Major DNA Repair Pathways Research Articles

Mismatch repair-dependent mutagenesis in nondividing cells

Mismatch repair (MMR) is a major DNA repair pathway in cells from all branches of life that removes replication errors in a strand-specific manner, such that mismatched nucleotides are preferentially removed from the newly replicated strand of DNA. Here we demonstrate a role for MMR in helping create new phenotypes in nondividing cells. We show that mispairs in yeast that escape MMR during replication can later be subject to MMR activity in a replication strand-independent manner in nondividing cells, resulting in either fully wild-type or mutant DNA sequence. In one case, this activity is responsible for what appears to be adaptive mutation. This replication strand-independent MMR activity could contribute to the formation of tumors arising in nondividing cells and could also contribute to mutagenesis observed during somatic hypermutation of Ig genes.

The effect of calorie restriction on the genome stability in DNA repair defective cells

It has been found that calorie restriction (CR) can extend the average and maximum life span from yeast to primates and delay the onset of age‐associated pathologies. Mismatch repair and double strand break repair systems are major DNA repair pathways whose function are critical for maintaining genome stability. The inactivation of these systems in human pathway has been reported to cause cancer predisposition such as Hereditary Nonpolyposis Colorectal Cancer, Werner and Bloom syndromes. By using Saccharomyces cerevisiae model system for CR and aging studies, DNA repair defective cells (msh2Δ, msh3Δ, msh6Δ and sgs1Δ) were cultured with normal (2% glucose) and CR (0.5% glucose) medium. Genome stability studies were analyzed by CANr, Homr and Lysr mutation assay. Results show that CR can significantly extend the lifespan of all wild‐type and mutants. Besides, CR shows significant decrease in HOMr mutation frequency by 2.7 × 104‐fold in msh2Δ mutant, whereas by 2.4 × 106‐fold and 1.4 × 106‐fold reduction in msh3Δ mutant for HOMr and LYSr assay respectively. In addition, for sgs1Δ mutant, there are significant decreases in HOMr and LYSr assay by two‐fold. Under normal condition, msh2Δ, msh3Δ and sgs1Δ cells found to have regrow phenotype upon aging. However, this phenotype was diminished under CR condition. Thus, CR promotes genome stability in DNA repair defective cells especially in msh2Δ, msh3Δ and sgs1Δ mutants.

ATP-dependent chromatin remodeling in the DNA-damage response

The integrity of DNA is continuously challenged by metabolism-derived and environmental genotoxic agents that cause a variety of DNA lesions, including base alterations and breaks. DNA damage interferes with vital processes such as transcription and replication, and if not repaired properly, can ultimately lead to premature aging and cancer. Multiple DNA pathways signaling for DNA repair and DNA damage collectively safeguard the integrity of DNA. Chromatin plays a pivotal role in regulating DNA-associated processes, and is itself subject to regulation by the DNA-damage response. Chromatin influences access to DNA, and often serves as a docking or signaling site for repair and signaling proteins. Its structure can be adapted by post-translational histone modifications and nucleosome remodeling, catalyzed by the activity of ATP-dependent chromatin-remodeling complexes. In recent years, accumulating evidence has suggested that ATP-dependent chromatin-remodeling complexes play important, although poorly characterized, roles in facilitating the effectiveness of the DNA-damage response. In this review, we summarize the current knowledge on the involvement of ATP-dependent chromatin remodeling in three major DNA repair pathways: nucleotide excision repair, homologous recombination, and non-homologous end-joining. This shows that a surprisingly large number of different remodeling complexes display pleiotropic functions during different stages of the DNA-damage response. Moreover, several complexes seem to have multiple functions, and are implicated in various mechanistically distinct repair pathways.

