Variation in DNA repair is a factor in cancer susceptibility: a paradigm for the promises and perils of individual and population risk estimation?

The repair of DNA damage protects the genome of the cell from the insults of cancer causing agents. This was originally demonstrated in individuals with the rare genetic disease, xeroderma pigmentosum, the prototype of cancer genes, and subsequently in the relationship of mismatch repair to colon cancer. Recent studies suggests that individuals with less dramatic reductions in the capacity to repair DNA damage are observed at polymorphic frequency and these individuals have an increased susceptibility to several types of cancer. Screening of individuals for DNA sequence variation in the exons of 9 DNA repair genes has resulted in identification of 15 different polymorphic amino acid substitution variants. Although the studies to relate these variants to reduced DNA repair capacity and cancer status have not been completed, the available information is sufficient to suggest that DNA repair genes should be incorporated into molecular epidemiology and cancer susceptibility studies. The availability of molecular epidemiology data presents exciting opportunities for refinement of risk estimation models and identification of individuals at increased risk of disease, with resultant opportunities for effective surveillance and early intervention and treatment. The opportunities to acquire susceptibility data are associated with possible perils for establishment of regulations for permissible exposures to carcinogenic agents and also stigmatization of ‘at risk’ individuals that may result in decreased access to employment opportunities and health care.     

Coding variants in human double-strand break DNA repair genes

DNA repair is essential for the maintenance of genomic integrity. Consequently, altered repair capacity may impact individual health in such areas as aging and susceptibility to certain diseases. Defects in some DNA repair genes, for example, have been shown to increase cancer risk, accelerate aging and impair neurological functions. Now that over 115 genes directly involved in human DNA repair have been characterized at the DNA sequence level, the identification of single nucleotide polymorphisms (SNPs) in DNA repair genes is becoming a reality. This information will likely lead to the identification of alleles, or combinations of alleles that affect disease predisposition. This communication summarizes SNPs identified to date in the coding region of 24 human double-strand break repair (DSBR) genes. SNP data for four of these genes were obtained by screening at least 100 individuals in our laboratory. For each SNP, the codon number, amino acid substitution, allele frequency and population information is supplied.     

DNA repair gene polymorphisms and risk of early onset colorectal cancer in Lynch syndrome

DNA repair plays a pivotal role in maintaining genomic integrity with over 130 genes involved in various repair pathways that include base excision repair, nucleotide excision repair, double strand break repair and DNA mismatch repair. Polymorphisms within genes that are involved in these processes have been widely reported to be associated with cancer susceptibility in an extensive range of malignancies that include colorectal cancer (CRC). Lynch syndrome is caused by inherited germline mutations in DNA mismatch repair genes, predominantly in MLH1 and MSH2, that predispose to a variety of epithelial malignancies, most notably CRC. Despite being a relatively well understood hereditary cancer syndrome there remain several questions in relation to genetic influences on disease expression. Since Lynch syndrome is associated with a breakdown in DNA mismatch repair variation in other DNA repair genes may influence disease expression. In this report we have genotyped 424 Australian and Polish Lynch syndrome participants for eight common DNA repair gene polymorphisms to assess any association with the age of CRC onset. The DNA repair gene SNPs included in the study were: BRCA2 (rs11571653), MSH3 (rs26279), Lig4 (rs1805386), OGG1 (rs1052133), XRCC1 (rs25487), XRCC2 (rs3218536 and rs1799793) and XRCC3 (rs861539). Cox multi-variant regression modelling failed to provide any convincing evidence of an effect in any of the polymorphisms analysed. The data suggest that polymorphisms in DNA repair genes do not contribute to cancer risk in a population of CRC patients who are at increased risk of disease as a result in a deficiency of DNA mismatch repair.     

