The roles of glutathione S‑transferase pi 1 (GSTP1), glutathione S‑transferase mu 2 (GSTM2) and glutathione S‑transferase alpha 1 (GSTA1) in cisplatin (DDP)‑resistance of solid cancer cells (A549/DDP, SKOV3/DDP and SGC7901/DDP) were compared following expression downregulation with small interfering RNAs (siRNAs). DDP cytotoxicity was reflected by its half maximal inhibition concentration (IC50) calculated from data using a Cell Counting Kit‑8 assay; cell apoptosis was examined using flow cytometry and Hoechst 33342 staining. Higher activities of GST were detected in the cytosol of DDP‑resistant cells, compared with those in the parental DDP‑susceptible cells. The silencing efficacy of each positive siRNA was supported by western blot analysis. GSTP1 silencing resulted in a 4‑fold sensitization of SGC7901/DDP cells to DDP cytotoxicity, but negligible sensitization of SKOV3/DDP and A549/DDP cells. GSTM2 silencing sensitized SKOV3/DDP and A549/DDP cells to DDP cytotoxicity by ~2‑fold, but did not sensitize SGC7901/DDP cells. Notably, GSTA1 silencing enhanced DDP cytotoxicity in SGC7901/DDP cells by 6‑fold, in A549/DDP cells by 5‑fold and in SKOV3/DDP cells by 2‑fold. The combined actions of positive siRNAs and DDP increased the percentages of apoptotic cells in the DDP‑resistant solid cancer cells compared with the combined actions of DDP and the negative siRNAs. The present findings indicated that GSTA1 is a predominant GST isozyme associated with DDP resistance of SGC7901/DDP, A549/DDP and SKOV3/DDP cells; GSTA1‑specific inhibitors may be general sensitizers of SGC7901/DDP, A549/DDP and SKOV3/DDP cells to DDP cytotoxicity through the promotion of cell apoptosis.

Context: KCNJ5 mutation is a major cause of aldosterone-producing adenomas (APAs). The development of APA apart from KCNJ5 mutation is less investigated.
Objective: To investigate other mechanisms affecting aldosterone secretion apart from KCNJ5.
Patients and Methods: Six pairs of KCNJ5-mutated, high and low aldosterone-secreting APAs, five non-KCNJ5-mutated APAs, and four normal adrenal glands were assayed by Affymetrix GeneChip Human Transcriptome Array 2.0. A total of 113 APA samples were investigated to explore the expression of glutathione-S-transferase A1 (GSTA1). H295R cells were used to verify the function of GSTA1.
Results: GSTA1 was the top gene downregulated in high-aldosterone KCNJ5-mutated APAs. GSTA1 was also downregulated in KCNJ5-mutated APAs compared with wild-type KCNJ5 APAs. Accordingly, mutant KCNJ5 decreased GSTA1 messenger RNA and protein expression levels. GSTA1 overexpression suppressed aldosterone secretion whether in wild-type or mutant KCNJ5 H295R cells. Adding ethacrynic acid or silencing of GSTA1 increased aldosterone secretion by increasing reactive oxygen species (ROS), superoxide, H2O2 levels, and Ca2+ influx. The expression of the transcription factors NR4A1, NR4A2, and CAMK1 and intracellular Ca2+ were significantly upregulated by GSTA1 inhibition. The reduced form of NAD phosphate oxidase inhibitor or H2O2 scavenger or blocking calmodulin or calcium channels could significantly reduce aldosterone secretion in GSTA1-inhibited cells.
Conclusions: (1) GSTA1 expression is reversely correlated with aldosterone level in KCNJ5-mutated APAs, (2) GSTA1 regulates aldosterone secretion by ROS and Ca2+ signaling, and (3) KCNJ5 mutation downregulates GSTA1 expression, and overexpression of GSTA1 reverses increased aldosterone in KCNJ5-mutated adrenal cells.

Cytochrome P450 (CYP), glutathione-S-transferase (GST), and N-acetyltransferase (NAT) are crucial for metabolism and clearance of xenobiotics. This study investigated whether CYP, GST, and NAT single nucleotide polymorphisms (SNPs) are associated with colorectal cancer (CRC) in patients with Lynch syndrome. The interaction between these SNPs and cigarette smoking or meat consumption was also explored. We identified 270 patients with Lynch syndrome from the Taiwan Hereditary Nonpolyposis Colorectal Cancer Consortium. A weighted Cox proportional hazard model was used to calculate the hazard ratios (HRs) and 95% confidence interval (CIs). The GSTA1 rs3957356 TT (HR = 5.36, 95% CI = 2.39-12.0) and CYP1B1 rs1056836 CC (HR = 7.24, 95% CI = 3.51-14.9) were significantly associated with CRC risk when compared to wild-type CC and GG genotypes, respectively. However, the CYP1A1 rs4646903 CC genotype significantly reduced the risk of CRC (HR = 0.33, 95% CI = 0.12-0.89) when compared to TT genotype. Moreover, significant interactions were observed between NAT1 acetylation and CYP1B1 rs1056827 and meat consumption.Our results suggest that xenobiotic-metabolizing SNPs are not only associated with CRC risk in patients with Lynch syndrome in Taiwan but also interact with meat consumption to modify the disease risk. Environ. Mol. Mutagen. 59:69-78, 2018. © 2017 Wiley Periodicals, Inc.

