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Background: Non-human primates, such as Rhesus macaques, are a powerful model for studies of the cellular and physiological effects of radiation, development of radiation biodosimetry, and for understanding the impact of radiation on human health. Here, we study the effects of 4 Gy total body irradiation (TBI) at the molecular level out to 28 days and at the cytogenetic level out to 56 days after exposure. We combine the global transcriptomic and proteomic responses in peripheral whole blood to assess the impact of acute TBI exposure at extended times post irradiation. Results: The overall mRNA response in the first week reflects a strong inflammatory reaction, infection response with neutrophil and platelet activation. At 1 week, cell cycle arrest and re-entry processes were enriched among mRNA changes, oncogene-induced senescence and MAPK signaling among the proteome changes. Influenza life cycle and infection pathways initiate earlier in mRNA and are reflected among the proteomic changes during the first week. Transcription factor proteins SRC, TGFb and NFATC2 were immediately induced at 1 day after irradiation with increased transcriptional activity as predicted by mRNA changes persisting up to 1 week. Cell counts revealed a mild / moderate hematopoietic acute radiation syndrome (H-ARS) reaction to irradiation with expected lymphopenia, neutropenia and thrombocytopenia that resolved within 30 days. Measurements of micronuclei per binucleated cell levels in cytokinesis-blocked T-lymphocytes remained high in the range 0.27-0.33 up to 28 days and declined to 0.1 by day 56. Conclusions: Overall, we show that the TBI 4 Gy dose in NHPs induces many cellular changes that persist up to 1 month after exposure, consistent with damage, death, and repopulation of blood cells.

PURPOSE: To identify metabolomic biomarkers of acute radiation exposure in saliva that show time-dependent changes. METHODS AND MATERIALS: Non-human primates (NHPs) were exposed to 4 Gy of total body irradiation with γ-rays. Saliva was collected from seven animals twice prior to and at days 1, 3, 5, 7, 15, 21,28, and 60 after irradiation. Profiling was conducted with liquid chromatography time-of-flight mass spectrometry. Multivariate data analysis and potential biomarker identification was conducted through Random Forests and the software MetaboAnalyst. Candidate biomarkers were validated through tandem mass spectrometry (MS/MS) and receiver operating characteristic (ROC) curves were constructed to show the diagnostic ability of the signature over time. RESULTS AND CONCLUSIONS: Untargeted metabolomic analysis revealed significant and persistent effects up to the 60 days evaluated in this study. Biomarkers spanning primarily amino acids and nucleotides were identified with a significant number of them showing long term responses. Fifteen biomarkers showed high statistical significance in the first week after irradiation, and sixteen at >7 days after irradiation [false discovery rate (FDR) adjusted p

Introduction Genome-wide association studies consistently show that single nucleotide polymorphisms (SNPs) in the complement receptor 1 ( CR1 ) gene modestly but significantly alter Alzheimer's disease (AD) risk. Follow-up research has assumed that CR1 is expressed in the human brain despite a paucity of evidence for its function there. Alternatively, erythrocytes contain >80% of the body's CR1, where, in primates, it is known to bind circulating pathogens. Methods Multidisciplinary methods were employed. Results Conventional Western blots and quantitative polymerase chain reaction failed to detect CR1 in the human brain. Brain immunohistochemistry revealed only vascular CR1. By contrast, erythrocyte CR1 immunoreactivity was readily observed and was significantly deficient in AD, as was CR1-mediated erythrocyte capture of circulating amyloid β peptide. CR1 SNPs associated with decreased erythrocyte CR1 increased AD risk, whereas a CR1 SNP associated with increased erythrocyte CR1 decreased AD risk. Discussion SNP effects on erythrocyte CR1 likely underlie the association of CR1 polymorphisms with AD risk.

