| Partâ… :A Functional Polymorphism in the Promoter Region of GSTM1 Implies a Novel Two-sided Role for GSTM1 in Breast Cancer SusceptibilityEstrogen-quinone can form unstable adducts with adenine and guanine in DNA,leading to depurination and mutation in vitro and in vivo.It seems that reactive equine estrogen metabolites mainly contribute to breast cancer.However,protective phaseâ…¡enzymes are active in breast tissue for protection against damage caused by reactive metabolites of endogenous and exogenous chemicals.The estrogen-quinone could be reduced by quinone oxidoreductases,or be conjugated by glutathione S-transferases.Therefore,quinone oxidoreductases and glutathione S-transferases are two kinds of important estrogen-quinone metabolizing related enzymes. Although a number of studies have been conducted to address the relationship between a gene deletion-polymorphism of glutathione-S-transferase M1(GSTM1) and breast cancer,no definite conclusion has been made and no clear risk pattern has yet to emerge for GSTM1.We first conducted case-control studies that included 1920 subjects using a true genotyping method.The results show that GSTM1-null(GSTM1-/-) confers an increased risk for breast cancer development compared with GSTM1-present individuals[homozygous wild-type(+/+) and heterozygous(+/-)],which was subsequently confirmed by a meta-analysis of all of the 41 relevant studies(odds ratio:1.10,P<0.001).Unexpectedly,we found that GSTM1+/+ is also a risk genotype compared with GSTM1+/-.Furthermore,we identified a functional polymorphism in the GSTM1 promoter region associated with breast cancer.The variant allele modifies the DNA-binding to the AP-2αtranscription factor,resulting in reduced promoter activity and mRNA expression.However,this low-activity allele is associated with reduced breast cancer risk. It seems that approximately 60-70%expression from one allele of GSTM1 could suffice for protection against breast cancer;null- and over-activity of GSTM1 are both disadvantageous. These results indicate a U-shaped association of GSTM1 with breast cancer,which challenges the linear gene-dosage effect of GSTM1 that was previously proposed.We recommend that a more complicated role for GSTM1 should be considered in breast cancer risk prediction.Partâ…¡:Genetic Variants in M3-like Gene within GSTM4-GSTM2-GSTM1-GSTM5-GSTM3 Cluster Influence Breast Cancer Susceptibility Depending on GSTM1Mu class GSTMs genes arrange in a tandem on chromosome 1p13.3.The previous single-variant-based findings warrant more comprehensive study to explore the relationship between genetic variants in GSTM1-5 gene cluster and breast cancer.In the present study, seventeen tagging single-nucleotide polymorphisms(SNPs) covering 64-kb of GSTMs cluster (excluding recombination-regions) are originally selected and genotyped in a pilot population. Eleven validated-SNPs enter large sample size genotyping in 847-cases and 676-controls.The association analyses are performed according to the absence or presence of GSTM1.In GSTM1-/- group,the allele frequency of one SNP in M3-like loci is significantly different between cases and controls(P=2.0×10-4,corrected P=0.001),with odds ratio of 1.75(95% confidence interval:1.26-2.44) for variant genotype compared with wild-type genotype.Two haplotypes are also significantly linked with breast cancer(corrected P=0.044 and 0.010).In contrast,no susceptible allele/haplotype is identified when GSTM1 is present.Based on epidemiological observations,we further conduct in vitro and ex vivo studies and identify two genetic variants in M3-like loci accounting for differential expression of GSTM3 in normal breast tissues by such means as altering binding of RNA-pol-â…¡.Protective genotypes demonstrated in association study stage are correlated with higher GSTM3 expression phenotypes.In conclusion,the variants/haplotypes in M3-like loci rather than in M1-like loci within GSTMs cluster are likely to contribute to breast cancer risk when GSTM1 is absent.Our study also indicates that increased GSTM3 catalyzing ability in normal breast tissue might protect against breast oncogenesis.Partâ…¢:Functional Polymorphisms,Altered Gene Expression and Genetic Association Link NRH:Quinone Oxidoreductase 2 to Breast CancerWe hypothesized that NRH:quinone oxidoreductase 2(NQO2) is a candidate susceptibility gene for breast cancer because of its known enzymatic activity on estrogen-derived quinones and its ability to stabilize p53.We performed case-control studies to investigate the contributions of genetic variants/haplotypes of the NQO2 gene to breast cancer risk.In the first hospital-based study(n=1,604),we observed significant associations between the incidence of breast cancer and a 29 bp-insertion/deletion polymorphism(29bp-I/D) and the rs2071002(+237A＞C) polymorphism,both of which are located within the NQO2 promoter region.Decreased risk was associated with the D-allele of 29bp-I/D(odds ratio(OR),0.76;P=0.0027) and the +237C-allele of rs2071002(OR,0.80;P=0.0031).Specifically,the susceptibility variants within NQO2 were notably associated with breast carcinomas with wild-type p53(the most significant P-value: 3.3×10-6).The associations were successfully replicated in an independent population set (familial/early-onset breast cancer cases and community-based controls,n=1,442).The combined P-values of the two studies(n=3,046) are 3.8×10-7 for 29bp-I/D and 2.3×10-6 for rs2071002. Furthermore,we revealed potential mechanisms of pathogenesis of the two susceptibility polymorphisms.Previous work has demonstrated that the risk-allele I-29 of 29bp-I/D introduces transcriptional-repressor Sp3 binding sites.Using promoter reporter-gene assays and electrophoretic-mobility-shift assays,our present work demonstrated that the other risk-allele, +237A-allele of rs2071002,abolishes a transcriptional-activator Sp1 binding site.Furthermore, an ex vivo study showed that normal breast tissues harboring protective genotypes expressed significantly higher levels of NQO2 mRNA than those in normal breast tissues harboring risk genotypes.Taken together,the data presented here strongly suggest that NQO2 is a susceptibility gene for breast carcinogenesis.
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