Because acute infection and inflammation affect drug metabolism and drug-metabolizing enzymes, the effect of the acute-phase response on the expression of glutathione S-transferase (GST) isoenzymes, glutathione synthesis, and several antioxidant enzymes was investigated. Hepatic expression of GST isozymes, positive and negative acute-phase reactants, and antioxidant enzymes were determined by Northern blotting and hybridization with gene-specific oligonucleotide probes after lipopolysaccharide treatment of rats. Lipopolysaccharide caused the expected acute-phase response as judged by the increased expression of positive and decreased expression of negative acute-phase proteins. The messenger RNA (mRNA) expression of the major hepatic rat GST isozymes A1, A2, A3, M1, and M2 was decreased 50% to 90%. Total hepatic GST activity toward 1-chloro-2,4-dinitrobenzene was also significantly decreased. mRNA expression of gamma-glutamylcysteine synthetase (GCS) large subunit and catalase was reduced by approximately 60%. GCS enzyme activity was also decreased, resulting in a 35% decrease in the hepatic content of reduced glutathione 4 days after lipopolysaccharide challenge. Mn-Superoxide dismutase expression was increased 13-fold, and thioredoxin level was elevated 3-fold after lipopolysaccharide challenge. The expression of all parameters determined returned to near control levels 7 days after treatment. Together, these data show that GSTs and GCS are negative acute-phase proteins and that decreased GCS activity results in a decrease in hepatic glutathione content. Thus, in addition to the phase I drug-metabolizing enzymes known to be decreased during the acute-phase response, some phase II enzymes involved in the elimination of xenobiotics and carcinogens are also decreased.

OBJECTIVE

In cystic fibrosis (CF) children, we investigated the predictive impact of glutathione S-transferases (GST) activity and genotypes P1, M1 and T1, and antioxidant levels on stage-severity of Pseudomonas aeruginosa lung infection.

METHODS

GST activity was determined in whole blood by spectrophotometry, and GST genotypes by multiplex PCR RFLP for 36 CF and 9 control children. Levels of glutathione in erythrocyte and vitamins A, E and C in plasma were measured by HPLC.

RESULTS

No difference in GST activity and no relationship between GST activity and antioxidant levels were observed in CF children as compared to controls. However, GST activity was lower in CF children with severe clinical status and infection, and the frequency of GSTP1 wild type genotype AA, prevalent in uninfected CF children (75%), decreased in infected ones (33%).

CONCLUSIONS

GST activity and genotype could play an important role in modulating P. aeruginosa lung infection in CF patients.

Monogenic forms of Parkinson's disease (PD) provide an opportunity to examine mechanisms underlying phenotypic variation. Glutathione S-transferase (GST) has detoxification and antioxidative functions. To screen genetic variations in GST for an effect on the onset age (OA) of PD, we typed seven common genetic polymorphisms in five GST isoenzymes, M1, M3, P1, T1, and Z1, in 36 affected individuals of Italian or Greek origin with the alpha-synuclein A53T (PARK1) mutation. Mean OA was 45.2 years with a wide SD of 11.03 years, similar to that of idiopathic PD. Our allelic analysis showed that the subjects homozygous for the GSTP1 G-for-A nucleotide substitution at position 313 had a mean OA acceleration of 15.2 years (31.3 +/- 7.09 years, n = 3 vs. 46.5 +/- 10.50 years, n = 33, P = 0.020). The GSTP1 C341T substitution was associated with a 9.7-year acceleration of OA, but the significance was borderline (36.4 +/- 8.35 years vs. 46.7 +/- 10.85 years, P = 0.0519). After correction for the five genes examined, both results lose statistical significance. Nevertheless, our results suggest that further investigation in GSTP1 variants and PD pathogenesis is warranted in sporadic PD and that a search for toxins that accelerate PD OA should pay particular attention to GST-P1 substrates.

