Determination of glutathione S-transferase mu and theta polymorphisms in neurological disease

1. Correlations between deletions in two glutathione S-transferase (GST) genes, GSTM1 and GSTT1 and susceptibility to Alzheimer's disease (AD), motor neuron disease (MND) and Parkinson's disease (PD) have been investigated by PCR, using primers specific for both genes. 2. It was found that males with a deletion of the GSTM1 gene were more susceptible to PD and males with a deletion of the GSTT1 gene more susceptible to MND and PD, possibly implying that environmental factors which specifically target men may be involved. Furthermore, subjects with a deletion of the GSTT1 gene were more susceptible to AD.     

Binding of pesticides to alpha,mu and pi class glutathione transferase

The binding of fluorodifen, fenarimol, acifluorfen, 2,4-DES, methyl parathion, paraquat and pyrazophos by alpha, mu and pi class glutathione transferases (GST) was determined by the 2-p-toluidinylnaphthalene-6-sulphonate (TNS) binding fluorescence inhibition technique. Although all the 3 GST classes appear to be capable of binding the pesticides investigated, mu class exhibited somewhat higher affinity than the alpha and pi classes.     

Inactivation of the genotoxic aldehyde acrolein by human glutathione transferases of classes alpha, mu, and pi

Acrolein, a genotoxic aldehyde released in the metabolic activation of the cytostatic drug cyclophosphamide, is inactivated by glutathione transferases either by conjugation with reduced glutathione or by covalent binding to the enzymes in the absence of glutathione. The catalytic efficiency (kcat/Km) with acrolein as a substrate was determined for representatives of the three classes Alpha, Mu, and Pi of human glutathione transferases. Transferase pi exhibited the highest and transferase epsilon the lowest catalytic efficiencies, respectively. As measured by the kcat/Km value, acrolein ranks among the most active substrates known for transferase pi. The irreversible binding of acrolein to the enzymes was monitored as the inactivation of the enzyme activity. Transferase pi reacted significantly more rapidly with acrolein than did transferases mu and epsilon.     

Alcohol consumption, glutathione S-transferase M1 and T1 genetic polymorphisms and breast cancer risk

To evaluate the potential association between GSTM1 and GSTT1 genotypes and development of breast cancer, a hospital based case-control study was conducted in a South Korean study population consisting of 189 histologically confirmed incident breast cancer cases and their 189 age-matched control subjects with no present or previous history of cancer. A multiplex polymerase chain reaction method was used for the genotyping analyses and statistical evaluations were performed by unconditional logistic regression model. The GSTM1 null genotype was significantly associated with breast cancer risk in premenopausal women [odds ratio (OR) = 2.0, 95% confidence interval (CI) = 1-3.7], but not in the postmenopausal women (OR = 0.9, 95% CI = 0.5-1.9), nor in all women grouped together (OR = 1.3, 95% CI = 0.8-1.1). The GSTT1 null genotype posed a similar risk of breast cancer with an OR of 1.6 (95% CI = 1.0-2.5) for the total breast cancer group, OR of 1.7 (95% CI = 0.9-3.2) for pre-menopausal women, and OR of 1.3 (95% CI = 0.6-2.8) for post-menopausal women. The breast cancer risk associated with concurrent lack of both GSTM1 and GSTT1 genes was 2.2 (95% CI = 1.1-4.5), and the risk increased as the number of null genotype increased (P for trend = 0.03). When the data were stratified by the known risk factors of breast cancer, a significant interaction was observed between the GSTM1 genotypes and alcohol consumption (P for interaction = 0.03). An especially remarkable risk of breast cancer was observed for alcohol-consuming premenopausal women lacking both the GSTM1 and GSTT1 genes (OR = 5.3, 95% CI = 1.0-27.8) compared to those with both of the genes. Our findings thus suggest a novel gene-environment interaction which plays an important role in the individual susceptibility to breast cancer. p6     

Ramucirumab for the treatment of gastric cancers,colorectal adenocarcinomas,and other gastrointestinal malignancies

Introduction: The use of antiangiogenic strategy in the treatment of advanced colorectal cancers has been largely evidence-based. More recently, novel vascular endothelial growth factor receptor (VEGFR) inhibitors have been studied in other gastrointestinal diseases. Ramucirumab, a recombinant monoclonal antibody that binds to VEGFR2 extracellular domain with a much greater affinity compared to its natural ligand, showed second-line effectiveness for patients with gastric or colorectal carcinomas.

