ABT-751, a novel orally available antitubulin agent, is mainly eliminated as inactive glucuronide (ABT-751G) and sulfate (ABT-751S) conjugates. We performed a pharmacogenetic investigation of ABT-751 pharmacokinetics using in-vitro data to guide the selection of genes for genotyping in a phase I trial of ABT-751.
UDP-glucuronosyltransferase (UGT) and sulfotransferase (SULT) enzymes were screened for ABT-751 metabolite formation in vitro. Forty-seven cancer patients treated with ABT-751 were genotyped for 21 variants in these genes.
UGT1A1, UGT1A4, UGT1A8, UGT2B7, and SULT1A1 were found to be involved in the formation of inactive ABT-751 glucuronide (ABT-751G) and sulfate (ABT-751S). SULT1A1 copy number (> 2) was associated with an average 34% increase in ABT-751 clearance (P= 0.044), an 18% reduction in ABT-751 AUC (P = 0.045), and a 50% increase in sulfation metabolic ratios (P=0.025). UGT1A8 rs6431558 was associated with a 28% increase in glucuronidation metabolic ratios (P =0.022), and UGT1A4*2 was associated with a 65% decrease in ABT-751 Ctrough (P = 0.009).
These results might represent the first example of a clinical pharmacokinetic effect of the SULT1A1 copy number variant on the clearance of a SULT1A1 substrate. A-priori selection of candidate genes guided by in-vitro metabolic screening enhanced our ability to identify genetic determinants of interpatient pharmacokinetic variability.
ABT-751; drug development; drug metabolism; pharmacogenetics; phase I; sulfotransferase; UDP-glucuronosyltransferase
Prior candidate gene studies have associated CYP2B6 516G→T [rs3745274] and 983T→C [rs28399499] with increased plasma efavirenz exposure. We sought to identify novel variants associated with efavirenz pharmacokinetics.
Materials and methods
Antiretroviral therapy-naive AIDS Clinical Trials Group studies A5202, A5095, and ACTG 384 included plasma sampling for efavirenz pharmacokinetics. Log-transformed trough efavirenz concentrations (Cmin) were previously estimated by population pharmacokinetic modeling. Stored DNA was genotyped with Illumina HumanHap 650Y or 1MDuo platforms, complemented by additional targeted genotyping of CYP2B6 and CYP2A6 with MassARRAY iPLEX Gold. Associations were identified by linear regression, which included principal component vectors to adjust for genetic ancestry.
Among 856 individuals, CYP2B6 516G→T was associated with efavirenz estimated Cmin (P = 8.5 × 10−41). After adjusting for CYP2B6 516G→T, CYP2B6 983T→C was associated (P = 9.9 × 10−11). After adjusting for both CYP2B6 516G→T and 983T→C, a CYP2B6 variant (rs4803419) in intron 3 was associated (P = 4.4 × 10−15). After adjusting for all the three variants, non-CYP2B6 polymorphisms were associated at P-value less than 5× 10−8. In a separate cohort of 240 individuals, only the three CYP2B6 polymorphisms replicated. These three polymorphisms explained 34% of interindividual variability in efavirenz estimated Cmin. The extensive metabolizer phenotype was best defined by the absence of all three polymorphisms.
Three CYP2B6 polymorphisms were independently associated with efavirenz estimated Cmin at genome-wide significance, and explained one-third of interindividual variability. These data will inform continued efforts to translate pharmacogenomic knowledge into optimal efavirenz utilization.
CYP2B6; efavirenz; HIV; pharmacogenomics; pharmacokinetics
ABCC4; ABCB1; HIV infection; UGT2B7; zidovudine
In a previous analysis involving protocol ANRS 12154, interindividual variability in steady-state nevirapine clearance among HIV-infected Cambodians was partially explained by CYP2B6 516G→T (CYP2B6*6). Here, we examine whether additional genetic variants predict nevirapine clearance in this cohort.
Analyses included Phnom Penh ESTHER (Ensemble pour une Solidarité Thérapeutique Hospitalière en Réseau) cohort participants who had consented for genetic testing. All participants were receiving nevirapine plus two nucleoside analogs. The mean individual nevirapine clearance estimates were derived from a population model developed on nevirapine concentrations at 18 and 36 months of therapy. Polymorphisms were assayed in ABCB1, CYP2A6, CYP2B6, CYP2C19, CYP3A4, CYP3A5, and NR1I2.
