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Query: UMLS:C0699790 (colon cancer)
 28,837 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mutations in KRAS exon 1 oncogene are frequently found in colon carcinomas. A correlation between the mutated KRAS and the prognosis and outcome of treatment of colon cancer patients was reported in the literature. The object of our work was to establish a high-throughput method with high sensitivity to enable screening of tumor mutation status of KRAS exon 1 in large groups of colon cancer patients. KRAS exon 1 sequences from DNA isolated from 191 sporadic colon cancers were PCR amplified using one primer labeled with fluorescein and a second primer extended by a GC-clamp. After PCR amplification samples were subjected to automated 96-array constant denaturant capillary electrophoresis using a modified MegaBACE 1000 sequencing instrument. Mutant samples were identified by characteristic peak patterns. The sensitivity of detection of a mutant allele in a background of the wild-type alleles was 0.3%. Using the 96-array instrument a typical screening of 191 samples for KRAS mutation status could be performed within 2 h. A KRAS exon 1 mutation was found in 66 of 191 (34.6%) of the samples. The 96-array constant denaturant capillary electrophoresis provides an opportunity for the high-sensitivity screening of large cancer populations for KRAS exon 1 mutations.
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PMID:Mutation detection in KRAS Exon 1 by constant denaturant capillary electrophoresis in 96 parallel capillaries. 1200 96

Over the past few decades, advances in genetics and molecular biology have revolutionized our understanding of cancer initiation and progression. Molecular progression models outlining genetic events have been developed for many solid tumors, including colon cancer. Previous reports in the literature have shown a relationship between different KRAS mutations and prognosis and response to medical treatment in colon cancer patients. Furthermore, the presence of a mutated KRAS has been correlated with different clinicopathological variables including age and gender of patients and tumor location. To our knowledge, few institutions screen for KRAS mutations on regular basis in colon cancer patients despite such evidence that knowledge of KRAS exon 1 status is informative. Here, we report on a mutation analysis method adapted to a 96-capillary electrophoresis instrument that allows identification of all 12 oncogenic mutations in KRAS exon 1 under denaturing conditions. To determine the optimal parameters, a series of DNA constructs generated by site-directed mutagenesis was analyzed and the migration times of all mutant peaks were measured. A classification tree was then made based on the differences in migration time between the mutants and an internal standard. A randomized series of 500 samples constructed with mutagenesis as well as 60 blind samples from sporadic colon carcinomas was analyzed to test the method. No wild-type samples were scored as mutants and all mutants were correctly identified. Post polymerase chain reaction (PCR) analysis time of 96 samples was performed within 40 min.
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PMID:Direct identification of all oncogenic mutants in KRAS exon 1 by cycling temperature capillary electrophoresis. 1265 73

Both benign and malignant tumors represent heterogenous tissue containing tumor cells and non-neoplastic mesenchymal and inflammatory cells. To detect a minority of mutant KRAS alleles among abundant wild-type alleles, we developed a sensitive DNA sequencing assay using Pyrosequencing, ie, nucleotide extension sequencing with an allele quantification capability. We designed our Pyrosequencing assay for use with whole-genome-amplified DNA from paraffin-embedded tissue. Assessing various mixtures of DNA from mutant KRAS cell lines and DNA from a wild-type KRAS cell line, we found that mutation detection rates for Pyrosequencing were superior to dideoxy sequencing. In addition, Pyrosequencing proved superior to dideoxy sequencing in the detection of KRAS mutations from DNA mixtures of paraffin-embedded colon cancer and normal tissue as well as from paraffin-embedded pancreatic cancers. Quantification of mutant alleles by Pyrosequencing was precise and useful for assay validation, monitoring, and quality assurance. Our Pyrosequencing method is simple, robust, and sensitive, with a detection limit of approximately 5% mutant alleles. It is particularly useful for tumors containing abundant non-neoplastic cells. In addition, the applicability of this assay for DNA amplified by whole-genome amplification technique provides an expanded source of DNA for large-scale studies.
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PMID:Sensitive sequencing method for KRAS mutation detection by Pyrosequencing. 1604 14