The aim of the present study was to investigate the prognostic value of genes that participate in the development of gastric adenocarcinoma, via exploring gene cross talk in disease‑related pathways. Differentially expressed genes (DEGs) in the gastric samples were identified by analyzing the expression data downloaded from the GEO database. The DEGs were subjected to the human protein‑protein interaction (PPI) network to construct the PPI network of DEGs, which was then used for the identification of key genes in cancer samples via the expression deviation score and degree in the network. A total of 635 DEGs, including 432 downregulated and 203 upregulated ones were screened in the gastric adenocarcinomas samples. The PPI network of DEGs comprised 590 DEGs and 4,299 interaction pairs. A total of 200 key genes were obtained, which were significantly enriched in six downregulated and six upregulated pathways. Cross talk genes in the connected pathways were analyzed, and the Kyoto Encyclopedia of Genes and Genomes pathways hsa00980 (Metabolism of xenobiotics by cytochrome P450) and hsa00982 (Drug metabolism) were reported to share 8 cross talk genes: ADH7, ALDH3A1, GSTA1, GSTA2, UGT2B17, UGT2B10, ADH1B and CYP2C18. Among all cross talk genes, ADH7, ALDH3A1 and CLDN3 were the most specific genes. The high‑ and low‑risk samples identified by the prognosis model presented a remarkable difference in total survival time, indicating its robustness and sensitivity as the prognosis genes for gastric adenocarcinoma. ADH7, ALDH3A1, GSTA1, GSTA2, UGT2B17, UGT2B10, ADH1B, CYP2C18ADH7, ALDH3A1 and CLDN3 may be used as the prognosis markers and target biomarkers for chemotherapies in gastric adenocarcinoma.

PURPOSE: Since several studies have proposed that epithelial ovarian cancer should not be considered as a single disease entity and that it results from an accumulation of genetic changes, we aimed to assess the polymorphic expression of major cytosolic glutathione S-transferases (GSTM1, T1, A1 and P1) with respect to ovarian cancer susceptibility and aggressiveness.
METHODS: This case-control study was conducted on 93 newly diagnosed epithelial ovarian cancer patients and 178 healthy matched controls. The multiplex polymerase chain reaction (PCR) was used to detect homozygous deletions of GSTM1 and GSTT1 genes. Analysis of the single nucleotide polymorphism (SNP) GSTA1 C69T was performed using PCR-restriction fragment length polymorphism (RFLP), while for SNP GSTP1 Ile105Val real-time PCR was used.
RESULTS: No significant association to ovarian cancer risk was found for individual GSTM1, GSTA1 and GSTP1 genotypes (p>0.05). However, the carriers of GSTT1-active genotype were at 2-fold higher risk of ovarian cancer development (95%CI: 1.00-4.01, p=0.049), which was even more elevated in the subgroup of patients with positive family history of cancer. Moreover, the frequency of all three GST genotypes that might be associated to ovarian cancer risk (GSTT1-active, GSTA1-active and GSTP1-referent) was significantly higher in patients than in the control group (p=0.042). Even more, patients who were carriers of combination of these three genotypes represented over 64% of the total number of patients within any of the International Federation of Gynecology and Obstetrics (FIGO) stages of ovarian cancer.
CONCLUSIONS: This study provides supportive evidence that GSTs might affect both susceptibility and progression of ovarian cancer.

The advent of imatinib mesylate (IM) has dramatically improved the outcome of patients with chronic myeloid leukemia, but drug resistance, particularly in advanced stage of disease, portents eventual relapse and progression. To identify the candidate molecule responsible for resistance during IM treatment, an IM-resistant K562 cell line was generated by culturing in gradually increasing dose of IM. The expression of Nrf-2 and its downstream target, Gst-α, were significantly induced in these cells. GST-α, in turn, mediated cell survival by maintaining intracellular low level of 4-HNE. Inhibition of Nrf-2 effectively reduced the expression of Gst-α, resulting in accumulation of 4-HNE and elevated sensitiveness to IM. Moreover, in IM-sensitive K562 cells enforced Gst-α expression strikingly protected cells from the insult of IM. Finally, we also examined the levels of Nrf-2 in clinical bone morrow samples. Nrf-2 and Gst-α were more abundant in bone morrow of CML patients compared with that of healthy donors. In addition, Nrf-2 and Gst-α were further up-regulated in samples of patients with weak response to IM. In conclusion, our study shows that rapid clearance of 4-HNE by Nrf-2/GST may represents a novel molecular basis of IM resistance in CML.

Reactive oxygen species (ROS), formed as an indirect production of radiotherapy (RT), could cause DNA damage of normal tissues. Meanwhile, our body possesses the ability to restore the damage by DNA repair pathways. The imbalance between the two systems could finally result in radiation injury. Therefore, in this prospective cohort study, we explored the association of genetic variants in ROS metabolism and DNA repair pathway-related genes with radiation pneumonitis (RP). A total of 265 locally advanced esophageal squamous cell carcinoma (ESCC) patients receiving RT in Chinese Han population were enrolled. Five functional single nucleotide polymorphisms (SNPs) (rs1695 in GSTP1; rs4880 in SOD2; rs3957356 in GSTA1; and rs1801131, rs1801133 in MTHFR) were genotyped using the MassArray system, and rs1801131 was found to be a predictor of ≥ 2 RP. Our results showed that, compared with TT genotype, patients with GG/GT genotypes of rs1801131 had a notably lower risk of developing ≥ 2 RP (HR = 0.339, 95% CI = 0.137-0.839, P = 0.019). Further independent studies are required to confirm this findings.