We conducted a hospital-based case-control study to evaluate the interactive effect of reproductive factors and glutathione S-transferase (GST) M1 and T1 genetic polymorphisms in individual susceptibility to breast cancer. The study population consisted of 189 incident breast cancer cases and 189 age-matched controls with no known malignant diseases. GSTM1/T1 genotypes were determined by a multiplex polymerase chain reaction (PCR) method, and odds ratios (ORs) and 95% confidence intervals (CIs) were calculated by conditional logistic regression model. The parity factors were grouped as (1) high-risk status defined as nullipara or para with experience of first full-term pregnancy (FFTP) at or over 30 years, and (2) low-risk status defined as para with experience of FFTP under 30 years. A significant multiplicative interaction was observed between GSTM1 and GSTT1 null genotypes and high-risk status of parity factor in all women and in premenopausal women (P < or = 0.01), but not in postmenopausal women (P>> 0.05). The interaction between the combined genotypes of GSTM1 and GSTT1 and status of parity factor was also significant in all women and in premenopausal women (P < 0.01). Our findings suggest that genetic polymorphisms GSTM1/T1 could modify estrogen-related breast cancer risk.

Breast cancer is the most common cancer among women worldwide and second in Thailand. Glutathione S-transferase (GST) enzymes involved in the detoxification of reactive metabolites of carcinogens may be important in modulating susceptibility to cancers. This study aimed to determine the influence of genetic polymorphisms of glutathione S-transferase T1, M1, P1 and A1 on breast cancer in Thai patients. Links with clinico-pathological characteristics were also analyzed. The results showed no association between GSTs polymorphism and overall susceptibility to breast cancer in Thai patients (P < 0.05). However, the data pointed to a relation of the GSTP1(Ile105Val) polymorphism with progesterone receptor status (P = 0.04) and age at diagnosis (P = 0.03) of breast cancer cases. In summary, this is the first study to report associations between glutathione S-transferase T1, M1, P1 and A1 gene polymorphisms and breast cancer in Thai patients. While GST genotypes may not be associated with susceptibility, a GSTP1 polymorphism (Ile105Val) may be related to progression of breast cancer.

OBJECTIVE

To analyse the association genetic polymorphisms of glutathione S-transferase (GST) M1 and T1 with genetic susceptibility to sporadic colorectal adenocarcinoma (SCRAC).

METHODS

All subjects are unrelated the Han people in Hubei province of China. Using multiple PCR, we studied the genetic polymorphisms of the GSTM1, GSTT1 genes of 101 healthy controls and 104 SCRAC patients.

RESULTS

(1) All the differences of the frequency of GSTM1 null genotype between SCRACs and the controls, between proximal and distal SCRACs and between the elder and the younger SCRACs did not reach statistical significance. (2) The differences of the frequency of GSTT1 null genotype between SCRACs and the controls did not reach statistical significance too; But the null genotype for GSTT1 was significantly more common among distal SCRACs when compared with the proximals (66.2%:44.4%, chi(2) = 3.97, P < 0.05) and more common among the elder SCRACs when compared with younger SCRACs (70.9%:49.0%, chi(2) = 5.21, P < 0.05). (3) Subjects carring both of the null genotypes for GSTM1 and GSTT1 had more than 4.33-fold risk for developing SCRACs compared with the subjects caring both of the nonull genotypes for GSTM1 and GSTT1 (OR = 4.33, 95% confidence interval, 1.56 - 12.04).

CONCLUSIONS

(1) There was no association the susceptibility to SCRAC with GSTM1 or GSTT1 null genotype alone, but the date suggests GSTT1 null genotype may influence the distal SCRAC, especially the elder; (2) The individual with both of the null genotypes of GSTM1 and GSTT1 increases markedly the risk of SCRAC, and this genotype is the susceptibility gene to SCRAC.