A special issue on DNA damage response and genome stability

The fidelity of the genome is under constant threat by exogenous and endogenous reactive species, including toxic chemicals, ionizing radiation and byproducts of normal cellular metabolism. These species cause damage to DNA by modifying DNA bases, breaking DNA strands and/or altering DNA structures. When this happens, the consequences to the cell can be disastrous, ranging from single gene mutations to massive chromosomal breakdown and rearrangements. These instabilities lead to severe human diseases including cancer. Fortunately, humans and other eukaryotes have evolved multiple mechanisms to ensure genome stability. These mechanisms can be classified into two major systems, one that deals with replication fidelity and the other that takes care of various DNA lesions. The replication fidelity maintenance system involves DNA polymerases and the DNA mismatch repair pathway. First, the 3'-5' proofreading nuclease activity of replicative DNA polymerases is responsible for removing the majority of mis-incorporated nucleotides during DNA replication [1]. However, there are always cases where certain mis-incorporations escape from the proofreading activity. In the latter situation, the mismatch repair machinery [2] is recruited to the site to correct the mismatch. If a DNA template contains a bulky DNA adduct that blocks the polymerization activity of replicative polymerases, a specialized DNA polymerase, also called translesion synthesis polymerase, is employed to bypass the bulky DNA lesion, followed by a switch back to the replicative polymerase to continue normal replication [3]. The DNA damage processing system can be divided into two steps: DNA damage response and DNA repair. Extensive studies in the past 20 years have elucidated the mechanisms of several major DNA repair pathways that remove almost all kinds of DNA lesions. These pathways include base excision repair [4], nucleotide excision repair [5,6], mismatch repair [2] and double strand break repair [7,8]. DNA damage response is a complex signal transduction process that has the ability to sense DNA damage and transduce the information to the cell to direct cellular responses to the damage. An arsenal of protein activities has been identified, which function to be damage sensors, mediators, transducers and effectors during DNA damage response [9]. Unlike the DNA repair pathways, however, the mechanism of DNA damage response is much less known. In this issue, Cell & Bioscience presents a series of reviews attempting to define the most challenging questions in DNA damage response and repair, and to provide an overview of the latest breakthroughs and developments in the field. The article by Nan Wu and Hongtao Yu [10] explores the mechanism by which the structural maintenance of chromosomes (Smc) proteins and non-Smc proteins, which are required for chromatid cohesion, chromosomal segregation and condensation, regulate DNA damage response and repair. A second article by Bin Wang [11] describes how BRCA1, an important tumor suppressor and a critical DNA damage response mediator, interacts with its partners to regulate activities of multiple repair and checkpoint pathways for genome maintenance. Maintaining the stability of repetitive DNA sequences, particularly the (CAG)n and (CTG)n trinucleotide repeats, whose expansions cause numerous human disorders, is extremely challenging, as is the understanding of the mechanisms that regulate the stability/instability of the repetitive sequences. In their article, Guoqi Liu and Michael Leffak [12] debate how collaborative efforts by the replication machinery and multiple DNA repair pathways stabilize (CAG)n and (CTG)n trinucleotide repeats. It is our sincere hope that this special issue brings our readers enlightenment and offers sufficient introductory information to help them appreciate new breakthroughs in the field.

P1-06-22: Identification of Biomarkers in Breast Cancer for Prediction of Response to PARP Inhibitor Olaparib.

Abstract Background: Poly (ADP-ribose) polymerase (PARP) is an enzyme involved in DNA repair. PARP inhibitors operate on the principle of synthetic lethality in conjunction with DNA damaging agents, and are likely to be useful for treatment of BRCA-mutated cancers and triple negative breast cancers exhibiting ‘BRCA-ness’ or other signs of DNA repair deficiency. Multiple PARP inhibitors have been developed, such as Olaparib (AstraZeneca), BSI-201 (Sanofi-Aventis) and ABT-888 (Abbott Laboratories). Though some clinical trials have shown drugs in this class to be promising, not all results have been positive. As PARP inhibitors differ in mechanism of action, dosing interval and toxicities, trial results seem to depend on the specific combination of PARP inhibitor and patient population. To understand why some studies succeeded and others failed and to guide new clinical trials in patient selection, there is an urgent need for biomarker identification, both for PARP inhibitors in general and for the specific idiosyncratic mechanisms of each drug. Material and Methods: Thirty-three in vitro breast cancer cell lines were administered the PARP inhibitor Olaparib, with sensitivity to the compound summarized as the dose necessary to kill 50% of each culture. mRNA expression (Affymetrix U133A, Exon 1.0ST array) and transcriptome sequence (Illumina GAII) were available for 22/33 cell lines, among which 9 were sensitive and 13 resistant. To obtain robust predictive markers that are minimally dependent on the specific PARP inhibitor and expression platform, a bottom-up approach was opted for, restricted to genes in the major DNA repair pathways. Logistic regression with forward selection was used to determine the most important markers, further reduced based on consistency across platforms. The weighted voting algorithm was used to build the final predictor. Eight U133A data sets with number of tumor samples varying from 61 to 289 were subsequently used to verify prevalence and to identify the subpopulations that are likely to respond according to the predictor. To verify cross-platform generalizability, the signature was additionally tested in 430 TCGA samples with custom Agilent 244K gene expression. Results: For the development of a genomic signature that might work for multiple PARP inhibitors and expression platforms, prior knowledge of DNA repair pathways was incorporated and stringent criteria for marker inclusion were applied using three different platforms. Eight genes fulfilled the criteria, of which 5 were resistance markers and 3 sensitivity markers. When testing the 8-gene signature in eight U133A data sets, 40–48% of patients were predicted to be responsive to Olaparib. In addition, well-known markers (ER, PR, ERBB2, KRT5/17) were examined in patients expressing the 8-gene sensitivity signature. A higher percentage of these patients were ERBB2-negative and did overexpress KRT5/17, indicative for the basal subtype. Prevalence and relationship with these markers were confirmed in 430 samples on a distinct platform (Agilent). Conclusion: Cell line exposure to Olaparib has yielded an 8-gene predictor of sensitivity. This signature was observed in a substantial fraction of primary breast tumors predicted to benefit from Olaparib. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-06-22.

