Decreased DNA repair gene expression among individuals exposed to arsenic in United States drinking water

Arsenic is well established as a human carcinogen, but its precise mechanism of action remains unknown. Arsenic does not directly damage DNA, but may act as a carcinogen through inhibition of DNA repair mechanisms, leading indirectly to increased mutations from other DNA damaging agents. The molecular mechanism underlying arsenic inhibition of nucleotide excision repair after UV irradiation (Hartwig et al., Carcinogenesis 1997;18:399-405) is unknown, but could be due to decreased expression of critical genes involved in nucleotide excision repair of damaged DNA. This hypothesis was tested by analyzing expression of repair genes and arsenic exposure in a subset of 16 individuals enrolled in a population based case-control study investigating arsenic exposure and cancer risk in New Hampshire. Toenail arsenic levels were inversely correlated with expression of critical members of the nucleotide excision repair complex, ERCC1 (r(2) = 0.82, p < 0.0001), XPF (r(2) = 0.56, p < 0.002), and XPB (r(2) = 0.75, p < 0.0001). The internal dose marker, toenail arsenic level, was more strongly associated with changes in expression of these genes than drinking water arsenic concentration. Our findings, based on human exposure to arsenic in a US population, show an association between biomarkers of arsenic exposure and expression of DNA repair genes. Although our findings need verification in a larger study group, they are consistent with the hypothesis that inhibition of DNA repair capacity is a potential mechanism for the co-carcinogenic activity of arsenic.     

XPF基因多态性与晚期非小细胞肺癌铂类联合化疗效果及不良反应的相关性研究

目的探讨XPF基因多态性与NSCLC铂类联合化疗效果及不良反应的相关性。方法选取NSCLC患者100例作为研究对象,铂类药物联合化疗后评价临床疗效和不良反应。应用Taqman检测技术对3个多态性位点的基因型分布进行分析。采用Logistic回归模型分析各基因型与NSCLC患者铂类联合化疗疗效及不良反应的相关性。结果XPF的rs28360317,相对于Ⅱ基因型患者,ID基因型的患者疗效较好(P<0.05);携带Ⅱ+ID基因型的患者疗效明显优于DD基因型(P<0.05)。对于rs1805377,GG基因型患者的疗效显著优于AA基因型(P<0.05)。对于rs2836007,TT基因型的患者疗效显著优于CC基因型患者(P<0.05)。对于rs28360317,ID、DD发生Ⅲ~Ⅳ级血液学毒性的几率显著高于Ⅱ级(P<0.05);对于rs2836007,CT患者发生Ⅲ~Ⅳ级血液学毒性的概率较CC患者低(P<0.05)。rs1805377位点的AA基因型发生Ⅲ~Ⅳ级胃肠道毒性的几率显著低于AG基因型(P<0.05)。结论XPF基因多态性与晚期NSCLC患者铂类联合化疗的疗效及不良反应相关,有一定的临床预测价值。     

Polymorphisms in ERCC1 and XPF gene and response to chemotherapy and overall survival of non-small cell lung cancer

We conducted a perspective study to assess the association between ERCC1 and XPF polymorphisms and response to chemotherapy and clinical outcome of NSCLC receiving chemotherapy. Between May 2009 and May 2011, a prospective study was conducted on 240 NSCLC cases. Genotypes of ERCC1 (rs11615, rs3212986 and rs2298881) and XPF (rs2276465 and rs6498486) were performed by Polymerase Chain Reaction Restriction Fragment Length Polymorphism (PCR-RFLP) assay. By conditional logistic regression analysis, patients carrying AA genotype of ERCC1 rs11615 showed more CR+PR to chemotherapy when compared with GG genotype, and the adjusted OR (95% CI) was 2.73 (1.21-6.18). By Cox regression analysis, AA genotype of ERCC1 rs11615 was associated with longer overall survival of NSCLC, and the adjusted HR (95% CI) was 0.38 (0.14-0.96). In conclusion, our study found that ERCC1 rs11615 polymorphism can influence the chemotherapy response and overall survival of NSCLC patients receiving cisplatin-based chemotherapy.     

The age-related expression decline of ERCC1 and XPF for forensic age estimation: A preliminary study

The age-related capacity decline of DNA damage repair in human peripheral blood has been demonstrated. Excision repair cross-complementation group1 (ERCC1) and Xeroderma pigmentosum complementation group F (XPF) were rate-limiting enzyme in nucleotide excision repair (NER) which was known as the most important DNA damage repair system. Consequently, we hypothesized that the expression and/or activity of ERCC1 and XPF may be associated with age. However, little was known about the quantitative relationship of ERCC1 and XPF expression levels with age. The aim of the present study was to analyze the correlation of ERCC1 and XPF expression levels with age by detecting the ERCC1 and XPF mRNA levels in peripheral blood mononuclear cells(PBMCs) and protein levels in plasma in healthy ethnic Han Chinese individuals, and finally find new molecular markers for forensic age estimation by establishing the mathematical model between ERCC1 and XPF expression levels and age. The results showed that the ERCC1 and XPF mRNA relative expression levels in PBMCs declined in an age-dependent manner (r = −0.578/−0.844, respectively, P < 0.01). The formula for age estimation based on the ERCC1 and XPF mRNA relative expression levels decline in PBMCs were Y = 3.3E−5x²−0.0261x+1.9175 (R² = 0.3244, P < 0.01) and Y = 0.0003x²−0.0459x+2.0439 (R² = 0.729, P < 0.01), respectively. There were no significant differences of the ERCC1 or XPF protein expression levels in plasma between age groups (P > 0.05). Furthermore, there were no significant differences of the ERCC1 or XPF mRNA and/or protein expression levels between males and females(P > 0.05). It suggested that the ERCC1 and XPF mRNA expression levels could be considered as valuable additional tool in individual age estimation, especially in cases where traditional morphologic method was inefficient or absent in forensic practice.     

