Glucose-induced gradual phenotypic modulation of cultured human glomerular epithelial cells may be independent of Wilms’ tumor 1 (WT1)

Background

Melatonin, a hormone-like substance involved in the regulation of the circadian rhythm, has been demonstrated to protect cells against oxidative DNA damage and to inhibit tumorigenesis.

Results

In the current study, we investigated the effect of melatonin on DNA strand breaks using the alkaline DNA comet assay in breast cancer (MCF-7) and colon cancer (HCT-15) cell lines. Our results demonstrated that cells pretreated with melatonin had significantly shorter Olive tail moments compared to non-melatonin treated cells upon mutagen (methyl methanesulfonate, MMS) exposure, indicating an increased DNA repair capacity after melatonin treatment. We further examined the genome-wide gene expression in melatonin pretreated MCF-7 cells upon carcinogen exposure and detected altered expression of many genes involved in multiple DNA damage responsive pathways. Genes exhibiting altered expression were further analyzed for functional interrelatedness using network- and pathway-based bioinformatics analysis. The top functional network was defined as having relevance for “DNA Replication, Recombination, and Repair, Gene Expression, [and] Cancer”.

Conclusions

These findings suggest that melatonin may enhance DNA repair capacity by affecting several key genes involved in DNA damage responsive pathways.     

Resveratrol retards progression of diabetic nephropathy through modulations of oxidative stress, proinflammatory cytokines, and AMP-activated protein kinase

Background

The Cdc42-interacting protein-4, Trip10 (also known as CIP4), is a multi-domain adaptor protein involved in diverse cellular processes, which functions in a tissue-specific and cell lineage-specific manner. We previously found that Trip10 is highly expressed in estrogen receptor-expressing (ER⁺) breast cancer cells. Estrogen receptor depletion reduced Trip10 expression by progressively increasing DNA methylation. We hypothesized that Trip10 functions as a tumor suppressor and may be involved in the malignancy of ER-negative (ER^-) breast cancer. To test this hypothesis and evaluate whether Trip10 is epigenetically regulated by DNA methylation in other cancers, we evaluated DNA methylation of Trip10 in liver cancer, brain tumor, ovarian cancer, and breast cancer.

Methods

We applied methylation-specific polymerase chain reaction and bisulfite sequencing to determine the DNA methylation of Trip10 in various cancer cell lines and tumor specimens. We also overexpressed Trip10 to observe its effect on colony formation and in vivo tumorigenesis.

Results

We found that Trip10 is hypermethylated in brain tumor and breast cancer, but hypomethylated in liver cancer. Overexpressed Trip10 was associated with endogenous Cdc42 and huntingtin in IMR-32 brain tumor cells and CP70 ovarian cancer cells. However, overexpression of Trip10 promoted colony formation in IMR-32 cells and tumorigenesis in mice inoculated with IMR-32 cells, whereas overexpressed Trip10 substantially suppressed colony formation in CP70 cells and tumorigenesis in mice inoculated with CP70 cells.

Conclusions

Trip10 regulates cancer cell growth and death in a cancer type-specific manner. Differential DNA methylation of Trip10 can either promote cell survival or cell death in a cell type-dependent manner.     

Induction of nephroblastoma by myeloblastosis-associated virus type 1: state of proviral DNAs in tumor cells

Myeloblastosis-associated virus type 1 (MAV1) derived from a molecular clone of infectious proviral DNA (B. Perbal, J. S. Lipsick, J. Svoboda, R. F. Silva, and M. A. Baluda, J. Virol. 56:240-244, 1985) was shown to specifically induce nephroblastoma in chickens and therefore belongs to the MAV-N class. We show that nephroblastomas are polyclonal tumors containing rearranged proviral genomes. Rearrangements occur preferentially in the gag-pol region of the MAV1 proviral genome, and similar rearrangements can be detected in well-developed independent tumors.     