DNA Repair: From Genome Maintenance to Biomarker and Therapeutic Target

A critical link exists between an individual's ability to repair cellular DNA damage and cancer development, progression, and response to therapy. Knowledge gained about the proteins involved and types of damage repaired by the individual DNA repair pathways has led to the development of a variety of assays aimed at determining an individual's DNA repair capacity. These assays and their use in the analysis of clinical samples have yielded useful though somewhat conflicting data. In this review article, we discuss the major DNA repair pathways, the proteins and genes required for each, assays used to analyze activity, and the relevant clinical studies to date. With the recent results from clinical trials targeting specific DNA repair proteins for the treatment of cancer, accurate, reproducible, and relevant analysis of DNA repair takes on an even greater significance. We highlight the strengths and limitations of these DNA repair studies and assays, with respect to the clinical assessment of DNA repair capacity to determine cancer development and response to therapy.

Abstract CN05-02: Targeting the Fanconi anemia/BRCA pathway in cancer therapy.

Abstract CN05-02: Targeting the Fanconi anemia/BRCA pathway in cancer therapy.

Abstract IA9: Targeting the Fanconi anemia/BRCA pathway in cancer therapy

Abstract IA9: Targeting the Fanconi anemia/BRCA pathway in cancer therapy

Histone deacetylases (HDACs) in XPC gene silencing and bladder cancer

Bladder cancer is one of the most common malignancies and causes hundreds of thousands of deaths worldwide each year. Bladder cancer is strongly associated with exposure to environmental carcinogens. It is believed that DNA damage generated by environmental carcinogens and their metabolites causes development of bladder cancer. Nucleotide excision repair (NER) is the major DNA repair pathway for repairing bulk DNA damage generated by most environmental carcinogens, and XPC is a DNA damage recognition protein required for initiation of the NER process. Recent studies demonstrate reduced levels of XPC protein in tumors for a majority of bladder cancer patients. In this work we investigated the role of histone deacetylases (HDACs) in XPC gene silencing and bladder cancer development. The results of our HDAC inhibition study revealed that the treatment of HTB4 and HTB9 bladder cancer cells with the HDAC inhibitor valproic acid (VPA) caused an increase in transcription of the XPC gene in these cells. The results of our chromatin immunoprecipitation (ChIP) studies indicated that the VPA treatment caused increased binding of both CREB1 and Sp1 transcription factors at the promoter region of the XPC gene for both HTB4 and HTB9 cells. The results of our immunohistochemistry (IHC) staining studies further revealed a strong correlation between the over-expression of HDAC4 and increased bladder cancer occurrence (p < 0.001) as well as a marginal significance of increasing incidence of HDAC4 positivity seen with an increase in severity of bladder cancer (p = 0.08). In addition, the results of our caspase 3 activation studies demonstrated that prior treatment with VPA increased the anticancer drug cisplatin-induced activation of caspase 3 in both HTB4 and HTB9 cells. All of these results suggest that the HDACs negatively regulate transcription of the XPC gene in bladder cancer cells and contribute to the severity of bladder tumors.