Of 198 assayed loci, 130 were polymorphic. Among 129 individuals with evaluable genetic data, nevirapine clearance ranged from 1.06 to 5.00 l/h in 128 individuals and was 7.81 l/h in one individual. In bivariate linear regression, CYP2B6 516G→T (CYP2B6*6) was associated with lower nevirapine clearances (P = 3.5 × 10–6). In a multivariate linear regression model conditioned on CYP2B6 516G→T, independent associations were identified with CYP2B6 rs7251950, CYP2B6 rs2279343, and CYP3A4 rs2687116. The CYP3A4 association disappeared after censoring the outlier clearance value. A model that included CYP2B6 516G→T (P = 1.0 × 10–9), rs7251950 (P = 4.8 × 10–5), and rs2279343 (P = 7.1 × 10–5) explained 11% of interindividual variability in nevirapine clearance.
Among HIV-infected Cambodians, several CYP2B6 polymorphisms were associated independently with steady-state nevirapine clearance. The prediction of nevirapine clearance was improved by considering several polymorphisms in combination.
Cambodia; CYP2B6; nevirapine pharmacokinetics; pharmacogenetics; population approach
AMP-activated protein kinase; diabetes mellitus; metformin; multidrug and toxin extrusion 1; OCT1; OCT2; pathway; pharmacodynamics; pharmacogenomic; pharmacokinetics; type 2 diabetes
Recently, lymphoblastoid cell lines (LCLs) have emerged as an innovative model system for mapping gene variants that predict dose response to chemotherapy drugs. In the current study, this strategy was expanded to the in vitro genome-wide association approach, using 516 LCLs derived from a Caucasian cohort to assess cytotoxic response to temozolomide. Genome-wide association analysis using approximately 2.1 million quality controlled single-nucleotide polymorphisms (SNPs) identified a statistically significant association (p < 10−8) with SNPs in the O6-methylguanine–DNA methyltransferase (MGMT) gene. We also demonstrate that the primary SNP in this region is significantly associated with differential gene expression of MGMT (p< 10−26) in LCLs, and differential methylation in glioblastoma samples from The Cancer Genome Atlas. The previously documented clinical and functional relationships between MGMT and temozolomide response highlight the potential of well-powered GWAS of the LCL model system to identify meaningful genetic associations.
lymphoblastoid cell lines; temozolomide; GWAS; MGMT; pharmacogenetics
drug response; genetic variants; pharmacogenomics; vitamin D receptor
A shift towards overall larger very low-density lipoprotein (VLDL), and smaller low-density lipoprotein and high-density lipoprotein (HDL) diameters occurs in insulin resistance (IR), which reflects shifts in the distribution of the subfraction concentrations. Fenofibrate, indicated for hypertriglyceridemia, simultaneously reduces IR and shifts in lipoprotein diameter. Individual responses to fenofibrate vary, and we conducted a genome-wide association study to identify genetic differences that could contribute to such differences.
Association analysis was conducted between single nucleotide polymorphisms (SNPs) on the Affymetrix 6.0 array and fasting particle diameter responses to a 12-week fenofibrate trial, in 817 related Caucasian participants of the Genetics of Lipid Lowering Drugs and Diet Network. Linear models were conducted, which adjusted for age, sex and study center as fixed effects, and pedigree as a random effect. The top three SNPs associated with each fraction were examined subsequently for associations with changes in subfraction concentrations.
SNPs in AHCYL2 and CD36 genes reached, or closely approached, genome-wide levels of significance with VLDL and HDL diameter responses to fenofibrate, respectively (P=4 × 10−9 and 8 × 10−8). SNPs in AHCYL2 were associated with a decrease in the concentration of the large VLDL subfraction only (P = 0.002). SNPs associated with HDL diameter change were not associated with a single subfraction concentration change (P > 0.05) indicating small shifts across all subfractions.
We report novel associations between lipoprotein diameter responses to fenofibrate and the AHCYL2 and CD36 genes. Previous associations of these genes with IR emphasize the role of IR in mediating lipoprotein response to fenofibrate.