Areas covered: We perform a narrative literature review. The aims of our work are to recall the current evidence of its efficacy in the treatment of gastric, hepatocellular and colorectal cancers and to present the ongoing studies enrolling gastrointestinal cancer patients in which ramucirumab is being tested.

Expert commentary: The landscape of angiogenesis-inhibition for the treatment of GI malignancies is rapidly evolving. The results of the REGARD and RAINBOW trials renewed the interest for antiangiogenic agents in gastric cancer and determined a swift change in the treating paradigm for this disease. Accordingly, ramucirumab was shown to be effective in pretreated colorectal cancer patients and it is being tested in other gastrointestinal malignancies.     

Glutathione S-transferase genetic polymorphisms and individual sensitivity to the ototoxic effect of cisplatin

One of the side effects of cisplatin therapy in malignant neoplasms is ototoxicity. This effect shows a wide inter-individual range which is more variable than the pharmacokinetic parameters. Oxidative stress has been implicated in cisplatin ototoxicity. The glutathione S-transferase (GST) supergene family encodes isoenzymes that appear to be critical in protection against oxidative stress. Certain GST loci are polymorphic, demonstrating alleles that are null (GSTM1 and GSTT1), encode low-activity variants (GSTP1) or are associated with variable inducibility (GSTM3). The aim of our study was to investigate genetic risk factors involved in the ototoxicity of cisplatin and to determine whether the polymorphisms in five GST genes affect the individual risk of ototoxicity by cisplatin. Two groups of patients were analyzed in this study: group H, 20 patients early and highly sensitive to the ototoxicity of cisplatin; and group N, 19 patients with no hearing impairment under comparable doses of the drug. We found a protective effect for the GSTM3*B allele with a frequency of 0.18 in the group with normal hearing after therapy versus 0.025 in the group with hearing impairment. (chi2=5.37; p=0.02).     

Glutathione S-transferases M1 and T1 polymorphisms and arsenic content in hair and urine in two ethnic clans exposed to indoor combustion of high arsenic coal in Southwest Guizhou, China

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.     

Risks of interleukin-1 genetic polymorphisms and Helicobacter pylori infection in the development of gastric cancer

BACKGROUND: The host genetic factors that determine the clinical outcomes of Helicobacter pylori-infected individuals remain unclear. AIM: To elucidate the risks of host interleukin-1 (IL-1) genetic polymorphisms and H. pylori infection in the development of gastric cancer. METHODS: In a case-control study of 164 controls and 142 patients with gastric cancer, the IL-1B-511 biallelic polymorphisms and the IL-1RN penta-allelic variable number of tandem repeats were genotyped. RESULTS: The carriage of IL-1RN*2, male gender, old age and H. pylori infection independently increased the risk of gastric cancer, with odds ratios of 3.3 [95% confidence interval (CI), 1.4-7.7], 2.1 (95% CI, 1.2-3.8), 5.3 (95% CI, 3.1-9.0) and 2.2 (95% CI, 1.3-3.8), respectively. H. pylori-infected individuals who were carriers of IL-1RN*2 showed increased risks of both intestinal and diffuse types of gastric cancer, with odds ratios of 11.0 and 8.7, respectively. In addition, these individuals also had a higher score of intestinal metaplasia in the corpus than did uninfected non-carriers. CONCLUSIONS: This study is the first to verify IL-1RN*2 as an independent factor governing the development of gastric cancer in Asian individuals. A combination of H. pylori testing and host genotyping may target the eradication of H. pylori to high-risk individuals.     