Glutathione S-transferases (GSTs) detoxify toxic molecules by conjugation with reduced glutathione and regulate cell signaling. Single nucleotide polymorphisms (SNPs) of GST genes have been suggested to affect GST functions and thus to increase the risk of human hepatocellular carcinoma (HCC). As GSTA1 is expressed in hepatocytes and the rs3957357C>T (TT) SNP is known to downregulate GSTA1 mRNA expression, the aims of this study were: (i) to explore the relationship between the TT SNP in GSTA1 and the occurrence of HCC; (ii) to measure GSTA1 mRNA expression in HCCs. For that purpose, we genotyped non-tumor-tissue-derived DNA from 48 HCC patients and white-blood-cell-derived DNA from 37 healthy individuals by restriction fragment length polymorphism (RFLP). In addition, expression of GSTA1 mRNA was assessed by real-time PCR in 18 matching pairs of HCCs and non-tumor livers. Survival analysis was performed on an annotated microarray dataset containing 247 HCC patients (GSE14520). The GSTA1 TT genotype was more frequent in HCC than in non-HCC patients (27% versus 5%, respectively), suggesting that individuals carrying this genotype could be associated with 2-fold higher risk of developing HCCs (odds ratio = 2.1; p = 0.02). Also, we found that GSTA1 mRNA expression was lower in HCCs than in non-tumor livers. HCCs expressing the highest GSTA1 mRNA levels were the smallest in size (R = -0.67; p = 0.007), expressed the highest levels of liver-enriched genes such as ALB (albumin, R = -0.67; p = 0.007) and COL18A1 (procollagen type XVIII, R = -0.50; p = 0.03) and showed the most favorable disease-free (OR = 0.54; p<0.001) and overall (OR = 0.56; p = 0.006) outcomes. Moreover, GSTA1 was found within a 263-gene network involved in well-differentiated hepatocyte functions. In conclusion, HCCs are characterized by two GSTA1 features: the TT SNP and reduced GSTA1 gene expression in a context of hepatocyte de-differentiation.

Doxorubicin (DOX), one of the most effective anticancer drugs, acts in a variety of ways including DNA damage, enzyme inhibition and generation of reactive oxygen species. Glutathione (GSH) and glutathione-related enzymes including: glutathione peroxidase (GPX), glutathione reductase (GSR) and glutathione S-transferases (GST) may play a role in adaptive detoxification processes in response to the oxidative stress, thus contributing to drug resistance phenotype. In this study, we investigated effects of DOX treatment on expression and activity of GSH-related enzymes and multidrug resistance-associated proteins in cultured human cervical cancer cells displaying different resistance against this drug (HeLa and KB-V1). Determination of expression level of genes encoding GST isoforms and MRP proteins (GCS, GPX, GSR, GSTA1-3, GSTM1, GSTP1, ABCC1-3, MGST1-3) was performed using StellARray™ Technology. Enzymatic activities of GPX and GSR were measured using biochemical methods. Expression of MRP1 was examined by immunofluorescence microscopy. This study showed that native expression levels of GSTM1 and GSTA3 were markedly higher in KB-V1 cells (2000-fold and 200-fold) compared to HeLa cells. Resistant cells have also shown significantly elevated expression of GSTA1 and GSTA2 genes (200-fold and 50-fold) as a result of DOX treatment. In HeLa cells, exposure to DOX increased expression of all genes: GSTM1 (7-fold) and GSTA1-3 (550-fold, 150-fold and 300-fold). Exposure to DOX led to the slight increase of GCS expression as well as GPX activity in KB-V1 cells, while in HeLa cells it did not. Expression of ABCC1 (MRP1) was not increased in any of the tested cell lines. Our results indicate that expression of GSTM1 and GSTA1-3 genes is up-regulated by DOX treatment and suggest that activity of these genes may be associated with drug resistance of the tested cells. At the same time, involvement of MRP1 in DOX resistance in the given experimental conditions is unlikely.

The aim of this study was to evaluate specific glutathione S-transferase (GST) gene variants as determinants of risk in patients with clear cell renal cell carcinoma (cRCC), independently or simultaneously with established RCC risk factors, as well as to discern whether phenotype changes reflect genotype-associated risk. GSTA1, GSTM1, GSTP1 and GSTT1 genotypes were determined in 199 cRCC patients and 274 matched controls. Benzo(a)pyrene diolepoxide (BPDE)-DNA adducts were determined in DNA samples obtained from cRCC patients by ELISA method. Significant association between GST genotype and risk of cRCC development was found for the GSTM1-null and GSTP1-variant genotype (p = 0.02 and p<0.001, respectively). Furthermore, 22% of all recruited cRCC patients were carriers of combined GSTM1-null, GSTT1-active, GSTA1-low activity and GSTP1-variant genotype, exhibiting 9.32-fold elevated cRCC risk compared to the reference genotype combination (p = 0.04). Significant association between GST genotype and cRCC risk in smokers was found only for the GSTP1 genotype, while GSTM1-null/GSTP1-variant/GSTA1 low-activity genotype combination was present in 94% of smokers with cRCC, increasing the risk of cRCC up to 7.57 (p = 0.02). Furthermore, cRCC smokers with GSTM1-null genotype had significantly higher concentration of BPDE-DNA adducts in comparison with GSTM1-active cRCC smokers (p = 0.05). GSTM1, GSTT1, GSTA1 and GSTP1 polymorphisms might be associated with the risk of cRCC, with special emphasis on GSTM1-null and GSTP1-variant genotypes. Combined GSTM1-null, GSTT1-active, GSTA1 low activity and GSTP1-variant genotypes might be considered as "risk-carrying genotype combination" in cRCC.