Analysis of the Oxidative Damage Repair GenesNUDT1,OGG1, andMUTYHin Patients from Mismatch Repair Proficient HNPCC Families (MSS-HNPCC)

Several studies have described molecular differences between microsatellite stable hereditary nonpolyposis colorectal cancer (MSS-HNPCC) and microsatellite unstable Lynch syndrome tumors (MSI-HNPCC). These differences highlight the possibility that other instability forms could explain cancer susceptibility in this group of families. The base excision repair (BER) pathway is the major DNA repair pathway for oxidative DNA damage. A defect in this pathway can result in DNA transversion mutations and a subsequent increased cancer risk. Mutations in MUTYH have been associated with increased colorectal cancer (CRC) risk while no association has been described for OGG1 or NUDT1. We performed mutational screening of the three genes involved in defense against oxidative DNA damage in a set of 42 MSS-HNPCC families. Eight rare variants and 5 frequent variants were found in MSS-HNPCC patients. All variants were previously described by other authors except variant c.285C>T in OGG1. Segregation studies were done and in silico programs were used to estimate the level of amino acid conservation, protein damage prediction, and possible splicing alterations. Variants OGG1 c.137G>A; MUTYH c.1187G>A were detected in Amsterdam I families and cosegregate with cancer. Analysis of OGG1 c.137G>A transcripts showed an inactivation of the splicing donor of exon 1. Two rare variants (OGG1 c.137G>A; MUTYH c.1187G>A) and one common polymorphism (NUDT1 c.426C>T) were associated with CRC risk. We show that the BER pathway can play a significant role in a number of MSS-HNPCC colorectal cancers. More studies could be of interest in order to gain further understanding of yet unexplained CRC susceptibility cases.

Nucleotide excision repair: DNA damage recognition and preincision complex assembly

Nucleotide excision repair (NER) is one of the major DNA repair pathways in eukaryotic cells counteracting genetic changes caused by DNA damage. NER removes a wide set of structurally diverse lesions such as pyrimidine dimers arising upon UV irradiation and bulky chemical adducts arising upon exposure to carcinogens or chemotherapeutic drugs. NER defects lead to severe diseases including some forms of cancer. In view of the broad substrate specificity of NER, it is of interest to understand how a certain set of proteins recognizes various DNA lesions in the context of a large excess of intact DNA. This review focuses on DNA damage recognition and following stages resulting in preincision complex assembly, the key and still most unclear steps of NER. The major models of primary damage recognition and preincision complex assembly are considered. The contribution of affinity labeling techniques in study of this process is discussed.

Abstract SY28-01: Targeting DNA repair pathways in cancer therapy

Abstract SY28-01: Targeting DNA repair pathways in cancer therapy

Abstract 4861: The role of histone deacetylases (HDACs) in XPC gene silencing and bladder cancer development

Abstract The role of histone deacetylases (HDACs) in XPC gene silencing and bladder cancer development Xiaoxin S. Xu, Le Wang, and Gan Wang Institute of Environmental Health Sciences, Wayne State University, 259 Mack Ave., Detroit, MI 48202 Bladder cancer is one of the most common malignancies and causes hundreds thousands of death worldwide each year. Bladder cancer is strongly associated with exposure to environmental carcinogens. It is believed that DNA damage generated by environmental carcinogens and their metabolites causes development of bladder cancer. Nucleotide excision repair (NER) is the major DNA repair pathway for repairing bulk DNA damage generated by most environmental carcinogens and XPC is a DNA damage recognition protein required for initiation of the NER process. Recent studies demonstrate reduced levels of XPC protein in tumors for a majority of bladder cancer patients. In this work we investigated the role of histone deacetylases (HDACs) in XPC gene silencing and bladder cancer development. The results of our HDAC inhibitor study revealed that the treatment of HTB4 and HTB9 bladder cancer cells with HDAC inhibitor valproic acid (VPA) caused an increase in transcription of the XPC gene in these cells. The results of our chromatin immunoprecipitation (ChIP) studies indicated that the VPA treatment caused increased bindings of both CREB1 and Sp1 transcription factors at the promoter of XPC gene for both HTB4 and HTB9 cells. The results of our immunohistochemistry (IHC) staining studies further revealed a strong correlation between the presence of HDAC4 and the occurrence of bladder tumors. In addition, the results of our caspase 3 activation studies demonstrated that the prior treatment with VPA increased the anticancer drug cisplatin-induced activation of caspase 3 in both HTB4 and HTB9 cells. All these results suggest that the HDACs negatively regulate transcription of the XPC gene in bladder cancer cells and contribute to the development of bladder cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4861.

