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.     

Convergence of mutation and epigenetic alterations identifies common genes in cancer that predict for poor prognosis

Background

The identification and characterization of tumor suppressor genes has enhanced our understanding of the biology of cancer and enabled the development of new diagnostic and therapeutic modalities. Whereas in past decades, a handful of tumor suppressors have been slowly identified using techniques such as linkage analysis, large-scale sequencing of the cancer genome has enabled the rapid identification of a large number of genes that are mutated in cancer. However, determining which of these many genes play key roles in cancer development has proven challenging. Specifically, recent sequencing of human breast and colon cancers has revealed a large number of somatic gene mutations, but virtually all are heterozygous, occur at low frequency, and are tumor-type specific. We hypothesize that key tumor suppressor genes in cancer may be subject to mutation or hypermethylation.

Methods and Findings

Here, we show that combined genetic and epigenetic analysis of these genes reveals many with a higher putative tumor suppressor status than would otherwise be appreciated. At least 36 of the 189 genes newly recognized to be mutated are targets of promoter CpG island hypermethylation, often in both colon and breast cancer cell lines. Analyses of primary tumors show that 18 of these genes are hypermethylated strictly in primary cancers and often with an incidence that is much higher than for the mutations and which is not restricted to a single tumor-type. In the identical breast cancer cell lines in which the mutations were identified, hypermethylation is usually, but not always, mutually exclusive from genetic changes for a given tumor, and there is a high incidence of concomitant loss of expression. Sixteen out of 18 (89%) of these genes map to loci deleted in human cancers. Lastly, and most importantly, the reduced expression of a subset of these genes strongly correlates with poor clinical outcome.

Conclusions

Using an unbiased genome-wide approach, our analysis has enabled the discovery of a number of clinically significant genes targeted by multiple modes of inactivation in breast and colon cancer. Importantly, we demonstrate that a subset of these genes predict strongly for poor clinical outcome. Our data define a set of genes that are targeted by both genetic and epigenetic events, predict for clinical prognosis, and are likely fundamentally important for cancer initiation or progression.     

Characterization of a novel human breast cancer associated gene (BCA3) encoding an alternatively spliced proline-rich protein

As part of an integrated study of breast cancer gene expression, partial cDNAs were cloned from normal and tumor breast cells by subtractive-hybridization and differential display cloning. The DNA sequence for one of these breast cancer associated genes was used to construct the larger 1319 bp BCA3 cDNA sequence using ESTs without assigned names or functions. High-level BCA3 mRNA expression was found in breast and prostate tumor cell lines whereas normal breast and prostate tissues have low-level expression. Further analysis revealed possible functional domains and alternative splicing of BCA3 that we confirmed by RT-PCR analysis. Immunohistochemistry revealed that the protein is expressed in breast tumor cells in vivo, and not in surrounding stromal tissue.     

Splicing factor poly(rC)-binding protein 1 is a novel and distinctive tumor suppressor

A lot of evidence has been found on the link between tumorigenesis and the aberrant expression of splicing factors. A number of splicing factors have been reported to be either oncogenic or overexpressed in cancer cells. However, splicing factors can also play negative roles in tumorigenesis. In the current review, we focus on splicing factor poly(rC)-binding protein 1 (PCBP1), a novel tumor suppressor that is characterized by downregulation in many cancer types and shows inhibition of tumor formation and metastasis. Notably, the messenger RNA levels of PCBP1 are not significantly decreased in most cancer types. In fact, PCBP1 protein is often degraded or shows a loss-of-function through phosphorylation in cancer cells. PCBP1 is highly homologous to its family member, PCBP2. Interestingly, PCBP2 appears to be an oncogenic splicing factor. A growing body of evidence has shown that PCBP1 regulates alternative splicing, translation, and RNA stability of many cancer-related genes. Taking together, PCBP1 has distinctive tumor suppressive functions, and increasing PCBP1 expression may represent a new approach for cancer treatment.     

Evolutionary dynamics of BRCA1 alterations in breast tumorigenesis

Cancer results from the accumulation of alterations in oncogenes and tumor suppressor genes. Tumor suppressors are classically defined as genes which contribute to tumorigenesis if their function is lost. Genetic or epigenetic alterations inactivating such genes may arise during somatic cell divisions or alternatively may be inherited from a parent. One notable exception to this rule is the BRCA1 tumor suppressor that predisposes to hereditary breast cancer when lost. Genetic alterations of this gene are hardly ever observed in sporadic breast cancer, while individuals harboring a germline mutation readily accumulate a second alteration inactivating the remaining allele—a finding which represents a conundrum in cancer genetics. In this paper, we present a novel mathematical framework of sporadic and hereditary breast tumorigenesis. We study the dynamics of genetic alterations driving breast tumorigenesis and explore those scenarios which can explain the absence of somatic BRCA1 alterations while replicating all other disease statistics. Our results support the existence of a heterozygous phenotype of BRCA1 and suggest that the loss of one BRCA1 allele may suppress the fitness advantage caused by the inactivation of other tumor suppressor genes. This paper contributes to the mathematical investigation of breast tumorigenesis.     