Whole body exposure to ionizing radiation (IR) (> 0.7 Gy) damages tissues leading to a range of physical symptoms contributing to acute radiation syndrome (ARS). Radiation biodosimetry aims to determine characteristic early biomarkers indicative of radiation exposure (generally at doses > 2 Gy) and is a necessity for effective triage in the event of an unanticipated radiological incident and emergency preparedness. Radiation metabolomics can address this aim by assessing metabolic perturbations following various emergency scenarios (e.g., elapsed time to medical care, absorbed dose, combined injury). Gas chromatography – mass spectrometry (GC-MS) is a standardized platform ideal for chromatographic separation, identification, and quantification of metabolites to discriminate molecular signatures that can be utilized in assessing radiation injury. We performed GC time-of-flight (TOF) MS for global profiling of nonhuman primate (NHP) urine and serum samples up to 60 d after a single 4 Gy γ-ray total body exposure. Multivariate statistical analysis showed a higher separation of groups from urine signatures vs. serum signatures. We identified biofluid markers involved in amino acid, lipid, purine, and serotonin metabolism, some of which may indicate host microbiome dysbiosis. Sex differences were observed amino acid fold changes in serum samples. Additionally, we explored mitochondrial dysfunction by analysis of tricarboxylic acid (TCA) intermediates with a GC tandem quadrupole (TQ) MS platform in samples collected in a time course during the first week (1, 3, 5, and 7 d) after exposure. By adding this temporal component to our previous work exploring dose effects at a single time point of 7 d, we observed the highest fold changes occurring at 3 d, returning closer to basal levels by 7 d. These results emphasize the utility of both MS-based metabolomics for biodosimetry and complementary analytical platforms for increased metabolome coverage.

Rapid assessment of radiation signatures in non-invasive biofluids may aid in assigning proper medical treatments for acute radiation syndrome (ARS) and delegating limited resources after a nuclear disaster. Metabolomic platforms allow for rapid screening of biofluid signatures and show promise in differentiating radiation quality and time postexposure. Here, we use global metabolomics to differentiate temporal effects (1–60 d) found in nonhuman primate (NHP) urine and serum small molecule signatures after a 4 Gy total body irradiation. Random Forests analysis differentially classifies biofluid signatures according to days post 4 Gy exposure. Eight compounds involved in protein metabolism, fatty acid β oxidation, DNA base deamination, and general energy metabolism were identified in each urine and serum sample and validated through tandem MS. The greatest perturbations were seen at 1 d in urine and 1–21 d in serum. Furthermore, we developed a targeted liquid chromatography tandem mass spectrometry (LC–MS/MS) with multiple reaction monitoring (MRM) method to quantify a six compound panel (hypoxanthine, carnitine, acetylcarnitine, proline, taurine, and citrulline) identified in a previous training cohort at 7 d after a 4 Gy exposure. The highest sensitivity and specificity for classifying exposure at 7 d after a 4 Gy exposure included carnitine and acetylcarnitine in urine and taurine, carnitine, and hypoxanthine in serum. Receiver operator characteristic (ROC) curve analysis using combined compounds show excellent sensitivity and specificity in urine (area under the curve [AUC] = 0.99) and serum (AUC = 0.95). These results highlight the utility of MS platforms to differentiate time postexposure and acquire reliable quantitative biomarker panels for classifying exposed individuals.