Relevance of the deletion polymorphisms of the glutathione S-transferases GSTT1 and GSTM1 in pharmacology and toxicology

Although cytosolic glutathione S-transferase (GST) enzymes occupy a key position in biological detoxification processes, two of the most relevant human isoenzymes, GSTT1-1 and GSTM1-1, are genetically deleted (non-functional alleles GSTT1*0 and GSTM1*0) in a high percentage of the human population, with major ethnic differences. The structures of the GSTT and GSTM gene areas explain the underlying genetic processes. GSTT1-1 is highly conserved during evolution and plays a major role in phase-II biotransformation of a number of drugs and industrial chemicals, e.g. cytostatic drugs, hydrocarbons and halogenated hydrocarbons. GSTM1-1 is particularly relevant in the deactivation of carcinogenic intermediates of polycyclic aromatic hydrocarbons. Several lines of evidence suggest that hGSTT1-1 and/or hGSTM1-1 play a role in the deactivation of reactive oxygen species that are likely to be involved in cellular processes of inflammation, ageing and degenerative diseases. There is cumulating evidence that combinations of the GSTM1*0 state with other genetic traits affecting the metabolism of carcinogens (CYP1A1, GSTP1) may predispose the aero-digestive tract and lung, especially in smokers, to a higher risk of cancer. The GSTM1*0 status appears also associated with a modest increase in the risk of bladder cancer, consistent with a GSTM1 interaction with carcinogenic tobacco smoke constituents. Both human GST deletions, although largely counterbalanced by overlapping substrate affinities within the GST superfamily, have consequences when the organism comes into contact with distinct man-made chemicals. This appears relevant in industrial toxicology and in drug metabolism.     

The influence of polymorphisms of glutathione S-transferases M1 and M3 on the development of human urothelial cancer

Cigarette smoking is the most important risk factor for development of transitional cell carcinoma of the urinary bladder. The effect of polymorphisms of glutathione S-transferases M1 (GSTM1) and M3 (GSTM3) on the influence of cigarette smoking on urinary bladder carcinogenesis was investigated. In total, 293 bladder cancer patients from hospitals in Dortmund and Wittenberg as well as 176 patients without any malignancy from a Department of Surgery from Dortmund were genotyped for GSTM1 and GSTM3 according to standard PCR/RFLP methods. Smoking habits were quantified by a standardized interview. The proportion of GSTM1 negative cases was 63% in the entire bladder cancer cases group compared to 50% in controls. The GSTM3*A/*A genotype was 76% in cancer cases versus 74% in controls. Smokers and ex-smokers were overrepresented in bladder cancer cases. A significant association between smoking status and GSTM1 or GSTM3 genotype was not detected. The elevated proportion of GSTM1 negative bladder cancer cases shows an effect of this polymorphic enzyme on development of bladder cancer. In contrast to other studies, an influence of GSTM1 on the risk due to cigarette smoking was not observed.     

Genetic polymorphisms in heterocyclic amine metabolism and risk of colorectal adenomas

High red meat intake has been linked with an increased risk of colorectal cancer and adenomas. During high temperature cooking of red meats, heterocyclic amines (HCAs) are generated; however, to be carcinogenic, they must be metabolized by enzymes including cytochrome P450 1A2 (CYP1A2) and N-acetyltransferase 1 (NAT1) and/or N-acetyltransferase 2 (NAT2). We have conducted a clinic-based case-control study of colorectal adenomas that focused on assessment of exposure to HCAs (estimated by use of a HCA database and meat cooking module) and modification of these exposures by genetic factors. We have previously reported that intake of MeIQx was associated with an increased risk of colorectal adenomas [overall association at 80th percentile, > 27.00 ng/day: odds ratio (OR) = 2.68, 95% confidence interval (CI) 1.58-4.55]. Here, we report our evaluation of whether variation in CYP1A2, NAT1 and/or NAT2 modify the association between HCAs and colorectal adenoma formation in 146 cases and 228 frequency-matched controls. The NAT1*10 allele was associated with a nonsignificant increased risk of colorectal adenomas (OR = 1.43; 95% CI 0.86-2.36). Further, when we analysed 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) intake as a categorical variable, we observed a six-fold increase in adenoma risk among rapid NAT1 acetylators who consumed more than 27 ng a day (OR = 6.50; 95% CI 2.16-19.7), whereas among slow NAT1 acetylators, the increase in risk was two-fold (OR = 2.32; 95% CI 1.12-4.81). While suggestive, the results were not significantly different from each other on either an additive or multiplicative scale. In contrast, NAT2 genotype and CYP1A2 and NAT2 hepatic activity measured by caffeine urinary metabolites were not associated with adenoma risk, although an increase in risk with rapid CYP1A2 activity could not be ruled out (OR = 1.46; 95% CI 0.76-2.81). Moreover, there was no evidence that the effect of MeIQx was enhanced among subjects in any subgroup defined by variation in these measures. These results are compatible with the hypothesis that high HCA exposure is associated with an increased risk of colorectal adenomas, particularly in genetically susceptible subgroups. Further study of larger populations is needed to confirm and extend these observations.     

