Kyoung-Yeon Kim

SNP–SNP interactions between DNA repair genes were associated with breast cancer risk in a Korean population

Single nucleotide polymorphisms (SNPs) in nucleotide excision repair (NER) pathway genes may modulate DNA repair capacity and increase susceptibility to breast cancer (BC). A case‐control study was conducted by evaluating genes involved in DNA repair to identify polymorphisms associated with BC.

Impact of genetic polymorphisms in base excision repair genes on the risk of breast cancer in a Korean population.

The contribution of single nucleotide polymorphisms (SNPs) in base excision repair (BER) genes to the risk of breast cancer (BC) was evaluated by focusing on two key genes: apurinic/apyrimidinic endonuclease 1 (APEX1) and 8-oxoguanine DNA glycosylase (OGG1). Genetic variations in the genes encoding these DNA repair enzymes may alter their functions and increase susceptibility to carcinogenesis. The aim of this study was to analyze polymorphisms in two BER genes, exploring their associations and particularly the combined effects of these variants on BC risk in a Korean population. Three SNPs of two BER genes were genotyped using the Illumina GoldenGate™ method. In total, 346 BC patients and 361 cancer-free controls were genotyped for these BER gene polymorphisms and analyzed for their correlation with BC risk in multiple logistic regression models. Multiple logistic regression models adjusted for age, family history of BC, and body mass index were used. The APEX1 Asp148Glu polymorphism was weakly associated with BC risk. The combined analysis among the BER genes, however, showed significant effects on BC risk. The APEX1 Asp148Glu carrier, in combination with OGG1 rs2072668 and OGG1 Ser326Cys, was strongly associated with an increased risk of BC. Moreover, the combination of the C-C haplotype of OGG1 with the APEX1 Asp148Glu genotype was also associated with an additive risk effect of BC [ORs=2.44, 2.87, and 3.50, respectively]. The combined effect of APEX1 Asp148Glu was found to be associated with an increased risk of BC. These results suggest that the combined effect of different SNPs within BER genes may be useful in predicting BC risk.

Association of GNLY Genetic Polymorphisms with Chronic Liver Disease in a Korean Population

Granulysin (GNLY) is found in cytotoxic granules of cytolytic T lymphocytes and natural killer (NK) cells, which are critical for hepatitis B virus (HBV) clearance. GNLY cytotoxicity plays an important role in the defense against viruses or intracellular bacteria. We hypothesized that genetic variation in the GNLY gene could affect the resistance of hosts against HBV infection. We compared the distribution frequencies of GNLY polymorphisms between an HBV-induced chronic liver disease (CLD) group and a spontaneous recovery (SR) control group to determine whether GNLY polymorphisms play a role in HBV clearance. A total of 117 patients in the SR group and 230 patients in the CLD group were enrolled. Samples derived from complex infections, including hepatitis C and human immunodeficiency virus, and those associated with insufficient clinical information (10 samples in SR and 24 samples in CLD) were excluded from the study. The final analysis included 107 SR and 206 CLD samples. DNA was extracted from peripheral blood, and GNLY genotypes were determined by the GoldenGate(®) method. The genotype distribution of the single-nucleotide polymorphisms (SNPs) rs2886767 (C>T), rs1561285 (G>C), and rs11127 (T>C) were significantly different between the SR and CLD groups in a recessive model (p<0.015). These three SNPs were in a complete linkage disequilibrium (LD) block. Diplotype distributions of haplotype (HT) 1 (C-G-T) and HT2 (T-C-C) were significantly different between the SR and CLD groups in a recessive model (p=0.025) and a dominant model (p=0.008). All p-values remained significant after multiple comparisons. GNLY polymorphism genotypes and diplotypes were associated with the chronicity of HBV. These data suggested that genetic variation of GNLY may be an important factor in HBV clearance through the CD8+ T or NK cell-mediated removal of HBV-infected cells from the host.

Association between Interferon-Inducible Protein 6 (IFI6) Polymorphisms and Hepatitis B Virus Clearance

CD8+ T cells are key factors mediating hepatitis B virus (HBV) clearance. However, these cells are killed through HBV-induced apoptosis during the antigen-presenting period in HBV-induced chronic liver disease (CLD) patients. Interferon-inducible protein 6 (IFI6) delays type I interferon-induced apoptosis in cells. We hypothesized that single nucleotide polymorphisms (SNPs) in the IFI6 could affect the chronicity of CLD. The present study included a discovery stage, in which 195 CLD patients, including chronic hepatitis B (HEP) and cirrhosis patients and 107 spontaneous recovery (SR) controls, were analyzed. The genotype distributions of rs2808426 (C > T) and rs10902662 (C > T) were significantly different between the SR and HEP groups (odds ratio [OR], 6.60; 95% confidence interval [CI], 1.64 to 26.52, p = 0.008 for both SNPs) and between the SR and CLD groups (OR, 4.38; 95% CI, 1.25 to 15.26; p = 0.021 and OR, 4.12; 95% CI, 1.18 to 14.44; p = 0.027, respectively). The distribution of diplotypes that contained these SNPs was significantly different between the SR and HEP groups (OR, 6.58; 95% CI, 1.63 to 25.59; p = 0.008 and OR, 0.15; 95% CI, 0.04 to 0.61; p = 0.008, respectively) and between the SR and CLD groups (OR, 4.38; 95% CI, 1.25 to 15.26; p = 0.021 and OR, 4.12; 95% CI, 1.18 to 14.44; p = 0.027, respectively). We were unable to replicate the association shown by secondary enrolled samples. A large-scale validation study should be performed to confirm the association between IFI6 and HBV clearance.