Changes in gene expression during the development of mammary tumors in MMTV-Wnt-1 transgenic mice

Background

In human breast cancer normal mammary cells typically develop into hyperplasia, ductal carcinoma in situ, invasive cancer, and metastasis. The changes in gene expression associated with this stepwise progression are unclear. Mice transgenic for mouse mammary tumor virus (MMTV)-Wnt-1 exhibit discrete steps of mammary tumorigenesis, including hyperplasia, invasive ductal carcinoma, and distant metastasis. These mice might therefore be useful models for discovering changes in gene expression during cancer development.

Results

We used cDNA microarrays to determine the expression profiles of five normal mammary glands, seven hyperplastic mammary glands and 23 mammary tumors from MMTV-Wnt-1 transgenic mice, and 12 mammary tumors from MMTV-Neu transgenic mice. Adipose tissues were used to control for fat cells in the vicinity of the mammary glands. In these analyses, we found that the progression of normal virgin mammary glands to hyperplastic tissues and to mammary tumors is accompanied by differences in the expression of several hundred genes at each step. Some of these differences appear to be unique to the effects of Wnt signaling; others seem to be common to tumors induced by both Neu and Wnt-1 oncogenes.

Conclusion

We described gene-expression patterns associated with breast-cancer development in mice, and identified genes that may be significant targets for oncogenic events. The expression data developed provide a resource for illuminating the molecular mechanisms involved in breast cancer development, especially through the identification of genes that are critical in cancer initiation and progression.     

An inhibitor of K+ channels modulates human endometrial tumor-initiating cells

Background

Many potassium ion (K⁺) channels function as oncogenes to sustain growth of solid tumors, but their role in cancer progression is not well understood. Emerging evidence suggests that the early progenitor cancer cell subpopulation, termed tumor initiating cells (TIC), are critical to cancer progression.

Results

A non-selective antagonist of multiple types of K⁺ channels, tetraethylammonium (TEA), was found to suppress colony formation in endometrial cancer cells via inhibition of putative TIC. The data also indicated that withdrawal of TEA results in a significant enhancement of tumorigenesis. When the TIC-enriched subpopulation was isolated from the endometrial cancer cells, TEA was also found to inhibit growth in vitro.

Conclusions

These studies suggest that the activity of potassium channels significantly contributes to the progression of endometrial tumors, and the antagonists of potassium channels are candidate anti-cancer drugs to specifically target tumor initiating cells in endometrial cancer therapy.     

Multiscale Hy3S: Hybrid stochastic simulation for supercomputers

Background

DNA copy number alterations are one of the main characteristics of the cancer cell karyotype and can contribute to the complex phenotype of these cells. These alterations can lead to gains in cellular oncogenes as well as losses in tumor suppressor genes and can span small intervals as well as involve entire chromosomes. The ability to accurately detect these changes is central to understanding how they impact the biology of the cell.

Results

We describe a novel algorithm called CARAT (Copy Number Analysis with Regression And Tree) that uses probe intensity information to infer copy number in an allele-specific manner from high density DNA oligonuceotide arrays designed to genotype over 100, 000 SNPs. Total and allele-specific copy number estimations using CARAT are independently evaluated for a subset of SNPs using quantitative PCR and allelic TaqMan reactions with several human breast cancer cell lines. The sensitivity and specificity of the algorithm are characterized using DNA samples containing differing numbers of X chromosomes as well as a test set of normal individuals. Results from the algorithm show a high degree of agreement with results from independent verification methods.

Conclusion

Overall, CARAT automatically detects regions with copy number variations and assigns a significance score to each alteration as well as generating allele-specific output. When coupled with SNP genotype calls from the same array, CARAT provides additional detail into the structure of genome wide alterations that can contribute to allelic imbalance.     

Signatures of selection in natural populations adapted to chronic pollution

Background

Ectocarpus siliculosus virus-1 (EsV-1) is a lysogenic dsDNA virus belonging to the super family of nucleocytoplasmic large DNA viruses (NCLDV) that infect Ectocarpus siliculosus, a marine filamentous brown alga. Previous studies indicated that the viral genome is integrated into the host DNA. In order to find the integration sites of the viral genome, a genomic library from EsV-1-infected algae was screened using labelled EsV-1 DNA. Several fragments were isolated and some of them were sequenced and analyzed in detail.

