Chromosomal abnormalities in patients with familial polyposis and colorectal cancer

A search for chromosomal (cytogenetic, karyotypic) changes with specificity for familial polyposis coli (FPC) and related conditions has not been successful. The cytogenetic anomalies described to date (chromosomal instability, tetraploidy, structural anomalies) are not only nonspecific and inconsistent but may also be seen in other congenital conditions predisposing to cancer of various organs. However, this does not mean that some (eg, tetraploidy) of the changes cannot be used as indices in the study and evaluation of affected individuals and their family members. Future application of DNA probes may be more successful in establishing genetic anomalies unique to FPC and related disorders.     

Effect of N-acetyl cysteine on oxidative DNA damage and the frequency of DNA deletions in atm-deficient mice

Ataxia telangiectasia (AT) is a hereditary human disorder resulting in a wide variety of clinical manifestations, including progressive neurodegeneration, immunodeficiency, and high incidence of lymphoid tumors. Cells from patients with AT show genetic instability, hypersensitivity to radiation, and a continuous state of oxidative stress. Oxidative stress and genetic instability, including DNA deletions, are involved in carcinogenesis. We examined the effect of dietary supplementation with the thiol-containing antioxidant N-acetyl-l-cysteine (NAC) on levels of oxidative DNA damage and the frequency of DNA deletions in Atm-deficient (AT-mutated) mice. We confirmed that Atm-deficient mice display an increased frequency of DNA deletions (Bishop et al., Cancer Res 2000;60:395). Furthermore, we found that Atm-deficient mice have significantly increased levels of 8-OH deoxyguanosine, an indication of oxidative DNA damage. Dietary supplementation with NAC significantly reduced 8-OH deoxyguanosine level and the frequency of DNA deletions in Atm-deficient mice. These levels were similar to the levels in wild-type mice. Our findings demonstrate that NAC counteracts genetic instability and suggest that genetic instability may be a consequence of oxidative stress in Atm-deficient mice.     

Mouse models for induced genetic instability at endogenous loci

Exposure to environmental factors and genetic predisposition of an individual may lead individually or in combination to various genetic diseases including cancer. These diseases may be a consequence of genetic instability resulting in large-scale genomic rearrangements, such as DNA deletions, duplications, and translocations. This review focuses on mouse assays detecting genetic instability at endogenous loci. The frequency of DNA deletions by homologous recombination at the pink-eyed unstable (p(un)) locus is elevated in mice with mutations in ATM, Trp53, Gadd45, and WRN genes and after exposure to carcinogens. Other quantitative in vivo assays detecting loss of heterozygosity events, such as the mammalian spot assay, Dlb-1 mouse and Aprt mouse assays, are also reviewed. These in vivo test systems may predict hazardous effects of an environmental agent and/or genetic predisposition to cancer.     

Myelodysplastic syndromes: the complexity of stem-cell diseases

The prevalence of patients with myelodysplastic syndromes (MDS) is increasing owing to an ageing population and increased awareness of these diseases. MDS represent many different conditions, not just a single disease, that are grouped together by several clinical characteristics. A striking feature of MDS is genetic instability, and a large proportion of cases result in acute myeloid leukaemia (AML). We Review three emerging principles of MDS biology: stem-cell dysfunction and the overlap with AML, genetic instability and the deregulation of apoptosis, in the context of inherited bone marrow-failure syndromes, and treatment-related MDS and AML.     

Genetic alterations in locally advanced stage II/III colon cancer: a search for prognostic markers

Heterogeneity in advanced colon cancer leads to different results from adjuvant chemotherapy. To identify groups of patients who may need adjuvant treatment, molecular staging and correlation with clinical data may be helpful in classifying histologically similar tumors. Colon cancer develops through a multistep process with an accumulation of multiple genetic alterations that are often the cause of a form of genomic instability. The 2 best known mechanisms of genomic instability are chromosomal instability (CIN) and microsatellite instability (MSI). The CIN phenotype is found in approximately 85% of sporadic colon cancers and is characterized by aneuploidy, multiple chromosomal rearrangements, and an accumulation of somatic mutations in oncogenes such as K-ras and tumor suppressor genes such as TP53 and APC. The MSI phenotype is associated with small insertions and deletions mainly in repetitive sequences (microsatellites) and is found in approximately 15% of cases. This instability, often referred to as high-frequency MSI (MSI-H), is caused by defects of the mismatch repair system, which is involved in repairing DNA errors that arise during DNA replication. Clear-cut correlations between the somatic genetic alterations in tumors and the clinical behavior of the tumor are rare. Only a few markers, such as MSI-H and TP53, seem to have a prognostic value. Mutations in the TP53 gene are associated with an aggressive tumor growth and subsequent reduced survival, whereas MSI-H seems to be correlated with a favorable outcome. In general, predicting biologic behavior of in particular stage III colon cancers is difficult and remains a great clinical problem.     

