Two new ArrayTrack libraries for personalized biomedical research.

BACKGROUND: Recent advances in high-throughput genotyping technology are paving the way for research in personalized medicine and nutrition. However, most of the genetic markers identified from association studies account for a small contribution to the total risk/benefit of the studied phenotypic trait. Testing whether the candidate genes identified by association studies are causal is critically important to the development of personalized medicine and nutrition. An efficient data mining strategy and a set of sophisticated tools are necessary to help better understand and utilize the findings from genetic association studies. DESCRIPTION: SNP (single nucleotide polymorphism) and QTL (quantitative trait locus) libraries were constructed and incorporated into ArrayTrack, with user-friendly interfaces and powerful search features. Data from several public repositories were collected in the SNP and QTL libraries and connected to other domain libraries (genes, proteins, metabolites, and pathways) in ArrayTrack. Linking the data sets within ArrayTrack allows searching of SNP and QTL data as well as their relationships to other biological molecules. The SNP library includes approximately 15 million human SNPs and their annotations, while the QTL library contains publically available QTLs identified in mouse, rat, and human. The QTL library was developed for finding the overlap between the map position of a candidate or metabolic gene and QTLs from these species. Two use cases were included to demonstrate the utility of these tools. The SNP and QTL libraries are freely available to the public through ArrayTrack at http://www.fda.gov/ArrayTrack. CONCLUSIONS: These libraries developed in ArrayTrack contain comprehensive information on SNPs and QTLs and are further cross-linked to other libraries. Connecting domain specific knowledge is a cornerstone of systems biology strategies and allows for a better understanding of the genetic and biological context of the findings from genetic association studies.

Cystine-glutamate transporter SLC7A11 mediates resistance to geldanamycin but not to 17-(allylamino)-17-demethoxygeldanamycin.

The cystine-glutamate transporter SLC7A11 has been implicated in chemoresistance, by supplying cystine to the cell for glutathione maintenance. In the NCI-60 cell panel, SLC7A11 expression shows negative correlation with growth inhibitory potency of geldanamycin but not with its analog 17-(allylamino)-17-demethoxygeldanamycin (17-AAG), which differs in the C-17 substituent in that the the methoxy moiety of geldanamycin is replaced by an amino group. Structure and potency analysis classified 18 geldanamycin analogs into two subgroups, "17-O/H" (C-17 methoxy or unsubstituted) and "17-N" (C-17 amino), showing distinct SLC7A11 correlation. We used three 17-O/H analogs and four 17-N analogs to test the role of the 17-substituents in susceptibility to SLC7A11-mediated resistance. In A549 cells, which are resistant to geldanamycin and strongly express SLC7A11, inhibition of SLC7A11 by (S)-4-carboxyphenylglycine or small interfering RNA increased sensitivity to 17-O/H, but had no effect on 17-N analogs. Ectopic expression of SLC7A11 in HepG2 cells, which are sensitive to geldanamycin and express low SLC7A11, confers resistance to geldanamycin, but not to 17-AAG. Antioxidant N-acetylcysteine, a precursor for glutathione synthesis, completely suppressed cytotoxic effects of 17-O/H but had no effect on 17-N analogs, whereas the prooxidant ascorbic acid had the opposite effect. Compared with 17-AAG, geldanamycin led to significantly more intracellular reactive oxygen species (ROS) production, which was quenched by addition of N-acetylcysteine. We conclude that SLC7A11 confers resistance selectively to 17-O/H (e.g., geldanamycin) but not to 17-N (e.g., 17-AAG) analogs partly as a result of differential dependence on ROS for cytotoxicity. Distinct mechanisms could significantly affect antitumor response and organ toxicity of these compounds in vivo.

Irreversible global DNA hypomethylation as a key step in hepatocarcinogenesis induced by dietary methyl deficiency.

Dietary methyl group deprivation is now well recognized as a model of hepatocarcinogenesis in rodents. In the present study, we examined the effects of feeding a methyl-deficient diet followed by a methyl-adequate diet on the extent of methylation of liver DNA and on the formation and evolution of altered hepatic foci. Male F344 rats were fed a methyl-deficient diet for 9, 18, 24, and 36 weeks, followed by re-feeding a methyl-adequate diet for a total of 54 weeks. Similar to previous findings, the methyl-deficient diet resulted in decreased levels of S-adenosylmethionine (SAM), SAM/SAH ratios, and global DNA hypomethylation. Feeding the methyl-adequate diet restored the liver SAM levels and SAM/SAH ratios to control levels in all experimental groups. In contrast, re-feeding the complete diet restored DNA methylation to normal level only in the group that had been fed the methyl-deficient diet for 9 weeks; in animals exposed to methyl deprivation longer, the methyl-adequate diet failed to reverse the hypomethylation of DNA. Liver tissue of rats exposed to methyl deficiency for 9, 18, 24, or 36 weeks was characterized by the persistent presence of placental isoform of glutathione-S-transferase (GSTpi)-positive lesions despite re-feeding the methyl-adequate diet. The persistence of altered hepatic foci in liver after withdrawal of methyl-deficient diet serves as an indication of the carcinogenic potential of a methyl-deficient diet. Substitution of the methyl-deficient diet with complete diet failed to prevent the expansion of initiated foci and restore DNA methylation in animals exposed to deficiency for 18, 24, or 36 weeks. The association between DNA hypomethylation and expansion of foci suggests that stable DNA hypomethylation is a promoting factor for clonal expansion of initiated cells. These results provide an experimental evidence and a mechanistic basis by which epigenetic alterations may contribute to the initiation and promotion steps of carcinogenesis.