A total of 2,402 cases of arsenic-related skin lesions (as of 2002) in a few villages of China's Southwest Guizhou Autonomous Prefecture represent a unique case of endemic arseniasis related with indoor combustion of high arsenic coal. A significant difference of skin lesion prevalence was observed between two clans of different ethnicities (Hmong and Han) in one of the hyperendemic villages in this prefecture. This study was focused on a possible involvement of GST T1 and M1 polymorphisms in risk modulation of skin lesions and in the body burden of As in this unique case of As exposure. GST T1 and M1 polymorphisms were genotyped by an allele-specific PCR-based procedure. Total As contents in hair and urine samples as well as environmental samples of the homes of the two ethnic clans were analyzed. No significant deviations in the population frequencies of GST T1 and M1 0/0 genotypes or their combination were recorded between diagnosed skin lesion patients and asymptomatic individuals in both clans. Significantly higher As contents in hair and urine were observed in GSTM1 0/0 carriers, not in GSTT1 0/0 carriers. After stratified by ethnicity and gender, a statistically significant association of the GSTM1 0/0 genotype and higher As content in hair was only confirmed in the subgroups of ethnic Han clan members and all male villagers, not in ethnic Hmong clan members or in females. GST T1 and M1 homozygous deletions were not associated with an increased susceptibility to skin lesions in long-term exposure to indoor combustion of high As coal. The polymorphic status at the locus of GSTM1 might modulate individual's body burden of total As in some Chinese ethnic groups.

Glutathione S-transferases (GSTs), including glutathione S-transferase M1 (GSTM1) and glutathione S-transferase T1 (GSTT1), are multifunctional enzymes which play vital roles in the detoxification of a variety of carcinogens. The genetic polymorphisms of GSTM1 and GSTT1 have been implicated in pancreatic cancer risk, but the results of published studies remain conflicting. Thus, a meta-analysis was conducted to estimate the effect of GSTM1 and GSTT1 polymorphisms on the risk of developing pancreatic cancer. A comprehensive search was performed in the PubMed, Embase, Web of Science, and Wanfang databases to identify the available studies on the associations of GSTM1 and GSTT1 polymorphisms with pancreatic cancer risk. The pooled odds ratio (OR) with its corresponding 95 % confidence interval (95 % CI) was used to estimate the associations. Stratified analyses by ethnicity and sensitivity analyses were performed to further identify the relationships. Overall, the null genotype of GSTT1 was associated with an increased risk of pancreatic cancer (OR = 1.61, 95 % CI 1.06-2.44, P OR = 0.025), but similar association was not found between the null genotype of GSTM1 and pancreatic cancer risk. Besides, a significant association of GSTT1 polymorphism with pancreatic cancer risk was identified in Asians (OR = 2.58, 95 % CI 1.67-3.98, P OR < 0.001), but not in Caucasians (OR = 1.16, 95 % CI 0.94-1.43, P OR = 0.170). Sensitivity analyses by sequential omission of individual study confirmed the stability of our results. Meta-analysis of available data thus far shows that the null genotype of GSTT1 is a risk factor for pancreatic cancer, particularly in the Asian population. The currently available data are not sufficient enough to identify the association between the GSTM1 polymorphism and pancreatic cancer risk.

OBJECTIVE

To investigate, in young patients with inflammatory bowel disease (IBD) treated with azathioprine, the association between genetic polymorphisms of thiopurine-S-methyl-transferase (TPMT), inosine-triphosphate-pyrophosphatase (ITPA), and glutathione-S-transferases (GST), involved in azathioprine metabolism, the concentration of the main metabolites of azathioprine, thioguanine nucleotides (TGNs) and the methylated nucleotides (MMPN), and the dose of the medication.

BACKGROUND

Azathioprine is widely used in IBD as an immunosuppressive agent, particularly to maintain remission in patients with steroid refractory disease. Azathioprine is a prodrug and requires conversion to its active form mercaptopurine, which has no intrinsic activity, and is activated by the enzymes of the purine salvage pathway to TGNs. Polymorphisms in genes of enzymes involved in azathioprine metabolism influence the efficacy and toxicity of treatment.

METHODS

Seventy-five young patients with IBD treated with azathioprine at least for 3 months were enrolled and genotyped for the selected genes; for these patients, TGN and MMPN metabolites were measured by high performance liquid chromatography in erythrocytes.

RESULTS

GST-M1 deletion was associated with lower TGN/dose ratio (P=0.0030), higher azathioprine dose requirement (P=0.022), and reduced response to therapy (P=0.0022). TPMT variant genotype was associated with lower MMPN concentration (P=0.0064) and increased TGN/dose ratio (P=0.0035). ITPA C94A polymorphism resulted in an increased MMPN concentration (P=0.037).