The role of DNA repair in benzene-induced carcinogenesis

Benzene is a well-known human carcinogen, but the ultimate mode of action is still not known. Several reactive metabolites have been identified, including benzene oxide, phenol, hydrochinone, catechol and benzoquinones, generating different types of DNA lesions. Furthermore, the latter three metabolites may lead to the formation of reactive oxygen species (ROS) due to redox cycling, which give rise to oxidative DNA lesions and altered signaling pathways. Also, the inhibition of DNA topoisomerase II may result in DNA double strand breaks. Even though the exact contribution of the respective metabolites to benzene-induced carcinogenicity is not yet resolved, the major DNA repair pathways such as base excision repair (BER), nucleotide excision repair (NER) and double strand break (DSB) repair are involved in the removal of benzene-induced DNA lesions. The observed target organ specificity may result from increased adduct formation, but also from poor repair in bone marrow progenitor cells. While especially excision repair pathways are predominantly error-free and thus protective, DSB repair is largely error prone and may contribute to benzene-induced genomic instability.

True Lies: The Double Life of the Nucleotide Excision Repair Factors in Transcription and DNA Repair

Nucleotide excision repair (NER) is a major DNA repair pathway in eukaryotic cells. NER removes structurally diverse lesions such as pyrimidine dimers, arising upon UV irradiation or bulky chemical adducts, arising upon exposure to carcinogens and some chemotherapeutic drugs. NER defects lead to three genetic disorders that result in predisposition to cancers, accelerated aging, neurological and developmental defects. During NER, more than 30 polypeptides cooperate to recognize, incise, and excise a damaged oligonucleotide from the genomic DNA. Recent papers reveal an additional and unexpected role for the NER factors. In the absence of a genotoxic attack, the promoters of RNA polymerases I- and II-dependent genes recruit XPA, XPC, XPG, and XPF to initiate gene expression. A model that includes the growth arrest and DNA damage 45α protein (Gadd45α) and the NER factors, in order to maintain the promoter of active genes under a hypomethylated state, has been proposed but remains controversial. This paper focuses on the double life of the NER factors in DNA repair and transcription and describes the possible roles of these factors in the RNA synthesis process.

DNA double-strand break signaling and human disorders

DNA double-strand breaks are among the most serious types of DNA damage and their signaling and repair is critical for all cells and organisms. The repair of both induced and programmed DNA breaks is fundamental as demonstrated by the many human syndromes, neurodegenerative diseases, immunodeficiency and cancer associated with defective repair of these DNA lesions. Homologous recombination and non-homologous end-joining pathways are the two major DNA repair pathways responsible for mediating the repair of DNA double-strand breaks. The signaling of DNA double-strand breaks is critical for cells to orchestrate the repair pathways and maintain genomic integrity. This signaling network is highly regulated and involves a growing number of proteins and elaborated posttranslational modifications including phosphorylation and ubiquitylation. Here, we highlight the recent progress in the signaling of DNA double-strand breaks, the major proteins and posttranslational modifications involved and the diseases and syndromes associated with impaired signaling of these breaks.

Unlocking the Molecular Mechanisms of DNA Repair and Platinum Drug Resistance in Cancer Chemotherapy

This review article is devoted exclusively to DNA repair and acquired resistance to platinum compounds, with an emphasis on research needs and clinical application. It focuses on the role of genetic variants (gene mutations and SNPs) and epigenetic alterations (methylation and acetylation). Four major DNA repair pathways and one enzymecorrection mechanism are presented: Base Excision Repair, Nucleotide Excision Repair, DNA Double-Strand Break Repair, Mismatch Repair and Direct Damage Reversal. It is suggested that one cause of platinum resistance is more accurately described as alterations of DNA repair system rather than activation of DNA repair mechanism, as this cause of resistance is brought about by changes in genetic and epigenetic regulation. Given what is known, research on DNA repair and platinum resistance might best be directed at 1) transcriptional cis-elements (activators/repressors) within promoters of essential DNA repair genes, 2) effects of epigenetic alterations, and 3) connections between gene expression and DNA methylation or protein acetylation. Special attention should be directed at three interrelationships: between the different DNA repair pathways; between DNA methylation and protein acetylation; and between DNA repair pathways and DNA methylation or protein acetylation. Improved clinical outcomes may be achieved by restoring wild type p53 with small molecule drugs, the use of gene demethylation strategies, individual-targeted treatment of BRCA mutation carrier, and combining platinum compounds with molecularly-targeted drugs such as EGFR inhibitors. Keywords: DNA repair, platinum resistance, genetic alteration, methylation, acetylation, epigenetic regulation