Results

Analysis revealed that the algal genome is split by a copy of viral sequences that have a high identity to EsV-1 DNA sequences. These fragments are interspersed with DNA repeats, pseudogenes and genes coding for products involved in DNA replication, integration and transposition. Some of these gene products are not encoded by EsV-1 but are present in the genome of other members of the NCLDV family. Further analysis suggests that the Ectocarpus algal genome contains traces of the integration of a large dsDNA viral genome; this genome could be the ancestor of the extant NCLDV genomes. Furthermore, several lines of evidence indicate that the EsV-1 genome might have originated in these viral DNA pieces, implying the existence of a complex integration and recombination system. A protein similar to a new class of tyrosine recombinases might be a key enzyme of this system.

Conclusion

Our results support the hypothesis that some dsDNA viruses are monophyletic and evolved principally through genome reduction. Moreover, we hypothesize that phaeoviruses have probably developed an original replication system.     

Preclinical evaluation of the antineoplastic action of 5-aza-2'-deoxycytidine and different histone deacetylase inhibitors on human Ewing's sarcoma cells

Background

Most patients with advanced Ewing's sarcoma (EWS) respond poorly to conventional chemotherapy, indicating the need for new treatment approaches. Epigenetic events, such as promoter hypermethylation and chromatin histone deacetylation, silence the expression of tumor suppressor genes (TSGs) and play an important role in tumorigenesis. These epigenetic changes can be reversed by using 5-aza-2'-deoxycytidine (5AZA-CdR), a potent inhibitor of DNA methylation, in combination with an inhibitor of histone deacetylase (HDAC).

Results

Here, we used a clonogenic assay to evaluate the in vitro antineoplastic activity of 5AZA-CdR in combination with different HDAC inhibitors on EWS cells. We observed that the HDAC inhibitors, MS-275, trichostatin-A, phenylbutyrate, LAQ824 and depsipeptide, enhanced the antineoplastic action of 5AZA-CdR on EWS cells. The combination of 5AZA-CdR and MS-275 showed marked synergy, and was correlated with significant reactivation of the expression of two TSGs, E-cadherin and tumor suppressor lung cancer-1 (TSLC1), in a EWS cell line.

Conclusion

These results suggest the value of future clinical studies investigating the combination of 5AZA-CdR and MS-275 in patients with advanced EWS.     

Gene duplication and fragmentation in the zebra finch major histocompatibility complex

Background

Due to its high polymorphism and importance for disease resistance, the major histocompatibility complex (MHC) has been an important focus of many vertebrate genome projects. Avian MHC organization is of particular interest because the chicken Gallus gallus, the avian species with the best characterized MHC, possesses a highly streamlined minimal essential MHC, which is linked to resistance against specific pathogens. It remains unclear the extent to which this organization describes the situation in other birds and whether it represents a derived or ancestral condition. The sequencing of the zebra finch Taeniopygia guttata genome, in combination with targeted bacterial artificial chromosome (BAC) sequencing, has allowed us to characterize an MHC from a highly divergent and diverse avian lineage, the passerines.

Results

The zebra finch MHC exhibits a complex structure and history involving gene duplication and fragmentation. The zebra finch MHC includes multiple Class I and Class II genes, some of which appear to be pseudogenes, and spans a much more extensive genomic region than the chicken MHC, as evidenced by the presence of MHC genes on each of seven BACs spanning 739 kb. Cytogenetic (FISH) evidence and the genome assembly itself place core MHC genes on as many as four chromosomes with TAP and Class I genes mapping to different chromosomes. MHC Class II regions are further characterized by high endogenous retroviral content. Lastly, we find strong evidence of selection acting on sites within passerine MHC Class I and Class II genes.

Conclusion

The zebra finch MHC differs markedly from that of the chicken, the only other bird species with a complete genome sequence. The apparent lack of synteny between TAP and the expressed MHC Class I locus is in fact reminiscent of a pattern seen in some mammalian lineages and may represent convergent evolution. Our analyses of the zebra finch MHC suggest a complex history involving chromosomal fission, gene duplication and translocation in the history of the MHC in birds, and highlight striking differences in MHC structure and organization among avian lineages.     

