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Quantitative DNA Methylation Profiling on a High-Density Oligonucleotide MicroarrayEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Recently, the analysis and functional elucidation of CpG island methylation has become a focus area of genomic research. Deviations from the normal parental imprinting pattern have been shown to cause developmental defects associated with serious symptoms. Aberrant DNA methylation of tumor suppressor and other functional genes, especially when found in 5′ untranslated regions and early exons, has been associated with tumorigenesis. In the context of applying DNA methylation analysis for the molecular characterization of cancer and other diseases, standardized protocols enabling parallel genome-wide methylation profil...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Manual Microdissection Combined with Antisense RNA–LongSAGE for the Analysis of Limited Cell NumbersEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Establishing a gene expression profile of defined subtypes of cells within an organ is still challenging because it frequently requires microdissection and subsequent amplification of the limited amount of messenger RNA (mRNA) isolated from the microdissected tissue in order to be able to proceed with comprehensive gene expression analyses via microarray or serial analysis of gene expression (SAGE) technology. Here we describe a manual microdissection strategy for the isolation of high-quality RNA. Furthermore, a strategy for combining linear amplification of RNA with longSAGE is described that allows the use of antisense ...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Automated Fluorescent Differential Display for Cancer Gene ProfilingEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Since its invention in 1992, differential display (DD) has become the most commonly used technique for identifying differentially expressed genes because of its many advantages over competing technologies such as DNA microarray, serial analysis of gene expression (SAGE), and subtractive hybridization. A large number of these publications have been in the field of cancer, specifically on p53 target genes. Despite the great impact of the method on biomedical research, there had been a lack of automation of DD technology to increase its throughput and accuracy for systematic gene expression analysis. Many previous DD work has...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Mining Expressed Sequence Tag (EST) Libraries for Cancer-Associated GenesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Originally established in the beginning of the 1990s as a direct route to gene finding, expressed sequence tags (ESTs) still lend themselves as a means to analyze gene expression in almost all human tissues. The type of questions that can be addressed using public EST libraries ranges from tissue-specific gene profiling to the comparison between tissues in diseased and healthy states. Thanks to a multitude of web-based online bioinformatics resources, mining in EST libraries is not restricted to experts in the field of data analysis, but can readily be performed by the medical or life scientist. In this chapter, a couple o...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