Reduced levels of XPA, ERCC1 and XPF DNA repair proteins in testis tumor cell lines

Over 80% of patients with advanced metastatic testis tumors can be cured using cisplatin-based combination chemotherapy. This is unusual as metastatic cancer in adults is usually incurable. Cell lines derived from testis tumors retain sensitivity to cisplatin in vitro. We previously investigated 2 testis tumor cell lines with a low capacity to remove cisplatin-induced DNA damage and found that they had low levels of the DNA nucleotide excision repair proteins XPA, ERCC1 and XPF. To determine whether low levels of XPA, ERCC1 and XPF proteins are characteristic of testis tumor cell lines, we investigated 35 cell lines derived from cancers to determine whether groups of cell lines from diverse tissue origins differ from one another in constitutive levels of these NER proteins. Quantitative immunoblotting was used to compare groups of cell lines representing prostate, bladder, breast, lung, cervical, ovarian and testis cancers. Only the 6 testis tumor cell lines showed significantly lower mean levels of XPA (p = 0.001), XPF (p = 0.001) and ERCC1 (p = 0.004) proteins from the other groups. Our results encourage further investigation of the possibility that low levels of these nucleotide excision repair proteins could be related to the favorable response of testis tumors to cisplatin-based chemotherapy.     

Computer‐aided drug design of small molecule inhibitors of the ERCC1‐XPF protein–protein interaction

The heterodimer of DNA excision repair protein ERCC‐1 and DNA repair endonuclease XPF (ERCC1‐XPF) is a 5′–3′ structure‐specific endonuclease essential for the nucleotide excision repair (NER) pathway, and it is also involved in other DNA repair pathways. In cancer cells, ERCC1‐XPF plays a central role in repairing DNA damage induced by chemotherapeutics including platinum‐based and cross‐linking agents; thus, its inhibition is a promising strategy to enhance the effect of these therapies. In this study, we rationally modified the structure of F06, a small molecule inhibitor of the ERCC1‐XPF interaction (Molecular Pharmacology, 84 , 2013 and 12), to improve its binding to the target. We followed a multi‐step computational approach to investigate potential modification sites of F06, rationally design and rank a library of analogues, and identify candidates for chemical synthesis and in vitro testing. Our top compound, B5 , showed an improved half‐maximum inhibitory concentration (IC₅₀) value of 0.49 µM for the inhibition of ERCC1‐XPF endonuclease activit, and lays the foundation for further testing and optimization. Also, the computational approach reported here can be used to develop DNA repair inhibitors targeting the ERCC1‐XPF complex.     

XPF基因多态性与晚期非小细胞肺癌铂类联合化疗效果及不良反应的相关性研究

目的探讨XPF基因多态性与NSCLC铂类联合化疗效果及不良反应的相关性。方法选取NSCLC患者100例作为研究对象,铂类药物联合化疗后评价临床疗效和不良反应。应用Taqman检测技术对3个多态性位点的基因型分布进行分析。采用Logistic回归模型分析各基因型与NSCLC患者铂类联合化疗疗效及不良反应的相关性。结果XPF的rs28360317,相对于Ⅱ基因型患者,ID基因型的患者疗效较好(P<0.05);携带Ⅱ+ID基因型的患者疗效明显优于DD基因型(P<0.05)。对于rs1805377,GG基因型患者的疗效显著优于AA基因型(P<0.05)。对于rs2836007,TT基因型的患者疗效显著优于CC基因型患者(P<0.05)。对于rs28360317,ID、DD发生Ⅲ~Ⅳ级血液学毒性的几率显著高于Ⅱ级(P<0.05);对于rs2836007,CT患者发生Ⅲ~Ⅳ级血液学毒性的概率较CC患者低(P<0.05)。rs1805377位点的AA基因型发生Ⅲ~Ⅳ级胃肠道毒性的几率显著低于AG基因型(P<0.05)。结论XPF基因多态性与晚期NSCLC患者铂类联合化疗的疗效及不良反应相关,有一定的临床预测价值。     

Expression of domains for protein–protein interaction of nucleotide excision repair proteins modifies cancer cell sensitivity to platinum derivatives and genomic stability

Nucleotide excision repair (NER) is involved in the repair of DNA damage caused by platinum derivatives and has been shown to decrease the cytotoxic activity of these drugs. Because protein–protein interactions are essential for NER activity, we transfected human cancer cell lines (A549 and HCT116) with plasmids coding the amino acid sequences corresponding to the interacting domains between excision repair cross‐complementation group 1 (ERCC1) and xeroderma pigmentosum, complementation group A (XPA), as well as ERCC1 and xeroderma pigmentosum, complementation group F (XPF), all NER proteins. Using the 3‐(4,5‐dimethyl‐2 thiazoyl)‐2,5‐diphenyl‐2H‐tetrazolium bromide (MTT) assay and annexin V staining, we showed that transfected A549 cells were sensitized 1.2–2.2‐fold to carboplatin and that transfected HCT116 cells were sensitized 1.4–5.4‐fold to oxaliplatin in vitro. In addition, transfected cells exhibited modified in vivo sensitivity to the same drugs. Finally, in particular cell models of the interaction between ERCC1 and XPF, DNA repair was decreased, as evidenced by increased phosphorylation of the histone 2AX after exposure to mitomycin C, and genomic instability was increased, as determined by comparative genomic hybridization studies. The results indicate that the interacting peptides act as dominant negatives and decrease NER activity through inhibition of protein–protein interactions.