Complex segregation analysis reveals a multigene model for lung cancer

Lung cancer risk is largely attributed to tobacco exposure, but genetic predisposition also plays an etiologic role. Several studies have investigated the involvement of genetic predisposition in lung cancer aggregation in affected families, although with inconsistent results. Some studies have provided evidence for Mendelian inheritance, whereas others have suggested that environmental models are most appropriate for lung cancer aggregation in families. To examine the genetic basis of lung cancer, we performed segregation analysis on 14,378 individuals from 1,561 lung cancer case families, allowing for the effects of smoking, sex, and age. Both a Mendelian decreasing model and a Mendelian codominant model were found to be the best fitting models for susceptibility. However, when we modeled age-of-onset, all Mendelian models and the environmental model were rejected suggesting that multiple genetic factors (possibly multiple genetic loci and interactions) contribute to the age-of-onset of lung cancer. The results provide evidence that multiple genetic factors contribute to lung cancer and may act as a guide in further studies to localize susceptibility genes in lung cancer.     

Variation in base excision repair capacity

The major DNA repair pathway for coping with spontaneous forms of DNA damage, such as natural hydrolytic products or oxidative lesions, is base excision repair (BER). In particular, BER processes mutagenic and cytotoxic DNA lesions such as non-bulky base modifications, abasic sites, and a range of chemically distinct single-strand breaks. Defects in BER have been linked to cancer predisposition, neurodegenerative disorders, and immunodeficiency. Recent data indicate a large degree of sequence variability in DNA repair genes and several studies have associated BER gene polymorphisms with disease risk, including cancer of several sites. The intent of this review is to describe the range of BER capacity among individuals and the functional consequences of BER genetic variants. We also discuss studies that associate BER deficiency with disease risk and the current state of BER capacity measurement assays.     

Survey of polymorphic sequence variation in the immediate 5' region of human DNA repair genes

Systematic screens have revealed extensive DNA sequence variation existing in the human population. Studies of the role of polymorphic genetic variants in explaining the association of family history with risk of common disease have generally focused on variants predicted to disrupt protein structure and activity. Recent studies have identified genetic variation in the level of expression of many genes, variation that is potentially biologically relevant in explaining individual variation in disease risk. In a survey of data available for 108 DNA repair genes that have been systematically screened for sequence variation, an average of 3.3 SNPs per gene were found to exist at a variant allele frequency of at least 0.02 in the region 2kb upstream from the 5'-untranslated region. One-third of the genes harbored a SNP with an allele frequency of at least 0.02 within a predicted promotor element. These variants are distributed among promoter elements that average 20 elements per gene. The frequency of polymorphic SNPs in CpG islands was 0.8 per gene, while the frequency of SNPs in the 5'-UTR was 0.7 per gene. The recognition of extensive genetic variation with potential to impact levels of gene expression, and thereby exacerbate the impact of amino acid substitution variants on the activity of proteins, increases the complexity of analyses required to explain the molecular genetic basis for the familial contribution to the sporadic incidence of common disease.     

Exome Sequencing Identifies Rare Deleterious Mutations in DNA Repair Genes FANCC and BLM as Potential Breast Cancer Susceptibility Alleles

Despite intensive efforts using linkage and candidate gene approaches, the genetic etiology for the majority of families with a multi-generational breast cancer predisposition is unknown. In this study, we used whole-exome sequencing of thirty-three individuals from 15 breast cancer families to identify potential predisposing genes. Our analysis identified families with heterozygous, deleterious mutations in the DNA repair genes FANCC and BLM, which are responsible for the autosomal recessive disorders Fanconi Anemia and Bloom syndrome. In total, screening of all exons in these genes in 438 breast cancer families identified three with truncating mutations in FANCC and two with truncating mutations in BLM. Additional screening of FANCC mutation hotspot exons identified one pathogenic mutation among an additional 957 breast cancer families. Importantly, none of the deleterious mutations were identified among 464 healthy controls and are not reported in the 1,000 Genomes data. Given the rarity of Fanconi Anemia and Bloom syndrome disorders among Caucasian populations, the finding of multiple deleterious mutations in these critical DNA repair genes among high-risk breast cancer families is intriguing and suggestive of a predisposing role. Our data demonstrate the utility of intra-family exome-sequencing approaches to uncover cancer predisposition genes, but highlight the major challenge of definitively validating candidates where the incidence of sporadic disease is high, germline mutations are not fully penetrant, and individual predisposition genes may only account for a tiny proportion of breast cancer families.     