NRF2 stabilizes redox potential through genes for glutathione and thioredoxin antioxidant systems. Whether blockade of glutathione and thioredoxin is useful in eliminating cancer stem cells remain unknown. We used xenografts derived from colorectal carcinoma patients to investigate the pharmacological inhibition of glutathione and thioredoxin systems. Higher expression of five glutathione S-transferase isoforms (GSTA1, A2, M4, O2, and P1) was observed in xenograft-derived spheroids than in fibroblasts. Piperlongumine (2.5-10 μmol/L) and auranofin (0.25-4 μmol/L) were used to inhibit glutathione S-transferase π and thioredoxin reductase, respectively. Piperlongumine or auranofin alone up-regulated the expression of NRF2 target genes, but not TP53 targets. While piperlongumine showed modest cancer-specific cell killing (IC50 difference between cancer spheroids and fibroblasts: P = 0.052), auranofin appeared more toxic to fibroblasts (IC50 difference between cancer spheroids and fibroblasts: P = 0.002). The synergism of dual inhibition was evaluated by determining the Combination Index, based on the number of surviving cells with combination treatments. Molar ratios indicated synergism in cancer spheroids, but not in fibroblasts: (auranofin:piperlongumine) = 2:5, 1:5, 1:10, and 1:20. Cancer-specific cell killing was achieved at the following drug concentrations (auranofin:piperlongumine): 0.25:2.5 μmol/L, 0.5:2.5 μmol/L, or 0.25:5 μmol/L. The dual inhibition successfully decreased CD44v9 surface presentation and delayed tumor emergence in nude mouse. However, a small subpopulation persistently survived and accumulated phosphorylated histone H2A. Such "persisters" still retained lesser but significant tumorigenicity. Thus, dual inhibition of glutathione S-transferase π and thioredoxin reductase could be a feasible option for decreasing the tumor mass and CD44v9-positive fraction by disrupting redox regulation.

BACKGROUND: We carried out a test whether or not transplant drugs such as cyclosporine A, Rapamycin (Sirolimus), Tacrolimus, Everolimus and Mycophenolate mofetil affects the expression of phase II enzymes comprising of UDP-glucuronosyltransferases (UGTs) and glutathione-S-transferases (GSTs), and antioxidant enzymes that consist of glutathione reductase (GSR), glutathione peroxidase 1 (GPX1) and heme-oxygenase 1 (HMOX1).
METHODS: Experiments were performed in primary cultures of human hepatocytes and in human hepatocarcinoma HepG2 cells, the models of metabolically competent and incompetent cells, respectively. We used quantitative real-time PCR.
RESULTS: We found that none of the tested compounds affected the expression of investigated genes in human hepatocytes. On the other hand, Mycophenolate mofetil induced GPX1 mRNA, although it suppressed mRNA level of UGT1A4/1A9/2B7/2B10, GSTA1/O1/T1, GSR and HMOX1 in HepG2 cells.
CONCLUSION: We showed that the tested transplant drugs have no effect on the expression of selected phase II and antioxidant enzymes in human hepatocytes. Nevertheless, the experiments carried out in two common and frequently used in vitro cellular models, we emphasize that finding based solely on carcinoma cells must be taken with caution when transposing to in vivo situations.

Curzerene is a sesquiterpene and component used in oriental medicine. It was originally isolated from the traditional Chinese herbal medicine

PURPOSE: Reactive oxygen species (ROS) are generated as an indirect product of radiation therapy (RT). Genetic variation in genes related to ROS metabolism may influence the level of RT-induced adverse effects. We evaluated the potential association of single nucleotide polymorphism (SNP)-related response to radiotherapy injury in breast cancer patients undergoing RT.
MATERIALS AND METHODS: Eighty patients receiving conventional RT were included. Acute effects were evaluated according to the Radiation Therapy Oncology Group (RTOG) scores. DNA was extracted from blood and buccal swab samples. SNPs were genotyped for GSTP1, GSTA1, SOD2, and NOS3 genes by polymerase chain reaction-based restriction fragment length polymorphism. Univariate analysis (odds ratios [ORs] and 95% confidence interval [CI]) and principal component analysis were used for correlation of SNPs and factors related to risk of developing ≥ grade 2 acute effects.
RESULTS: Sixty-five patients (81.2%) showed side effects, 32 (40%) presented moderate to severe acute skin toxicity, and 33 (41.2%) manifested minimal acute skin reactions by the end of treatment. In both univariate and multivariate analyses, nominally significant associations were found among body mass index (OR, 3.14; 95% CI, 8.5338 to 1.1274; p=0.022), breast size (OR, 5.11; 95% CI, 17.04 to 1.54; p=0.004), and grade ≥ 2 acute radiation skin toxicity. A significant association was also observed between NOS3 G894T polymorphism (OR, 9.8; 95% CI, 211.6 to 0.45; p=0.041) and grade ≥ 2 acute radiation skin toxicity in patients with neo-adjuvant chemotherapy treatment.
CONCLUSION: The analysis of the factors involved in individual radiosensitivity contributed to the understanding of the mechanisms underlying this trait.

