Sara Piacentini

Human genetic variation of CYP450 superfamily: analysis of functional diversity in worldwide populations

AIM The present study aimed to investigate the human genetic diversity of the CYP450 superfamily in order to identify functional interethnic differences and analyze the role of CYP450 enzymes in human adaptation. MATERIALS & METHODS A computational analysis of genetic and functional differences of the 57 CYP450 genes was performed using the Human Genome Diversity Project and HapMap data; comprising approximately 1694 individuals belonging to 62 human populations. RESULTS Twenty-six CYP450 SNPs with F-statistics significantly different than the general distribution were identified. Some showed high differentiation among human populations, suggesting that functional interethnic differences may be present. Indeed, some of these are significantly associated with drug response or disease risk. Furthermore, our data highlighted that TBXAS1 and genes in CYP3A cluster may have a role in some processes of human adaptation. CONCLUSION Our study provided an analysis of genetic diversity of CYP450 superfamily, identifying functional differences among ethnic groups and their related clinical phenotypes.

HapMap-based study of human soluble glutathione S-transferase enzymes: the role of natural selection in shaping the single nucleotide polymorphism diversity of xenobiotic-metabolizing genes.

Objective Glutathione S-transferase enzymes (GSTs; EC: 2.5.1.18) constitute the principal phase II superfamily, which plays a key role in cellular detoxification. GST genes are organized in chromosomal clusters; most of these genes are polymorphic, mainly due to single nucleotide substitutions. Different studies proved significant interethnic differences in GST allelic frequencies but, at present, the role of natural selection in human genetic variability of GSTs is poorly understood. The aim of this study is to investigate the role of natural selection in shaping single nucleotide polymorphism (SNP) diversity of soluble GST genes. Methods Using the HapMap database, we analyzed the population differences in the soluble GST genes using the phasing data from unrelated individuals shared among 11 populations in the International HapMap project. A Fst-based selection test was applied to HapMap data to detect soluble GST loci under selection. Results Comparisons between GST gene polymorphisms among HapMap populations highlight that ethnicity is an influencing factor of GST genetic variability. By applying a genome scan based on F-statistics, we identified nine SNPs that present F-coefficients significantly more different than those expected under neutrality (rs2239892, rs3814309, rs7483, rs1571858, rs929166, rs11807, rs4715344, rs4715354, and rs3734431). Conclusion Our study confirms that GST gene variation reflects human demographic history, but it also demonstrates that natural selection could shape the genetic profile of some GST SNPs. Moreover, the identification of human genome regions and targets of natural selection may have detected candidate genes for complex diseases. In analyzing the literature, we provide complex disease hypothesis (male infertility, embryotoxicity) for the identified GST SNPs.

Glutathione S-transferase gene polymorphisms and air pollution as interactive risk factors for asthma in a multicentre Italian field study: A preliminary study.

Abstract Background: Asthma is one of the most common chronic diseases. Several studies have indicated that oxidative stress impairs pulmonary function. Glutathione S-transferases (GSTs) are believed to be critical in the protection of cells from reactive oxygen species. Aim: In this case-control study we analysed the possible association between polymorphism in several cytosolic GST genes, air pollution and asthma development. Methods: Genotyping of GSTM1 and GSTT1 genes was carried out by a multiplex PCR; GSTA1, GSTO1, GSTO2, GSTP1 polymorphisms were determined using the PCR-RFLP method. Data on atmospheric pollutants were collected by the regional air-quality monitoring network. Results: Among all the polymorphisms studied, the frequencies of GSTA1, GSTM1, GSTO2 and GSTT1 genotypes found in the group of asthmatic patients seem to differ from the frequencies of those found in the control group. Air pollutants were analysed and the air quality parameters considered proved to be significantly different, and therefore suitable for this study. Conclusion: The final result of this research should hopefully lead to a better understanding of gene–environment interactions, so allowing earlier prediction and diagnosis of asthma disease and providing an efficient means of prevention.