Polymorphisms in DNA repair and environmental interactions

The repair of damage to DNA is critical to the survival of a cell. However, not all organisms nor all individuals express a similar response to challenges to their genetic material. Numerous polymorphisms in genes involved in DNA repair have been found in individuals with DNA repair-related disease as well as in the general population. Studies of these variants are critical in understanding the response of the cell to DNA damage. In some cases, these changes predispose the carrier to a greatly increased risk of cancer. In other cases, the effects are subtler and depend on interactions between the alleles of several genes, or with environmental factors. Consequently, the health effects of exposure to genotoxic or carcinogenic compounds or agents can depend on the variations in these genes. This review will highlight some of the effects that variants, found in many of the genes involved in human DNA repair pathways, have on the response to damage, and their role in susceptibility of the cell and organism to environmental genotoxins. This review will concentrate on the mismatch repair, nucleotide repair, base excision repair, strand break repair, and direct alkyl repair pathways.     

DNA Repair Deficiency as a Susceptibility Marker for Spontaneous Lymphoma in Golden Retriever Dogs: A Case-Control Study

There is accumulating evidence that an individual’s inability to accurately repair DNA damage in a timely fashion may in part dictate a predisposition to cancer. Dogs spontaneously develop lymphoproliferative diseases such as lymphoma, with the golden retriever (GR) breed being at especially high risk. Mechanisms underlying such breed susceptibility are largely unknown; however, studies of heritable cancer predisposition in dogs may be much more straightforward than similar studies in humans, owing to a high degree of inbreeding and more limited genetic heterogeneity. Here, we conducted a pilot study with 21 GR with lymphoma, 20 age-matched healthy GR and 20 age-matched healthy mixed-breed dogs (MBD) to evaluate DNA repair capability following exposure to either ionizing radiation (IR) or the chemical mutagen bleomycin. Inter-individual variation in DNA repair capacity was evaluated in stimulated canine lymphoctyes exposed in vitro utilizing the G2 chromosomal radiosensitivity assay to quantify clastogen-induced chromatid-type aberrations (gaps and breaks). Golden retrievers with lymphoma demonstrated elevated sensitivity to induction of chromosome damage following either challenge compared to either healthy GR or MBD at multiple doses and time points. Using the 75^th percentile of chromatid breaks per 1,000 chromosomes in the MBD population at 4 hours post 1.0 Gy IR exposure as a benchmark to compare cases and controls, GR with lymphoma were more likely than healthy GR to be classified as “sensitive” (odds ratio = 21.2, 95% confidence interval 2.3-195.8). Furthermore, our preliminary findings imply individual (rather than breed) susceptibility, and suggest that deficiencies in heritable factors related to DNA repair capabilities may be involved in the development of canine lymphoma. These studies set the stage for larger confirmatory studies, as well as candidate-based approaches to probe specific genetic susceptibility factors.     

Exploration of methods to identify polymorphisms associated with variation in DNA repair capacity phenotypes

Elucidating the relationship between polymorphic sequences and risk of common disease is a challenge. For example, although it is clear that variation in DNA repair genes is associated with familial cancer, aging and neurological disease, progress toward identifying polymorphisms associated with elevated risk of sporadic disease has been slow. This is partly due to the complexity of the genetic variation, the existence of large numbers of mostly low frequency variants and the contribution of many genes to variation in susceptibility. There has been limited development of methods to find associations between genotypes having many polymorphisms and pathway function or health outcome. We have explored several statistical methods for identifying polymorphisms associated with variation in DNA repair phenotypes. The model system used was 80 cell lines that had been resequenced to identify variation; 191 single nucleotide substitution polymorphisms (SNPs) are included, of which 172 are in 31 base excision repair pathway genes, 19 in 5 anti-oxidation genes, and DNA repair phenotypes based on single strand breaks measured by the alkaline Comet assay. Univariate analyses were of limited value in identifying SNPs associated with phenotype variation. Of the multivariable model selection methods tested: the easiest that provided reduced error of prediction of phenotype was simple counting of the variant alleles predicted to encode proteins with reduced activity, which led to a genotype including 52 SNPs; the best and most parsimonious model was achieved using a two-step analysis without regard to potential functional relevance: first SNPs were ranked by importance determined by random forests regression (RFR), followed by cross-validation in a second round of RFR modeling that included ever more SNPs in declining order of importance. With this approach six SNPs were found to minimize prediction error. The results should encourage research into utilization of multivariate analytical methods for epidemiological studies of the association of genetic variation in complex genotypes with risk of common diseases.     