Multiple myeloma: Implementing signaling pathways and molecular biology in clinical trials

Multiple Myeloma is a molecularly heterogeneous disease with a high degree of genomic instability in which specific genetic changes can be linked to clinical presentation and prognosis. Despite recent improvements in event-free survival and overall survival with the use of high dose chemotherapy and stem cell support as well as the development of novel agents such as thalidomide, lenalidomide and Bortezomib, MM remains an incurable disease. The development of effective targeted therapies requires a detailed knowledge of various genetic and signaling pathways governing MM genesis. This review will focus on the current understanding of the molecular pathogenesis of MM and the intracellular signaling pathways and their regulations, with emphasis on the rationale for identifying therapeutic targets that can be applied in the clinic.     

A gene expression signature of genetic instability in colon cancer

Genetic instability plays a central role in the development and progression of human cancer. Two major classes of genetic instability, microsatellite instability (MSI) and chromosome instability (microsatellite stable; MSS), are best understood in the context of colon cancer, where MSI tumors represent approximately 15% of cases, and compared with MSS tumors, more often arise in the proximal colon and display favorable clinical outcome. To further explore molecular differences, we profiled gene expression in a set of 18 colon cancer cell lines using cDNA microarrays representing approximately 21,000 different genes. Supervised analysis identified a robust expression signature distinguishing MSI and MSS samples. As few as eight genes predicted with high accuracy the underlying genetic instability in the original and in three independent sample sets, comprising 13 colon cancer cell lines, 61 colorectal tumors, and 87 gastric tumors. Notably, the MSI signature was retained despite genetically correcting the underlying instability, suggesting the signature reflects a legacy of the tumor having arisen from MSI, rather than sensing the ongoing state of MSI. Our findings support a model in which MSI and MSS preferentially target different genes and pathways in cancer. Further, among the MSI signature genes, our findings implicate a role of elevated metallothionein expression in the clinical behavior of MSI cancers.     

Increased cancer risk as a genetic effect of ionizing radiation

The well known genetic effects of ionizing radiation include severe developmental disorders in the progeny of irradiated parents resulting in embryonic death, stillbirth and early postnatal mortality, congenital abnormalities, malformations and fertility disturbances in live-born organisms. These effects are considered to be due to gross mutations (genomic, chromosomal and those of essential genes). Physiological inferiority and an increased cancer risk in phenotypically normal offspring of irradiated parents appear to be two further types of genetic effect of radiation. The genetic background of these effects is suggested to be induced recessive polygene mutations and regulatory DNA alterations, which may lead to instability of the hereditary apparatus of cells, activation of protooncogenes and other inducible processes. A comparison of somatic and genetic effects of radiation shows certain similarities, not only in phenomenology, but probably also in pathogenetic mechanisms.     

Microsatellite analysis at 10q25-q26 in Sardinian patients with sporadic endometrial carcinoma: identification of specification patterns of genetic alteration

BACKGROUND: Loss of heterozygosity (LOH) at chromosome 10q25-q26 has been reported previously in endometrial carcinoma (EC), suggesting the presence of tumor suppressor gene(s). Nevertheless, frequency of genome-wide microsatellite instability (MSI) has been demonstrated higher in EC than in other common malignancy, mostly due to defective DNA mismatch repair. The authors further evaluated the role of the chromosome 10q25-q26 in endometrial tumorigenesis as well as the clinical significance of any observed genetic alteration in sporadic EC. METHODS: Paired normal and tumor samples from 94 Sardinian patients with sporadic EC at various stages of disease were screened by polymerase chain reaction (PCR)-based microsatellite analysis. Genomic DNA was isolated from paraffin embedded tissues and amplified by PCR using microsatellite markers spanning approximately 14 cM at 10q25-q26. Microsatellite instability was studied at four loci mapping to different chromosomal locations. RESULTS: Thirty-two (34%) EC patients were found negative for genetic alterations within the 10q25-q26 region. Among the remaining 62 (66%) EC cases, the authors identified 1) a minimum consensus region of LOH of approximately 1 cM, between D10S610 and D10S542 markers; and 2) a subset of tumors with prevalence of instability at 10q25-q26 (10qMI+), as expression of the presence of a MSI+ phenotype. CONCLUSIONS: The authors' data establish the existence of significant correlations between disease stages and 10qMI+ (with or without MSI+). However, longer follow-up and additional studies are required to define the clinical significance of these findings as prognostic factors. Moreover, the minimum region of LOH at 10q25-q26 will be further analyzed for identifying the putative tumor suppressor gene involved in EC pathogenesis.     

