Meeting report: mitochondrial DNA and cancer epidemiology.
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DNA, Mitochondrial↗
Biomedical subjects
Mukesh Verma
Publications and source records attributed to Mukesh Verma.
Genetic and epigenetic biomarkers in cancer diagnosis and identifying high risk populations.
Biomarkers present the normal and/or disease state in humans. Genetic and epigenetic biomarkers assessed in easily accessible biological materials are useful in diagnosis, early onset or risk of developing cancer or to predict the treatment efficacy or clinical outcome of different human malignancies. Moreover, some of these markers are expressed during early stages of the tumor development and hence provide an opportunity to develop intervention and treatment strategies. Attempts are being made to validate cancer biomarkers in non-invasively collected samples. Multiplexing of clinically validated markers is still a challenge. Once validated, these markers can be utilized in clinical settings and to identify high risk populations. In this review, the current status of the clinical genetic and epigenetic biomarkers and their implication in cancer diagnosis and risk assessment are discussed.
Biomarkers, Tumor↗
Genetic and epigenetic biomarkers in cancer : improving diagnosis, risk assessment, and disease stratification.
Gene expression patterns change during the initiation, progression, and development of cancer, as a result of both genetic and epigenetic mechanisms. Genetic changes arise due to irreversible changes in the nucleotide sequence, whereas epigenetic changes occur due to changes in chromatin conformation, histone acetylation, and methylation of the CpG islands located primarily in the promoter region of a gene. Both genetic and epigenetic markers can potentially be utilized to identify different stages of tumor development. Several such markers exhibit high sensitivity and specificity for different tumor types and can be assayed in biofluids and other specimens collected by noninvasive technologies. In spite of the availability of large numbers of diagnostic markers, only a few have been clinically validated so far. The current status and the challenges in the field of genetic and epigenetic markers in cancer diagnosis, risk assessment, and disease stratification are discussed.
Biomarkers, Tumor↗
Use of nanotechnology for the development of novel cancer biomarkers.
Novel nanotechnologies can complement and augment existing genomic and proteomic techniques employed to analyze variations across different tumor types, thus offering the potential to distinguish between normal and malignant cells. Sensitive biosensors constructed out of nanoscale components (e.g., nanocantilevers, nanowires and nanochannels) can recognize genetic and molecular events and have reporting capabilities, thereby offering the potential to detect rare molecular signals associated with malignancy. Such signals may then be collected for analysis by nanoscale harvesters that selectively isolate cancer-related molecules from tissues. Another area with near-term potential for the early detection of cancer is the identification of mutations and genomic instability.
Animals↗
WT1 and WT1-AS genes are inactivated by promoter methylation in ovarian clear cell adenocarcinoma.
BACKGROUND: Ovarian clear cell adenocarcinoma is associated with one of the poorest prognoses among human epithelial ovarian cancers. The authors hypothesized that Wilms tumor suppressor 1 gene (WT1) sense and antisense (WT1-AS) expression and their promoter methylation status could characterize ovarian clear cell adenocarcinoma from ovarian serous adenocarcinoma. METHODS: To test this hypothesis, ovarian cancer cell lines and 42 cancer tissues (17 clear cell and 25 serous adenocarcinoma) were analyzed for expression and methylation of WT1 and WT1-AS genes. RESULTS: These experiments demonstrated that all serous adenocarcinoma tissues expressed both WT1 and WT1-AS genes, although expression of these genes was lacking in clear cell adenocarcinoma. The WT1 and WT1-AS promoter were significantly methylated in clear cell adenocarcinoma (88.2% and 88.2%, respectively) compared with serous adenocarcinoma (24.0% and 20.0%, respectively). Significant correlation between methylation and mRNA expression status was observed for each gene. Also in agreement with these data, WT1 and WT1-AS negative ovarian cancer cell lines reexpressed these genes after treatment with the demethylating agent, 5-aza-2'-deoxycytidine. CONCLUSIONS: The current study shows that CpG hypermethylation is an important mechanism of WT1 and WT1-AS gene inactivation in ovarian clear cell adenocarcinoma. This is the first report that has demonstrated differential expression and methylation of WT1-AS in ovarian clear cell and serous adenocarcinomas. This study presents new molecular characterizations between these two types of adenocarcinoma and may provide insight as to why clear cell adenocarcinoma has a poorer prognosis than serous adenocarcinoma of the ovary.
Adenocarcinoma, Clear Cell↗
Pancreatic cancer epidemiology.