GSTO1*E155del polymorphism associated with increased risk for late-onset Alzheimer's disease: association hypothesis for an uncommon genetic variant

Glutathione S-transferases are multifunctional enzymes involved in cellular detoxification. A genetic linkage was found between Alzheimers Disease (AD) and the chromosome 10q, where the GSTO1 and GSTO2 genes are located, leading to the hypothesis that GST Omega class (GSTO) genes may be an AD risk factor. Since it is still controversial, we decided to explore GSTO polymorphisms in Italian cohorts. We analyzed 119 AD patients and 114 healthy controls for the GSTO gene polymorphisms. In particular we investigated two common polymorphisms (GSTO1*A140D, GSTO2*N142D) and two uncommon variants (GSTO1*E155del, GSTO1*E208K) to find loci associated with AD risk. Detection of GSTO1*A140D and GSTO2*N142D was performed by PCR-RFLP, while GSTO1*E155del and GSTO1*E208K were detected using confronting two-pair primer and allele specific PCR, respectively. While GSTO1*A140D, GSTO1*E208K and GSTO2*N142D polymorphisms did not show significant outcomes, the GSTO1*E155del polymorphism is associated with AD [P=0.003; adjusted OR=3.70 (1.57-8.75)]. Our results suggest that GSTO1-1 plays a role in AD since the GSTO1*del155 variant is involved in changes in GSTO1-1 activities decreasing in enzyme stability. Specifically, three hypotheses may explain the role of GSTO1-1 in the pathophysiology of AD: the antioxidant activity of GSTO1-1 may protect brain tissue against oxidative stress; GSTO1-1 activity regulate interleukin-1β activation and its genetic variation may act to modulate inflammation in AD; GSTO1-1 is involved in the arsenic biotransformation pathway and gene polymorphisms may be implicated in the modulation of arsenic neurotoxicity. In conclusion, we hypothesized that GSTO1*E155del is an uncommon genetic variant associated with AD risk.

GSTM1 null genotype as risk factor for late-onset Alzheimer's disease in Italian patients

Alzheimers disease (AD) is the most common form of dementia in the elderly. The causes of AD are very complex but there is general agreement about the existence of a link between Alzheimers disease and oxidative stress. The Glutathione S-transferases (GSTs) act to detoxify products of oxidation that cause damage to macromolecules. Particular attention has been focused on GST genes because polymorphisms are important determinants of disease risk. To evaluate if GSTA1, GSTM1, GSTP1, and GSTT1 genes are associated with LOAD we screened a case-control population (n=311). Differences in genotype distributions between AD patients and controls were found only for the GSTM1 null genotype (P<0.001). In addition, a logistic regression analysis also conferred a positive association between the GSTM1 null genotype and LOAD after adjustment for age and gender (OR=2.09; 95%CI=1.31-3.35). The GSTM1 enzyme detoxifies substances such as exogenous and endogenous metabolites and plays a regulatory role in cellular signaling. Previous studies have highlighted that GSTM1 has a role in neurodegenerative disorders, but no data have associated the GSTM1 gene with AD risk. Our outcome suggests that the GSTM1 null genotype is a risk factor for AD in Italian patients.

Genetic variability of glutathione S-transferase enzymes in human populations: Functional inter-ethnic differences in detoxification systems

Glutathione S-Transferase enzymes (GSTs) constitute the principal Phase II superfamily which plays a key role in cellular detoxification and in other biological processes. Studies of GSTs have revealed that genetic polymorphisms are present in these enzymes and that some of these are Loss-of-Function (LoF) variants, which affect enzymatic functions and are related to different aspects of human health. The aim of this study was to analyze functional genetic differences in GST enzymes among human populations. Attention was focused on LoF polymorphisms of GSTA1, GSTM1, GSTO1, GSTO2, GSTP1 and GSTT1 genes. These LoF variants were analyzed in 668 individuals belonging to six human groups with different ethnic backgrounds: Amhara and Oromo from Ethiopia; Colorado and Cayapa Amerindians and African Ecuadorians from Ecuador; and one sample from central Italy. The HapMap database was used to compare our data with reference populations and to analyze the haplotype and Linkage Disequilibrium diversity in different ethnic groups. Our results highlighted that ethnicity strongly affects the genetic variability of GST enzymes. In particular, GST haplotypes/variants with functional impact showed significant differences in human populations, according to their ethnic background. These data underline that human populations have different structures in detoxification genes, suggesting that these ethnic differences influence disease risk or response to drugs and therefore have implications for genetic association studies involving GST enzymes. In conclusion, our investigation provides data about the distribution of important LoF variants in GST genes in human populations. This information may be useful for designing and interpreting genetic association studies.

Glutathione S-transferase ω class (GSTO) polymorphisms in a sample from Rome (Central Italy).