Chromosome Instability in Malignant Astrocytic Tumors Detected by Fluorescence in situ Hybridization

Malignant tumors intrinsically manifest genetic instability, and consequently genetic aberrations successively accumulate in tumor cells as the tumor progresses. However, the relationship of genetic instability and biological behavior still remains to be investigated in malignant tumors. In the present investigation, the relationship between chromosomal instability and patient prognosis was studied in 19 malignant gliomas. Chromosomal instability was estimated by numerical variation in chromosomes 7, 10 and 17 which was measured by fluorescence in situ hybridization (FISH), and DNA ploidy was determined by laser scanning cytometry. The mean number of fractions was significantly higher in cases with DNA aneuploidy than in those with DNA diploidy. The tendency toward higher fractions in glioblastomas existed, although it did not reach statistical significance. Kaplan–Meier survival rate analysis demonstrated significantly lower survival rates in patients with higher fractions of chromosome 7 (5) than others. Our results suggest that DNA aneuploidy in malignant gliomas reflects an underlying chromosomal instability, and that this instability is associated with clinical behavior.     

Somatic evolution of cancer cells

Cancers develop through a process called genomic instability, which generates diversity, from which clonal evolution may occur. In colorectal cancers, this process has been extensively studied, and there are three identifiable processes involved in generating diversity at the genetic or epigenetic level. Colorectal cancers may have chromosomal instability (CIN), microsatellite instability (MSI), or the CpG island methylator phenotype (CIMP). Each of these processes is associated with a unique mutational or epigenetic "signature" identifiable in the tumor cells, and there are important conceptual and clinical implications of each.     

Mismatch repair expression in testicular cancer predicts recurrence and survival

We investigated mismatch repair (MMR) gene expression in testicular cancer as a molecular marker for clinical outcome (recurrence, response to chemotherapy and death) using protein expression and specific genetic alterations associated with the presence or absence of MMR activity. One hundred sixty-two cases of paraffin-embedded testis cancer specimens were subjected to immunohistochemical analysis using monoclonal antibody for MLH1 and MSH2 MMR proteins and genetic analysis using specific polymorphic markers. The degree of MMR immunoreactivity and genetic instability in the form of loss of heterozygosity (LOH) and/or microsatellite instability (MSI) were determined by comparing matched normal and tumor tissue. The degree of immunohistochemical staining for MMR expression was associated with a shorter time to tumor recurrence, resistance to chemotherapy and death. Furthermore, clinical relapse and cancer specific death was also associated with tumors exhibiting a high degree of MSI, p = 0.01 and 0.04, respectively. In contrast, LOH was not associated with recurrence, resistance to chemotherapy or death. Therefore, MMR expression defines testis cancers with distinct molecular properties and clinical behavior, such that tumors with decreased MMR immunostaining and/or increased frequency of MSI have a shorter time to recurrence and death despite chemotherapy.     

Epigenetic inactivation of IkappaB Kinase-alpha in oral carcinomas and tumor progression.

PURPOSE: The loss of epithelial phenotypes in the process of carcinoma progression correlates with clinical outcome, and genetic/epigenetic changes accumulate aggressive clones toward uncurable disease. IkappaB kinase-alpha (IKKalpha) has a decisive role in the development of the skin and establishes keratinocyte phenotypes. We assessed clinical implications of IKKalpha expression in oral carcinomas and epigenetic aberrations for the loss of expression. EXPERIMENTAL DESIGN: We examined IKKalpha expression in oral carcinomas by immunostaining (n = 64) and genetic instability by microsatellite PCR (n = 46). Promoter methylation status was analyzed by bisulfite-modified sequence (n = 11). RESULTS: IKKalpha was expressed in the nucleus of basal cells of normal oral epithelium, but not or marginally detected in 32.8% of carcinomas. The immunoreactivity was significantly decreased in less differentiated carcinomas (P < 0.05) and correlated to long-term survival of patients (P < 0.01) with an independent prognostic value (P < 0.05). Although allelic/biallelic loss of the gene was limited to four cases, we detected microsatellite instability in 63.0% cases in which the immunoreactivities were decreased and the promoter was hypermethylated. CONCLUSION: These results showed that oral carcinomas exhibiting genetic instability and promoter hypermethylation down-regulate expression of IKK and suggest that the epigenetic loss of the expression closely associates with disease progression toward unfavorable prognosis.     