A Decade of Cancer Gene Profiling: From Molecular Portraits to Molecular FunctionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Cancer gene profiling has greatly profited from the progress in high-throughput technologies, including microarray-, sequencing-, and bioinformatics-based methods. The flood of data generated during the last decade has provoked a panel of “-omics” fields that significantly changed our understanding of malignant diseases. However, while the terms “-omics” and “-ome” in principle refer to the completeness of a genetic approach, we are in fact far from a complete understanding of cancer progression. We may understand gene expression patterns better and successfully use gene signatures for o...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Tissue MicroarraysEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Modern array technologies allow for the simultaneous screening of virtually all human genes on the DNA and RNA level. Studies using such techniques have lead to the identification of hundreds of genes with a potential role in cancer or other diseases. The validation of all of these candidate genes requires in situ analysis of high numbers of clinical tissues samples. The tissue microarray (TMA) technology greatly facilitates such analysis. In this method, minute tissue samples (0.6 mm in diameter) from up to 1,000 different tissues can be analyzed on one microscope glass slide. All in situ methods suitable for histological...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Laser MicrodissectionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Gene expression analysis requires a sound basis of cell material to obtain realistic results. Tissue, however, consists of diverse types of cells, which often differentially express target genes, so that cell populations need to be selected. If tissue diversity is moderate and negligible, manual microdissection can be the cost-efficient method of choice. In contrast, the advantage of laser microdissection is a very exact selection down to the level of a single cell, but often with a considerable time needed to get enough material for the following analyses. The latter issue and the method of tissue preparation needed for l...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Manual MicrodissectionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The new opportunities of modern assays of molecular biology can only be exploited fully if the results can be accurately correlated to the tissue phenotype under investigation. This is a general problem of non-in situ techniques, whereas results from in situ techniques are often difficult to quantitate. The use of bulk tissue, which is not precisely characterized in terms of histology, has long been the basis for molecular analysis. It has, however, become apparent, that this simple approach is not sufficient for a detailed analysis of molecular alterations, which might be restricted to a specific tissue phenotype (e.g., t...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Target Gene Discovery for Novel Therapeutic Agents in Cancer TreatmentEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Target identification of novel therapeutic drugs is pivotal for the establishment of (1) new anticancer regimens, (2) to control side effects of the drugs, and (3) to identify appropriate combinations with established drugs. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Meta-analysis of Cancer Gene-Profiling DataEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
DNA microarray profiles are plagued by the issue of large number of variables but small number of samples and are often notorious for their low signal-to-noise ratio for clinical applications. Therefore, a great need for meta-analysis techniques is emerging to yield more valid and informative results than each experiment separately. By exploring the power of several studies in one single analysis, meta-analysis of many cancer gene-profiling data increases the statistical power to detect differentially expressed genes and allows assessment of heterogeneity. OrderedList is such a method that was specially proposed for cancer...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Calibration of Microarray Gene-Expression DataEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We present our hook method, a new calibration approach that is based on a graphical summary of the actual hybridization characteristics of a particular microarray. Although single-chip related, hook performs as well as the multi-chip-related gcRMA, presently one of the best state-of-the-art methods for estimating expression values. The hook method, in addition, provides a set of chip summary characteristics that evaluate the performance of a given hybridization. The algorithm of the method is briefly described and its performance is exemplified. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Organizational Issues in Providing High-Quality Human Tissues and Clinical Information for the Support of Biomedical ResearchEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The objectives of tissue repositories also vary based on the types of tissues provided (e.g., fresh tissue aliquots, fixed paraffin-embedded tissue, paraffin tissue sections, etc.) and how the tissues are to be used in research. For example, the potential use of tissues affects the need for extensive annotation of the specimen including both clinical information (e.g., clinical outcomes) and demographics. Specifically, if the tissues are to be used for extraction of proteins or basic studies of disease processes, less clinical information, if any, may be needed than if the tissues are to be used for the correlation of an a...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Functional Profiling Methods in CancerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The introduction of new high-throughput methodologies such as DNA microarrays constitutes a major breakthrough in cancer research. The unprecedented amount of data produced by such technologies has opened new avenues for interrogating living systems although, at the same time, it has demanded of the development of new data analytical methods as well as new strategies for testing hypotheses. A history of early successful applications in cancer boosted the use of microarrays and fostered further applications in other fields. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Quantitation of CD39 Gene Expression in Pancreatic Tissue by Real-Time Polymerase Chain ReactionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Within the past decade, the field of gene expression analysis has constantly evolved, with numerous technologies being available for RNA quantification, including differential display, serial analysis of gene expression (SAGE), quantitative real-time (qRT) polymerase chain reaction (PCR), and microarrays. Although every technique has its specific application, the high levels of accuracy, reproducibility, sensitivity, and specificity have established qRT-PCR as a standard method for detection and quantification of gene expression. In this chapter, all steps of the qRT-PCR procedure, including purification of total RNA from ...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