What is already known about this subject Initial evidence from recent studies indicated a relationship between the high-activity MAOA uVNTR polymorphism and lower risk of obesity even though findings were varied across populations, and hence replication studies with population-based samples are clearly warranted. What this study adds Our findings confirmed the genetic effects of MAOA uVNTR polymorphism on body mass index in a Chinese adolescent population. This study demonstrated a significant gene–environment interaction with negative familial stressors in girls only, which provides a new insight on the causes of obesity, and thus, potential avenues of prevention. Summary Objectives Monoamine oxidase A (MAOA) modulates metabolism of serotonin and dopamine metabolism, neurotransmitters involved in regulation of appetite and food intake. The gene coding for MAOA contains a 30-bp tandem repeat (uVNTR) polymorphism in its promoter region that has been previously identified to be associated with obesity with mixed findings in the literature. Our goals were to replicate the population effects of this functional polymorphism on obesity risk, and to further explore gender differences and interaction effects with negative stressors. Methods Analyses were conducted with data on genotypes, measured weight and height, and self-reported behavioural characteristics among 1101 Chinese adolescents 11–15 years old living in Wuhan, China. Results Girls with the high-activity allele had significantly lower body mass index (BMI; β = −0.25 ± 0.98, P = 0.011) compared to those with the low activity allele. Experience of negative familial stressors (e.g., death or illness of family members, hit or scolded by parents and increased quarrelling with parents, parents argued frequently) significantly weakened this protective genetic effect on BMI (P for interaction = 0.043). Stratified analyses showed a significant protective genetic effect on BMI only within the stratum of low stress level (β = −0.44 ± 0.14, P = 0.002). No similar effect was observed among boys. Conclusions Our findings confirm the genetic effects of MAOA uVNTR polymorphism on BMI in a Chinese adolescent population and suggest potential genetic interactions with negative familial stressors.

Introduction There is increasing evidence that response to pharmacological treatment for nicotine dependence may be moderated by genetic polymorphisms. However, the feasibility, acceptability, and impact of genetically tailoring treatment in real-world clinical settings are unknown. Methods We conducted a multiphased, mixed-methods feasibility study with current smokers to develop and evaluate a patient-centered, theoretically grounded personalized medicine treatment protocol. The initial research phase included formative work to develop intervention materials. The second phase included a randomized pilot trial to evaluate the intervention. Trial participants (n = 36) were genotyped for ANKK1 rs1800497 and were randomized to receive genetic feedback (GF) plus standard behavioral counseling (BC) for smoking cessation or BC without GF. All participants received genetically tailored pharmacotherapy (nicotine patch or bupropion). Results The intervention was feasible to implement and was acceptable to participants based on satisfaction ratings and objective measures of participation. There was no evidence that the GF resulted in adverse psychological outcomes (e.g., depression, fatalism, reduced perceived control over quitting, differential motivation for quitting) based on quantitative or qualitative outcomes. Conclusions Study results suggest that it is feasible to offer treatment within a health care setting that includes genetically tailored pharmacotherapy and doing so had no apparent adverse psychological impacts. Further evaluation of pharmacogenetically tailored smoking cessation interventions appears warranted.

We investigated whole saliva as a source of biomarkers to distinguish individuals who have, and who have not, been chronically exposed to severe and threatening life difficulties. We evaluated RNA and DNA metrics, expression of 37 candidate genes, and cortisol release in response to the Trier Social Stress Test, as well as clinical characteristics, from 48 individuals stratified on chronic exposure to psychosocial stressors within the last year as measured by the Life Events and Difficulties Schedule. Candidate genes were selected based on their differential gene expression ratio in circulating monocytes from a published genome-wide analysis of adults experiencing different levels of exposure to a chronic stressor. In univariate analyses, we observed significantly decreased RNA integrity (RIN) score (P = 0.04), and reduced expression of glucocorticoid receptor-regulated genes (Ps < 0.05) in whole saliva RNA from individuals exposed to chronic stressors, as compared to those with no exposure. In those exposed, we observed significantly decreased BMI (P < 0.001), increased ever-smoking and increased lifetime alcohol abuse or dependence (P ≤ 0.03), and a reduction of cortisol release. In post hoc multivariate analyses including clinical and biospecimen-derived variables, we consistently observed significantly decreased expression of IL8 (Ps