Current approaches in hereditary nonpolyposis colorectal cancer

This article emphasizes the central role of tumor-based testing for microsatellite instability followed by performance of genetic counselor-driven germline mutation testing in hereditary nonpolyposis colorectal cancer (HNPCC). Suitably aggressive colorectal neoplasm surveillance is shown to be critical. Limitations of the evidentiary base for extracolonic screening are conceded, with some cautious suggestions for possible strategies notwithstanding the lack of data. Advances in chemoprevention have been made in both familial adenomatous polyposis (clinical trial data favoring eicosapentaenoic acid) and HNPCC (controversial aspirin data). For various reasons, however, no agent or combination of agents has yet come into routine use in either condition, with further trials underway or being designed for both conditions.     

Pituitary tumour transforming gene (PTTG) induces genetic instability in thyroid cells

Cancer reflects the progressive accumulation of genetic alterations and subsequent genetic instability of cells. Cytogenetic studies have demonstrated the importance of aneuploidy in differentiated thyroid cancer development. The pituitary tumour transforming gene (PTTG), also known as securin, is a mitotic checkpoint protein which inhibits sister chromatid separation during mitosis. PTTG is highly expressed in many cancers and overexpression of PTTG induces aneuploidy in vitro. Using fluorescent intersimple sequence repeat PCR (FISSR-PCR), we investigated the relationship between PTTG expression and the degree of genetic instability in normal and tumorous thyroid samples. The genomic instability index (GI index) was 6.7-72.7% higher in cancers than normal thyroid tissues. Follicular thyroid tumours exhibited greater genetic instability than papillary tumours (27.6% (n=9) versus 14.5% (n=10), P=0.03). We also demonstrated a strong relationship between PTTG expression and the degree of genetic instability in thyroid cancers (R2=0.80, P=0.007). To further investigate PTTG's role in genetic instability, we transfected FTC133 thyroid follicular cells and observed increased genetic instability in cells overexpressing PTTG compared with vector-only-transfected controls (n=3, GI Index VO=29.7+/-5.2 versus PTTG=63.7+/-6.4, P=0.013). Further, we observed a dose response in genetic instability and PTTG expression (GI Index low dose (0.5 microg DNA/ six-well plate) PTTG=15.3%+/-1.7 versus high dose (3 microg DNA) PTTG=50.8%+/-3.3, P=0.006). Overall, we describe the first use of FISSR-PCR in human cancers, and demonstrate that PTTG expression correlates with genetic instability in vivo, and induces genetic instability in vitro. We conclude that PTTG may be an important gene in the mutator phenotype development in thyroid cancer.     

Evolving molecular classification by genomic and proteomic biomarkers in colorectal cancer: potential implications for the surgical oncologist

Colorectal cancer (CRC) is one of the most frequent cancers in the Western world and represents a major health burden. CRC development is a multi-step process that spans 10-15years, thereby providing an opportunity for early detection and even prevention. As almost half of all patients undergoing surgery develop recurrent disease, surveillance is advocated, albeit with various means and intervals. Current screening and surveillance efforts have so far only had limited impact due to suboptimal compliance. Currently, CEA is the only biomarker in clinical use for CRC, but has suboptimal sensitivity and specificity. New and better biomarkers are therefore strongly needed. Non-invasive biomarkers may develop through the understanding of colorectal carcinogenesis. Three main pathways occur in CRC, including chromosomal instability (CIN), microsatellite instability (MSI) and epigenetic silencing through the CpG Island Methylator Phenotype (CIMP). These pathways have distinct clinical, pathological, and genetic characteristics, which can be used for molecular classification and comprehensive tumour profiling for improved diagnostics, prognosis and treatment in CRC. Molecular-biological research has advanced with the sequencing of the human genome and the availability of genomic and proteomic high-throughput technologies using different chip platforms, such as tissue microarrays, DNA microarrays, and mass spectrometry. This review aims to give an overview of the evolving biomarker concepts in CRC, with concerns on methods, and potential for clinical implications for the surgical oncologist.     