The EGFR Pathway as an Example for Genotype: Phenotype Correlation in Tumor GenesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We describe the correlation between the EGFR genotype, including the KRAS mutation, and the consequences of the resulting genotype for anti-EGFR therapy in colorectal cancer. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Cancer Gene Profiling for Response PredictionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Preoperative treatment strategies are now recommended for a variety of human cancers. Unfortunately, the response of individual tumors to a preoperative treatment is not uniform, and ranges from complete regression to resistance. This poses a considerable clinical dilemma, because patients with a priori resistant tumors could either be spared exposure to radiation or DNA-damaging drugs, i.e., they could be referred to primary surgery or dose-intensified protocols could be pursued. Because the response of an individual tumor as well as therapy-induced side effects represent the major limiting factors of current treatment st...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Cancer Gene Profiling in Prostate CancerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Gene profiling and expression analysis using microarrays have made a significant impact on our biological understanding of prostate cancer. The procedures for generating high-quality expression data from prostate cancer cell lines and tumors are not trivial. However, during the past 9 years, methods by which to process samples for gene profiling have been developed. In this chapter, techniques to process prostate cancer specimens either en bloc (macrodissection) or using laser capture microdissection are presented in detail along with extensive technical notes. Although we focus on prostate cancer and discuss the specific ...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Cancer Gene Profiling in Pancreatic CancerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
High levels of RNases present in the normal pancreas and the abundance of desmoplastic stroma of most pancreatic cancers have traditionally caused difficulty in the extraction of high-quality RNA and gene expression profiling from pancreatic tissues. However, a variety of innovative strategies have made it possible to successfully perform a molecular analysis of pancreatic cancer, and the expression profiles that have been generated have provided tremendous insight into the nature of this aggressive disease. Here, we describe some of these techniques. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Integrative Oncogenomic Analysis of Microarray Data in Hematologic MalignanciesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
During the last decade, gene expression microarrays and array-based comparative genomic hybridization (array–CGH) have unraveled the complexity of human tumor genomes more precisely and comprehensively than ever before. More recently, the simultaneous assessment of global changes in messenger RNA (mRNA) expression and in DNA copy number through “integrative oncogenomic” analyses has allowed researchers the access to results uncovered through the analysis of one-dimensional data sets, thus accelerating cancer gene discovery. In this chapter, we discuss the major contributions of DNA microarrays to the stud...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Search for and Identification of Novel Tumor-Associated AutoantigensEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
During the development of tumors, autoantibodies against aberrant or overexpressed autoantigens can be induced. Several hundreds of tumor-associated autoantibodies (TAAB) with more or less specificity for tumors have been found until now by molecular cloning and proteomics technologies. Many TAAB are detectable in preclinical stages of the disease and may be indicators of tumor development. The screening for autoantibody responses in tumor patients may lead to new diagnostic tumor markers and may be a simple and effective way to identify concomitantly cytotoxic T-lymphocyte (CTL) reactivity. However, most of the TAAB lack ...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Application of Proteomics in Cancer Gene Profiling: Two-Dimensional Difference in Gel Electrophoresis (2D-DIGE)Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
In the post-genomic era, proteomic strategies are at the forefront of cancer research. By studying the complement of all expressed genes, proteomics aims to provide knowledge of biomarkers indicative of the physiological state of cancer cells at a specific time, enabling screening, early diagnosis, monitoring the course of cancer development/progression, and gauging the efficacy and safety of novel therapeutic agents. Onco-proteomics thus has the ability to revolutionise oncology practice by delivering highly selective and individualised clinical care. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Single-Nucleotide Polymorphism (SNP) Analysis to Associate Cancer RiskEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Identification of hereditary factors that predispose to cancer allows targeted cancer screening and better quantification of environmental risk factors. The ability to identify which single nucleotide polymorphisms (SNPs) are associated with cancer or segregate with disease in families allows high-risk loci to be identified. In this chapter, two platforms for analysing SNPs are discussed, the Affymetrix and Illumina systems. Application of both platforms requires the same principles of good laboratory practice but there are important differences in materials and methods, which will be discussed. (Source: Springer protocols...
Source: Springer protocols feed by Cancer Research - September 15, 2009 Category: Cancer & Oncology Source Type: info

Hormones as Biomarkers: Practical Guide to Utilizing Luminex Technologies for Biomarker ResearchEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Hormones are chemical messengers produced in one part of the body and released into the blood to trigger or regulate particular functions of the body in another part. Hormone actions vary widely, but can include stimulation or inhibition of growth, induction or suppression of apoptosis, activation or inhibition of the immune system, regulating metabolism, and preparation for a new activity or phase of life. There is a growing interest in the role that hormones may play in the development and progression of various cancers. Recent research suggests that hormone levels may explain differences in risk for some of the most com...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Serum Proteomics Using Mass SpectrometryEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The identification and eventual application of tumor markers in cancer screening, early detection, diagnosis, and prognosis is a continuing focus of significant translational cancer research. While many new candidate markers have been discovered and at least partly characterized, very few have found widespread clinical application limited presently to the use of CA-125 in ovarian cancer, CEA, primarily in colon cancer, and PSA in prostate cancer screening and patient monitoring. The rapidly emerging field of cancer genomics and proteomics, and their clinical translation as “molecular diagnosis” and “molec...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Reverse-Phase Protein Microarrays for Theranostics and Patient Tailored TherapyEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Analysis of the genome provides important information about the somatic genetic changes existing in the tissue; however, it is the proteins that do the work of the cell. Diseases such as cancer are caused by derangements in cellular protein molecular networks and cell signaling pathways. These pathways contain a large and growing collection drug targets, governing cellular survival, proliferation, invasion, and cell death. The clinical utility of reverse-phase protein microarrays (RPPA), a new technology invented in our laboratory, lies in its ability to generate a functional map of known cell signaling networks or pathway...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