Although previous investigations have indicated a role for genetic factors in smoking initiation, the underlying genetic mechanisms are still unknown. In 2,339 adolescents from a Chinese Han population in the Wuhan Smoking Prevention Trial (Wuhan, China, 1998-1999), the authors explored the association of 57 genes in the dopamine pathway with smoking initiation. Using a conservative approach for declaring significance, positive findings were further examined in an independent sample of 603 Caucasian adolescents followed for up to 10 years as part of the Children's Health Study (Southern California, 1993-2009). The authors identified 1 single nucleotide polymorphism (rs2298122) in the calcyon neuron-specific vesicular protein gene (CALY) that was positively associated with smoking initiation in females (odds ratio = 2.21, 95% confidence interval: 1.49, 3.27; P = 8.4 × 10(-5)) in the Wuhan Smoking Prevention Trial cohort, and they replicated the association in females from the Children's Health Study cohort (hazard rate ratio = 2.05, 95% confidence interval: 1.27, 3.31; P = 0.003). These results suggest that the CALY gene may influence smoking initiation in adolescents, although the potential roles of underlying psychological characteristics that may be components of the smoking-initiation phenotype, such as impulsivity or novelty-seeking, remain to be explored.

We utilized a cohort of 828 treatment-seeking self-identified white cigarette smokers (50% female) to rank candidate gene single nucleotide polymorphisms (SNPs) associated with the Fagerström Test for Nicotine Dependence (FTND), a measure of nicotine dependence which assesses quantity of cigarettes smoked and time- and place-dependent characteristics of the respondent's smoking behavior. A total of 1123 SNPs at 55 autosomal candidate genes, nicotinic acetylcholine receptors and genes involved in dopaminergic function, were tested for association to baseline FTND scores adjusted for age, depression, education, sex, and study site. SNP P-values were adjusted for the number of transmission models, the number of SNPs tested per candidate gene, and their intragenic correlation. DRD2, SLC6A3, and NR4A2 SNPs with adjusted P-values

CYP2A6 is the major enzyme involved in nicotine metabolism, yet large interindividual variations in the rate of nicotine metabolism exist within groups of individuals having the same CYP2A6 genotype. We investigated the influence of genetic variation in another potential nicotine-metabolizing enzyme, CYP2B6, and its interaction with CYP2A6, on the metabolism of nicotine.Two hundred and twelve healthy Caucasian adult twin volunteers underwent an intravenous infusion of stable isotope-labeled nicotine and its major metabolite, cotinine, for characterization of pharmacokinetic and metabolism phenotypes. Five CYP2B6 genetic polymorphisms causing amino acid substitutions (R22C, Q172 H, S259R, K262R, and R487C) were analyzed.We observed that the CYP2B6*6 haplotype (defined as having both Q172 H and K262R variants) was associated with faster nicotine and cotinine clearance, and that such associations were more prominent among individuals having decreased-activity CYP2A6 genotypes. Statistically significant interactions between CYP2B6 and CYP2A6 genotypes were observed (P0.003 for nicotine clearance and P0.002 for cotinine clearance).Our results indicate that CYP2B6 genetic variation is associated with the metabolism of nicotine and cotinine among individuals with decreased CYP2A6 activity. Further investigation of the roles of CYP2B6 and the interaction between CYP2B6 and CYP2A6 genotypes in mediating nicotine dependence and tobacco-related diseases is merited.