Carcinoma of the exocrine pancreas remains a challenging disease mainly due to advanced stage diagnosis, the early systemic dissemination, aggressive local tumor progression, and subsequent short patient survival. Thus, assessment of incidence markers instead of prevalence indicators is recommended because the time between diagnosis and death is usually very short. Investigations to date have led to the discovery of many rare genes and environmental factors that contribute to pancreatic cancer. However, common genes involved in genetic polymorphisms, and specific risk factors have not been identified. Furthermore, the role of gene-environment interactions on the tumorogenesis and progression of pancreatic cancer need to be further investigated. Given current therapeutic modalities, only early detection of cancer of the pancreas, followed by surgical resection, offers the possibility of lengthening survival time.
Environment↗
CpG hypermethylation of MDR1 gene contributes to the pathogenesis and progression of human prostate cancer.
Multidrug resistance 1 (MDR1) gene encodes for P-glycoprotein (P-gp), a Mr 170,000 transmembrane calcium-dependent efflux pump that is inactivated in prostate cancer. We hypothesize that inactivation of the MDR1 gene through CpG methylation contributes to the pathogenesis and progression of prostate cancer. To test this hypothesis, CpG methylation status of the MDR1 promoter and its correlation with clinicopathological findings were evaluated in 177 prostate cancer samples and 69 benign prostate hypertrophy (BPH) samples. Cellular proliferation index and apoptotic index were determined by proliferating cell nuclear antigen (PCNA) and single-strand DNA immunostaining, respectively. After 5-aza-2'-deoxycytidine treatment, increased expression of MDR1 mRNA transcript was found in prostate cancer cell lines (DU145, DuPro, and ND1). MDR1 methylation frequency was significantly higher in prostate cancer samples compared with BPH samples (54.8 versus 11.6%, respectively, P < 0.001). Logistic regression analysis revealed that PC patients are 11.5 times more likely to have MDR1 methylation than BPH patients (95% confidence interval 4.87-27.0) and that MDR1 methylation is independent of the age. Significant correlation of MDR1 methylation was observed with high pT category (P < 0.001), high Gleason sum (P = 0.008), high preoperative prostate-specific antigen (P = 0.01), and advancing pathological features. In addition, PCNA-labeling index were significantly higher in methylation-specific PCR (MSP)-positive than in MSP-negative prostate cancer samples (P = 0.048). In contrast, no significant difference in apoptotic index was found between MSP-positive and -negative prostate cancer samples. These findings suggest that CpG hypermethylation of MDR1 promoter is a frequent event in prostate cancer and is related to disease progression via increased cell proliferation in prostate cancer cells.
Aged↗
Expression and methylation status of 14-3-3 sigma gene can characterize the different histological features of ovarian cancer.
We hypothesize that 14-3-3 sigma gene expression and its regulation by methylation can characterize histological types of primary human epithelial ovarian cancer. To test this hypothesis, ovarian cancer cell lines and 54 ovarian cancer tissue samples were analyzed for expression and methylation of 14-3-3 sigma gene using methylation specific PCR. The results of our experiments demonstrate that 14-3-3 sigma gene was methylated and inactivated in ES-2 ovarian cell line, which was derived from clear cell adenocarcinoma. Treatment of this cell line with demethylating agent 5-aza-2'-deoxycytidine restored the expression of 14-3-3 sigma gene. In human ovarian cancer tissues, the expression of 14-3-3 sigma protein was inactivated in most of the ovarian clear cell carcinoma tissues. Interestingly, 14-3-3 sigma protein expression was positive in significantly higher percentages of serous (89.5%), endometrioid (90%), and mucinous (81.8%) ovarian adenocarcinoma tissues. The ovarian clear cell carcinoma samples with inactivated 14-3-3 sigma protein were highly methylated, suggesting that inactivation of 14-3-3 sigma gene is through DNA methylation. Using direct DNA sequencing, 14-3-3 sigma gene methylation on all the 17 CpG sites was significantly higher in ovarian clear cell carcinoma as compared to other histological types of ovarian cancer (serous, endometrioid, and mucinous). This is the first report suggesting that 14-3-3 sigma gene expression and methylation status can characterize histological features of different types of ovarian cancer.
14-3-3 Proteins↗
Data standards for proteomics: mitochondrial two-dimensional polyacrylamide gel electrophoresis data as a model system.
