Diseases, College of Dentistry, 3Protein Research Laboratory, Research Resource Center, and 7Graduate College, UIC Cancer Center, University of Illinois at Chicago, Chicago, IL, U.S.A.; 2Department of Oral and Maxillofacial Surgery, and 5Department of Pathology, The First Affiliated Hospital, 8Guanghua School and Research Institute of Stomatology, Sun Yat-Sen University, Guangzhou, China; 4Department of Biostatistics, Rollins School of Public Health, Emory University, Atlanta, GA; 6School of Dentistry, Dental Research Institute, University of California Los Angeles, Los Angeles, CA, USA;
Abstract: Metastasis is a critical event in oral squamous cell carcinoma (OSCC) progression. To identify proteomic biomarkers for OSCC metastasis, 3 paired OSCC cell lines (UM1/UM2, 1386Tu/1386Ln, 686Tu/686Ln) with different metastatic potential were examined. Among those 3 cell lines, UM1, 1386Ln and 686Ln exhibited a higher degree of metastatic potential than their paired cell lines UM2, 1386Tu and 686Tu, respectively, as measured using an in vitro cell invasion assay. A total of 40 differentially expressed proteins were identified using 2D-PAGE/MS proteomic approach. Selected protein candidates (superoxide dismutase 2 and heat shock protein 27) were further investigated by immuno-histochemistry (IHC) method using independent OSCC patient tissue samples. The statistically significantly increases in IHC staining for manganese superoxide dismutase 2 (SOD2) were observed in lymph node metastatic disease when compared with the paired primary OSCC. Our results thus indicated that elevated SOD2 levels is associated with lymph node metastasis in OSCC and may provide predictive values for diagnosis of metastasis.
Feasibility and Relevance of Global Expression Profiling of Gene Transcripts in Serum from Breast Cancer Patients Using Whole Genome Microarrays and Quantitative RT-PCR
LORRAINE O'DRISCOLL1, ELAINE KENNY1, JAI PRAKASH MEHTA1, PADRAIG DOOLAN1, HELENA JOYCE1, PATRICK GAMMELL1, ARNOLD HILL2, BRENDAN O'DALY2, DONAL O'GORMAN3, MARTIN CLYNES1
1National Institute for Cellular Biotechnology and 3School of Health and Human Performance, Dublin City University, Dublin 9; 2St. Vincent's University Hospital, Dublin 4, Ireland
Abstract: Background: Previous studies, by ourselves and others, have indicated that gene transcripts are detectable extracellularly. Advancing on this work, in order to investigate the feasibility of analysing global gene expression profiles and so the possibility in the future of identifying panels of circulating mRNA biomarkers that may be diagnostic, prognostic or predictive for cancer, here we performed the first whole genome microarray analysis of human serum. Patients and Methods: RNA was isolated from pre-surgery serum and corresponding breast tumour and normal tissue biopsies, and from post-surgery and normal control serum. Specimens were examined using Affymetrix whole genome microarrays and quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR). Results: Of the 54,675 mRNAs/variants analysed, approximately 8% and 45% were called Present in serum and breast tissue specimens, respectively. Differentially expressed genes were identified for each group of specimens analysed. Analysis, by qRT-PCR, of 3 selected transcripts further indicated that the nucleic acids detected were mRNA, not DNA. mRNAs are apparently present in serum and their global detection and identification can be successfully achieved using microarray technologies. Conclusion: The potential implication of this novel finding is that using microarrays it may be possible to identify a panel of extracellular mRNAs that are diagnostic, prognostic and/or predictive of outcome for cancer patients.
1Chemicals Assessment Center, Chemicals Evaluation and Research Institute, Saitama; 2Environmental Health Science Laboratory, Sumitomo Chemical Co. Ltd, Osaka; 3Kashima Laboratory, Mitsubishi Chemical Safety Institute Ltd., Ibaraki; 4Tokyo University of Agriculture and Technology, Tokyo; 5Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan
Abstract: A global quantitative analysis of post-translational modifications (PTMs) of distinct proteins was executed at the proteomic level using two-dimensional fluorescence differential gel electrophoresis. We evaluated the effects of 66 chemical compounds, including 15 genotoxic carcinogens, 28 non-genotoxic carcinogens, and 23 non-carcinogens, in the male F344 rat liver in a 28-day repeated dose study. In the master gel of rat liver protein, we identified 728 spots by hybrid quadrupole time-of-flight mass spectrometry. They collapsed into 356 distinct proteins. Of these, 126 were represented by two or more spots in the 2-D gel. We calculated the logarithmic ratio of volume changes of all 1028 combinations generated from 126 proteins and investigated the relevance to carcinogenicity. This quantitative proteomic study revealed the existence of several PTMs characteristic of carcinogens that may play an important role in early stage of carcinogenicity. Prediction of carcinogenicity from PTM data gave a higher concordance (92.4%) than prediction from protein expression data (74.2%). This novel approach holds great promise as a way of revealing the roles of charge modifications and molecular weight variations of proteins in biological processes.