© 2015 Bergen et al. The Nicotine Metabolite Ratio (NMR, ratio of trans-3'-hydroxycotinine and cotinine), has previously been associated with CYP2A6 activity, response to smoking cessation treatments, and cigarette consumption. We searched for drug metabolizing enzyme and transporter (DMET) gene variation associated with the NMR and prospective abstinence in 2,946 participants of laboratory studies of nicotine metabolism and of clinical trials of smoking cessation therapies. Stage I was a meta-analysis of the association of 507 common single nucleotide polymorphisms (SNPs) at 173 DMET genes with the NMR in 449 participants of two laboratory studies. Nominally significant associations were identified in ten genes after adjustment for intragenic SNPs; CYP2A6 and two CYP2A6 SNPs attained experiment-wide significance adjusted for correlated SNPs (CYP2A6 PACT=4.1E-7, rs4803381 PACT=4.5E-5, rs1137115, PACT=1.2E-3). Stage II was mega-regression analyses of 10 DMET SNPs with pretreatment NMR and prospective abstinence in up to 2,497 participants from eight trials. rs4803381 and rs1137115 SNPs were associated with pretreatment NMR at genome-wide significance. In post-hoc analyses of CYP2A6 SNPs, we observed nominally significant association with: abstinence in one pharmacotherapy arm; cigarette consumption among all trial participants; and lung cancer in four case:control studies. CYP2A6 minor alleles were associated with reduced NMR, CPD, and lung cancer risk. We confirmed the major role that CYP2A6 plays in nicotine metabolism, and made novel findings with respect to genome-wide significance and associations with CPD, abstinence and lung cancer risk. Additional multivariate analyses with patient variables and genetic modeling will improve prediction of nicotine metabolism, disease risk and smoking cessation treatment prognosis.

The mu1 opioid receptor gene, OPRM1, has long been a high-priority candidate for human genetic studies of addiction. Because of its potential functional significance, the non-synonymous variant rs1799971 (A118G, Asn40Asp) in OPRM1 has been extensively studied, yet its role in addiction has remained unclear, with conflicting association findings. To resolve the question of what effect, if any, rs1799971 has on substance dependence risk, we conducted collaborative meta-analyses of 25 datasets with over 28,000 European-ancestry subjects. We investigated non-specific risk for “general” substance dependence, comparing cases dependent on any substance to controls who were non-dependent on all assessed substances. We also examined five specific substance dependence diagnoses: DSM-IV alcohol, opioid, cannabis, and cocaine dependence, and nicotine dependence defined by the proxy of heavy/light smoking (cigarettes-per-day >20 vs. ≤10). The G allele showed a modest protective effect on general substance dependence (OR = 0.90, 95 % C.I. [0.83–0.97], p value = 0.0095, N = 16,908). We observed similar effects for each individual substance, although these were not statistically significant, likely because of reduced sample sizes. We conclude that rs1799971 contributes to mechanisms of addiction liability that are shared across different addictive substances. This project highlights the benefits of examining addictive behaviors collectively and the power of collaborative data sharing and meta-analyses.

Author(s): Bergen, Andrew W; Javitz, Harold S; Krasnow, Ruth; Michel, Martha; Nishita, Denise; Conti, David V; Edlund, Christopher K; Kwok, Pui-Yan; McClure, Jennifer B; Kim, Richard B; Hall, Sharon M; Tyndale, Rachel F; Baker, Timothy B; Benowitz, Neal L; Swan, Gary E | Abstract: IntroductionWe evaluated chr6q25.3 organic cation transporter gene (SLC22A1, SLC22A2, SLC22A3) variation and response to smoking cessation therapies. The corresponding proteins are low-affinity transporters of choline, acetylcholine and monoamines, and smoking cessation pharmacotherapies expressed in multiple tissues.MethodsWe selected 7 common polymorphisms for mega-regression analysis. We assessed additive model association of polymorphisms with 7-day point prevalence abstinence overall and by assigned pharmacotherapy at end of treatment and at 6 months among European-ancestry participants of 7 randomized controlled trials adjusted for demographic, population genetic, and trial covariates.ResultsInitial results were obtained in 6 trials with 1,839 participants. Nominally statistically significant associations of 2 SLC22A2 polymorphisms were observed: (1) with rs316019 at 6 months, overall ([c.808TgG; p.Ser270Ala], OR = 1.306, 95% CI = 1.034-1.649, p = .025), and among those randomized to nicotine replacement therapy (NRT) (OR = 1.784, 95% CI = 1.072-2.970, p = .026); and (2) with rs316006 (c.1502-529AgT) among those randomized to varenicline (OR = 1.420, 95% CI = 1.038-1.944, p = .028, OR = 1.362, 95% CI = 1.001-1.853, p = .04) at end of treatment and 6 months. Individuals randomized to NRT from a seventh trial were genotyped for rs316019; rs316019 was associated with a nominally statistically significant effect on abstinence overall at 6 months among 2,233 participants (OR = 1.249, 95% CI = 1.007-1.550, p = .043).ConclusionsThe functional OCT2 Ser270Ala polymorphism is nominally statistically significantly associated with abstinence among European-ancestry treatment-seeking smokers after adjustments for pharmacotherapy, demographics, population genetics, and without adjustment for multiple testing of 7 SNPs. Replication of these preliminary findings in additional randomized controlled trials of smoking cessation therapies and from multiple continental populations would describe another pharmacogenetic role for SLC22A2/OCT2.

