Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Chronic hypoxia in patients with colon carcinoma

ABSTRACT: In Western countries, colorectal cancer (CRC) is the third most common cancer in both men and women and the second leading cause of cancer-related deaths (approximately 500,000 deaths annually). For CRC, the tumor stage is the main prognostic factor for survival or relapse after surgery. Current staging is based on the AJCC classification which takes into account tumor size/depth, lymph node involvement and distant metastases. Surgical intervention can cure a significant portion of the patients, especially those that are presented in stages I, II or III (around 75% of patients). Patients in stage II undergo surgery only, whereas stage III patients receive adjuvant chemotherapy. However, a major fraction of these patients either have no need for adjuvant treatment (40% stage III) or would benefit when they were given adjuvant therapy in stage II. Therefore, in the current practice, the majority of patients receive not the optimal treatment. An accurate and reliable method that identifies patients with low and high risk for recurrence could improve the selection for adjuvant therapies in these groups, reducing over- or under-treatment. Molecular profiling of mRNA expression by microarray is an approach to identify relevant genes and gene signatures and has been used already for different types of cancer, like breast and HCC. Most studies, however, resulted in their own classification with a specific separation into groups. Most of these microarray studies show remarkably little overlap and it is difficult to find a clear correlation between the molecular classes and prognosis. The results of the studies seem to be center-dependent because of the different microarray techniques used, the small heterogeneous cohorts that are studied and the different clinical parameters used for the evaluation. Recently, we used a mechanism-driven approach to find a correlation between gene expression in HCC and prognosis in which we tried to overcome most shortcomings (van Malenstein et al. CCR 2010). In this study, we want to apply the approach which we used for HCC now in CRC with the aim to improve the prognosis prediction for patients in AJCC stage II and III. We determined the gene expression by microarray in the colon cancer cell line Caco2 and identified the genes that are differentially expressed under hypoxia (72 hours, 2% O2). Using a bioinformatic approach, we used 5 sets of expression data with corresponding clinical information (training: 3 sets, 234 patients and validation: 2 sets, 322 patients). This resulted in a unique 21 gene set with good performance in retrospective analysis. A second improvement we wanted to achieve is the development of a method that can be applied on FFPE material, which would expand the use of the technique to samples outside the university or research setting. We used the nCounter technique that uses hybridization of labeled probes and quantification without an amplification step. We tested our 21 genes on a cohort of 384 well-characterized patients that were collected between 2004 and 2006 at the University Hospitals of Leuven. We compared cells under hypoxia 2% O2 versus 20% O2 conditions. This was done for two cell lines (Caco2 human colorectal adenocarcinoma cell line and HPAC pancreas adenocarcinoma cell line). Each sample has a biological replicate. Samples are hybridized in dye-swap, resulting in 4 hybridizations per cell line.

ORGANISM(S): Homo sapiens

SUBMITTER: Wouter Van Delm

PROVIDER: E-GEOD-31079 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Hypoxia-driven gene expression is an independent prognostic factor in stage II and III colon cancer patients.

Dekervel Jeroen J Hompes Daphne D van Malenstein Hannah H Popovic Dusan D Sagaert Xavier X De Moor Bart B Van Cutsem Eric E D'Hoore André A Verslype Chris C van Pelt Jos J

Clinical cancer research : an official journal of the American Association for Cancer Research 20140131 8

<h4>Purpose</h4>Hypoxia is considered a major microenvironmental factor influencing cancer behavior. Our aim was to develop a hypoxia-based gene score that could identify high and low risk within stage II and III colon cancer patients.<h4>Experimental design</h4>Differential gene expression of CaCo-2 colon cancer cells cultured in chronic hypoxia versus normoxia was tested for correlation with prognostic variables in published microarray datasets. These datasets were further used to downsize and ...[more]

PMID: 24486594

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Project description:In Western countries, colorectal cancer (CRC) is the third most common cancer in both men and women and the second leading cause of cancer-related deaths (approximately 500,000 deaths annually). For CRC, the tumor stage is the main prognostic factor for survival or relapse after surgery. Current staging is based on the AJCC classification which takes into account tumor size/depth, lymph node involvement and distant metastases. Surgical intervention can cure a significant portion of the patients, especially those that are presented in stages I, II or III (around 75% of patients). Patients in stage II undergo surgery only, whereas stage III patients receive adjuvant chemotherapy. However, a major fraction of these patients either have no need for adjuvant treatment (40% stage III) or would benefit when they were given adjuvant therapy in stage II. Therefore, in the current practice, the majority of patients receive not the optimal treatment. An accurate and reliable method that identifies patients with low and high risk for recurrence could improve the selection for adjuvant therapies in these groups, reducing over- or under-treatment. Molecular profiling of mRNA expression by microarray is an approach to identify relevant genes and gene signatures and has been used already for different types of cancer, like breast and HCC. Most studies, however, resulted in their own classification with a specific separation into groups. Most of these microarray studies show remarkably little overlap and it is difficult to find a clear correlation between the molecular classes and prognosis. The results of the studies seem to be center-dependent because of the different microarray techniques used, the small heterogeneous cohorts that are studied and the different clinical parameters used for the evaluation. Recently, we used a mechanism-driven approach to find a correlation between gene expression in HCC and prognosis in which we tried to overcome most shortcomings (van Malenstein et al. CCR 2010). In this study, we want to apply the approach which we used for HCC now in CRC with the aim to improve the prognosis prediction for patients in AJCC stage II and III. We determined the gene expression by microarray in the colon cancer cell line Caco2 and identified the genes that are differentially expressed under hypoxia (72 hours, 2% O2). Using a bioinformatic approach, we used 5 sets of expression data with corresponding clinical information (training: 3 sets, 234 patients and validation: 2 sets, 322 patients). This resulted in a unique 21 gene set with good performance in retrospective analysis. A second improvement we wanted to achieve is the development of a method that can be applied on FFPE material, which would expand the use of the technique to samples outside the university or research setting. We used the nCounter technique that uses hybridization of labeled probes and quantification without an amplification step. We tested our 21 genes on a cohort of 384 well-characterized patients that were collected between 2004 and 2006 at the University Hospitals of Leuven.