AIM: To investigate the effects of single nucleotide polymorphisms (SNPs) in glutathione S-transferase (GST) genes on survival of hepatocellular carcinoma (HCC) patients.
METHODS: Twelve tagging SNPs in GST genes (including GSTA1, GSTA4, GSTM2, GSTM3, GSTO1, GSTO2 and GSTP1) were genotyped using Sequenom MassARRAY iPLEX genotyping method in a cohort of 214 Chinese patients with resected HCC. The Cox proportional hazards model and log-rank test were performed to determine the SNPs related to outcome. Additionally, stratified analysis was performed at each level of the demographic and clinical variables. An SNP-gene expression association model was further established to investigate the correlation between SNP and gene expression.
RESULTS: Two SNPs (GSTO2: rs7085725 and GSTP1: rs4147581) were significantly associated with overall survival in HCC patients (P = 0.035 and 0.042, respectively). In stratified analysis, they were more significantly associated with overall survival in patients with younger age, male gender and cirrhosis. We further investigated cumulative effects of these two SNPs on overall survival in HCC patients. Compared with the patients carrying no unfavorable genotypes, those carrying 2 unfavorable genotypes had a 1.70-fold increased risk of death (P < 0.001). The cumulative effects were more significant in those patients with younger age, male gender and cirrhosis (HR = 2.00, 1.94 and 1.97, respectively; all P < 0.001). Additionally, we found that heavy smoking resulted in a significantly worse overall survival in those patients carrying variant alleles of rs7085725 (HR = 2.07, 95%CI: 1.13-3.76, P = 0.018). The distributions of GSTO2: rs7085725 and GSTP1: rs4147581 genotypes were associated with altered gene expression and contributed to influences on overall survival.
CONCLUSION: Our study provides the first evidence that GSTO2 and GSTP1 gene polymorphisms may serve as independent prognostic markers for HCC patients.

PURPOSE: Glutathione S-transferases (GSTs) are involved in the detoxification of carcinogens, and may be linked to carcinogenesis. As a vital component of GSTs, GSTA1 plays an important role in carcinogenesis. However, the studies about the effect of GSTA1 polymorphisms on cancer risk are limited and the conclusions are contradictory. This meta-analysis aimed to evaluate the association between GSTA1 polymorphisms (-567T>G, (69C>T and -52G>A) and cancer risk.
METHODS: A literature search of PubMed and Web of Science databases was conducted from their inception through December 2013. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated to assess the association of GSTA1 polymorphisms and cancer risk.
RESULTS: A total of 15 studies were enrolled, and the results indicated that GSTA1 BB genotype was associated with elevated cancer risk, especially in colorectal cancer. Further stratifications showed that GSTA1 BB genotype was associated with increased cancer risk in Caucasian populations and in the study with population-based controls.
CONCLUSIONS: This meta-analysis suggested that GSTA1 BB genotype was a risk factor for colorectal cancer, especially in Caucasian populations.

I.v. BU has been proven to have better bioavailability, reliable systemic drug exposure with more predictable blood levels and lower toxicity than oral BU when used as part of conditioning regimens before hematopoietic SCT (HSCT). Some studies have shown that once-daily i.v. BU had the same clinical efficacy as i.v. BU administered four times daily. To observe the clinical efficacy and pharmacokinetics (PK) of once-daily i.v. BU and to evaluate the influence of glutathione S-transferase (GST) gene polymorphisms on once-daily i.v. BU PK in adult Chinese patients with allogeneic HSCT, we analyzed 25 patients receiving related or unrelated donor transplant conditioned with i.v. BU-based regimens. With a median follow-up of 32.7 months, the 2-year OS and EFS were 64 and 63.8% for all the patients, respectively, and the 2-year cumulative incidence of relapse for all patients was 18.3%. On the basis of HPLC analysis, the mean clearance and mean daily area under the curve (AUC) of i.v. BU were calculated as 4.02 mL/min per kg and 3380.77 μM/min, respectively. The estimated Cmax was 1.031±0.0325 μg/mL. The estimated t1/2 and Vd values were 3.618±0.1932 h and 1.212±0.0352 L/kg. The once-daily i.v. BU-based conditioning regimen was very well tolerated with minor toxicity in patients, most likely because of dose assurance with predictable PK. There was no GSTA1 *B/*B homozygous patient in our Chinese patients. A significant association between BU metabolism and GSTA1 polymorphism was observed. The GSTA1 *A/*B genotype group showed a significantly higher AUC (P<0.0001), higher Cmax (P=0.0003) and lower clearance (P=0.0007) than the GSTA1 *A/*A genotype group. AUC was lower in GSTP1 *A/*A genotypes compared with*A/*G (P=0.0283) and *G/*G genotypes (P=0.0111). The BU clearance in GSTP1 *A/*A genotype was shown to be higher than *A/*G (P=0.0255) and *G/*G genotypes (P=0.0111). In addition, the differences of PK in BU among different ethnic groups existed because of the different distribution frequencies of GST gene polymorphism in Chinese patients and Caucasian patients.

BACKGROUND: The etiology of myeloproliferative neoplasms (MPN) (polycythemia vera; essential thrombocythemia; primary myelofibrosis) is unknown, however they are associated with a somatic mutation--JAK2 V617F--suggesting a potential role for environmental mutagens.
METHODS: We conducted a population-based case-control study in three rural Pennsylvania counties of persons born 1921-1968 and residing in the area between 2000-2008. Twenty seven MPN cases and 292 controls were recruited through random digit dialing. Subjects were genotyped and odds ratios estimated for a select set of polymorphisms in environmentally sensitive genes that might implicate specific environmental mutagens if found to be associated with a disease.
RESULTS: The presence of NAT2 slow acetylator genotype, and CYP1A2, GSTA1, and GSTM3 variants were associated with an average 3-5 fold increased risk.
CONCLUSIONS: Exposures, such as to aromatic compounds, whose toxicity is modified by genotypes associated with outcome in our analysis may play a role in the environmental etiology of MPNs.