OBJECTIVE: To evaluate the association of nicotinic acetylcholine receptor (nAChR) single nucleotide polymorphism (SNP) with 7-day point prevalence abstinence (abstinence) in randomized clinical trials of smoking cessation therapies in individuals grouped by pharmacotherapy randomization to inform the development of personalized smoking cessation therapy. MATERIALS AND METHODS: We quantified association of four SNPs at three nAChRs with abstinence in eight randomized clinical trials. Participants were 2633 outpatient treatment-seeking, self-identified European ancestry individuals smoking at least 10 cigarettes/day, recruited through advertisement, prescribed pharmacotherapy, and provided with behavioral therapy. Interventions included nicotine replacement therapy (NRT), bupropion, varenicline, placebo (PLA), or combined NRT and bupropion, and five modes of group and individual behavioral therapy. Outcome measures tested in multivariate logistic regression were end of treatment and 6 month (6MO) abstinence, with demographic, behavioral, and genetic covariates. RESULTS: 'Risk' alleles previously associated with smoking heaviness were significantly (P

Background Clinical trial and epidemiological studies need high quality biospecimens from a representative sample of participants to investigate genetic influences on treatment response and disease. Obtaining blood biospecimens presents logistical and financial challenges. As a result, saliva biospecimen collection is becoming more frequent because of the ease of collection and lower cost. This article describes an assessment of saliva biospecimen samples collected through the mail, trial participant demographic and behavioral characteristics, and their association with saliva and DNA quantity and quality. Methods Saliva biospecimens were collected using the Oragene® DNA Self-Collection Kits from participants in a National Cancer Institute funded smoking cessation trial. Saliva biospecimens from 565 individuals were visually inspected for clarity prior to and after DNA extraction. DNA samples were then quantified by UV absorbance, PicoGreen®, and qPCR. Genotyping was performed on 11 SNPs using TaqMan® SNP assays and two VNTR assays. Univariate, correlation, and analysis of variance analyses were conducted to observe the relationship between saliva sample and participant characteristics. Results The biospecimen kit return rate was 58.5% among those invited to participate (n = 967) and 47.1% among all possible COMPASS participants (n = 1202). Significant gender differences were observed with males providing larger saliva volume (4.7 vs. 4.5 ml, p = 0.019), samples that were more likely to be judged as cloudy (39.5% vs. 24.9%, p < 0.001), and samples with greater DNA yield as measured by UV (190.0 vs. 138.5, p = 0.002), but reduced % human DNA content (73.2 vs. 77.6 p = 0.005) than females. Other participant characteristics (age, self-identified ethnicity, baseline cigarettes per day) were associated with saliva clarity. Saliva volume and saliva and DNA clarity were positively correlated with total DNA yield by all three quantification measurements (all r > 0.21, P < 0.001), but negatively correlated with % human DNA content (saliva volume r = -0.148 and all P < 0.010). Genotyping completion rate was not influenced by saliva or DNA clarity. Conclusion Findings from this study show that demographic and behavioral characteristics of smoking cessation trial participants have significant associations with saliva and DNA metrics, but not with the performance of TaqMan® SNP or VNTR genotyping assays. Trial registration COMPASS; registered as NCT00301145 at clinicaltrials.gov.