Hypoxia inducible factor-1 (HIF-1) is considered a crucial mediator of the cellular response to hypoxia through its regulation of genes that control angiogenesis. It represents an attractive therapeutic target in colon cancer, one of the few tumor types that shows a clinical response to antiangiogenic therapy. But it is unclear whether inhibition of HIF-1 alone is sufficient to block tumor angiogenesis. In HIF-1alpha knockdown DLD-1 colon cancer cells (DLD-1(HIF-kd)), the hypoxic induction of vascular endothelial growth factor (VEGF) was only partially blocked. Xenografts remained highly vascularized with microvessel densities identical to DLD-1 tumors that had wild-type HIF-1alpha (DLD-1(HIF-wt)). In addition to the preserved expression of VEGF, the proangiogenic cytokine interleukin (IL)-8 was induced by hypoxia in DLD-1(HIF-kd) but not DLD-1(HIF-wt) cells. This induction was mediated by the production of hydrogen peroxide and subsequent activation of NF-kappaB. Furthermore, the KRAS oncogene, which is commonly mutated in colon cancer, enhanced the hypoxic induction of IL-8. A neutralizing antibody to IL-8 substantially inhibited angiogenesis and tumor growth in DLD-1(HIF-kd) but not DLD-1(HIF-wt) xenografts, verifying the functional significance of this IL-8 response. Thus, compensatory pathways can be activated to preserve the tumor angiogenic response, and strategies that inhibit HIF-1alpha may be most effective when IL-8 is simultaneously targeted.
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PMID:Induction of interleukin-8 preserves the angiogenic response in HIF-1alpha-deficient colon cancer cells. 1614 72

Early events involved in the pathogenesis of colorectal cancer include mutations in the Adenomatous Polyposis Coli tumor-suppressor gene and oncogenic KRAS mutations. Later events include deletions on chromosome 18q, which are observed in a high proportion of colorectal cancers. However, the important tumor suppressor genes targeted by these deletions have not been fully defined. A previous study found Cables is located on human chromosome 18q11-12. Loss of Cables expression as determined by immunohistochemical staining (IHC) occurred in 60-70% of sporadic colorectal cancers that were usually correlated to loss of heterozygosity at 18q. To determine if Cables is an important target for the chromosome 18q deletions, the susceptibility of Cables-/- mice to develop colon tumors was studied. A well characterized colonic carcinogen, 1,2-dimethylhydrazine (DMH) was used as a tumor initiator. Cables-/- mice (n = 25) and the Cables+/+ littermates (n = 25) were treated with subcutaneous DMH injections over 20 weeks to initiate tumorigenesis. The median survival after DMH injections was significantly shorter for the Cables-/- mice compared to Cables+/+ littermates. The total number of colorectal tumors that developed in the Cables-/- mice was 46 tumors versus 21 tumors. The increased numbers of colorectal tumors, as well as shorter survival of the Cables-/- mice provides compelling evidence that Cables could play an important role in the pathogenesis and progression of colon cancer in mice. These data coupled with previous observations support the hypothesis that Cables is a relevant target of the chromosome 18q deletions frequently seen in human colorectal cancer.
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PMID:The Cables gene on chromosome 18Q regulates colon cancer progression in vivo. 1612 80

The gene that encodes the alpha-isoform of phosphatidylinositol 3-kinase (PIK3Ca) is frequently mutated in human cancers. We profiled the mutation status of the PIK3Ca gene in the National Cancer Institute (NCI)-60 panel of human cancer cell lines maintained by the Developmental Therapeutics Program of the NCI. Mutation hotspots on the gene were PCR amplified and sequenced, and the trace data were analyzed with software designed to detect mutations. Seven of the cell lines tested have PIK3Ca mutations: two lines derived from breast cancer, two from colon cancer, two from ovarian cancer, and one from lung cancer. BRAF and EGFR genes were normal in the PIK3Ca mutant lines. Two of the cell lines with mutant PIK3Ca also have a mutant version of the KRAS gene. The mutation status was correlated with array-based gene expression that is publicly available for the NCI-60 cell lines. We found increased expression levels for estrogen receptor (ER) and ERBB2 in PIK3Ca mutant lines. The PIK3Ca mutation status was also correlated with compound screening data for the cell lines. PIK3Ca-mutant cell lines were relatively more sensitive than PIK3Ca-normal cell lines to the ER inhibitor tamoxifen and the AKT inhibitor triciribine, among other compounds. The results provide insights into the role of mutant PIK3Ca in oncogenic signaling and allow preliminary identification of novel targets for therapeutic intervention in cancers harboring PIK3Ca mutations.
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PMID:Correlation of PIK3Ca mutations with gene expression and drug sensitivity in NCI-60 cell lines. 1637 1

Novel activating mutations in sporadic colorectal cancer (CRC) have recently been identified on major kinase encoding genes such as BRAF and PI3KCA. The presence of these activating point mutations, including the well characterized KRAS oncogene mutations, represent up to 75% of cases in CRC. These genes, that have been implicated in the adenoma-carcinoma transition, cause deregulation and constitutive activation of the MAP AKT/kinase pathways, rendering growth advantages to colon tumor cells. This review focuses on the key genetic alterations underlying the cumulative effect of multiple mutations within the colon cancer cell. Moreover, the currently available and alternative treatment approaches that may target these different genetic alterations are discussed, such as the novel BRAF inhibitor. Identification of novel mutations as well as differential gene expression analyzed by microarray reveal potential targets for combined therapeutic protocols which will result in personalized treatments in the near future.
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PMID:Cancer genetics of sporadic colorectal cancer: BRAF and PI3KCA mutations, their impact on signaling and novel targeted therapies. 1661 9

