Polymorphisms of NRF2 and NRF2 target genes in urinary bladder cancer patients

Edyta Reszka 0 Zbigniew Jablonowski 0 Edyta Wieczorek 0 Ewa Jablonska 0 Magdalena Beata Krol 0 Jolanta Gromadzinska 0 Adam Grzegorczyk 0 Marek Sosnowski 0 Wojciech Wasowicz 0 0 Z. Jablonowski a. grzegorczyk M. sosnowski I Department of Urology, Medical University , Zeromskiego st. 113, 90-549 lodz, Poland 1 ) e. Wieczorek e. Jablonska M. B. Krol J. gromadzinska W. Wasowicz Department of t oxicology and Carcinogenesis, nofer Institute of Occupational Medicine , t eresy st. 8, 91-348 lodz, Poland Purpose nRF2 transcription factor is involved in modulation of various antioxidant and metabolic genes and, therefore, may modulate anti-carcinogenic potential. association between polymorphisms of NRF2 and five nRF2regulated genes and urinary bladder cancer (BC) risk was analyzed. Methods the study group included 244 BC patients, while the control group comprised 365 individuals with no evidence of malignancy. genotyping of GSTM1 (deletion), GSTT1 (deletion), GSTA1 69C/t (rs3957357), GSTP1 Ile105Val (rs1695), SOD2 ala16Val (rs4880) and NRF2 617C/a (rs6721961) in blood genomic Dna was performed by means of real-time PCR assays. the associations between gene polymorphism and BC risk were computed by logistic regression. Results the frequency of GSTA1, GSTP1, SOD2 and NRF2 genotypes did not differ in both groups. a significantly higher BC risk was associated with GSTM1 null genotype after adjusting to age, sex and smoking habit (OR 1.85, 95 % CI 1.30-2.62; P = 0.001). GSTT1 null (OR 0.50, 95 % CI 0.31-0.81; P = 0.005) and GSTP1 Val105Val (OR 0.52, 95 % CI 0.27-0.98; P = 0.04) genotypes were associated with reduced BC risk separately or in combination (OR 0.24, 95 % CI 0.11-0.51; P < 0.0001) (P heterogeneity = 0.01). Combined GSTT1 null and SOD2 with at least one 16Val allele among never smokers encompass reduced BC risk (OR 0.14, 95 % CI 0.03-0.63; P = 0.01) (P heterogeneity = 0.04). Conclusions this study supports hypothesis that GSTM1 null genotype may be a moderate BC risk factor. the genegene and gene-environment interactions associated with combined GSTP1/GSTT1 and combined GSTT1/SOD2 genetic polymorphisms along with cigarette smoking habit may play a significant role in BC risk modulation. - Urinary bladder cancer (BC) is a common disease with high prevalence in the developed countries in comparison with the rest of the world. BC occurs more frequently in men than in women, making it the fourth most common cancer among men and the eighth among women in europe (glOBOCan 2008). Incidence of BC in Poland is slightly lower than the average incidence in Western and southern europe, but it has been increasing rapidly. the majority (9095 %) of BCtransitional cell carcinomacomprises superficial tumors (70 %), which are usually low-grade and non-muscle-invasive bladder cancer (nMIBC) at the stage t a/t1, and the second one has the form of muscle-invasive disease (MIBC) at the stages from t2 to t4 (30 %). the overall rate of recurrence for nMIBC ranges from 60 to 70 %, and the overall rate of progression to a higher stage or grade and metastasis from 20 to 30 % (grotenhuis et al. 2010). several well-defined BC risk factors have been identified, including tobacco smoke (encompassing approximately 3050 % BC risk, similar BC in men and women), aromatic amines, polycyclic aromatic hydrocarbons (Pahs), dietary nitrites and nitrates, chlorinated hydrocarbons, coal, alkylating agents, arsenic, diesel engine exhaust. In addition, BC was one of the first cancers shown to be industrially associated with aniline dye industry (Cohen 1998; IaRC 2012; Volanis et al. 2010). BC risk has also been linked with coffee consumption and overall fluid consumption, including chlorinated water. the protective effect on BC risk was attributed to fruit and vegetables consumption and to high selenium status (Brinkman and Zeegers 2008). Chemically induced urinary bladder carcinogenesis in rodents showed the importance of nuclear factor (erythroid-derived 2)-like 2 (nRF2 or nFe2l2)-regulated signaling pathway (Iida et al. 2004, 2007; Jiang et al. 2009). Moreover, urinary bladder in rats was the most sensitive organ to gst enzyme induction after administration of isothiocyanates, the chemical compounds which contribute well-known nRF2 inductors (Munday and Munday 2002). Various studies have shown that nRF2 signaling pathway is the major mechanism, which controls the expression of target genes with antioxidant response element (aRe) sequence in their promoters. Under basal conditions, transcription factor nRF2 is localized in the cytoplasm and regulated by Kelch-like eCh-associated protein 1 (KeaP1). alteration of redox balance leads to nRF2 translocation to the nucleus and activation of aRe-containing genes. nRF2-modulated antioxidant that involved in xenobiotic metabolism enzymes may protect against oxidants and electrophilic agents and therefore may contribute to the enhancement of anti-carcinogenic activity (Kensler and Wakabayashi 2010; Maher and Yamamoto 2010). the role of specific nRF2-regulated metabolic and antioxidant genes in urinary bladder tissue has been intensively investigated. It was found that highly metastatic human bladder cells displayed significantly higher mitochondrial superoxide dismutase (sOD2) levels and activities compared with the non-metastatic parental cell line (hempel et al. 2009). several studies have shown that glutathione S-transferase P1 (gstP1), gstM1 and gstt1 are highly expressed in urinary bladder tissues and showed significantly higher activity and expression of these enzymes in bladder tumors than in normal uroepithelium (Pljesa-ercegovac et al. 2011; savic-Radojevic et al. 2007; simic et al. 2005). the fact that human urinary bladder tumors are characterized by up-regulation of NRF2 expression in comparison with adjacent non-cancer tissues is also worth attention (Kawakami et al. 2006). Individual differences in biotransformation of BC carcinogens and in scavenging of reactive oxygen and nitrogen species are quoted as one of the proposed mechanisms in BC etiology. It was observed that cytoprotective genes very often possess aRe sequence and therefore can be modulated by nRF2 transcription factor. For the last two decades, functional polymorphisms of GSTM1 and GSTT1 (gene deletion), GSTP1 Ile105Val (rs1695) affecting gene and enzyme expression, enzyme activity or substrate affinity have been analyzed in relation to BC risk in various ethnic groups. lack of the enzyme due to gene deletion was observed in case of GSTM1 (Fryer et al. 1993) and GSTT1 (Pemble et al. 1994). Minor GSTP1 105Val alleles can be associated with lower enzymatic gst activity than major GSTP1 105Ile alleles, due to changes in hydrophobic substrate active center (Watson et al. 1998). Functional significance of GSTA1 69C/t (rs3957357) polymorphism in promoter region results in differential expression with lower transcriptional activation and lower gst activity of minor GSTA1 69t allele tha (...truncated)