RS-SNP: a random-set method for genome-wide association studies

Background

Common carp is one of the most important aquaculture teleost fish in the world. Common carp and other closely related Cyprinidae species provide over 30% aquaculture production in the world. However, common carp genomic resources are still relatively underdeveloped. BAC end sequences (BES) are important resources for genome research on BAC-anchored genetic marker development, linkage map and physical map integration, and whole genome sequence assembling and scaffolding.

Result

To develop such valuable resources in common carp (Cyprinus carpio), a total of 40,224 BAC clones were sequenced on both ends, generating 65,720 clean BES with an average read length of 647 bp after sequence processing, representing 42,522,168 bp or 2.5% of common carp genome. The first survey of common carp genome was conducted with various bioinformatics tools. The common carp genome contains over 17.3% of repetitive elements with GC content of 36.8% and 518 transposon ORFs. To identify and develop BAC-anchored microsatellite markers, a total of 13,581 microsatellites were detected from 10,355 BES. The coding region of 7,127 genes were recognized from 9,443 BES on 7,453 BACs, with 1,990 BACs have genes on both ends. To evaluate the similarity to the genome of closely related zebrafish, BES of common carp were aligned against zebrafish genome. A total of 39,335 BES of common carp have conserved homologs on zebrafish genome which demonstrated the high similarity between zebrafish and common carp genomes, indicating the feasibility of comparative mapping between zebrafish and common carp once we have physical map of common carp.

Conclusion

BAC end sequences are great resources for the first genome wide survey of common carp. The repetitive DNA was estimated to be approximate 28% of common carp genome, indicating the higher complexity of the genome. Comparative analysis had mapped around 40,000 BES to zebrafish genome and established over 3,100 microsyntenies, covering over 50% of the zebrafish genome. BES of common carp are tremendous tools for comparative mapping between the two closely related species, zebrafish and common carp, which should facilitate both structural and functional genome analysis in common carp.     

Tissue of origin determines cancer-associated CpG island promoter hypermethylation patterns

Background

Aberrant CpG island promoter DNA hypermethylation is frequently observed in cancer and is believed to contribute to tumor progression by silencing the expression of tumor suppressor genes. Previously, we observed that promoter hypermethylation in breast cancer reflects cell lineage rather than tumor progression and occurs at genes that are already repressed in a lineage-specific manner. To investigate the generality of our observation we analyzed the methylation profiles of 1,154 cancers from 7 different tissue types.

Results

We find that 1,009 genes are prone to hypermethylation in these 7 types of cancer. Nearly half of these genes varied in their susceptibility to hypermethylation between different cancer types. We show that the expression status of hypermethylation prone genes in the originator tissue determines their propensity to become hypermethylated in cancer; specifically, genes that are normally repressed in a tissue are prone to hypermethylation in cancers derived from that tissue. We also show that the promoter regions of hypermethylation-prone genes are depleted of repetitive elements and that DNA sequence around the same promoters is evolutionarily conserved. We propose that these two characteristics reflect tissue-specific gene promoter architecture regulating the expression of these hypermethylation prone genes in normal tissues.

Conclusions

As aberrantly hypermethylated genes are already repressed in pre-cancerous tissue, we suggest that their hypermethylation does not directly contribute to cancer development via silencing. Instead aberrant hypermethylation reflects developmental history and the perturbation of epigenetic mechanisms maintaining these repressed promoters in a hypomethylated state in normal cells.     

Transfection of Chicken Embryo Cells with DNA Extracted from Avian Virus-Producing Neoplastic Cells

DNA isolated from avian virus-producing leukemic myeloblasts induced the production of viruses, but not morphological transformation, in cultivated chicken fibroblasts. The recovered virus had the same biological characteristics as the original avian myeloblastosis virus (AMV) and produced myeloblastosis and nephroblastomas when injected into chickens. Neutralization experiments with chicken anti-AMV-BAI strain A sera showed an antigenic community between the DNA-transfected virus and the original virus. Virus induced in fibroblasts after treatment with DNA from a viral nephroblastic nephroblastoma line only gave nephroblastoma when injected into chicken. Treatment of chicken embryo cells with DNA extracted from normal chicken embryos did not induce viral production.     