Proteomics has emerged as a major discipline that led to a re-examination of the need for consensus and a nationally sanctioned set of proteomics technology standards. Such standards for databases and data reporting may be applied to two-dimensional polyacrylamide gel electrophoresis (2D PAGE) technology as a pilot project for assessing global and national needs in proteomics, and the role of the National Institute of Standards and Technology (NIST) and other similar standards and measurement organizations. The experience of harmonizing the heterogeneous data included in the Protein Data Bank (PDB) provides a paradigm for technology in an area where significant heterogeneity in technical detail and data storage has evolved. Here we propose an approach toward standardizing mitochondrial 2D PAGE data in support of a globally relevant proteomics consensus.
Journal Article↗
Epigenetics and cancer.
Both genetics and epigenetics regulate gene expression in cancer. Regulation by genetics involves a change in the DNA sequence, whereas epigenetic regulation involves alteration in chromatin structure and methylation of the promoter region. During the initiation, development, and progression of cancer, a number of genes undergo epigenetic changes. Some of these changes can be used as biomarkers for early detection of cancer as well as to follow treatment. A panel of epigenetic biomarkers is preferred to a single biomarker in clinical assays. Changes in gene expression due to epigenetic regulation can be reversed by chemicals, and this approach opens up a novel approach in cancer prevention and treatment.
Biomarkers, Tumor↗
Challenges for biomarkers in cancer detection.
Cancer remains the leading cause of death in the United States. Biomarkers can be used to detect cancer in different stages, initiation, development, and progression. The desirable property and utility of a biomarker lie in its ability to provide an early indication of disease progression. Biomarkers should be easy to detect, measurable across populations, and useful for detection of cancer at an early stage, identification of high-risk individuals, detection of recurrence, or monitoring endpoints in intervention studies. Recent technological advances have helped develop noninvasive, sensitive, and specific biomarkers to detect cancer at early stages of the disease.
Biomarkers, Tumor↗
Molecular quality of exfoliated cervical cells: implications for molecular epidemiology and biomarker discovery.
The biologic sample collected in molecular epidemiology studies must accurately reflect the disease being studied and have sufficient molecular quality for the intended assays. Noninvasive sampling methods, such as scrapes or brushes, are increasingly used. In this study, we evaluate the impact of sample collection media and extraction methods on the quality and yield of RNA from routine exfoliated cervical cytology. Excess cellular material remaining on the cytologic collection device after preparation of the routine screening Papanicolaou smear was placed in a variety of collection media and extracted using two commercial kits. The collection media had the largest impact on the yield and quality of RNA as evaluated by denaturing agarose gel electrophoresis and image analysis. Two collection media, PAXgene and PreservCyt, yielded RNA from most samples. The RNA showed some degree of degradation as evidenced by the reduced size of the higher molecular weight ribosomal band. However, with a sensitive gold particle-based detection method, reproducible microarray results were obtained using this RNA.
Biomarkers, Tumor↗
New cancer biomarkers deriving from NCI early detection research.
Cancer is not a single disease but an accumulation of several events, genetic and epigenetic, arising in a single cell over a long time interval. A high priority in the cancer field is to identify these events. This can be achieved by characterizing cancer-associated genes and their protein products. Identifying the molecular alterations that distinguish any particular cancer cell from a normal cell will ultimately help to define the nature and predict the pathologic behavior of that cancer cell. It will also indicate the responsiveness to treatment of that particular tumor. Understanding the profile of molecular changes in any particular cancer will be extremely useful as it will become possible to correlate the resulting phenotype of that cancer with molecular events. Achieving these goals and knowledge will provide an opportunity for discovering new biomarkers for early cancer detection and developing prevention approaches. This will also help us identify new targets for therapeutic development. Advancement in technology includes methods and tools that enable research including, but not limited to, instrumentation, techniques, devices, and analysis tools (e.g., computer software). Resources such as databases, reagents, and tissue repositories are different than technologies. The identification and definition of the molecular profiles of cancer will require the development and dissemination of high-throughput molecular analysis technologies, as well as elucidation of all of the molecular species embedded in the genome of cancer and normal cells. The main challenge in cancer control and prevention is to detect the cancer early. This could then enable effective interventions and therapies contributing to reduction in mortality and morbidity. At a specific time, biomarkers serve as molecular signposts of the physiologic state of a cell. These signposts are the result of genes, their products (proteins) and other organic chemicals made by the cell. Biomarkers could prove to be vital for the identification of early cancer and subjects at risk of developing cancer as a normal cell progresses through the complex process of transformation to a cancerous state. This chapter discusses ongoing research in genetic and proteomic approaches to identify molecular signatures such as protein profiles, microsatellite instability, hypermethylation, and single nucleotide polymorphisms. Other topics covered here include the use of genomics and proteomics as high-throughput technology platforms to facilitate biomarker-aided detection of early cancer. Other areas covered include issues surrounding the analysis, validation, and predictive value of biomarkers using such technologies. Recent advances in noninvasive techniques, such as buccal cell isolates serving as viable sources of biomarkers, complementary to traditional sources such as serum or plasma, are also presented. The review also brings attention to the efforts of the Early Detection Research Network (EDRN) at the National Cancer Institute (NCI), in bringing together scientific expertise from leading national and international institutions, to identify and validate biomarkers for the detection of precancerous and cancerous cells in determining risk for developing cancer. The network's serious determined efforts in linking discovery to process development, resulting in early detection tests and clinical assessment, are also discussed.