S-adenosyl-L-methionine is able to reverse micronucleus formation induced by sodium arsenite and other cytoskeleton disrupting agents in cultured human cells.

Deficiencies of folic acid and methionine, two of the major components of the methyl metabolism, correlate with an increment of chromosome breaks and micronuclei. It has been proposed that these effects may arise from a decrease of S-adenosyl-L-methionine (SAM), the universal methyl donor. Some xenobiotics, such as arsenic, originate a reduction of SAM levels, and this is believed to alter some methylation processes (e.g. DNA methylation). The aim of the present work was to analyze the effects of exogenous SAM on the micronucleus (MN) frequency induced by sodium arsenite in human lymphocytes treated in vitro and to investigate whether these effects are related to DNA methylation. Results showed a reduction in the MN frequency in cultures treated with sodium arsenite and SAM compared to those treated with arsenite alone. To understand the mechanism by which SAM reduced the number of micronucleated cells, its effect on MN induced by other xenobiotics was also analyzed. Results showed that SAM did not have any effect on the increase in MN frequency caused by alkylating (mitomycin C or cisplatin) or demethylating agents (5-azacytidine, hydralazine, ethionine and procainamide), but it reduced the number of micronucleated cells in those treated with agents that inhibit microtubule polymerization (albendazole sulphoxide and colcemid). Since albendazole sulphoxide and colcemid inhibit microtubule polymerization, we decided to evaluate the effect of SAM on microtubule integrity. Data obtained from these evaluations showed that sodium arsenite, albendazole sulphoxide, and colcemid affect the integrity and organization of microtubules and that these effects are significantly reduced when cultures were treated at the same time with SAM. The data taken all together point out that the positive effects of SAM could be due to its ability to protect microtubules through an unknown mechanism.

Genetic associations with micronutrient levels identified in immune and gastrointestinal networks.

The discovery of vitamins and clarification of their role in preventing frank essential nutrient deficiencies occurred in the early 1900s. Much vitamin research has understandably focused on public health and the effects of single nutrients to alleviate acute conditions. The physiological processes for maintaining health, however, are complex systems that depend upon interactions between multiple nutrients, environmental factors, and genetic makeup. To analyze the relationship between these factors and nutritional health, data were obtained from an observational, community-based participatory research program of children and teens (age 6-14) enrolled in a summer day camp in the Delta region of Arkansas. Assessments of erythrocyte S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), plasma homocysteine (Hcy) and 6 organic micronutrients (retinol, 25-hydroxy vitamin D3, pyridoxal, thiamin, riboflavin, and vitamin E), and 1,129 plasma proteins were performed at 3 time points in each of 2 years. Genetic makeup was analyzed with 1 M SNP genotyping arrays, and nutrient status was assessed with 24-h dietary intake questionnaires. A pattern of metabolites (met_PC1) that included the ratio of erythrocyte SAM/SAH, Hcy, and 5 vitamins were identified by principal component analysis. Met_PC1 levels were significantly associated with (1) single-nucleotide polymorphisms, (2) levels of plasma proteins, and (3) multilocus genotypes coding for gastrointestinal and immune functions, as identified in a global network of metabolic/protein-protein interactions. Subsequent mining of data from curated pathway, network, and genome-wide association studies identified genetic and functional relationships that may be explained by gene-nutrient interactions. The systems nutrition strategy described here has thus associated a multivariate metabolite pattern in blood with genes involved in immune and gastrointestinal functions.

Methylation potential associated with diet, genotype, protein, and metabolite levels in the Delta Obesity Vitamin Study.

Micronutrient research typically focuses on analyzing the effects of single or a few nutrients on health by analyzing a limited number of biomarkers. The observational study described here analyzed micronutrients, plasma proteins, dietary intakes, and genotype using a systems approach. Participants attended a community-based summer day program for 6-14 year old in 2 years. Genetic makeup, blood metabolite and protein levels, and dietary differences were measured in each individual. Twenty-four-hour dietary intakes, eight micronutrients (vitamins A, D, E, thiamin, folic acid, riboflavin, pyridoxal, and pyridoxine) and 3 one-carbon metabolites [homocysteine (Hcy), S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH)], and 1,129 plasma proteins were analyzed as a function of diet at metabolite level, plasma protein level, age, and sex. Cluster analysis identified two groups differing in SAM/SAH and differing in dietary intake patterns indicating that SAM/SAH was a potential marker of nutritional status. The approach used to analyze genetic association with the SAM/SAH metabolites is called middle-out: SNPs in 275 genes involved in the one-carbon pathway (folate, pyridoxal/pyridoxine, thiamin) or were correlated with SAM/SAH (vitamin A, E, Hcy) were analyzed instead of the entire 1M SNP data set. This procedure identified 46 SNPs in 25 genes associated with SAM/SAH demonstrating a genetic contribution to the methylation potential. Individual plasma metabolites correlated with 99 plasma proteins. Fourteen proteins correlated with body mass index, 49 with group age, and 30 with sex. The analytical strategy described here identified subgroups for targeted nutritional interventions.