CONCLUSIONS

This study describes the effect of candidate genetic polymorphisms in TPMT, ITPA, and GST-M1 on azathioprine pharmacokinetics in IBD patients, showing, for the first time, relevant effects of GST-M1 genotype on azathioprine metabolites concentration.

P450 (CYP) and glutathione S-transferase (GST) are involved in the activation and detoxification of many potential carcinogens. Although, the interaction between environmental exposure and genetic polymorphisms of cytochrome P450 2E1 (CYP2E1) and glutathione S-transferase M1 (GSTM1) in breast cancer has been assessed, the gene-gene interactions between CYP2E1 and GSTM1 related to breast cancer have not been focused on and reported. We conducted a hospital-based case-control study to investigate whether the genetic interaction effects of CYP2E1 and GSTM1 modify the risk of developing breast cancer independent of the effect of cigarette smoking and alcohol consumption. Individuals with the C2/C2 genotype of CYP2E1 had a lower risk (OR = 0.24, 95% CI = 0.08-0.74) when compared with those with the C1/C1 genotype. However, there was no significant difference (OR = 1.05, 95% CI = 0.73-1.50) in the GSTM1 genotype frequency between the cases with breast cancer and that of the controls. When individuals with the genotype of C1/C1 or C1/C2 of CYP2E1 and the wild-type of GSTM1 were compared with those of C2/C2 of CYP2E1 and the null-type of GSTM1 however, we found a significantly increased risk (OR = 3.50, 95% CI = 1.01-16.55) in the breast cancer patients. Our findings indicated a gene-gene interaction between CYP2E1 and GSTM1 was accessible to developing breast cancer in Taiwanese women without the habits of cigarette smoking and alcohol consumption even though independent effects of CYP2E1 and GSTM1 were weak or non-significant and suggest that environmental carcinogen besides cigarette and alcohol consumption could induce breast cancer.

The association between alcohol-induced disorders of human spermatogenesis and glutathione S-transferase-M1 genotype was investigated in an autopsy study comprising 271 subjects, including interviews with relatives or close acquaintances on the alcohol consumption of the deceased. Of the 50 moderate drinking men (reported mean daily alcohol consumption < 40 g), 21 (42.0%) had normal spermatogenesis, whereas 24 men (48.0%) had partial, and five (10.0%) complete arrest of spermatogenesis. Of the 21 men with normal spermatogenesis, nine (42.9%) had GST M1 'null' genotype, while this genotype was found in 13 (44.8%) of the 29 men with partial or complete spermatogenic arrest. Of 212 heavy-drinking men (reported mean daily alcohol consumption>> 80 g), only 45 (21.2%) had normal spermatogenesis, whereas 77 (36.3%) had partial spermatogenic arrest. Complete arrest of spermatogenesis was found in 81 men (38.2%) and nine men had Sertoli-cell-only syndrome (4.2%). Of the 45 heavy drinkers with normal spermatogenesis, 27 (60%) men had GST M1 'null' genotype (OR 2.7 with 95% confidence intervals: 1.0-4.0, when compared to those with disorders of spermatogenesis). The frequency of GST M1 'null' genotype in heavy drinkers with normal spermatogenesis also differed from that of corresponding moderate drinkers, whereas the frequency of GST M1 'null' genotype in heavy drinkers with disorders of spermatogenesis was similar to moderate drinkers with or without disorders of spermatogenesis. The finding that>> 20% of men in the heavy-drinking group had normal spermatogenesis suggests individual variations in sensitivity to alcohol-induced disorders of this process. Heavy drinkers with GST M1 'null' genotype were slightly less prone to develop disorders of spermatogenesis. Thus, the GST M1 locus may be associated with susceptibility to develop alcohol-induced disorders of spermatogenesis.