Serum composition is linked to metabolic diseases not only to understand their pathogenesis but also for diagnostic purposes. Quality and quantity of nutritional intake can affect disease risk and serum composition. It is then possible that diet derived serum components directly affect pathogenetic mechanisms. To identify involved factors, we evaluated the effect on gene expression of direct addition of dyslipidemic human serum samples to cultured human hepatoma cells (HepG2). Sera were selected on the basis of cholesterol level, considering this parameter as mostly linked to dietary intake. Cells were treated with 32 sera from hypercholesterolemic and normocholesterolemic subjects to identify differentially regulated mRNAs using DNA microarray analysis. We identified several mRNAs with the highest modulations in cells treated with dyslipidemic sera versus cells treated with normal sera. Since the two serum groups had variable polyunsaturated fatty acids (PUFAs) contents, selected mRNAs were further assessed for their regulation by docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA) and arachidonic acid (AA). Four genes resulted both affected by serum composition and PUFAs: 3-hydroxy-3-methylglutaryl-CoenzymeA synthase 2 (HMGCS2), glutathione S-transferase alpha 1 (GSTA1), liver expressed antimicrobial peptide 2 (LEAP2) and apolipoprotein M (ApoM). HMGCS2 expression appears the most relevant and was also found modulated via transcription factors peroxysome proliferator activated receptor α (PPARα) and forkhead box O1 (FoxO1). Our data indicate that expression levels of the selected mRNAs, primarily of HMGCS2, could represent a reference of nutritional intake, PUFAs effects and dyslipidemic diseases pathogenesis.

Glutathione S-transferase A1 (GSTA1) appears to be primarily involved in detoxification processes, but possible roles in lung cancer remain unclear. The objective of this study was to investigate the expression and function of GSTA1 in lung cancer cells. Real-time PCR and Western blotting were performed to assess expression in cancer cell lines and the normal lung cells, then verify the A549 cells line with stable overexpression. Localization of GSTA1 proteins was assessed by cytoimmunofluorescence. Three double-strand DNA oligoRNAs (SiRNAs) were synthesized prior to being transfected into A549 cells with Lipofectamine 2000, and then the most efficient SiRNA was selected. Expression of the GSTA1 gene in the transfected cells was determined by real-time PCR and Western blotting. The viability of the transfected cells were assessed by MTT. Results showed that the mRNA and protein expression of A549 cancer cells was higher than in MRC-5 normal cells. Cytoimmunofluorescence demonstrated GSTA1 localization in the cell cytoplasm and/or membranes. Transfection into A549 cells demonstrated that down-regulated expression could inhibit cell viability. Our data indicated that GSTA1 expression may be a target molecule in early diagnosis and treatment of lung cancer.

OBJECTIVE: To evaluate the influence of polymorphisms in GSTA1, GSTM1, GSTT1, and GSTP1 in the risk of developing Prostate Cancer (PCa) in a population of Rio de Janeiro and compare the distribution of allele and genotype frequencies of the polymorphisms analyzed in the present study population with other regions in the country and different ethnic groups.
MATERIALS AND METHODS: We analyzed a sample of the Brazilian population, comprising 196 patients with PCa treated by the urology services of the Brazilian National Cancer Institute (INCA) and Mario Kroeff Hospital (HMK), and 208 male blood donors from the Clementino Fraga Filho Hospital, Federal University of Rio de Janeiro (UFRJ). The polymorphisms were determined in DNA, extracted from peripheral blood leucocytes using the Polymerase Chain Reaction and Restriction Fragment Length Polymorphism (PCR-RFLP).
RESULTS: Our results showed that the distribution of polymorphisms can vary significantly according to the Brazilian region and ethnic groups. The distribution of allele and genotype frequencies of the polymorphism GSTA1 was statistically different between cases and controls. Genotypes (A / B + B / B) were associated with protection (OR = 0.61, 95% CI = 0.40-0.92) for PCa in comparison to genotype A / A.
CONCLUSION: The distribution of genotype frequencies of the polymorphism GSTA1 was statistically different between the case and control groups (p = 0.023), and the presence of genotypes A / B and B / B suggests a protective role against the risk of PCa compared to genotype A / A. This is the first study that reports the genotypic frequency of this polymorphism and its association with PCa in a Brazilian population sample.

The aryl hydrocarbon receptor (AhR) mediates a variety of biological responses to ubiquitous environmental pollutants. In this study the effect of administration of β-naphthoflavone (BNF), potent AhR ligand, on the expression of AhR, AhRR, CYP1A1, CYP1A2, CYP1B1, NQO1, GSTA1, ALDH3A1 and UGT1A genes encoding the enzymes controlled by AhR were examined in thirteen laryngeal tumor cell lines and in HepaRG cell line. The analyzed cell lines were derived from patients with squamous laryngeal cancer, with history of cigarette smoking and without signs of human papillomavirus types 16 and 18 infection in investigated cells. Quantitative real-time RT-PCR analysis revealed huge interindividual differences in expression of genes from AhR regulatory network. Our results strongly suggest predominant effect of DNA methylation on induction of CYP1A1 expression by AhR ligands as well. Our results indicate that differentiated HepaRG cell line appeared to be very good substitute for human liver in studies on xenobiotic metabolism by AhR regulated enzymes.

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 NRF2-regulated 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 gene-gene 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.