Project description:Distant metastasis is the major causes of death in colorectal cancer (CRC) patients. In order to identify genes influencing the prognosis of patients with CRC, we compared gene expression in primary tumors with and without distant metastasis using an oligonucleotide microarray. We also examined the expression of the candidate gene in 100 CRC patients by quantitative real-time reverse transcription PCR and studied the relationship between its expression and the prognosis of patients with CRC. As a result, we identified MUC12 as a candidate gene involved in metastasis processes by microarray analysis. Quantitative real-time reverse transcription PCR showed that MUC12 expression was significantly lower in cancer tissues than in adjacent normal tissues (P < 0.001). In stage II and stage III CRC, patients with low expression showed worse disease-free survival (P = 0.038). Multivariate analysis disclosed that MUC12 expression status was an independent prognostic factor in stage II and stage III CRC (relative risk, 9.532; 95% confidence interval, 2.303-41.905; P = 0.002). This study revealed the prognostic value of MUC12 expression in CRC patients. Moreover, our result suggests MUC12 expression is a possible candidate gene for assessing postoperative adjuvant therapy for CRC patients. Total of 111 microarray datasets (77 for LCM samples, and 17 pairs for homogenized samples from tumor and adjacent tissues) were normalized using robust multi-array average (RMA) method under R 2.6.2 statistical software together with BioConductor package, as described previously. Then, the gene expression levels were log2-transformed, and 62 control probe sets were removed for further analysis. In order to identify a set of genes associated with development of metastatic recurrence, we performed Wilcoxon rank-sum test for gene expression differences of 54,613 probe sets between recurrence and non-recurrence groups. Similarly, Wilcoxon singed-rank test was conducted to select genes which showed significant expression difference between tumor and adjacent tissue. Then, we selected a set of genes that satisfied both of above two criteria.

Project description:Background - Hepatocellular carcinomas (HCCs) are heterogeneous tumors with respect to etiology, cell of origin and biology. The course of the disease is unpredictable and is in part dependent on the tumor microenvironment. One of the microenvironmental factors is hypoxia, which is known to promote aggressiveness in other malignant tumors. We hypothesized that certain regions in HCC exist with chronic hypoxia and a characteristic gene expression pattern. Moreover, during the development of HCC there is an important contribution of this chronic hypoxia on prognosis via this gene expression. Until now, most research has been performed in acute hypoxic models (< 24 hours). Methods – Human hepatoblastoma cells HepG2 were cultured in either normoxic (20% O2) or hypoxic (2% O2) conditions for 72 hrs, the time it takes to adapt to chronic hypoxia. After 3 days the cells were harvested and analyzed by microarray technology. The highly significant differentially expressed genes were selected and used to assess the clinical value of our in vitro chronic hypoxia gene signature in four published patient studies. Three of these independent microarray studies on HCC patients were used as training sets to determine a minimal prognostic gene set and one study was used for validation. Gene expression analysis and correlation with clinical outcome was assessed with the bioinfomatic method of Goeman et al (). Results – In the HepG2 cells, 3592 genes were differentially expressed in cells cultured at 2% oxygen for 72 hrs. Out of these, 265 showed a high significant change (2-fold change and p=0.0001). The level of gene expression after 72 hrs was different from the acute hypoxic response (during the first 24 hours) and represented chronicity. Using computational methods we identified 7 out of the 265 highly significant genes that showed correlation with prognosis in all three different training sets and this was independently validated in a 4th dataset. With our approach we could include the largest number of HCC patients in one single study. Conclusion – We identified a 7-gene signature, which is associated with chronic hypoxia and predicts prognosis in patients with HCC. In the future this signature could be used as a diagnostic tool. In addition, chronic hypoxia gene expression information can be used in the search for new therapeutic targets. Two conditions were compared and each sample has a biological replicate. Samples are hybridized in dye-swap, resulting in 4 hybridizations.

Dataset Information

Chronic hypoxia in patients with colon carcinoma

Publications

Hypoxia-driven gene expression is an independent prognostic factor in stage II and III colon cancer patients.

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