Heterozygosity for the alpha1-antitrypsin Z allele may confer genetic risk of cholangiocarcinoma

Aliment Pharmacol Ther 2011; 33: 389–394

Summary

Background Alpha1‐antitrypsin (α1AT) deficiency caused by Z allele homozygosity represents a well‐established risk factor for hepatocellular carcinoma. Previous studies have also implicated α1AT Z heterozygosity in cholangiocarcinogenesis. Aim To assess the ‘common’ Z and S alleles as well as the promoter variant rs8004738 for association with cholangiocarcinoma. Methods We genotyped 182 Caucasian patients and 350 controls for rs28929474 (Z), rs17580 (S) and the variant rs8004738. Exploratory analyses were performed in relation to gender and cholangiocarcinoma localisation. Results rs28929474 was significantly enriched in the cholangiocarcinoma group (4.1 vs. 1.7%; OR 2.46, 95% CI 1.14–5.32; Bonferroni corrected p_c = 0.036), reinforced by Armitage trend testing (OR 2.53; p_c = 0.032). The rs8004738 (promoter) minor allele tended to be overrepresented in Z heterozygotes (30.0 vs. 16.7%: P = 0.13). Exploratory data analyses suggested a high genetic risk for extrahepatic tumour localisation (OR 3.0; p_c = 0.016) and potentially female Z allele carriers (OR 3.37; unadjusted P = 0.022, p_c = 0.088). Conclusions These data point to a novel role of α1AT Z heterozygosity as a potential genetic susceptibility factor for cholangiocarcinoma formation and suggest a contribution of aberrant α1AT function in biliary carcinogenesis. However, given the overall low rs28929474 minor allele frequency, larger studies are warranted to confirm and extend our findings.     

Disposition of 3H-enisoprost,a gastric antisecretory prostaglandin,in healthy humans

1. After oral administration of ³H-enisoprost (450 μg) to five healthy men, as a solution in capsules, peak ³H levels of 5624 ± 566 pg equiv./ml (mean ± S.E.M.) were reached within one hour. No unchanged drug was detected in plasma.

2. Enisoprost was rapidly de-esterified to SC-36067 [(±)11α, 16ζ-dihydroxy-16-methyl-9-oxoprost-4Z, 13E-dien-1-oic acid], a pharmacologically active analogue, which reached peak concentrations of 651±200 pg/ml within 20 min of dosing. SC-36067 was eliminated metabolically, with a half-life of 1.61 h, by a combination of β-oxidation, β-oxidation and 9-keto-reduction.

3. After nine days 59.0±2.98% and 17.4±1.57% of the dose was excreted in urine and faeces respectively. The majority of this excretion was complete in two days.

4. Five urinary metabolites were identified by GC-MS. These were (±)3-[2β-(4-hydroxy-4-methyl-1E-octenyl)-3α-hydroxy-5-oxo-1α-cyclopentanyl]propanoic acid (SC-41411; 3.6% dose), (±)3-[3α,5-dihydroxy-2β-(4-hydroxy-4-methyl-1E-octenyl)-1α-cyclopentanyl]propanoic acid (SC-41411 PGF analogue; 4.8% dose), (±)3-[2β-(8-carboxy-4-hydroxy-4-methyl-1E-octenyl)-3α-hydroxy-5-oxo-1α-cyclopentanyl] propanoic acid (SC-41411-16-carboxylic acid; 22% dose), (±)3-[2β-(8-carboxy-4-hydroxy-4-methyl-1E-octenyl)-3α,5-dihydroxy-1α-cylopentanyl]propanoic acid (SC-41411 PGF analogue-16-carboxylic acid; 8.5% dose) and its γ lactone (2.6% dose).