OBJECTIVE: We investigated the role of the glutathione S-transferase A1, M1, P1 and T1 gene polymorphisms and potential effect modification by occupational exposure to different chemicals in Serbian bladder cancer male patients.
PATIENTS AND METHODS: A hospital-based case-control study of bladder cancer in men comprised 143 histologically confirmed cases and 114 age-matched male controls. Deletion polymorphism of glutathione S-transferase M1 and T1 was identified by polymerase chain reaction method. Single nucleotide polymorphism of glutathione S-transferase A1 and P1 was identified by restriction fragment length polymorphism method. As a measure of effect size, odds ratio (OR) with corresponding 95% confidence interval (95%CI) was calculated.
RESULTS: The glutathione S-transferase A1, T1 and P1 genotypes did not contribute independently toward the risk of bladder cancer, while the glutathione S-transferase M1-null genotype was overrepresented among cases (OR = 2.1, 95% CI = 1.1-4.2, p = 0.032). The most pronounced effect regarding occupational exposure to solvents and glutathione S-transferase genotype on bladder cancer risk was observed for the low activity glutathione S-transferase A1 genotype (OR = 9.2, 95% CI = 2.4-34.7, p = 0.001). The glutathione S-transferase M1-null genotype also enhanced the risk of bladder cancer among subjects exposed to solvents (OR = 6,5, 95% CI = 2.1-19.7, p = 0.001). The risk of bladder cancer development was 5.3-fold elevated among glutathione S-transferase T1-active patients exposed to solvents in comparison with glutathione S-transferase T1-active unexposed patients (95% CI = 1.9-15.1, p = 0.002). Moreover, men with glutathione S-transferase T1-active genotype exposed to pesticides exhibited 4.5 times higher risk in comparison with unexposed glutathione S-transferase T1-active subjects (95% CI = 0.9-22.5, p = 0.067).
CONCLUSION: Null or low-activity genotypes of the glutathione S-transferase A1, T1, and P1 did not contribute independently towards the risk of bladder cancer in males. However, in association with occupational exposure, low activity glutathione S-transferase A1 and glutathione S-transferase M1-null as well as glutathione S-transferase T1-active genotypes increase individual susceptibility to bladder cancer.

Microarray data were collected from bile duct samples from subjects with malignant biliary strictures by endoscopic retrograde cholangiopancreatography to screen for key genes associated with this disease. A predicted interaction network was constructed for these genes to interpret their functions. The gene expression dataset GSE34166 (10 samples: 6 malignant and 4 benign control samples) was downloaded from the Gene Expression Omnibus database. R package scripts were used to process the data and screen for differentially expressed genes. Genes identified were uploaded to the analysis tool String 8.3 to generate a gene interaction network. A hub gene was identified by calculating the node degree. The interaction network of the hub gene with other genes in the human genome was constructed and screened (score >0.9), and pathway-enrichment analysis was performed to elucidate the hub gene function. In total, 377 differentially expressed genes were identified and a network comprising 209 pairs of interactions was constructed. The most critical hub gene was identified as GSTA1, and a GSTA1-based interaction network was constructed consisting of 25 genes (containing the differentially expressed gene GSTA3). The cytochrome P450 (CYP450)-metabolic pathway displayed the most significant enrichment. Additionally, 4 transcription factors and their binding sites were also identified. In conclusion, we have identified the differentially expressed genes GSTA1 (a hub gene) and GSTA3, which may cause abnormal gene expression and tumorigenesis through CYP450-metabolic pathways. The transcription factors and their binding sites in the promoter of the hub gene provide potential directions for future drug design.

PURPOSE: To identify clinical, dosimetric, and genetic factors that are associated with late urinary toxicity after a (125)I prostate brachytherapy implant.
METHODS AND MATERIALS: Genomic DNA from 296 men treated with (125)I prostate brachytherapy monotherapy was extracted from saliva samples for this study. A retrospective database was compiled including clinical, dosimetric, and toxicity data for this cohort of patients. Fourteen candidate single-nucleotide polymorphism (SNPs) from 13 genes (TP53, ERCC2, GSTP1, NOS, TGFβ1, MSH6, RAD51, ATM, LIG4, XRCC1, XRCC3, GSTA1, and SOD2) were tested in this cohort for correlations with toxicity.
RESULTS: This study identified 217 men with at least 2 years of followup. Of these, 39 patients developed Grade ≥2 late urinary complications with a transurethral resection of prostate, urethral stricture, gross hematuria, or a sustained increase in their International Prostate Symptom Score. The only clinical or dosimetric factor that was associated with late urinary toxicity was age (p = 0.02). None of the 14 SNPs tested in this study were associated with late urinary toxicity in the univariate analysis.
CONCLUSIONS: This study identified age as the only variable being associated with late urinary toxicity. However, the small sample size and the candidate gene approach used in this study mean that further investigations are essential. Genome-wide association studies are emerging as the preferred approach for future radiogenomic studies to overcome the limitations from a candidate gene approach.

BACKGROUND: Therapy-induced leukemia is a well-known clinical syndrome occurring as a late complication in patients treated with cytotoxic therapy.
OBSERVATION: We herein present results of analysis of common gene polymorphisms in methylenetetrahydrofolate reductase (MTHFR) and glutathione S-transferase (GST) genes in a 10-year-old boy who developed very rare type of cancer, mixed phenotype acute leukemia, 6 years after treatment of acute lymphoblastic leukemia.
CONCLUSIONS: Impairment in function of GST and MTHFR enzymes found in our patient may have contributed to the development of secondary mixed phenotype acute leukemia, although precise mechanism remains elusive.