An immune-active tumor microenvironment favors clinical response to ipilimumab

Purpose

Ipilimumab, a fully human monoclonal antibody specific to CTLA-4, has been shown to improve overall survival in metastatic melanoma patients. As a consequence of CTLA-4 blockade, ipilimumab treatment is associated with proliferation and activation of peripheral T cells. To better understand various tumor-associated components that may influence the clinical outcome of ipilimumab treatment, gene expression profiles of tumors from patients treated with ipilimumab were characterized.

Experimental design

Gene expression profiling was performed on tumor biopsies collected from 45 melanoma patients before and 3?weeks after the start of treatment in a phase II clinical trial.

Results

Analysis of pre-treatment tumors indicated that patients with high baseline expression levels of immune-related genes were more likely to respond favorably to ipilimumab. Furthermore, ipilimumab appeared to induce two major changes in tumors from patients who exhibited clinical activity: genes involved in immune response showed increased expression, whereas expression of genes for melanoma-specific antigens and genes involved in cell proliferation decreased. These changes were associated with the total lymphocyte infiltrate in tumors, and there was a suggestion of association with prolonged overall survival in these patients. Many IFN-γ-inducible genes and Th1-associated markers showed increased expression after ipilimumab treatment, suggesting an accumulation of this particular type of T cell at the tumor sites, which might play an important role in mediating the antitumor activity of ipilimumab.

Conclusions

These results support the proposed mechanism of action of ipilimumab, suggesting that cell-mediated immune responses play an important role in the antitumor activity of ipilimumab.     

Oncogenes and Tumor Suppressor Genes

Breast cancer progression involves multiple genetic events, which can activate dominant-acting oncogenes and disrupt the function of specific tumor suppressor genes. This article describes several key oncogene and tumor suppressor signaling networks that have been implicated in breast cancer progression. Among the tumor suppressors, the article emphasizes BRCA1/2 and p53 tumor suppressors. In addition to these well characterized tumor suppressors, the article highlights the importance of PTEN tumor suppressor in counteracting PI3K signaling from activated oncogenes such as ErbB2. This article discusses the use of mouse models of human breast that recapitulate the key genetic events involved in the initiation and progression of breast cancer. Finally, the therapeutic potential of targeting these key tumor suppressor and oncogene signaling networks is discussed.Karyotypic and epidemiological analyses of mammary tumors at various stages suggest that breast carcinomas become increasingly aggressive through the stepwise accumulation of genetic changes. The majority of genetic changes found in human breast cancer fall into two categories: gain-of-function mutations in proto-oncogenes, which stimulate cell growth, division, and survival; and loss-of-function mutations in tumor suppressor genes that normally help prevent unrestrained cellular growth and promote DNA repair and cell cycle checkpoint activation. Epigenetic deregulation also contributes to the abnormal expression of these genes. For example, genes that encode enzymes involved in histone modification are mutated in primary renal cell carcinoma (Dalgliesh et al. 2010; van Haaften et al. 2009). In addition, the involvement of noncoding RNAs in tumorigenesis and tumor metastasis has been recently documented (Croce 2009; Shimono et al. 2009). These can act as oncogenes or tumor suppressor genes, depending on the context. Here, we discuss genes that are frequently altered in breast cancer, focusing on ErbB2, PI3K (phosphatidylinositol 3 kinase) pathways, TP53, BRCA1/2, and PTEN (phosphatase and tensin homolog deleted on chromosome 10). Genetically engineered mouse models are emphasized because these provide a wealth of biological information. We consider in detail genetic and biochemical studies that have shown that oncogenic proteins and tumor suppressors provide a critical balance in regulation of key pathways that control cell number and cell behavior.     

Predictive genes in adjacent normal tissue are preferentially altered by sCNV during tumorigenesis in liver cancer and may rate limiting

Background

In hepatocellular carcinoma (HCC) genes predictive of survival have been found in both adjacent normal (AN) and tumor (TU) tissues. The relationships between these two sets of predictive genes and the general process of tumorigenesis and disease progression remains unclear.