Abstract Glutathione S-transferases are a superfamily of enzymes that are involved in biotransformation of drugs, xenobiotics and play a fundamental role in the protection of cells from oxidative stress. In humans, the recently described GST Omega class contains two expressed genes GSTO1 and GSTO2, located on chromosome 10 (10q24.3). Four polymorphisms in GSTO genes have been identified in ethnic groups: GSTO1*A140D (rs4925), GSTO1*E155del (rs56204475), GSTO1*E208K (rs11509438) and GSTO2*N142D (rs156697). This study provides the allele frequencies of GSTO polymorphism in a sample consisting of 116 apparently healthy individuals of both sexes from Rome (Central Italy). Detection of GSTO1*A140D and GSTO2*N142D alleles was performed by PCR-RFLP analysis, while GSTO1*E155del and GSTO1*E208K alleles were detected using the Confronting Two-Pair Primers analysis (PCR-CTPP) and allele specific PCR, respectively. The GSTO allele frequencies found in the Italian sample were included in the variability range observed in European populations. Comparison between the data presented in this study and data in previous studies showed different patterns among European, Asian and African populations.

Glutathione S-transferase genes and the risk of recurrent miscarriage in Italian women.

OBJECTIVE To investigate the role of glutathione S-transferases (GSTs) in the pathogenesis of recurrent miscarriage (RM). DESIGN Genetic association study. SETTING University of Rome, Tor Vergata and San Giovanni Calibita, Fatebenefratelli Hospital. PATIENT(S) One hundred twenty-one women with RM and 113 women without pregnancy complications. INTERVENTION(S) Genomic DNA extracted from buccal cells and screening of positive/null genotypes of GSTM1 and GSTT1 genes and single nucleotide polymorphisms of GSTA1, GSTO2, and GSTP1 genes. MAIN OUTCOME MEASURE(S) Occurrence of GST polymorphisms. RESULT(S) Women with at least one GSTA1*-69T allele are more frequent in the RM group than in the control group: 67% vs. 48%, respectively. Significant outcomes were obtained considering different genetic models: codominant, dominant, and log-additive. In addition, the combined analysis suggests that GSTA1 and GSTM1 variants have a significant interaction in RM risk. CONCLUSION(S) Our study highlighted a significant association between the GSTA1 gene and an increased risk of RM. In particular, the -69T allele in the GSTA1 gene may be considered as a predisposing factor of RM.

Modulation of the GSTT1 activity by the GSTM1 phenotype in a sample of Italian farm-workers

Glutathione S-transferase (GST) isozymes catalyze nucleophilic attack by reduced Glutathione (GSH) on a variety of electrophilic compounds and play a central role in biotransformation of xenobiotics (Hayes et al., Annu Rev Pharmacol Toxicol 45:51–88, 2005). We performed a case–control study to evaluate the GSTM1 and GSTT1 polymorphisms and to investigate if exposure to pesticides conditions the GSTT1 activity level in 115 healthy controls and 90 farm-workers exposed to pesticides. Polymorphisms were investigated using a GSTM1 or a GSTT1-specific PCR. Enzyme activity was measured by means of DCM as co-substrate, as described by Bruhn et al. (Biochem Pharmacol 56:1189–1193, 1998). There was no significant difference between the farm-workers and the healthy controls regarding the distribution of various alleles of the GSTM1 and GSTT1 genes and the GSTT1 enzyme activity. In farm-workers, the GSTM1 null genotype was associated with a significant increase of GSTT1 activity, suggesting a regulative mechanism common to GSTM1 and GSTT1 enzymes after exposure to xenobiotics.

Human GST loci as markers of evolutionary forces: GSTO1*E155del and GSTO1*E208K polymorphisms may be under natural selection induced by environmental arsenic

Over the last two decades, significant data has been accumulated linking Glutatione S-Transferases (GSTs) with the development of several diseases. Contemporary studies have demonstrated the impact of ethnicity on GST allele frequencies. The aim is to verify if the variability of GST genes reflects population demographic history or rather selective pressures. GST genes (GSTM1, GSTO1 GSTO2, GSTT1) were analysed in three Ecuadorian populations (Cayapas, n = 114; Colorados, n = 104; African-Ecuadorian, n = 77) and compared with HapMap data. GST SNPs were determined using the PCR-RFLP method while GST null phenotype was determined using a Multiplex PCR. The population relationship achieved using GSTM1 positive/null, GSTO1*A140D, GSTO2*N142D and GSTT1 positive/null are in agreement with the data obtained using neutral polymorphisms: Amerindians are close to Asian populations and African-Ecuadorians to African populations. To what concerns GSTO1*del155 and GSTO1*K208 variants, allele frequencies never exceeded 10%, showing no significant differences in the Ecuadorian groups and in worldwide populations. The features of GSTO1*del155 and GSTO1*K208 variants and their association with arsenic biotransformation deficiency suggest the presence of a selection mechanism towards these loci. In particular, this hypothesis is strengthened by a possible linkage between these alleles and the susceptibility of arsenic-induced male infertility.