All-Liquid Separations, Protein Microarrays, and Mass Spectrometry to Interrogate Serum Proteomes: An Application to Serum GlycoproteomicsEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Disease-related changes in serum proteins are reasonable targets for early detection particularly due to the noninvasive approach in obtaining samples. Glycoproteins specifically have been implicated in a variety of disease types ranging from immune diseases to cancers. High-throughput screening methods that can assess glycosylation states of all serum proteins in normal and diseased sample groups can facilitate early detection as well as shed light on disease progression mechanisms. Outlined here is a combination of liquid separation, protein microarray, and mass spectrometry approach to highlight candidate proteins invol...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Glycoproteomic Analysis by Two-Dimensional ElectrophoresisEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Changes in N-linked glycosylation are known to occur during the development of cancer. For example, increased branching of oligosaccharides has been associated with metastasis and has been correlated to tumor progression in human cancers of the breast, colon, and melanomas. Increases in core fucosylation have also been associated with the development of hepatocellular carcinoma (HCC). To a large extent, the proteins to which these N-linked glycans are attached have been unknown. However, with the advent of sensitive glycan analysis and proteomic technologies, the ability to comprehensively identify all the fucosylated prot...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Analysis of Glycans on Serum Proteins Using Antibody MicroarraysEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We describe those methods along with the methods for preparing and treating serum samples, running the experiments, and designing and interpreting the experiments. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Discovery of Antibody Biomarkers Using Protein Microarrays of Tumor Antigens Cloned in High ThroughputEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Development of humoral and cellular immunity against self-cellular proteins in cancer patients is a phenomenal observation. The ability of immune system to sense the presence of the disease and to fight of the disease by generating autoantibodies against tumor antigens makes it a natural biosensor. Several screening technologies have been employed for the identification of tumor-specific antibodies in cancer patients. We have developed a multidimensional approach for the identification of diagnostic antigens that utilizes a combination of high-throughput antigen cloning and protein microarray-based serological detection of...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Autoantibodies Against Cancer AntigensEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We describe in this chapter a simple implementation of a serological monitoring platform for autoantibody measurement in cancer patients, from production of recombinant antigens to ELISA testing and interpretation. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Magnetic Resonance Spectroscopy of Living TissuesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The comprehensive work of both clinical and basic science colleagues has demonstrated a clear proof of concept for “in vitro discovered- in vivo validated” biomarkers in translational metabolic profiling research using magnetic resonance techniques. Major tissue metabolites (initially discovered by high-resolution in vitro techniques on cancer specimens) can be translated into in vivo protocols based on noninvasive magnetic resonance spectroscopy (MRS). Using 1H- and 31P-MRS on living animals or patients, a decrease in citrate and polyamines in prostate cancer, an increase of cholines in breast cancer, as well ...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

MRI and MRS of Human Brain TumorsEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The purpose of this chapter is to provide an introduction to magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) of human brain tumors, including the primary applications and basic terminology involved. Readers who wish to know more about this broad subject should seek out the referenced books (1. Tofts (2003) Quantitative MRI of the brain. Measuring changes caused by disease. Wiley; Bradley and Stark (1999) 2. Magnetic resonance imaging, 3rd Edition. Mosby Inc; Brown and Semelka (2003) 3. MRI basic principles and applications, 3rd Edition. Wiley-Liss) or reviews (4. Top Magn Reson Imaging 17:127&nda...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Metabolomics of CancerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Metabolomics, one of the “omic” sciences in systems biology, is the global assessment and validation of endogenous small-molecule biochemicals (metabolites) within a biologic system. Initially, putative quantitative metabolic biomarkers for cancer detection and/or assessment of efficacy of anticancer treatment are usually discovered in a preclinical setting (using animal and human cell cultures), followed by translational validation of these biomarkers in biofluid or tumor tissue. Based on the tumor origin, various biofluids, such as blood, urine, and expressed prostatic secretions, can be used for validating m...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Identification of Tumor-Associated Antigens as Diagnostic and Predictive Biomarkers in CancerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Many studies demonstrated that cancer sera contain antibodies which react with autologous cellular antigens generally known as tumor-associated antigens (TAAs). In our laboratories, the approach used in the identification of TAAs has involved initially examining the sera of cancer patients using extracts of tissue culture cells as source of antigens in Western blotting and by indirect immunofluorescence on whole cells. With these two techniques, we identify sera which have high-titer fluorescent staining or strong signals to cell extracts on Western blotting and subsequently use these sera as probes in immunoscreening cDNA...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Developing Classifiers for the Detection of Cancer Using Multi-AnalytesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The development of a successful classifier from multiple predictors (analytes) is a multistage process complicated typically by the paucity of the data samples when compared to the number of available predictors. Choosing an adequate validation strategy is key for drawing sound conclusions about the usefulness of the classifier. Other important decisions have to be made regarding the type of prediction model to be used and training algorithm, as well as the way in which the markers are selected. This chapter describes the principles of the classifier development and underlines the most common pitfalls. A simulated dataset ...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Study Designs in Genetic EpidemiologyEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Identification of germline mutations that may modulate individual risk of developing cancer is a rapidly developing field. Over the last few decades, germline mutations in p53, BRCA1, BRCA2, APC, MLH1, MSH2, and MSH6 have been identified in families with a large number of relatives who have been diagnosed with particular types of cancer. These mutations are rare but substantially increase the risk of cancer in carriers, and account for a small fraction of cancer cases diagnosed in the general population. The search for common mutations that correlate with a very modest increased risk of developing cancer is ongoing. With t...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

PharmacogenomicsEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
encompasses several major areas including the identification and analysis of variations of DNA and RNA that affect the efficacy and toxicity of drug therapy. It represents an integration of analytical approaches including DNA and RNA detection and quantitation which may be applied to either candidate genes or a global genome analysis. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Analysis of Loss of Heterozygosity in Circulating DNAEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Analysis of genetic altera tion in circulating DNA can have clinical utility in predicting disease outcome. Loss of heterozygosity (LOH) of DNA microsatellites has been shown to occur commonly among all chromosomes in various cancers, such as melanoma, breast cancer, and lung cancer. In this protocol, we focused on the utility of LOH of microsatellite biomarkers for detection of analyzing circulating DNA. The protocol describes how PCR is performed on each patient’s paired DNA samples (normal lymphocyte DNA and serum DNA) using specific microsatellite biomarkers followed by post-PCR product analysis using capillary a...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

DNA Sequencing of Cancer-Related Genes for Biomarker DiscoveryEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Dideoxy DNA sequencing is routinely used in research and, increasingly, in clinical care for the detection of DNA sequence variants, single nucleotide changes, or small insertions or deletions, when the spectrum of DNA variation is unknown. DNA sequence variation can be present in tumor tissue that is not present in the normal tissue from the same individual. This somatic DNA sequence variation is often the cause of abnormal cell growth and/or regulation and, ultimately, tumorigenesis. Identification of these oncogenic DNA sequence variants has successfully led to the development of cancer therapies, since the abnormal pro...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

High-Throughput Mutation Screening Using a Single Amplification ConditionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Numerous innovative and high-throughput techniques have been established to identify human disease genes. However, DNA sequencing of candidate genes still remains as a major limitation in the identification of causative mutations. Much of this limitation is due to the time and labor needed for the polymerase chain reaction (PCR) optimization and reaction setup. Toward this end, we have established a simplified protocol that utilizes a single PCR amplification condition. PCR purification is accomplished via enzymatic digestion and all products can be sequenced using universal primers. This combination of a single amplificat...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Tumor Marker Discovery by Expression Profiling RNA from Formalin Fixed Paraffin Embedded TissuesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Clear identification among early-stage cancer patients of those at highest risk of having metastatic disease would be of great benefit in treatment planning and management. Considerable additional benefit would accrue to high-risk patients if their responses to specific therapeutic alternatives could be predicted. Molecular biomarkers in the form of gene expression profiles are proving to be more effective tools for both prognostic and predictive patient stratification than more traditional methods such as patient demographics and histopathology indicators. Such biomarkers must be clinically validated before they can be ef...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Quantitative, Fluorescence-Based In-Situ Assessment of Protein ExpressionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
As companion diagnostics grow in prevalence and importance, the need for accurate assessment of in situ protein concentrations has increased. Traditional immunohistochemistry (IHC), while valuable for assessment of context of expression, is less valuable for quantification. The lack of rigorous quantitative potential of traditional IHC led to our development of an immunofluorescence-based method now commercialized as the AQUA® technology. Immunostaining of tissue samples, image acquisition, and use of AQUA software allow investigators to quickly, efficiently, and accurately measure levels of expression within user-defi...
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