Carbohydrate metabolic pathway genes associated with quantitative trait loci (QTL) for obesity and type 2 diabetes: identification by data mining.

Increasing consumption of refined carbohydrates is now being recognized as a primary contributor to the development of nutritionally related chronic diseases such as obesity and type 2 diabetes mellitus (T2DM). A data mining approach was used to evaluate the role of carbohydrate metabolic pathway genes in the development of obesity and T2DM. Data from public databases were used to map the position of the carbohydrate metabolic pathway genes to known quantitative trait loci (QTL) for obesity and T2DM and for examining the pathway genes for the presence of sequence and structural genetic variants such as single nucleotide polymorphisms (SNPs) and copy number variants (CNS), respectively. The results demonstrated that a majority of the genes of the carbohydrate metabolic pathways are associated with QTL for obesity and many for T2DM. In addition, some key genes of the pathways also encode non-synonymous SNPs that exhibit significant differences in population frequencies. This study emphasizes the significance of the metabolic pathways genes in the development of disease phenotypes, its differential occurrence across populations and between individuals, and a strategy for interpreting an individuals' risk for disease.

A strategy for analyzing gene-nutrient interactions in type 2 diabetes.

Type 2 diabetes mellitus (T2DM), like all chronic diseases, results from interactions between multiple genes and multiple environmental factors. Nevertheless, many research studies focus on either nutrition or genetic factors independently of each other. The challenges of analyzing gene-nutrient interactions in T2DM are the (i) genetic heterogeneity in humans, (ii) complexity of environmental factors, particularly dietary chemicals, and (iii) diverse physiologies that produce the same apparent disease. Many of these variables are not accounted for in the design or study of T2DM or, indeed, most chronic diseases, although exceptions are noteworthy. Establishing experimental paradigms to analyze the complexity of these interactions and physiologies is challenging, but possible. This article provides a strategy to extend nutrigenomic experimental strategies to include early environmental influences that may promote adult-onset disease.

Personalizing nutrigenomics research through community based participatory research and omics technologies.

Personal and public health information are often obtained from studies of large population groups. Risk factors for nutrients, toxins, genetic variation, and more recently, nutrient-gene interactions are statistical estimates of the percentage reduction in disease in the population if the risk were to be avoided or the gene variant were not present. Because individuals differ in genetic makeup, lifestyle, and dietary patterns than those individuals in the study population, these risk factors are valuable guidelines, but may not apply to individuals. Intervention studies are likewise limited by small sample sizes, short time frames to assess physiological changes, and variable experimental designs that often preclude comparative or consensus analyses. A fundamental challenge for nutrigenomics will be to develop a means to sort individuals into metabolic groups, and eventually, develop risk factors for individuals. To reach the goal of personalizing medicine and nutrition, new experimental strategies are needed for human study designs. A promising approach for more complete analyses of the interaction of genetic makeups and environment relies on community-based participatory research (CBPR) methodologies. CBPR's central focus is developing a partnership among researchers and individuals in a community that allows for more in depth lifestyle analyses but also translational research that simultaneously helps improve the health of individuals and communities. The USDA-ARS Delta Nutrition Intervention Research program exemplifies CBPR providing a foundation for expanded personalized nutrition and medicine research for communities and individuals.

Methamphetamine Treatment Affects Blood and Liver S-Adenosylmethionine (SAM) in Mice: Correlation with Dopamine Depletion in the Striatuma.

Methamphetamine (METH) is a major drug of abuse which causes neurotoxicity by depleting dopamine, its metabolites, high-affinity dopamine uptake sites and tyrosine hydroxylase activity in the striatum. Dopamine depletion and reduced dopamine transit are associated with depression. S-Adenosylmethionine (SAM) is the chief methyl donor used in dopamine and other neurotransmitter metabolism in mammals. Low SAM is associated with depression and other psychological and neurological disorders in humans. SAM is used to treat depression and some other neurological and psychiatric disorders. The present study was designed to determine if single or multiple doses of METH induce alterations in blood or liver SAM in mice and if these correlate with dopamine levels in the striatum. Adult male C57 mice were injected intraperitoneally with either single (1 × 40 mg/kg) or multiple (4 × 10 mg/kg) doses of METH. Animals were sacrificed at various intervals. A single injection of METH resulted in slightly higher blood SAM levels at 4 hr. Multiple doses of METH resulted in decreased hepatic and blood SAM levels at 72 hr. Blood SAM returned to control levels by 1 wk. Published work shows that dopamine levels increase hours after a single injection of METH, whereas dopamine decreases days after multiple injections of METH. These present data clearly demonstrate that METH dosing leads to significant alterations in liver and blood SAM and that these changes in SAM levels correlate with changes in striatal dopamine levels.