AIM: Brostallicin is a DNA minor groove binder that has shown activity in patients with soft tissue sarcoma (STS) failing first-line therapy. The present study assessed the safety and efficacy of first-line brostallicin in patients with advanced or metastatic STS >60 years or not fit enough to receive combination chemotherapy. A prospective explorative pharmacogenetic analysis was undertaken in parallel.
METHODS: Patients were randomised in a 2:1 ratio between IV brostallicin 10mg/m(2) and doxorubicin 75 mg/m(2) once every 3 weeks for a maximum of six cycles. Disease stabilisation at 26 weeks (primary end-point) was considered a 'success'. Further testing of brostallicin was warranted if ≥ 35 'successes' were observed in the first 72 eligible patients treated with brostallicin. In addition, patients were genotyped for glutathione S transferase (GST) polymorphisms.
RESULTS: One hundred and eighteen patients were included (79 brostallicin and 39 doxorubicin). Brostallicin was well tolerated in comparison to doxorubicin with less grade 3-4 neutropenia (67% versus 95%), grade 2-3 systolic dysfunction (0% versus 11%), alopecia (17% versus 61%) and grade 2-3 mucositis (0% versus 18%). For brostallicin versus doxorubicin, 'successes' were observed in 5/77 versus 10/36, progression free survival at 1 year was 6.5% versus 15.6%, objective response rate was 3.9% versus 22.2% and overall survival at 1 year was 50.5% versus 57.9%, respectively. Only GSTA1 genotype was significantly associated with success rate of doxorubicin treatment.
CONCLUSION: Brostallicin cannot be recommended at this dose and schedule in this patient population as first-line therapy. GSTA1 genotype may be predictive for doxorubicin efficacy but warrants further study.

Patients with localized high-risk soft tissue sarcomas (STS) of the limbs and trunk wall still have a considerable metastatic recurrence rate of more than 50%, in spite of adjuvant chemotherapy. This drug-ceiling effect of chemotherapy in sarcoma setting could be explained, at least partially, by multidrug resistance (MDR) mechanisms. The aim of this study was to ascertain whether mRNA and protein expression of ABCB1 (P-glycoprotein), ABCC1 (MRP1), and GSTA1 (glutathione S-transferase pi) was prognostic in localized high-risk STS. Immunohistochemistry and reverse transcriptase-PCR studies were performed from biopsies at the time of diagnosis. Patients of this series were prospectively enrolled into a phase III trial that compared three versus five cycles of epirubicin plus ifosfamide. The series of 102 patients found 41 events of recurrence and 37 of death with a median follow-up of 68 months. In univariate analysis, variables with a statistically significant relationship with relapse-free survival (RFS) were: MRP1 expression (5-year RFS rate of 23% in positive cases and 63% in negative cases, P = 0.029), histology (5-year RFS rate of 74% in undifferentiated pleomorphic sarcoma and 43% in synovial sarcoma, P = 0.028), and ABCC1 expression (5-year RFS rate of 33% in overexpression and 65% in downregulation, P = 0.012). Combined ABCC1/MRP1 was the only independent prognostic factor for both RFS (HR = 2.704, P = 0.005) and overall survival (HR = 2.208, P = 0.029). ABCC1/MRP1 expression shows robust prognostic relevance in patients with localized high-risk STS treated with anthracycline-based chemotherapy, which is the standard front line treatment in STS. This finding deserves attention as it points to a new targetable protein in STS.

OBJECTIVES: Glutathione S-transferases (GSTs) are a family of enzymes involved in detoxification. Genes encoding for GSTA1, GSTM1, GSTP1, and GSTT1 proteins are polymorphic, which can result in complete or partial loss of enzyme activity. Previous studies have associated polymorphisms of GSTA1, GSTM1, and GSTP1 genes with a higher risk of bladder cancer, but this is still controversial. Potential role of GSTA1 polymorphism in susceptibility to bladder cancer in Whites is lacking. We examined association between GSTA1, GSTM1, GSTP1, and GSTT1 gene variants and bladder cancer risk and evaluated whether they were modified by smoking.
MATERIALS AND METHODS: A hospital-based case-control study recruited 201 incidence cases and 122 age-matched controls. Deletion polymorphism of GSTM1 and GSTT1 was identified by polymerase chain reaction method. Single nucleotide polymorphism of GSTA1 and GSTP1 was identified by restriction fragment length polymorphism method. Uniconditional multivariate logistic regression was applied to model association between genetic polymorphisms and bladder cancer risk, as well as effect modification by smoking.
RESULTS: No significant difference was observed in the distributions of GSTM1, GSTT1, GSTA1, and GSTP1 gene variants between patients and controls. None of the examined polymorphisms was significantly associated with bladder cancer risk independently. The results of gene-smoking interaction analyses indicated a significant combined effect of smoking and all common GST polymorphisms tested (P for trend = 0.001). However, the most significant effect on bladder cancer risk was observed in smokers carrying lower activity GSTA1-AB/BB and GSTM-null genotype (OR = 3.5, P < 0.05) compared with GSTA1-AA and GSTM1-active non-smokers. Overall, the risk observed did not significantly differ with respect to quantity of cigarettes smoked. However, heavy smokers with GSTM1-null genotype had 2 times higher risk of bladder cancer than GSTM1-null light smokers (OR = 4.8 vs. OR = 2.0) when GSTM1-active non-smokers served as reference group. Smokers carrying both GSTM1-null and GSTA1-AB + BB genotypes exhibited the highest risk of bladder cancer (OR = 2.00, P = 0.123).
CONCLUSIONS: Null or low-activity genotypes of the GSTA1, GSTM1, GSTT1, and GSTP1 did not contribute independently towards the risk of bladder cancer in our patients. However, in association with smoking, both low activity GSTA1 and GSTM1-null genotype increase individual susceptibility to bladder cancer.