NBN Gene Polymorphisms and Cancer Susceptibility: A Systemic Review

The relationship between DNA repair failure and cancer is well established as in the case of rare, high penetrant genes in high cancer risk families. Beside this, in the last two decades, several studies have investigated a possible association between low penetrant polymorphic variants in genes devoted to DNA repair pathways and risk for developing cancer. This relationship would be also supported by the observation that DNA repair processes may be modulated by sequence variants in DNA repair genes, leading to susceptibility to environmental carcinogens. In this framework, the aim of this review is to provide the reader with the state of the art on the association between common genetic variants and cancer risk, limiting the attention to single nucleotide polymorphisms (SNPs) of the NBN gene and providing the various odd ratios (ORs). In this respect, the NBN protein, together with MRE11 and RAD50, is part of the MRN complex which is a central player in the very early steps of sensing and processing of DNA double-strand breaks (DSBs), in telomere maintenance, in cell cycle control, and in genomic integrity in general. So far, many papers were devoted to ascertain possible association between common synonymous and non-synonymous NBN gene polymorphisms and increased cancer risk. However, the results still remain inconsistent and inconclusive also in meta-analysis studies for the most investigated E185Q NBN miscoding variant.     

Sporadic colorectal cancer and individual susceptibility: a review of the association studies investigating the role of DNA repair genetic polymorphisms

Mutations in one of the DNA repair genes are one of the most common reasons for cancer, and it may be assumed that the individual genetic background modulating the DNA repair capacity may affect the susceptibility to cancer. Numerous polymorphisms (mainly SNPs) have been identified for DNA repair genes, although their functional outcome and phenotypic effect is often unknown. The aim of the present review is to evaluate the studies investigating a possible influence of DNA repair polymorphisms in the risk of sporadic colorectal cancer and/or adenoma. Overall, no relevant common findings emerge among the studies, except for some statistically significant associations between polymorphisms in the XRCC1 and XPD genes, mainly for colorectal adenoma risk. Other individual associations remain to be confirmed. This inconclusive data may suggest that the modulation of cancer risk depends not only on a single gene/SNP, but also on a joint effect of multiple polymorphisms (or haplotypes) within different genes or pathways, in close interaction with environmental factors. The relevance of many low-penetrance genes in cancer susceptibility is supposed to be very subtle. Several reviewed association studies revealed weaknesses in their design. However, there has been a progressive improvement over the years in aspects such as simultaneous genotyping and combined analyses of different polymorphisms in larger numbers of patients and controls, as well as stratification of results by ethnicity, gender, and tumor localization. This gained experience shows that only carefully designed studies of a sufficient statistical power may resolve the relationships between polymorphisms and colorectal cancer risk.     

The biological revolution - towards a mechanistic understanding of the impact of diet on cancer risk

There is strong epidemiological evidence to show that differences in diet explain a significant proportion of the variation in cancer incidence worldwide. However, because of the complex nature of eating behaviour and the chemical heterogeneity of foods, it remains very difficult to ascertain which aspects of diet, in what quantities and over what time-frames are responsible for modifying risk. In addition, there are few dietary intervention studies demonstrating reduction in cancer risk. Much faster progress has been made in understanding the biological basis of cancer. It is now clear that damage to the genome resulting in aberrant expression of genes (principally suppression of tumour suppressor genes (TSGs) and inappropriate expression of oncogenes) is fundamental to tumorigenesis. It is also becoming clear that much of the inter-individual variation in cancer experience is due to differences in the amount of damage experienced and/or the capacity to repair that damage. Both of these processes are influenced strongly by dietary factors and by genetic predisposition (polymorphisms in the requisite genes). It is possible that understanding diet:gene interactions in DNA damage and in repair will not only explain much of the inter-individual variation in risk but also offer opportunities to design better dietary intervention studies aimed at chemoprevention. The Human Genome maps and the SNPs databases, together with the rapid development of tools suitable for investigating genetic and epigenetic changes in small tissue biopsies provide the means to begin to test hypotheses about the mechanisms by which diet influences cancer risk directly in human subjects. This is likely to form a significant component of the emerging science of nutrigenomics.     