Functional Validation of Rare Human Genetic Variants Involved in Homologous Recombination Using Saccharomyces cerevisiae

Systems for the repair of DNA double-strand breaks (DSBs) are necessary to maintain genome integrity and normal functionality of cells in all organisms. Homologous recombination (HR) plays an important role in repairing accidental and programmed DSBs in mitotic and meiotic cells, respectively. Failure to repair these DSBs causes genome instability and can induce tumorigenesis. Rad51 and Rad52 are two key proteins in homologous pairing and strand exchange during DSB-induced HR; both are highly conserved in eukaryotes. In this study, we analyzed pathogenic single nucleotide polymorphisms (SNPs) in human RAD51 and RAD52 using the Polymorphism Phenotyping (PolyPhen) and Sorting Intolerant from Tolerant (SIFT) algorithms and observed the effect of mutations in highly conserved domains of RAD51 and RAD52 on DNA damage repair in a Saccharomyces cerevisiae-based system. We identified a number of rad51 and rad52 alleles that exhibited severe DNA repair defects. The functionally inactive SNPs were located near ATPase active site of Rad51 and the DNA binding domain of Rad52. The rad51-F317I, rad52-R52W, and rad52-G107C mutations conferred hypersensitivity to methyl methane sulfonate (MMS)-induced DNA damage and were defective in HR-mediated DSB repair. Our study provides a new approach for detecting functional and loss-of-function genetic polymorphisms and for identifying causal variants in human DNA repair genes that contribute to the initiation or progression of cancer.

Yeast-based assays for characterization of the functional effects of single nucleotide polymorphisms in human DNA repair genes

DNA repair mechanisms maintain genomic integrity upon exposure to various types of DNA damage, which cause either single- or double-strand breaks in the DNA. Here, we propose a strategy for the functional study of single nucleotide polymorphisms (SNPs) in the human DNA repair genes XPD/ERCC2, RAD18, and KU70/XRCC6 and the checkpoint activation gene ATR that are essentially involved in the cell cycle and DNA damage repair. We analyzed the mutational effects of the DNA repair genes under DNA-damaging conditions, including ultraviolet irradiation and treatment with genotoxic reagents, using a Saccharomyces cerevisiae system to overcome the limitations of the human cell-based assay. We identified causal variants from selected SNPs in the present analyses. (i) R594C SNP in RAD3 (human XPD/ERCC2) caused severe reductions in the growth rate of mutant cells upon short-wavelength UV irradiation or chemical reagent treatment. (ii) The growth rates of the selected variants in RAD18, YKU70, and MEC1 were similar to those of wild-type cells on methyl methanesulfonate and hydroxyurea treated media. (iii) We also assessed the structural impact of the SNPs by analyzing differences in the structural conformation and calculating the root mean square deviation, which is a measure of the discordance of the Cα atoms between protein structures. Based on the above results, we propose that these analytical approaches serve as efficient methods for the identification of causal variants of human disease-causing genes and elucidation of yeast-cell based molecular mechanisms.

Erythropoietin-producing Human Hepatocellular Carcinoma Receptor B1 Polymorphisms are Associated with HBV-infected Chronic Liver Disease and Hepatocellular Carcinoma in a Korean Population

Erythropoietin-producing human hepatocellular carcinoma receptor B1 (EPHB1) is a member of the Eph family of receptor tyrosine kinases that mediate vascular system development. Eph receptor overexpression has been observed in various cancers and is related to the malignant transformation, metastasis, and differentiation of cancers, including hepatocellular carcinoma (HCC). Eph receptors regulate cell migration and attachment to the extracellular matrix by modulating integrin activity. EphrinB1, the ligand of EPHB1, has been shown to regulate HCC carcinogenesis. Here, we sought to determine whether EPHB1 polymorphisms are associated with hepatitis B virus (HBV)-infected liver diseases, including chronic liver disease (CLD) and HCC. We genotyped 26 EPHB1 single nucleotide polymorphisms (SNPs) in 399 Korean CLD, HCC, and LD (CLD+HCC) cases and seroconverted controls (HBV clearance, CLE) using the GoldenGate assay. Two SNPs (rs6793828 and rs11717042) and 1 haplotype that were composed of these SNPs were associated with an increased risk for CLD, HCC, and LD (CLD+HCC) compared with CLE. Haplotypes that could be associated with HBV-infected liver diseases by affecting downstream signaling were located in the Eph tyrosine kinase domain of EPHB1. Therefore, we suggest that EPHB1 SNPs, haplotypes, and diplotypes may be genetic markers for the progression of HBV-associated acute hepatitis to CLD and HCC.