Integrative genomics revealed RAI3 is a cell growth-promoting gene and a novel P53 transcriptional target

In this study, differential gene expression between normal human mammary epithelial cells and their malignant counterparts (eight well established breast cancer cell lines) was studied using Incyte GeneAlbum 1-6, which contains 65,873 cDNA clones representing 33,515 individual genes. 3,152 cDNAs showed a > or =3.0-fold expression level change in at least one of the human breast cancer cell lines as compared with normal human mammary epithelial cells. Integration of breast tumor gene expression data with the genes in the tumor suppressor p53 signaling pathway yielded 128 genes whose expression is altered in breast tumor cell lines and in response to p53 expression. A hierarchical cluster analysis of the 128 genes revealed that a significant portion of genes demonstrate an opposing expression pattern, i.e. p53-activated genes are down-regulated in the breast tumor lines, whereas p53-repressed genes are up-regulated. Most of these genes are involved in cell cycle regulation and/or apoptosis, consistent with the tumor suppressor function of p53. Follow-up studies on one gene, RAI3, suggested that p53 interacts with the promoter of RAI3 and repressed its expression at the onset of apoptosis. The expression of RAI3 is elevated in most tumor cell lines expressing mutant p53, whereas RAI3 mRNA is relatively repressed in the tumor cell lines expressing wild-type p53. Furthermore, ectopic expression of RAI3 in 293 cells promotes anchorage-independent growth and small interfering RNA-mediated depletion of RAI3 in AsPc-1 pancreatic tumor cells induces cell morphological change. Taken together, these data suggest a role for RAI3 in tumor growth and demonstrate the predictive power of integrative genomics.     

ARLTS1, MDM2 and RAD51 gene variations are associated with familial breast cancer

Genetic factors that contribute to the risk of breast cancer are largely not known and association studies have revealed several genes with low penetrance risk alleles for breast cancer. Analysis of these genes may provide important information on the risk factors affecting carcinogenesis. Variations in the ARLTS1, RAD51 and MDM2 genes have been associated with increased risk of different cancer types but for breast cancer the results are not consistent. In this study we investigated the role of the allelic variants in candidate genes acting in the tumor suppressor, DNA repair and p53 pathways as risk factors for familial breast cancer in 147 patients displaying characteristics of familial disease. Presence of the polymorphic variants were investigated by amplification of the corresponding regions and restriction fragment length polymorphism analysis. Genotype and allele frequencies in the patients were significantly different for all three variants. Our results indicate that the polymorphic variants might affect individual susceptibility towards breast cancer.     

Analysis of the candidate tumor suppressor Ris-1 in primary human breast carcinomas

Frequent chromosome 3 losses have been described in several tumors types, which strongly suggest the presence of one or several tumor suppressor genes. Recently, a novel candidate tumor suppressor gene termed Ris-1 (for Ras-induced senescence 1) has been identified at chromosomal position 3p21.3. Ris-1 has been proposed to participate in anti-tumor responses that resemble cellular senescence and that are elicited by oncogenes such as Ras. To analyze the role of Ris-1 as a putative tumor suppressor gene in human breast cancer, we have performed a real-time quantitative analysis of its mRNA expression in 60 patients. Moreover, we carried out a first approach to evaluate the most common inactivation mechanism that can affect expression levels of tumor suppressor genes (mutation, promoter hypermethylation and allelic losses). Furthermore, a correlation study between expression as well as inactivating mechanisms of Ris-1 and several clinico-pathological parameters of the tumors was designed, with the objective of appraising the prognostic value of Ris-1 status. Decreased expression of Ris-1 was observed in 23% of the cases and overexpressed Ris-1 was detected in 15% of the primary breast tumors. Our data showed high frequency of LOH (30%) at one of the markers used. Nevertheless, a polymorphism related with the expression levels was described. Statistically significant correlations were found between decreased Ris-1 expression and negative progesterone receptors, as well as between overexpressing Ris-1 tumors and high histological grade. Despite all these data, we conclude that the suggested role of Ris-1 as tumor suppressor gene is not evident, at least in breast cancer. Future and larger series studies in different tumor types are necessary to clarify Ris-1 function in human cancer.     