Polymorphisms associated with the risk of lung cancer in a healthy Mexican Mestizo population: Application of the additive model for cancer

Lung cancer is the leading cause of cancer mortality in Mexico and worldwide. In the past decade, there has been an increase in the number of lung cancer cases in young people, which suggests an important role for genetic background in the etiology of this disease. In this study, we genetically characterized 16 polymorphisms in 12 low penetrance genes (AhR, CYP1A1, CYP2E1, EPHX1, GSTM1, GSTT1, GSTPI, XRCC1, ERCC2, MGMT, CCND1 and TP53) in 382 healthy Mexican Mestizos as the first step in elucidating the genetic structure of this population and identifying high risk individuals. All of the genotypes analyzed were in Hardy-Weinberg equilibrium, but different degrees of linkage were observed for polymorphisms in the CYP1A1 and EPHX1 genes. The genetic variability of this population was distributed in six clusters that were defined based on their genetic characteristics. The use of a polygenic model to assess the additive effect of low penetrance risk alleles identified combinations of risk genotypes that could be useful in predicting a predisposition to lung cancer. Estimation of the level of genetic susceptibility showed that the individual calculated risk value (iCRV) ranged from 1 to 16, with a higher iCRV indicating a greater genetic susceptibility to lung cancer.     

Molecular epidemiology, biomarkers and cancer prevention.

Molecular epidemiological studies within the field of cancer research provide the potential for elucidating the carcinogenic cascade at the molecular level. Identification of susceptible subsets of the population, based on polymorphisms in genes involved in carcinogenesis, has the potential to delineate more clearly those factors that might increase cancer risk among some, but not all, individuals. Rapid advances in human genomics are making it possible to develop detailed profiles of susceptible subgroups based upon genetic variants in multiple pathways. Here we discuss examples of recent susceptibility studies involving genes, such as those involved in carcinogen metabolism, DNA repair, cell cycle and immune status, that hold the promise of increasing our understanding of cancer etiology and possible prevention strategies.     

Familiäre Leukämien

Leukemias and other hematological neoplasias are frequently observed in association with different genetic disorders, such as DNA repair deficiency syndromes, tumor predisposition syndromes, immunodeficiency syndromes, familial cancer syndromes and bone marrow failure syndromes, as well as in connection with several constitutional chromosomal anomalies. Recently, in families with increased leukemia incidence, constitutional mutations have been identified in genes that are also affected by somatic mutations in sporadic leukemias. In addition to these high penetrance mutations, gene alterations with low penetrance and polymorphisms seem to predispose to leukemia and/or modify the clinical course of the disease. Predisposing and modifying polymorphisms can be found in genes involved in cell proliferation, apoptosis, DNA repair, detoxification, etc. The novel findings on constitutional genetic alterations predisposing to leukemia start to close the gap between inborn and acquired genetic diseases.     

Telomere Length and Genetic Anticipation in Lynch Syndrome

Telomere length variation has been associated with increased risk of several types of tumors, and telomere shortening, with genetic anticipation in a number of genetic diseases including hereditary cancer syndromes. No conclusive studies have been performed for Lynch syndrome, a hereditary colorectal cancer syndrome caused by germline mutations in the DNA mismatch repair genes. Here we evaluate telomere length in Lynch syndrome, both as a cancer risk factor and as a mechanism associated with anticipation in the age of cancer onset observed in successive generations of Lynch syndrome families. Leukocyte telomere length was measured in 244 mismatch repair gene mutation carriers from 96 Lynch syndrome families and in 234 controls using a monochrome multiplex quantitative PCR method. Cancer-affected mutation carriers showed significantly shorter telomeres than cancer-free mutation carriers. In addition, cancer-affected carriers showed the most pronounced shortening of telomere length with age, compared with unaffected carriers. The anticipation in the age of cancer onset observed in successive generations was not associated with telomere shortening, although, interestingly, all mother-son pairs showed telomere shortening. In conclusion, cancer-affected mismatch repair gene mutation carriers have distinct telomere-length pattern and dynamics. However, anticipation in the age of onset is not explained by telomere shortening. Pending further study, our findings suggest that telomere attrition might explain the previously reported dependence of cancer risk on the parent-of-origin of mismatch repair gene mutations.