Tissue Microarrays as a Tool in the Discovery and Validation of Tumor MarkersEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Tissue microarrays are a platform of condensed histopathology that has revolutionized the translation of basic science to clinical utility. Tissue microarrays have resulted in a paradigm shift from histopathology to immunopathology and moved analysis of small selected samples sets of tens of specimens to a high-throughput environment of hundreds of specimens. Tissue microarrays have influenced validation strategies, but have a role in discovery as well, allowing a pathways approach to analysis of tumors. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

High-Throughput Analysis of Serum Antigens Using Sandwich ELISAs on MicroarraysEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Enzyme-linked immunosorbent assay (ELISA) microarrays promise to be a powerful tool for the detection and validation of disease biomarkers. ELISA microarrays are capable of simultaneous detection of many proteins using a small sample volume. Although there are many potential pitfalls to the use of ELISA microarrays, these can be avoided by careful planning of experiments. In this chapter we describe a high-throughput protocol for processing ELISA microarrays that will result in reliable and reproducible data. (Source: Springer protocols feed by Cancer Research)
Source: Springer protocols feed by Cancer Research - February 3, 2009 Category: Cancer & Oncology Source Type: info

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Despite the current availability of an impressive in vitro assay battery developed to quantitatively analyze the broad panel of small compounds and macromolecules that possess the inflammatory potential, little methodology exists nowadays that affords a researcher or clinician to quantify the ultimate output on the level of cell signaling response caused by inflammatory pathway stimulation. As a matter of fact, majority of analytical tools measure bona fide inflammatory substances (e.g., cytokines or chemokines) by their direct binding to secondary reagents such as specific antibodies or other selectively affine substrates...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info

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Nitric oxide (NO) is a free radical that is involved in the inflammatory process and carcinogenesis. There are four nitric oxide synthase enzymes involved in NO production: induced nitric oxide synthase (iNOS), endothelial NO synthase (eNOS), neural NO synthase (nNOS), and mitochondrial NOS. iNOS is an inducible and key enzyme in the inflamed tissue. Recent literatures indicate that NO as well as iNOS and eNOS can modulate cancer-related events including nitro-oxidative stress, apoptosis, cell cycle, angio-genesis, invasion, and metastasis. This chapter focuses on linking NO/iNOS/eNOS to inflammation and carcinogenesis fro...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info

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Activation of the transcription factor, nuclear factor of κB (NF-κB), has recently emerged as the critical link between inflammation and cancer. NF-κB is activated by many signals including DNA damage and microbial pathogen recognition. Although microbial infections account for an estimated 15% of all cancer related deaths, NF-κB is constitutively activated in various types of cancer as a direct result of chromosomal translocations, deletions and mutations affecting both NF-κB and its regulators. NF-κB exerts the transcriptional activation and repression of inflammatory and immune respon...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info

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Global gene expression profiling studies conducted over the last couple of years have shown that molecular profiling of breast cancers can be used to identify clinically and genetically significant subtypes of breast carcinomas and subgroups of patients with different prognosis or disease outcome, and to predict therapeutic response to both endocrine and chemotherapeutic drugs. We studied one particularly aggressive form of locally advanced breast cancer, namely inflammatory breast cancer (IBC) using cDNA micro-arrays. We demonstrated that IBC is characterized by a different gene expression profile, not related to any of t...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info

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It is now apparent that epigenetic abnormalities, in particular altered DNA methylation, play a crucial role in the development and progression of human cancers. DNA hypermethylation at promoter CpG islands is now recognized as a third mechanism by which inactivation of tumor suppressor genes occurs. Aberrant CpG island hypermethylation is also frequently observed in chronic inflammation and precancerous lesions, which suggests that it is an early event in tumorigenesis that could serve as a useful tumor marker. A variety of screening techniques have been developed for genome-wide screening of methylation status. Of those,...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info

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NF-κB plays a pivotal role in immunity and inflammation and is considered to be a promising candidate for drug development. However, global suppression of NF-κB may have undesirable side-effects. Our data and the results of others suggest that each of the five NF-κB subunits may have a specific function in controlling the expression of inflammatory mediators in immune cells. Identifying the role for each NF-κB subunit in primary human immune cells will allow a more targeted approach to inhibiting NF-κB subunit-specific cellular functions. However, results obtained with primary human cells can ...
Source: Springer protocols feed by Cancer Research - January 1, 2009 Category: Cancer & Oncology Source Type: info