AHCYL2; CD36; fenofibrate; inflammation; insulin resistance; insulin signaling; lipoprotein diameter; methylation; PPARγ; subclass
FKBP51 (51 kDa immunophilin) acts as a modulator of the glucocorticoid receptor and a negative regulator of the Akt pathway. Genetic variation in FKBP5 plays a role in antidepressant response. The aim of this study was to comprehensively assess the role of genetic variation in FKBP5, identified by both Sanger and Next Generation DNA resequencing, as well as genome-wide single nucleotide polymorphisms (SNPs) associated with FKBP5 expression in the response to the selective serotonin reuptake inhibitor (SSRI) treatment of major depressive disorder.
We identified 657 SNPs in FKBP5 by Next Generation sequencing of 96 DNA samples from white patients, and 149 SNPs were selected for the genotyping together with 235 SNPs that were trans-associated with variation in FKBP5 expression in lymphoblastoid cells. A total of 529 DNA samples from the Mayo Clinic PGRN-SSRI Pharmacogenomic trial for which genome-wide SNPs had already been obtained were genotyped for these 384 SNPs, and associations with treatment outcomes were determined. The most significant SNPs were genotyped using 96 DNA samples from white non-Hispanic patients of the NIMH-supported Sequenced Treatment Alternatives to Relieve Depression (STAR*D) study to attempt replication, followed by functional genomic studies.
Genotype–phenotype association analysis indicated that rs352428 was associated with both 8-week treatment response in the Mayo study (odds ratio =0.49; P = 0.003) and 6-week response in the STAR*D replication study (odds ratio = 0.74; P =0.05). The electrophoresis mobility shift assay and the reporter gene assay confirmed the possible role of this SNP in transcription regulation.
This comprehensive FKBP5 sequence study provides insight into the role of common genetic polymorphisms that might influence SSRI treatment outcomes in major depressive disorder patients.
FKBP5; genotype–phenotype association; major depressive disorder; Next Generation DNA resequencing; selective serotonin reuptake inhibitor; single nucleotide polymorphism
CYP2A6; inter-individual variation; pharmacokinetics; genetic polymorphisms; drug metabolism; drug efficacy
CYP3A5; CYP3A5*2; CYP3A5*3; CYP3A5*6; CYP3A5*7; pharmacogenomics; rs10264272; rs28365083; rs76293380; rs776746
Oxidative stress is integral to the development of endothelial dysfunction and cardiovascular disease. As NRF2 is a key transcription factor in antioxidant defense, we aimed to determine whether polymorphisms within the promoter region of the gene encoding NRF2 (NFE2L2) would significantly modify vasodilator responses in humans.
Associations between the – 653A/G (rs35652124), – 651G/A (rs6706649), and – 617C/A (rs6721961) polymorphisms within the NFE2L2 promoter and vascular function were evaluated in healthy African-American (n= 64) and white (n= 184) individuals. Forearm blood flow (FBF) was measured by strain-gauge venous occlusion plethysmography at baseline and in response to incremental doses of bradykinin or sodium nitroprusside. Forearm vascular resistance (FVR) was calculated as the mean arterial pressure/FBF.
In African Americans, – 653G variant allele carriers had significantly lower FBF and higher FVR under basal conditions as well as in response to bradykinin or sodium nitroprusside compared with wild-type individuals (P< 0.05 for each comparison). In whites, although no significant associations were observed with the – 653A/G genotype, – 617A variant allele carriers had significantly higher FVR at baseline and in response to bradykinin or sodium nitroprusside compared with wild-type individuals (P < 0.05 for each comparison). The – 651G/A polymorphism was not associated with vasodilator responses in either racial group.