Hypermutability of Damaged Single-Strand DNA Formed at Double-Strand Breaks and Uncapped Telomeres in Yeast Saccharomyces cerevisiae

The major DNA repair pathways operate on damage in double-strand DNA because they use the intact strand as a template after damage removal. Therefore, lesions in transient single-strand stretches of chromosomal DNA are expected to be especially threatening to genome stability. To test this hypothesis, we designed systems in budding yeast that could generate many kilobases of persistent single-strand DNA next to double-strand breaks or uncapped telomeres. The systems allowed controlled restoration to the double-strand state after applying DNA damage. We found that lesions induced by UV-light and methyl methanesulfonate can be tolerated in long single-strand regions and are hypermutagenic. The hypermutability required PCNA monoubiquitination and was largely attributable to translesion synthesis by the error-prone DNA polymerase ζ. In support of multiple lesions in single-strand DNA being a source of hypermutability, analysis of the UV-induced mutants revealed strong strand-specific bias and unexpectedly high frequency of alleles with widely separated multiple mutations scattered over several kilobases. Hypermutability and multiple mutations associated with lesions in transient stretches of long single-strand DNA may be a source of carcinogenesis and provide selective advantage in adaptive evolution.

Interaction with RPA Is Necessary for Rad52 Repair Center Formation and for Its Mediator Activity

Homologous recombination (HR) is a major DNA repair pathway and therefore essential for maintaining the integrity of the genome. HR is catalyzed by proteins encoded by genes of the RAD52 epistasis group, including the recombinase Rad51 and its mediator Rad52. HR proteins fused with green fluorescent protein form foci at damaged DNA reflecting the assembly of repair centers that harbor a high concentration of repair proteins. Rad52 mediates the recruitment of Rad51 and other HR proteins to DNA damage. To understand the mechanism for the assembly of Rad52-dependent DNA repair centers, we used a mutational strategy to identify a Rad52 domain essential for its recruitment to DNA repair foci. We present evidence to implicate an acidic domain in Rad52 in DNA repair focus formation. Mutations in this domain confer marked DNA damage sensitivity and recombination deficiency. Importantly, these Rad52 mutants are specifically compromised for interaction with the single-stranded DNA-binding factor RPA. Based on these findings, we propose a model where Rad52 displaces RPA from single-stranded DNA using the acidic domain as a molecular lever.

Genetic Susceptibility to Renal Cell Carcinoma: The Role of DNA Double-Strand Break Repair Pathway

Alterations in DNA repair genes have been shown to cause a reduction in host DNA repair capacity and may influence host susceptibility to carcinogenesis. The double-strand break repair is a major DNA-repair pathway. This study tested the hypothesis that common sequence variants of the double-strand break pathway genes predispose susceptible individuals to an increased risk for renal cell carcinoma. Toward this end, we evaluated the associations of 13 single-nucleotide polymorphisms in 10 candidate genes involved in the double-strand break pathway with renal cell carcinoma risk in a population-based case-control study that included 326 Caucasian renal cell carcinoma patients and 335 controls. Using the homozygous wild type as the reference group, we observed a significantly increased renal cell carcinoma risk associated with the homozygous variant genotype of NBS1 (rs1805794; odds ratio, 2.13; 95% confidence interval (95% CI), 1.17-3.86). Carrying of at least one copy of the variant XRCC4 allele was also associated with a significantly increased risk (rs1805377; odds ratio, 1.56; 95% CI, 1.08-2.26). Importantly, in pathway analysis, compared with the reference group (1 or less adverse alleles), individuals with two (odds ratio, 1.26; 95% CI, 0.83-1.91), three (odds ratio, 1.00; 95% CI, 0.64-1.56), and more than three adverse alleles (odds ratio, 1.75; 95% CI, 1.03-2.98) were at increased risk for renal cell carcinoma with significant association in subjects carrying more than 3 adverse alleles. Results from this study provide evidence that individuals with a higher number of genetic variations in the DBS repair pathway are at an increased risk for renal cell carcinoma. These findings require further validation in independent populations.