Challenges and complexities in estimating both the functional impact and the disease risk associated with the extensive genetic variation in human DNA repair genes

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Abstract

Individual risk and the population incidence of disease result from the interaction of genetic susceptibility and exposure. DNA repair is an example of a cellular process where genetic variation in families with extreme predisposition is documented to be associated with high disease likelihood, including syndromes of premature aging and cancer. Although the identification and characterization of new genes or variants in cancer families continues to be important, the focus of this paper is the current status of efforts to define the impact of polymorphic amino acid substitutions in DNA repair genes on individual and population cancer risk. There is increasing evidence that mild reductions in DNA repair capacity, assumed to be the consequence of common genetic variation, affect cancer predisposition. The extensive variation being found in the coding regions of DNA repair genes and the large number of genes in each of the major repair pathways results in complex genotypes with potential to impact cancer risk in the general population. The implications of this complexity for molecular epidemiology studies, as well as concepts that may make these challenges more manageable, are discussed. The concepts include both experimental and computational approaches that could be employed to develop predictors of disease susceptibility based on DNA repair genotype, focusing initially on studies to assess functional impact on individual proteins and pathways and then on molecular epidemiology studies to assess exposure-dependent health risk. In closing, we raise some of the non-technical challenges to the utilization of the full richness of the genetic variation to reduce disease occurrence and ultimately improve health care.

Introduction

As many a mother has told her children, “you are different!” Genetic individualization is a critical element in determining individual risk of disease and the incidence of disease in the population. DNA repair is an example of a cellular process where genetic variation is documented to be associated with disease risk, including premature aging and cancer. This connection is most apparent in families with extreme predisposition, families that were the resources for the identification of many cancer genes. The focus of this paper is not on the genes or variants in cancer families. Rather, we focus on the current status of efforts to define the role of common (polymorphic) variants in DNA repair genes, and small to moderate reductions in repair capacity, in individual cancer susceptibility and the incidence of cancer in the population. As background we briefly highlight the current status of systematic screens for genetic variation in the human population, as well as studies that are revealing greater and greater complexity in the roles of both genetics and environmental exposures in cancer etiology (Section 2). Next, we summarize the DNA repair pathways and their genes (Section 3), the extent of polymorphic variation that has been identified in many of these genes (Section 4), and the evidence that mild reductions in repair capacity (that is, non-inactivating mutations) affect cancer predisposition (Section 5). We then discuss the complexity of the genotypes existing in the general population (Section 6) and the implications of the extensive variation among individuals for future studies, as well as concepts that may make the challenges of accounting for the role of the vast genetic variation more manageable (Section 7). Subsequently, we present experimental approaches that take into account these matters and that might be employed to develop predictors of disease susceptibility based on DNA repair pathway genotypes. We focus initially on studies to assess the functional impact of variants (Section 8) and then on molecular epidemiology studies to assess the associated health risk (Section 9). In closing we raise some of the non-technical challenges to utilization of the full richness of the genetic variation to reduce disease occurrence and ultimately improve health care (Section 10). Although we focus on DNA repair and cancer, similar stories are being built for other pathways and diseases, e.g. the immune response and cardiovascular disease.

Section snippets

Background

We did not need the Human Genome Program to tell us that individuals are unique. But, the completion of the sequencing of the human genome has provided the resources for initiating systematic screens for genetic variation over much of the genome, an obvious extension of the Human Genome Program. With the resources and data available, it is increasingly feasible to provide quantitative estimates of the extent of genetic differences among individuals of varying degrees of relationship. The most

The integrity of the genome is protected by DNA repair proteins working in pathways

Our genome integrity is continually threatened—by endogenous products produced during normal cellular respiration, by errors that arise during DNA replication and recombination, and by exogenous exposures. The generated DNA “damage” intermediates, if not repaired, can lead to the accumulation of nucleotide sequence changes (mutations). Thus, organisms have an array of defense mechanisms that restore DNA back to its original, unmodified state and avert the potential deleterious health effects of

There is extensive polymorphic variation in DNA repair genes

Reviewing the results from the systematic screening of DNA repair genes for sequence variation reveals the extent and complexity of genetic variation in the general population. Over 80 DNA repair and repair-related genes have been screened for variation, mostly in a sample set of 90 unrelated and presumably generally healthy individuals from the NIH DPDR [2]. Embracing the approach of the Human Genome Project, much of the screening for genetic variation is being conducted by a few laboratories

Evidence that “mildly” reduced DNA repair capacity affects susceptibility

There should be no question regarding the role of DNA repair in ameliorating the consequences of DNA damage and minimizing the incidence of cancer. Many of the cancer genes identified in family studies over the last several decades have roles in DNA replication and repair, and thus normal function is required for minimizing the transmission of unrepaired DNA damage during cell division. The more challenging question is the extent to which small to moderate reductions in the ability to repair

Implications of extensive polymorphic variation for analysis of the role of genetics in cancer susceptibility

An important observation from the data obtained in screening for polymorphic variants in DNA repair genes is the relatively limited number of high frequency amino acid substitution variants. Instead, a very large number of low frequency variants have been identified. The distribution of alleles by individual variant allele frequency is summarized in Table 3. This summary is based on data for 74 genes and 423 variants (Table 2). The 41 variant alleles existing at 10% allele frequency or greater

Thinking in terms of pathways and known environmental risk factors can guide analysis of the relationship of complex genotypes to repair function and disease

Adding to the complexity of cancer etiology are the non-genetic factors, often referred to as “the environment”. This is especially true for the cases of sporadic cancer, where genetics has a less dominant role in determining risk than in the relatively few cancer families. Viewed broadly, the environment of people includes endogenous factors, lifestyle factors such as smoking and reproductive history, as well as exposure to exogenous environmental pollutants. Endogenous factors include

How can the polymorphic variation most likely to impact function, and therefore risk, be identified in laboratory-based studies?

Although a large number of amino acid substitutions have been identified and catalogued, only some (currently unknown) fraction of these substitutions are likely to have at least a modest impact on protein function, repair capacity and genetic susceptibility. The currently available list of variant alleles in repair genes makes it possible to address many questions. (1) What fraction of the polymorphic variants have at least a modest impact on protein function and an associated impact on

How might knowledge of extensive genetic variation in repair pathways be utilized in population studies of health risk?

Molecular epidemiology studies are the ultimate “experiments” for evaluating the relationship of variant genotypes to human disease. The challenge is moving from assessing the effect of a single polymorphic variant on health risk in individuals with very divergent genetic backgrounds, to identifying the combinations of variants that are associated with different levels of risk of disease, given a particular exposure history. Studies of individual, high frequency variants have provided proof of

In closing

We have presented a fairly exhaustive discussion of our perspective of the challenges and complexities involved in estimating both the functional impact of genetic variants and the disease risk associated with the extensive genetic variation in human DNA repair genes. In retrospect, disciplines studying the physiological impact of the extensive genetic variation among individuals, e.g. molecular epidemiology and pharmacogenetics, are facing challenges similar to those faced by a number of other

Acknowledgements

The authors thank many colleagues and associates for fruitful discussions, in particular David O. Nelson. Given the scope of the topic, the literature cited is by no means comprehensive; many relevant studies have not been included. This work was performed under the auspices of the US Department of Energy by the University of California, Lawrence Livermore National Laboratory (LLNL) under Contract No. W-7405-Eng-48. This research was funded in part by the Laboratory Directed Research and

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