Methodology/Principal Findings

Here we have investigated HCC tumorigenesis by comparing gene expression, DNA copy number variation and survival using ∼250 AN and TU samples representing, respectively, the pre-cancer state, and the result of tumorigenesis. Genes that participate in tumorigenesis were defined using a gene-gene correlation meta-analysis procedure that compared AN versus TU tissues. Genes predictive of survival in AN (AN-survival genes) were found to be enriched in the differential gene-gene correlation gene set indicating that they directly participate in the process of tumorigenesis. Additionally the AN-survival genes were mostly not predictive after tumorigenesis in TU tissue and this transition was associated with and could largely be explained by the effect of somatic DNA copy number variation (sCNV) in cis and in trans. The data was consistent with the variance of AN-survival genes being rate-limiting steps in tumorigenesis and this was confirmed using a treatment that promotes HCC tumorigenesis that selectively altered AN-survival genes and genes differentially correlated between AN and TU.

Conclusions/Significance

This suggests that the process of tumor evolution involves rate-limiting steps related to the background from which the tumor evolved where these were frequently predictive of clinical outcome. Additionally treatments that alter the likelihood of tumorigenesis occurring may act by altering AN-survival genes, suggesting that the process can be manipulated. Further sCNV explains a substantial fraction of tumor specific expression and may therefore be a causal driver of tumor evolution in HCC and perhaps many solid tumor types.     

Anti-tumor effects of a human VEGFR-2-based DNA vaccine in mouse models

Background

Vascular endothelial growth factor (VEGF) and its receptor, VEGFR-2 (Flk-1/KDR), play a key role in tumor angiogenesis. Blocking the VEGF-VEGFR-2 pathway may inhibit tumor growth. Here, we used human VEGFR-2 as a model antigen to explore the feasibility of immunotherapy with a plasmid DNA vaccine based on a xenogeneic homologue of this receptor.

Methods

The protective effects and therapeutic anti-tumor immunity mediated by the DNA vaccine were investigated in mouse models. Anti-angiogenesis effects were detected by immunohistochemical staining and the alginate-encapsulate tumor cell assay. The mechanism of action of the DNA vaccine was primarily explored by detection of auto-antibodies and CTL activity.

Results

The DNA vaccine elicited a strong, protective and therapeutic anti-tumor immunity through an anti-angiogenesis mechanism in mouse models, mediated by the stimulation of an antigen-specific response against mFlk-1.

Conclusion

Our study shows that a DNA vaccine based on a xenogeneic homologue plasmid DNA induced autoimmunity against VEGFR-2, resulting in inhibition of tumor growth. Such vaccines may be clinically relevant for cancer immunotherapy.     

Silencing Agrobacterium oncogenes in transgenic grapevine results in strain-specific crown gall resistance

Key message

Grapevine rootstock transformed with an Agrobacterium oncogene-silencing transgene was resistant to certain Agrobacterium strains but sensitive to others. Thus, genetic diversity of Agrobacterium oncogenes may limit engineering crown gall resistance.

Abstract

Crown gall disease of grapevine induced by Agrobacterium vitis or Agrobacterium tumefaciens causes serious economic losses in viticulture. To establish crown gall-resistant lines, somatic proembryos of Vitis berlandieri × V. rupestris cv. ‘Richter 110’ rootstock were transformed with an oncogene-silencing transgene based on iaaM and ipt oncogene sequences from octopine-type, tumor-inducing (Ti) plasmid pTiA6. Twenty-one transgenic lines were selected, and their transgenic nature was confirmed by polymerase chain reaction (PCR). These lines were inoculated with two A. tumefaciens and three A. vitis strains. Eight lines showed resistance to octopine-type A. tumefaciens A348. Resistance correlated with the expression of the silencing genes. However, oncogene silencing was mostly sequence specific because these lines did not abolish tumorigenesis by A. vitis strains or nopaline-type A. tumefaciens C58.