Polymorphisms within the NFE2L2 promoter were associated with impaired forearm vasodilator responses in an endothelial-independent manner, suggesting an important role of NRF2 in the regulation of vascular function in humans.
bradykinin; NFE2L2; NRF2; polymorphism; sodium nitroprusside; vascular function
Nicotine metabolism and genetic variation have an impact on nicotine addiction and smoking abstinence, but further research is required. The nicotine metabolite ratio (NMR) is a robust biomarker of nicotine metabolism used to categorize slow and normal nicotine metabolizers (lower 25th quartile cutoff). In two randomized clinical trials of smoking abstinence treatments, we conducted NMR-stratified analyses on smoking abstinence across 13 regions coding for nicotinic acetylcholine receptors and proteins involved in the dopamine reward system. Gene × NMR interaction P-values were adjusted for multiple correlated tests, and we used a Bonferroni-corrected α-level of 0.004 to determine system-wide significance. Three SNPs in DRD1 (rs11746641, rs2168631, rs11749035) had significant interactions (0.001 ≤ adjusted P-values ≤ 0.004), with increased odds of abstinence within slow metabolizers (ORs=3.1–3.5, 95% CI 1.7–6.7). Our findings support the role of DRD1 in nicotine dependence, and identify genetic and nicotine metabolism profiles that may interact to impact nicotine dependence.
Genetic association studies; heterogeneity; smoking abstinence; nicotine metabolism; nicotine metabolite ratio; DRD1
Clinical use of paclitaxel is limited by variable responses and the potential for significant toxicity. To date, studies of association between variants in candidate genes and paclitaxel effects have yielded conflicting results. We sought to evaluate relationships between global gene expression and paclitaxel sensitivity.
We utilized well-genotyped lymphoblastoid cell lines derived from the International HapMap Project to evaluate relationships between cellular susceptibility to paclitaxel and global gene expression. Cells were exposed to varying concentrations of paclitaxel to evaluate paclitaxel-induced cytotoxicity and apoptosis. Among the top genes, we identified solute carrier (SLC) genes associated with paclitaxel sensitivity and narrowed down the list to those that had single nucleotide polymorphisms (SNPs) associated with both their expression level of the SLC gene and also with paclitaxel sensitivity. We performed independent validation in an independent set of cell lines and also conducted functional studies using RNA interference.
Of all genes associated with paclitaxel-induced cytotoxicity at p<0.05 (1713 genes), there was a significant enrichment of SLC genes (31 genes). A subset of SLC genes, namely SLC31A2, SLC43A1, SLC35A5, and SLC41A2, were associated with paclitaxel sensitivity and had regulating SNPs that were also associated with paclitaxel-induced cytotoxicity. Multivariate modeling demonstrated that those 4 SLC genes jointly explain 20% of the observed variability in paclitaxel susceptibility. Using RNA interference, we demonstrated increased paclitaxel susceptibility with knockdown of 3 SLC genes, SLC31A2, SLC35A5, and SLC41A2.
Our findings are novel and lend further support to the role of transporters, specifically solute carriers in mediating cellular susceptibility to paclitaxel.
paclitaxel; solute carrier genes; transporters; lymphoblastoid cell lines; pharmacogenomics
Aspirin, a major anti-platelet and cancer preventing drug, irreversibly blocks the cyclooxygenase activity of prostaglandin H synthase-1 (PGHS-1). Considerable differences in aspirin effectiveness are observed between individuals, and some of this variability may be due to PGHS-1 protein variants. Our overall aim is to determine which, if any, of the known variants in the mature PGHS-1 protein lead to functional alterations in cyclooxygenase catalysis or inhibition by aspirin. The present study targeted four PGHS-1 variants: R53H, R108Q, L237M and V481I.
Wildtype human PGHS-1 and the four polymorphic variants were expressed as histidine-tagged, homodimeric proteins in insect cells using baculovirus vectors, solubilized with detergent, and purified by affinity chromatography. The purified proteins were characterized in vitro to evaluate cyclooxygenase and peroxidase catalytic parameters and the kinetics of cyclooxygenase inhibition by aspirin and NS-398.