Characterization of breast cancer by array comparative genomic hybridization.

Cancer progression is due to the accumulation of recurrent genomic alterations that induce growth advantage and clonal expansion. Most of these genomic changes can be detected using the array comparative genomic hybridization (CGH) technique. The accurate classification of these genomic alterations is expected to have an important impact on translational and basic research. Here we review recent advances in CGH technology used in the characterization of different features of breast cancer. First, we present bioinformatics methods that have been developed for the analysis of CGH arrays; next, we discuss the use of array CGH technology to classify tumor stages and to identify and stratify subgroups of patients with different prognoses and clinical behaviors. We finish our review with a discussion of how CGH arrays are being used to identify oncogenes, tumor suppressor genes, and breast cancer susceptibility genes.     

Discovery of novel splice forms and functional analysis of cancer-specific alternative splicing in human expressed sequences

We report here a genome-wide analysis of alternative splicing in 2 million human expressed sequence tags (ESTs), to identify splice forms that are up-regulated in tumors relative to normal tissues. We found strong evidence (P < 0.01) of cancer-specific splice variants in 316 human genes. In total, 78% of the cancer-specific splice forms we detected are confirmed by human-curated mRNA sequences, indicating that our results are not due to random mis-splicing in tumors; 73% of the genes showed the same cancer-specific splicing changes in tissue-matched tumor versus normal datasets, indicating that the vast majority of these changes are associated with tumorigenesis, not tissue specificity. We have confirmed our EST results in an independent set of experimental data provided by human-curated mRNAs (P-value 10^–5.7). Moreover, the majority of the genes we detected have functions associated with cancer (P-value 0.0007), suggesting that their altered splicing may play a functional role in cancer. Analysis of the types of cancer-specific splicing shifts suggests that many of these shifts act by disrupting a tumor suppressor function. Sur prisingly, our data show that for a large number (190 in this study) of cancer-associated genes cloned originally from tumors, there exists a previously uncharacterized splice form of the gene that appears to be predominant in normal tissue.     

Loss of heterozygosity studies in tumors from families with breast-ovarian cancer syndrome

A gene (BRCA1) predisposing for familial breast and ovarian cancer has been mapped to chromosome band 17q12-21. Based on the observation that ovarian tumors from families with breast and ovarian cancer lose the wild-type allele in the region for the BRCA1 locus, it has been suggested that the gene functions as a tumor suppressor gene. We have studied chromosomal deletions in the BRCA1 region in seven breast tumors, three ovarian tumors, one bladder cancer, and one colon cancer from patients in six families with breast-ovarian cancer, in order to test the hypothesis of the tumor suppressor mechanism at this locus. We have found a low frequency of loss of heterozygosity at this region, and our results do not support the idea that BRCA1 is a tumor suppressor gene. Alternatively, the disease segregating in these families is linked to one or more different loci.     

Murine tumor suppressor models

Tumor suppressor genes have been shown to be necessary for proper maintenance of cell growth control. Inactivation of these genes in the germline of humans is linked to inherited cancer predisposition. Moreover, sporadically arising human tumors often have somatic mutations in tumor suppressor genes. During the past few years, advances in molecular and cellular biology have led to the creation of animal models that have germline mutations of various tumor suppressor genes. Such mice potentially represent important animal models for familial cancer predisposition syndromes, and the study of the tumorigenesis process has been greatly assisted by their development. Such models have also demonstrated the importance of tumor suppressor function in embryonic development. In this review, we describe mice with inactivated germline tumor suppressor genes that are genetically analogous to 10 different inherited cancer syndromes in humans. We describe the variable usefulness of the mutant mice as models for human disease.     

miR-126 调控前列腺癌机制研究

miR-126通过靶向作用于表皮生长因子域7(EGFL7)、同源框A9(HOXA9)、胰岛素受体底物-1(IlLS-1)、p85-B基因等,在转录后水平调控靶基因表达,在肿瘤形成中起重要作用。前列腺癌细胞中高表达miR-126,能明显下调VEGF-A、EGLF7、HOXA9、VCAM-l等与肿瘤生长、转移密切相关的蛋白分子。miR-126作为抑癌因子,在多种肿瘤中均下调。其抑癌作用及机制在肺癌、白血病、乳腺癌、宫颈癌等中均已得到证实。本课题拟对miR-126调控前列腺癌机制做一综述。