Biomarkers, Tumor↗
Global amplification of sense RNA: a novel method to replicate and archive mRNA for gene expression analysis.
We have developed a procedure to amplify mRNA into sense RNA (sRNA) so as to create a regenerating biorepository representing the complex mRNA profile in the original sample. The procedure exploits the template-switching activity of reverse transcriptase to incorporate RNA polymerase binding sites upstream of single-stranded cDNA (ss cDNA). Limited PCR was used for double-stranded DNA (dsDNA) synthesis. sRNA was synthesized from PCR products by in vitro transcription (IVT). sRNA was evaluated by real-time reverse transcription (RT)-PCR. sRNA synthesis was successful with RNA from human cell lines and tissues, yielding 2000- to 2500-fold amplification of glyceraldeyde-3 phosphate dehydrogenase (G3PDH). The size of sRNA ranged from 3.0 to 0.1 kb. sRNA synthesis preserved the relative differences in plant mRNAs spiked at abundance ranging over 5 orders of magnitude (0.00001-0.1%). This reflects the high fidelity of sRNA synthesis for mRNA as low as 0.3 copies/cell. sRNA is amplified synthetic mRNA in the 5'-->3' direction; the appropriate template for any gene expression analysis.
Cell Line↗
Proteomic analysis of cancer-cell mitochondria.
Mitochondrial dysfunction and mutations in mitochondrial DNA have been frequently reported in cancer cells. Mitochondrial gene-expression signatures of transformed cells have been identified; however, the phenotypic effects of these genetic alterations remain to be established. Identification of mitochondrial proteins that are aberrantly expressed in cancer cells has been made possible by the recent development of mitochondrial functional proteomics and could identify new markers for early detection and risk assessment, as well as targets for therapeutic intervention.
Humans↗
Epigenetics in cancer prevention: early detection and risk assessment: introduction.
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Animals↗
Viral genes and methylation.
Epigenetics represents a new frontier in cancer research. Methylation is the best studied of the epigenetic mechanisms that regulate gene expression. Regulation of gene expression by means of methylation has been reported for tumor suppressor genes, oncogenes, viral promoters, and age-related genes. In this review, the regulation of viral gene expression by methylation is discussed, with particular emphasis on: (1) the virus-specific factors that bind to promoter regions; (2) the implications of this knowledge for designing viral vectors that can be used to deliver genes for the purpose of gene therapy; and (3) the use of this knowledge for the early detection and prevention of cancer. Since methylation can be reversed by a variety of exogenous agents, great potential exists to develop interventions that target cancer-associated aberrant methylation in an effort to reverse or prevent carcinogenesis.
Animals↗
Early detection and risk assessment: proceedings and recommendations from the Workshop on Epigenetics in Cancer Prevention.
Recent advances in molecular biology that have provided a greater understanding of multistage carcinogenesis include the use of biomarkers of early detection and risk assessment. Prominent among such biomarkers are epigenetic changes. The field of epigenetics has seen a recent surge of interest among cancer researchers since alterations in DNA methylation have emerged as one of the most consistent molecular alterations in multiple neoplasms. Chromatin condensation, histone deacetylation, and promoter methylation are major steps in the epigenetic regulation of gene expression. Epigenetic changes may occur due to environmental factors, aging, and genomic imprinting. An important distinction between genetic and epigenetic alterations in cancer prevention is that the latter might be more easily reversed using therapeutic interventions. In the workshop the following areas of research were recognized for emphasis in future work: (1) basic epigenetic mechanisms in cancer need further investigation; (2) technology development in the area of epigenetics, such as high-throughput quantitative assays and increased sensitivity/specificity, is essential for the early detection and risk assessment of cancer; (3) the clinical application of epigenetic changes to cancer prevention and risk assessment needs further investigation. Further research will lead to the identification of new targets for cancer prevention.
Aging↗