Genotoxicity of tobacco smoke has long been investigated and tobacco smoke is considered to be one of the principal human carcinogens. Although its role in DNA-damage induction and cancer development has been documented, the mechanisms by which this happens are not well understood. Many chemical constituents of tobacco smoke are enzymatically metabolized by phase-I and phase-II enzymes, but modifications in coding and regulating sequences of these genes could influence their ability to detoxify these compounds. In this work, we studied several enzymes involved in the metabolism of xenobiotics, viz. the glutathione S-transferases (GST) M1, T1, P1 and A1, with respect to their influence on the genotoxic effects induced by cigarette smoking. We assessed the genotoxic effects of tobacco smoke on peripheral blood lymphocytes of 72 healthy caucasians by use of the chromosomal aberration (CA) assay and the micronucleus (MN) test. Genotypes of GST M1, T1, P1 and A1 were determined by means of the polymerase chain reaction and methods based on restriction fragment length polymorphism (RFLP). We found that smoke and gender are the two variables that most influence the DNA damage. In particular, we observed that female smokers seem to be more sensitive than male smokers, having a significantly higher frequency of CAs. Moreover, a significant increase in frequency of micronuclei in bi-nucleated cells (BNMN) was found in smokers, but not in non-smokers. This increase seems to be influenced not only by age and gender, but also by genetic constitution. Subjects carrying GSTM1-null genotype seemed to have an higher susceptibility to DNA damage induced by tobacco smoke than GSTM1-positive ones. When considering a combination of GST genotypes, we found a lower BNMN frequency in subjects with GSTP1 variant allele plus GSTM1-positive genotypes, while the most damaged cells are found in subjects bearing GSTM1-null plus GSTP1-wild type. Our results suggest that investigation of the association between several gene polymorphisms and important endpoints of DNA damage could contribute to better understanding the role of gene-gene interaction.

To identify new host factors that modulate the replication of influenza A virus, we performed a yeast two-hybrid screen using the cytoplasmic tail of matrix protein 2 from the highly pathogenic H5N1 strain. The screen revealed a high-score interaction with cyclin D3, a key regulator of cell cycle early G1 phase. M2-cyclin D3 interaction was validated through GST pull-down and recapitulated in influenza A/WSN/33-infected cells. Knockdown of Ccnd3 by small interfering RNA significantly enhanced virus progeny titers in cell culture supernatants. Interestingly, the increase in virus production was due to cyclin D3 deficiency per se and not merely a consequence of cell cycle deregulation. A combined knockdown of Ccnd3 and Rb1, which rescued cell cycle progression into S phase, failed to normalize virus production. Infection by influenza A virus triggered redistribution of cyclin D3 from the nucleus to the cytoplasm, followed by its proteasomal degradation. When overexpressed in HEK 293T cells, cyclin D3 impaired binding of M2 with M1, which is essential for proper assembly of progeny virions, lending further support to its role as a putative restriction factor. Our study describes the identification and characterization of cyclin D3 as a novel interactor of influenza A virus M2 protein. We hypothesize that competitive inhibition of M1-M2 interaction by cyclin D3 impairs infectious virion formation and results in attenuated virus production. In addition, we provide mechanistic insights into the dynamic interplay of influenza virus with the host cell cycle machinery during infection.

Glutathione S-transferases (GSTs) constitute a family of detoxification enzymes that catalyze the conjugation of glutathione with a variety of hydrophobic compounds, including drugs and their metabolites, to yield water-soluble derivatives that are excreted in urine or bile. Profiling the effect of small molecules on GST activity is an important component in the characterization of drug candidates and compound libraries. Additionally, specific GST isozymes have been implicated in drug resistance, especially in cancer, and thus represent potential targets for intervention. To date, there are no sensitive miniaturized high-throughput assays available for GST activity detection. A series of GST substrates containing a masked luciferin moiety have been described recently, offering the potential for configuring a sensitive screening assay via coupled luciferase reaction and standard luminescence detection. We report on the optimization and miniaturization of this homogeneous method to 1,536-well format using GSTs from 3 different species: mouse isozyme A4-4, human isozymes A1-1, M1-1, and P1-1, and the major GST from the parasitic worm Schistosoma japonicum.