Compared to wildtype, several variants exhibited a higher COX/POX ratio (up to 1.5-fold, for R108Q), an elevated arachidonate Km (up to 1.9-fold, for R108Q), and/or a lower aspirin reactivity (up to 60% less, for R108Q). The decreased aspirin reactivity in R108Q reflected both a 70% increase in the Ki for aspirin and a 30% decrease in the rate constant for acetyl group transfer to the protein. Computational modeling of the brief aspirin pulses experienced by PGHS-1 in circulating platelets during daily aspirin dosing predicted that the 60% lower aspirin reactivity in R108Q gives a 15-fold increase in surviving cyclooxygenase activity; smaller, ~2-fold increases in surviving cyclooxygenase activity were predicted for L237M and V481I. NS-398 competitively inhibited cyclooxygenase catalysis of the wildtype (Ki = 6 μM) and inhibited cyclooxygenase inactivation by 1.0 mM aspirin in both wildtype (IC50 = 0.8 μM) and R108Q (IC50 = 2.1 μM).
Of the four PGHS-1 variants examined, R108Q has the largest functional effects, with evidence for impaired interactions with cyclooxygenase substrate and inhibitors. As Arg108 is located on the protein surface and not in the active site, the effects of R108Q suggest a novel, unsuspected mechanism for modulation of the PGHS-1 active site structure. The lower intrinsic aspirin reactivity of R108Q, V481I and L237M, combined with the rapid hydrolysis of aspirin in the blood, suggests that these variants decrease the anti-platelet effectiveness of the drug. These PGHS-1 variants are uncommon but aspirin is very widely used, so a considerable number of individuals could b e affected. Further examination of these and other PGHS-1 variants will be needed to determine whether PGHS-1 genotyping can be used to personalize anti-cyclooxygenase therapy.
prostaglandin H synthase-1; polymorphisms; cyclooxygenase; aspirin
We aimed to define effects of PPARγ and PPARα agonist mono and combination therapy on adipose tissue and skeletal muscle gene expression in relation to insulin sensitivity. We further investigated the role of genetic polymorphisms in PPAR ligand-modulated genes in transcriptional regulation and glucose homeostasis.
Genome-wide transcript profiles of subcutaneous adipose and skeletal muscle and metabolic phenotypes were determined before and after 10 weeks of pioglitazone and fenofibrate mono or combination therapy in 26 subjects with impaired glucose tolerance. To establish the functional role of SNPs in genes modulated by pioglitazone alone or in combination with fenofibrate, we interrogated genome-wide association data of continuous glycemic phenotypes from the MAGIC study and adipose eQTL data from the MuTHER study.
PPARγ, alone or in combination with PPARα agonists, mediated up-regulation of genes involved in the TCA cycle, branched chain amino acid metabolism, fatty acid metabolism, PPAR signaling, AMPK and cAMP signaling, and insulin signaling pathways, and downregulation of genes in antigen processing and presentation, immune and inflammatory response in adipose tissue. Remarkably few changes were found in muscle. Strong enrichment of genes involved in propanoate metabolism, fatty acid elongation in mitochondria, and acetyl-CoA metabolic process were observed only in adipose tissue of the combined pioglitazone and fenofibrate treatment group. After interrogating MAGIC data, SNPs in 22 genes modulated by PPAR ligands were associated with fasting plasma glucose and the expression of 28 transcripts modulated by PPAR ligands was under control of local genetic regulators (cis-eQTLs) in adipose tissue of MuTHER study twins.
We found differences in transcriptional mechanisms that may describe insulin sensitizing effects of PPARγ agonist monotherapy or in combination with PPARα agonist. The regulatory and glucose homeostasis trait-associated SNPs in PPAR agonist-modulated genes are important candidates for future studies that may explain the inter-individual variability in response to thiazolidinedione and fenofibrate treatment.
insulin resistance; gene expression profile; muscle; adipose; pioglitazone; fenofibrate; eQTL
C-reactive protein (CRP) is a non-specific marker of inflammation linked to cardiovascular disease and possibly colon cancer. Polymorphisms in CRP have been associated with differential CRP concentrations among healthy adults, with some evidence for functional effects on CRP expression.
A linkage-disequilibrium-based tagSNP-selection algorithm identified six tagSNPs for Europeans (−821A>G, −390C>T/A 90A>T 838G>C 2043G>A and 4363C>A), defining 6 haplotypes >1% frequency. In a case-control study of adenomatous (n=491) or hyperplastic (n=184) polyps vs. polyp-free controls (n=583) we investigated these SNPs in relation to colorectal polyp risk.