5. These metabolites were also identified chromatographically in plasma, as were SC-36067, (±)3-[2β-(4-hydroxy-4-methyl-1E-octenyl)-5-oxo-1α-cyclopent-3-enyl]propaloic acid and (±)3-[2β-(4-hydroxy-4-methyl-1E-octenyl)-5-oxo-cyclopent-1-enyl]propanoic acid.

6. Some 5-10% of the dose was excreted in urine as tritiated water, indicating that oxidation of the 11α-hydroxy group in SC-36067 or its metabolites also occurred.     

Disposition of 3H-enisoprost, a gastric antisecretory prostaglandin, in healthy humans

1. After oral administration of 3H-enisoprost (450 micrograms) to five healthy men, as a solution in capsules, peak 3H levels of 5624 +/- 566 pg equiv./ml (mean +/- S.E.M.) were reached within one hour. No unchanged drug was detected in plasma. 2. Enisoprost was rapidly de-esterified to SC-36067 [(+/-)11 alpha, 16 zeta-dihydroxy-16-methyl-9-oxoprost-4Z, 13E-dien-1-oic acid], a pharmacologically active analogue, which reached peak concentrations of 651 +/- 200 pg/ml within 20 min of dosing. SC-36067 was eliminated metabolically, with a half-life of 1.61 h, by a combination of beta-oxidation, omega-oxidation and 9-keto-reduction. 3. After nine days 59.0 +/- 2.98% and 17.4 +/- 1.57% of the dose was excreted in urine and faeces respectively. The majority of this excretion was complete in two days. 4. Five urinary metabolites were identified by GC-MS. These were (+/-)3-[2 beta-(4-hydroxy-4-methyl-1E-octenyl)-3 alpha-hydroxy-5-oxo-1 alpha-cyclopentanyl]propanoic acid (SC-41411; 3.6% dose), (+/-)3-[3 alpha,5-dihydroxy-2 beta-(4-hydroxy-4-methyl-1E-octenyl)-1 alpha-cyclopentanyl]propanoic acid (SC-41411 PGF analogue; 4.8% dose), (+/-)3-[2 beta-(8-carboxy-4-hydroxy- 4-methyl-1E-octenyl)-3 alpha-hydroxy-5-oxo-1 alpha-cyclopentanyl] propanoic acid (SC-41411-16-carboxylic acid; 22% dose), (+/-)3-[2 beta-(8-carboxy-4- hydroxy-4-methyl-1E-octenyl)-3 alpha,5-dihydroxy-1 alpha-cyclopentanyl] propanoic acid (SC-41411 PGF analogue-16-carboxylic acid; 8.5% dose) and its gamma lactone (2.6% dose). 5. These metabolites were also identified chromatographically in plasma, as were SC-36067, (+/-)3-[2 beta-(4-hydroxy-4-methyl-1E-octenyl)-5-oxo-1 alpha- cyclopent-3-enyl]propanoic acid and (+/-)3-[2 beta-(4-hydroxy-4-methyl-1E-octenyl)-5-oxo-cyclopent-1- propanoic acid. 6. Some 5-10% of the dose was excreted in urine as tritiated water, indicating that oxidation of the 11 alpha-hydroxy group in SC-36067 or its metabolites also occurred.     

The mu1 and mu2 opioid receptor binding of ketobemidone,norketobemidone and 3-dimethylamino-1,1-diphenylbutene

Abstract: Ketobemidone is a mu receptor agonist, available in Scandinavia in a combined preparation (Ketogan$$), which contains ketobemidone and the spasmolytical agent 3-dimethylamino-1, 1-diphenylbutene (A29) in a one to five ratio. A29 is chemically related to methadone and experiments in rats have shown that A29 potentiates the analgesic effect of ketobemidone (Petersen 1951). We have recently shown that ketobemidone has lower binding affinity for the total mu receptor than morphine. At $$-binding sites the two opioids have equal affinity, whereas ketobemidone has much lower affinity than morphine to the $$ receptor (Christensen 1993). The existence of two subtypes of mu receptors has been suggested by Pasternak and co-workers: a high affinity, mu₁ receptor subtype that appears to mediate supraspinal analgesia and a low affinity, mu₂ subtype that appears to mediate spinal analgesia, respiratory depression and inhibition of gastrointestinal motility (Clark et al. 1988; Paul et al. 1989).