Galectin-3 Expression in Human Papillary Thyroid Carcinoma
PHANNEE SAWANGAREETRAKUL1, CHANTRAGAN SRISOMSAP1,2, DARANEE CHOKCHAICHAMNANKIT1 and JISNUSON SVASTI1,2,3
1Laboratory of Biochemistry, Chulabhorn Research Institute, and 2Chulabhorn Graduate Institute, Bangkok 10210; 3Department of Biochemistry and Center for Protein Structure and Function, Faculty of Science, Mahidol University, Bangkok 10400, Thailand
Abstract: Previous studies have suggested that galectin-3 expression was markedly elevated in papillary thyroid carcinoma compared to other thyroid diseases. In order to better understand this protein, galectin-3 from papillary thyroid carcinoma was partially purified by affinity chromatography on lactose-agarose. Proteins eluted from the column were separated by SDS-PAGE, and galectin-3 was detected with antibodies against the N-terminus and C-terminus of galectin-3. Some protein bands from the lactose binding fraction were also selected for identification by liquid chromatography/mass spectrometry/mass spectrometry (LC/MS/MS). Seven protein bands, with molecular weights ranging from 16 kDa to 31 kDa, were identified as galectin-3. The antibody raised against the C-terminus of galectin-3 gave a strong band for one of the bands detected by the N-terminal antibody and weak bands for the other three. One additional dark immunoreactive band with an approximate molecular weight of 20 kDa, was also detected by the C-terminal galectin-3 antibody. To determine the structural differences of each protein band, N-terminal amino acid sequencing of the seven protein bands was conducted. The three upper bands were N-terminally blocked, while the other bands had N-terminal amino acid sequences starting at positions Gly35, Gly65 (2 bands) and Ala100, respectively. Further studies are necessary to determine whether these are due to nonspecific proteolysis or post-translation modification
Protein Phosphatase and TRAIL Receptor Genes as New Candidate Tumor Genes on Chromosome 8p in Prostate Cancer
MAX HORNSTEIN1, MICHÈLE J. HOFFMANN1, ADRIAN ALEXA2, MASANORI YAMANAKA1, MIRKO MÜLLER1, VOLKER JUNG3, JÖRG RAHNENFÜHRER4, WOLFGANG A. SCHULZ1
1Department of Urology, Heinrich Heine University, Dusseldorf; 2Max-Planck Institute for Informatics, 66123 Saarbrucken; 3Department of Urology, Medical University of the Saarland, Homburg; 4Institute for Statistics, University of Dortmund, Dortmund, Germany
Abstract: Background: Allelic losses on chromosome 8p are common in prostate carcinoma, but it is not known exactly how they contribute to cancer development and progression. Materials and Methods: Expression of 12 genes located across chromosome 8p, including established tumor suppressor candidates (CSMD1, DLC1, NKX3.1), and others from a new microarray-based comparison was studied by quantitative RT-PCR in 45 M0 prostate carcinomas and 13 benign prostate tissues. Results: Significantly reduced expression was observed for two protein phosphatase subunit genes (PPP2CB, PPP3CC) and two TRAIL decoy receptors (TNFRSF10C/DcR1, TNFRSF10D/DcR2), but not for the three established candidates nor for TRAIL death receptor genes. Low expression of PPP3CC and TNFRSF10C located at 8p21.3 was highly significantly associated with tumor recurrence. In addition to allele loss, down-regulation of TNFRSF10C and TNFRSF10D was found to be associated with hypermethylation, although bisulfite sequencing usually revealed it to be partial. Conclusion: Our data strongly support a recent proposal that a segment at 8p21.3 contains crucial prostate cancer tumor suppressors. In addition, they raise the paradoxical issue of why TRAIL decoy receptors rather than death receptors are down-regulated in aggressive prostate cancer.