Molecular profiling of sporadic colorectal tumors by microsatellite analysis

To investigate the prognostic value of multiple genetic alterations, individual molecular tumor profiles were established in 79 sporadic colorectal carcinomas (41 stage II and 38 stage III). Tumors were analyzed for allelic loss (LOH) and genetic instability (MSI) using 14 microsatellites intragenic to or associated with tumor suppressor or DNA mismatch repair genes. Molecular profiling identified tumors with LOH at multiple loci without microsatellite instability (MSS), tumors with high levels of LOH and low level microsatellite marker instability (MSI-L), and tumors with high levels of MSI (MSI-H), but rare LOH. K-ras mutations occurred more frequently in MSS/MSI-L carcinomas (26%) than in MSI-H colorectal tumors (10%), the latter showing a high frequency of TGFbeta type II frameshift mutations (82%). Correlation of molecular and clinical data revealed a better prognosis for stage III tumor patients displaying 5q12 loss rather than retention of heterozygosity. Thus, molecular profiling allows the identification of new prognostic markers and might facilitate the stratification of colorectal cancer patients.     

A genome-wide study of microsatellite instability in advanced gastric carcinoma.

BACKGROUND: Microsatellite instability (MSI) has been described in many human carcinomas, including gastric carcinomas (GCs). There are inconsistent findings regarding the association of MSI with various subsets of GC with specific clinicopathologic features. The objective of this study was to define MSI in advanced GC at a genome-wide level and to evaluate the clinical relevance of MSI in these patients. METHODS: Forty-one gastric adenocarcinomas with serosa invasion (T3) were analyzed at 59 loci that detected at least one site per arm of each autosome in human genome. The expression patterns of mismatch repair proteins hMLH1 and hMSH2 were examined by immunohistochemistry. Comparisons were made by categorizing tumors into three groups: tumors with MSI at multiple loci (at more than three loci), tumors with MSI at low level (at one to three loci), and microsatellite-stable (MSS) tumors. Clinical significance of MSI in advanced GC was evaluated. The relative rates of hypermutability of the 59 markers also were determined. RESULTS: A significant association was found between tumors with MSI at multiple loci and the expanding type of tumor growth by Ming's histologic classification (P = 0.001), whereas tumors with MSI at low level and MSS tumors are clinicopathologically indistinguishable. The 59 dinucleotide repeat markers displayed varying degrees of susceptibility toward genetic instability. The relative rates of hypermutability of these markers were consistent with a normal distribution pattern in which the frequency of unstable tumors detected at different chromosomal loci varied from 0% to 20%. CONCLUSIONS: The authors' results showed that advanced GC with MSI at multiple loci progress preferentially in an expanding mode, supporting the notion that high MSI tumors and low MSI/MSS tumors evolve through different genetic pathways. Thus, microsatellite testing may have clinical utility as a favorable prognostic marker.     

Chromosome instability and progression in urothelial cell carcinoma of the bladder

Superficial urothelial cell carcinoma (UCC) is a heterogeneous group of neoplasia with an unpredictable clinical course. Numerical alterations of chromosomes 7, 9 and 17 in superficial and invasive UCCs were analysed to evaluate the importance of chromosome instability in the progression of these tumours. Our sample consisted of 75 patients (47 with superficial and 28 with invasive bladder tumours). In situ hybridization using centromeric probes for chromosomes 7, 9 and 17 was done for the chromosome analysis in paraffin-embedded tissues. From the results obtained it can be concluded that losses of genetic material seem to be important early events in the carcinogenesis of the urothelium, but during progression of UCCs there seems to be a selection of those cells with gains of genetic material. This chromosome instability may be due to the acquisition of mechanisms involved in aneuploidization, namely p53 function disorders.     

Identification of microsatellite length polymorphism by a polymerase chain reaction-based assay at IL-1beta gene locus in cancer cells

It has long been considered that genetic instability is an integral component of human neoplasia, however the frequency of microsatellite instability in human melanoma and non-melanoma skin cancer was less evidenced. We report here an alteration at the complex dinucleotide repeat (TC)n sequence in the interleukin 1-beta gene locus by polymerase chain reaction (PCR)-based assay in mouse melanoma as well as in various cancer cells, which may provide a simple, rapid, non-radioactive screening assay for cancer detection.