Molecular analyses of tumors are increasingly useful for prognosis and for guiding therapy. Colonoscopic biopsy provides the first source of tissue for most cases of colorectal carcinoma and therefore might become an important source for molecular analyses. We have addressed the question whether molecular analyses of colonoscopic biopsy yield results similar to the findings from the surgical specimen. Further, we analyzed 2 separate areas of the colectomy specimen to assess tumor heterogeneity. We evaluated 3 samples from each of 67 patients for point mutations in the KRAS gene, loss of heterozygosity (LOH) at the Adenomatous Polyposis Coli (APC) and Deleted in Colon Cancer (DCC) genes and for microsatellite instability (MSI) using polymerase chain reaction based techniques. The average time interval between biopsy and surgery was 2.2+/-0.15 weeks. Lesions were from all colon segments and all surgical stages. The degree of agreement between the biopsy and surgical sites was high for APC LOH, MSI, and KRAS mutations (kappa=0.85, 1.00, and 0.93, respectively) but less so for DCC LOH (kappa=0.62). Colonoscopic biopsies are an acceptable source of neoplastic DNA for studies of KRAS, APC LOH, and MSI, but less so for DCC LOH, primarily resulting from technical considerations.
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PMID:Adequacy of colonoscopic biopsy specimens for molecular analysis: a comparative study with colectomy tissue. 1693 72

Our objective was to study whether products of oxidative stress, such as hydrogen peroxide (H(2)O(2)), trans-2-hexenal, and 4-hydroxy-2-nonenal (HNE), cause DNA damage in genes, relevant for human colon cancer. For this, total DNA damage was measured in primary human colon cells and colon adenoma cells (LT97) using the single-cell gel electrophoresis assay, known as "Comet Assay." APC, KRAS, and TP53 were marked in the comet images using fluorescence in situ hybridization (Comet FISH). The migration of APC, KRAS, or TP53 signals into the comet tails was quantified and compared to total DNA damage. All three substances were clearly genotoxic for APC, KRAS, and TP53 genes and total DNA in both types of cells. In primary colon cells, TP53 gene was more sensitive toward H(2)O(2), trans-2-hexenal, and HNE than total DNA was. In LT97 cells, the TP53 gene was more sensitive only toward trans-2-hexenal and HNE. APC and KRAS genes were more susceptible than total DNA to both lipid peroxidation products but only in primary colon cells. This suggests genotoxic effects of lipid peroxidation products in APC, KRAS, and TP53 genes. In LT97 cells, TP53 was more susceptible than APC and KRAS toward HNE. Based on the reported gatekeeper properties of TP53, which in colon adenoma is frequently altered to yield carcinoma, this implies that HNE is likely to contribute to cancer progression. This new experimental approach facilitates studies on effects of nutrition-related carcinogens in relevant target genes.
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PMID:Comet fluorescence in situ hybridization analysis for oxidative stress-induced DNA damage in colon cancer relevant genes. 1719 41

Despite tremendous effort and progress in the diagnostics of pancreatic cancer with respect to imaging techniques and molecular genetics, only very few patients can be cured by surgery leading to a 5-year survival rate of only 3%. Especially the lack of chemotherapeutical options in this entity requires a better understanding of the molecular mechanisms leading to pancreatic carcinoma growth and progression in order to develop novel treatment regimens. To identify signaling pathways that are critical for this tumor entity, we compared six well-established pancreatic cancer cell lines (Capan-1, Capan-2, HUP-T3, HUP-T4, KCL-MOH, PaTu-8903) with colon cancer cell lines and tumor cell lines of non-epithelial origin by expression profiling. For this purpose we employed Human Genome Focus Arrays representing about 8500 well annotated human genes. We identified 353 genes with significantly high expression in the group of pancreatic carcinomas. Based on Gene Ontology annotations these genes are especially involved in Rho protein signal transduction, proteasome activator activity, cell motility, apoptotic program, and cell-cell adhesion processes indicating these pathways to be interesting candidates for the design of targeted therapies. Most pancreatic carcinomas are characterized by mutations in the TP53 and the KRAS genes and the absence of microsatellite instability, which could also be confirmed for our panel of pancreatic carcinoma cell lines. Looking for individual differences within this group that may be responsible for more or less aggressive behavior, we identified genomic amplifications at the 8q22.1 and the 8q24.22 loci to be associated with enhanced gene transcription. Because we have previously shown that gains of genomic material from the long arm of chromosome 8 have an adverse effect on the outcome of pancreatic carcinoma patients, we conclude that functional analysis of amplified genes at 8q22 and/or 8q24 may lead to an improved understanding of pancreatic carcinoma progression.
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PMID:Expression analysis of pancreatic cancer cell lines reveals association of enhanced gene transcription and genomic amplifications at the 8q22.1 and 8q24.22 loci. 1720 80


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