Glutathione S-transferases (GSTs) are major intracellular antioxidants, which, impaired in their function, are involved in the progress of schizophrenia (SCZ). The aim of this case-control study was to investigate the association between the polymorphism of glutathione S-transferases M1 (GSTM1), T1 (GSTT1), the glutathione S-transferase P1 gene (GSTP1) and SCZ. We isolated genomic DNA from peripheral blood of 93 individuals with SCZ and 99 healthy control subjects' genotypes analyzing them for GSTM1, GSTT1 and GSTP1 using polymerase chain reaction. The analysis of the gene-gene interaction between GSTs indicated that the magnitude of the association was greater for the combined AG/GSTT1 & GSTM1 genotypes (OR = 2.51; 95% CI: 1.13-5.63, P = 0.02). The AG and combined AG + GG genotypes of GSTP1 increased the risk of SCZ (OR = 1.83; 95% CI: 0.94-3.75 and OR = 1.71; 95% CI: 0.92-3.19, respectively). The genotypes of GSTT/NULL, NULL/GSTM and NULL/NULL increased the risk of SCZ (OR = 2.05; 95% CI: 0.9-4.74; OR = 2.0; 95% CI: 1.68-2.31; and OR = 1.8; 95% CI: 0.57-2.46, respectively). The present study supports previous data that suggest that impairment in the function of GSTs genes may increase the risk of SCZ.

Modeling methods allow the identification and analysis of determinants of reactivity and specificity in enzymes. The reaction between glutathione and 1-chloro-2,4-dinitrobenzene (CDNB) is widely used as a standard activity assay for glutathione S-transferases (GSTs). It is important to understand the causes of differences between catalytic GST isoenzymes and the effects of mutations and genetic polymorphisms. Quantum mechanical/molecular mechanical (QM/MM) molecular dynamics simulations have been performed here to investigate the addition of the glutathione anion to CDNB in the wild-type M1-1 GST isoenzyme from rat and in three single point mutant (Tyr6Phe, Tyr115Phe, and Met108Ala) M1-1 GST enzymes. We have developed a specifically parameterized QM/MM method (AM1-SRP/CHARMM22) to model this reaction by fitting to experimental heats of formation and ionization potentials. Free energy profiles were obtained from molecular dynamics simulations of the reaction using umbrella sampling and weighted histogram analysis techniques. The reaction in solution has also been simulated and is compared to the enzymatic reaction. The free energies are in excellent agreement with experimental results. Overall the results of the present study show that QM/MM reaction pathway analysis provides detailed insight into the chemistry of GST and can be used to obtain mechanistic insight into the effects of specific mutations on this catalytic process.

The aim of this study is to verify whether the combination of glutathione S-transferase (GST) M1 null and GSTT1 null genotypes, which is a candidate genetic risk factor for troglitazone-induced liver failure, is common to that for the carbamazepine-induced mild hepatotoxicity.

METHODS

The genotypes of GSTM1 and GSTT1, and microsomal epoxide hydrolase-3 and -4, were determined in 192 Japanese epileptics treated with carbamazepine.

RESULTS

The GSTM1 null (GSTM1-) and GSTT1 null (GSTT1-) genotypes in the subjects were 55.7 and 39.6%, respectively. The alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were elevated in 46 (24.0%) and 62 (32.3%) cases, and the mean values were approximately 2.3- and 1.8-times higher than the upper limit of normal levels, respectively. The levels of ALT and AST were significantly higher in GSTM1- than in GSTM1 present (GSTM1+) genotypes (p = 0.007 and 0.004, respectively). The level of ALT was significantly higher in GSTM1-/T1- than in GSTM1+/T1- and GSTM1+/T1+ (p = 0.01 and 0.01, respectively), and that of AST was significantly higher in GSTM1-/T1- and GSTM1-/T1+ than in GSTM1+/T1+ (p = 0.02 and 0.003, respectively). The microsomal epoxide hydrolase genotype did not influence the hepatotoxicity.

CONCLUSIONS

These findings suggested that GSTM1- rather than GSTM1-/T1- was a risk factor for carbamazepine-induced mild hepatotoxicity.

In order to find the effect of genetic polymorphisms of glutathione S-transferase M1 (GSTM1) and GSTT1 on hematological changes of individuals chronically exposed to natural sour gas, the present study was done. Study subjects (59 males, 55 females) were residents of contaminated areas of Masjid-i-Sulaiman (southwest of Iran). The GSTM1 and GSTT1 genotypes were determined using a polymerase chain reaction-based method. The multiple linear regression method was applied. There is significant association between GSTs genotypes and either hemoglobin (t=2.185, P=0.031) or hematocrit (t=2.454, P=0.016). Also there is weak association between GSTs genotypes and WBC counts (t=1.802, P=0.074). The hemoglobin and hematocrit levels and WBC counts increased in individuals who had null genotypes of GSTM1 and GSTT1 compared to subjects with one or two active genes. Also the levels of hemoglobin and hematocrit and WBC counts increased in persons with one active genotype compared to subjects who had two active genes. There is no significant association between neither platelet nor WBC differential parameters and GSTs genotypes.

Rat micro class glutathione transferases M1-1 and M2-2 are homodimers that share a 78% sequence identity but display differences in stability. M1-1 is more stable at the secondary and tertiary structural levels, whereas its quaternary structure is less stable. Each subunit in these proteins consists of two structurally distinct domains with intersubunit contacts occurring between domain 1 of one subunit and domain 2 of the other subunit. The chimeric subunit variants M(12), which has domain 1 of M1 and domain 2 of M2, and its complement M(21), were used to investigate the conformational stability of the chimeric homodimers M(12)-(12) and M(21)-(21) to determine the contribution of each domain toward stability. Exchanging entire domains between class micro GSTs is accommodated by the GST fold. Urea-induced equilibrium unfolding data indicate that whereas the class micro equilibrium unfolding mechanism (i.e., N(2) <->> 2I <->> 2U) is not altered, domain exchanges impact significantly on the conformational stability of the native dimers and monomeric folding intermediates. Data for the wild-type and chimeric proteins indicate that the order of stability for the native dimer (N(2)) is M2-2>> M(12)-(12) M1-1 approximately M(21)-(21), and that the order of stability of the monomeric intermediate (I) is M1>> M2 approximately M(12)>> M(21). Interactions involving Arg 77, which is topologically conserved in GSTs, appear to play an important role in the stability of both the native dimeric and folding monomeric structures.

The aim of this study was to assess if genetic variants in the glutathione-S-transferase genes GST-T1, M1, and P1 reflect risk factors in acetaminophen (APAP)-poisoned patients assessed by investigation of the relation to prothrombin time (PT), which is a sensitive marker of survival in these patients. A total of 104 APAP-poisoned patients were genotyped for deletion polymorphisms in the GSTT1 and GSTM1 genes and for the GSTP1 Ile105Val polymorphism. We found a borderline association (p = 0.05) between the GSTT1 homozygous deletion genotype and high trough PT (a marker of prognosis in APAP poisoning) compared to carrying two functioning copies of the gene. No significant association was found between any of the GSTM1 and GSTP1 genotypes and PT. The frequency of GSTP1 Val/Val genotypes was significantly lower in the patients than in the background population (p = 0.047). The results suggest that the GSTT1 homozygous deletion genotype may be associated with a better prognosis after APAP poisoning and that carriers of the GSTP1 homozygous variant genotype may have a decreased risk of being APAP poisoned.

Ischemic cerebrovascular disease (ICVD) is a multifactorial disease caused by the interactions of several genetic and environmental factors. Tobacco smoke is a major cause of both cancer and vascular disease. Although its carcinogenic role via induction of DNA damage and mutation is well established, the mechanisms involved in vascular disease remain unclear. One possibility is that DNA damage causes smooth muscle cell proliferation in the intima of arteries, thereby contributing to atherothrombotic processes. The binding of chemicals to DNAis modulated by detoxification enzymes, including glutathione S-transferase (GST). We examined whether polymorphisms in this gene, as well as the angiotensin-converting enzyme (ACE) gene influence the risk of ICVD on smoking status. DNA was analyzed for deletions in the GST M1, T1, and ACE genes by polymerase chain reaction (PCR). No significant association was observed between GST null genotype and ICVD, even in smokers. However, a significant association between ACE and ICVD was observed only in smokers (chi(2) = 0.023, p < 0.05). We conclude that GST polymorphism is not a risk factor for the development of ICVD through smoking and suggest a high probability that ACE polymorphism may contribute to the odds of ICVD in smokers.

The objective of this study was to examine the association between the genetic polymorphism of glutathione S-transferase (GST) M1, T1 and N-acetyltransferase 2 (NAT2) genes and urothelial cancer risk in relation to smoking status. In this study, 325 Japanese patients with urothelial transitional cell carcinoma and 325 healthy controls were compared for frequencies of GSTM1, T1 and NAT2 genotypes. The frequencies of GSTM1 null genotype and NAT2 slow genotype were significantly higher in the cases than in the controls (adjusted odds ratio (OR) 1.37, 95% confidence interval (CI) 1.01-1.87, adjusted OR 3.09, 95% CI 1.69-5.63, individually). Furthermore, the risk of GSTM1 null genotype and NAT2 slow genotype was higher among smokers (adjusted OR 1.48, 95% CI 1.01-2.15, adjusted OR 4.28, 95% CI 1.96-9.36, individually). The regression analysis of cancer risk as a function of the amount of smoking showed that the susceptibility of people who had GSTM1 null genotype increased from 45 pack-years, while the susceptibility of people with NAT2 intermediate or slow genotype increased rapidly from 25 pack-years, compared with non-smokers. A multiplicative interaction between NAT2 intermediate or slow genotype and pack-years of smoking was found (P<0.001), but GSTM1 null genotype was not (P=0.06). Our results indicate that the GSTM1 null genotype and NAT2 intermediate or slow genotype are associated with an increased risk of urothelial cancer in relation to smoking amounts. Furthermore, the interaction between NAT2 intermediate or slow genotype and pack-years of smoking has a strong impact on urothelial cancer.

Deficiency in the capability of xenobiotic detoxification and arsenic methylation may be correlated with individual susceptibility to arsenic-related skin cancers. We hypothesized that glutathione S-transferase (GST M1, T1, and P1), reactive oxygen species (ROS) related metabolic genes (NQO1, EPHX1, and HO-1), and DNA repair genes (XRCC1, XPD, hOGG1, and ATM) together may play a role in arsenic-induced skin carcinogenesis. We conducted a case-control study consisting of 70 pathologically confirmed skin cancer patients and 210 age and gender matched participants with genotyping of 12 selected polymorphisms. The skin cancer risks were estimated by odds ratio (OR) and 95% confidence interval (CI) using logistic regression. EPHX1 Tyr113His, XPD C156A, and GSTT1 null genotypes were associated with skin cancer risk (OR = 2.99, 95% CI = 1.01-8.83; OR = 2.04, 95% CI = 0.99-4.27; OR = 1.74, 95% CI = 1.00-3.02, resp.). However, none of these polymorphisms showed significant association after considering arsenic exposure status. Individuals carrying three risk polymorphisms of EPHX1 Tyr113His, XPD C156A, and GSTs presented a 400% increased skin cancer risk when compared to those with less than or equal to one polymorphism. In conclusion, GSTs, EPHX1, and XPD are potential genetic factors for arsenic-induced skin cancers. The roles of these genes for arsenic-induced skin carcinogenesis need to be further evaluated.

Cytochrome P450 1A1 (CYP1A1) is involved in the production of arachidonic acid-derived vasoactive substance. We hypothesized that CYP1A1 polymorphism might be related to pathological conditions associated with cerebral infarction (CI). We investigated the effect of genetic polymorphism in the 3'-flanking region (T6235C) of CYP1A1 gene in 353 patients with CI and 376 controls. The distributions of T6235C CYP1A1 genotypes in patients with (TT: 36.0%; TC/CT: 64.0%; n = 353) and without CI (TT: 44.7%; TC/CT: 55.3%; n = 376) indicate that the C allele is associated with CI (P = 0.017, odds ratio (O.R.) = 1.44; 95% confidence interval (C. I.) = 1.07-1.94). Furthermore, we examined whether the glutathione S-transferase (GST) gene, which is one of detoxification enzyme, influence the risk of CI. GST M1 null genotype increased the relative risk for the CI in the subjects with the CYP1A1 C allele (P = 0.015, O.R. = 1.47; C. I. = 1.08-2.00). We conclude that T6235C CYP1A1 polymorphism is a risk factor for the development of CI and suggest that GST polymorphism contribute to the odds of CI.