Individuals with 838 GC or CC genotypes had a modestly, although not statistically significantly, increased risk of adenomas (OR=1.4 95% CI 0.9-2.1) and a nearly 2-fold increased risk of concurrent adenomas and hyperplastic polyps (OR=2.0 95% CI 1.1-3.6). Increased risk for concurrent adenomas and hyperplastic polyps was also observed for haplotype ACACAC. No other main associations were detected. Risk of adenomas associated with 2043G>A differed with NSAID use. Among NSAID non-users, there was a suggestion that the GA or AA genotypes were associated with decreased risk of adenomas; this was not seen among NSAID users (p-interaction = 0.03). We also observed interactions between UGT1A1 [TA](7) promoter repeat polymorphism and CRP tagSNPs −390C>T/A and 90A>T, in which only the homozygous variant CRP genotype was associated with increased adenoma risk among those with the UGT1A1 6rpt/6rpt genotype (p-interaction= 0.02 and 0.04 for −390C>T/A and 90A>T, respectively).
These results provide limited support for associations between genetic variation in CRP and colorectal polyp risk. The observed interactions should be evaluated further.
CRP; UGT; CYP2C9; colorectal cancer; colorectal polyps; NSAIDs; aspirin
Carboxylesterase 1 (CES1) is the primary enzyme responsible for converting clopidogrel into biologically inactive carboxylic acid metabolites.
We genotyped a functional variant in CES1, G143E, in participants of the Pharmacogenomics of Anti-Platelet Intervention (PAPI) study (n=566) and in 350 patients with coronary heart disease treated with clopidogrel, and carried out an association analysis of bioactive metabolite levels, on-clopidogrel ADP-stimulated platelet aggregation, and cardiovascular outcomes.
The levels of clopidogrel active metabolite were significantly greater in CES1 143E-allele carriers (P = 0.001). Consistent with these findings, individuals who carried the CES1 143E-allele showed a better clopidogrel response as measured by ADP-stimulated platelet aggregation in both participants of the PAPI study (P = 0.003) and clopidogrel-treated coronary heart disease patients (P = 0.03). No association was found between this single nucleotide polymorphism and baseline measures of platelet aggregation in either cohort.
Taken together, these findings suggest, for the first time, that genetic variation in CES1 may be an important determinant of the efficacy of clopidogrel.
carboxylesterase 1; CES1; clopidogrel; percutaneous coronary intervention; pharmacogenetics
Alaska Native people (AN) have a high prevalence of tobacco use and associated morbidity and mortality when compared to the general U.S. population. Variation in the CYP2A6 and CYP2B6 genes, encoding enzymes responsible for nicotine metabolic inactivation and procarcinogen activation, has not been characterized in AN and may contribute to the increased risk.
AN people (n = 400) residing in the Bristol Bay region of South Western Alaska were recruited for a cross-sectional study on tobacco use. They were genotyped for CYP2A6*1X2A, *1X2B, *1B, *2, *4, *7, *8, *9, *10, *12, *17, *35 and CYP2B6*4, *6, *9 and provided plasma and urine samples for measurement of nicotine and metabolites.
CYP2A6 and CYP2B6 variant frequencies among the AN Yupik people (n=361) were significantly different from other ethnicities. Nicotine metabolism (as measured by the plasma and urinary ratio of metabolites trans-3’hydroxycotinine to cotinine [(3HC/COT)] was significantly associated with CYP2A6 (P< 0.001) but not CYP2B6 genotype (P = 0.95) when controlling for known covariates. Of note, plasma 3HC/COT ratios were high in the entire Yupik people, and among the Yupik CYP2A6 wild-type participants they were substantially higher than previously characterized racial/ethnic groups (P < 0.001 vs. Caucasians and African Americans).
Yupik AN people have a unique CYP2A6 genetic profile which associated strongly with in vivo nicotine metabolism. More rapid CYP2A6-mediated nicotine and nitrosamine metabolism in the Yupik people may modulate tobacco-related disease risk.
CYP2A6; CYP2B6; nicotine; tobacco; smoking; genetic variation; Alaska Native people
The goal of pharmacogenomics is the translation of genomic discoveries to individualized patient care. Recent advances in the means to survey human genetic variation are fundamentally transforming our understanding of the genetic basis of interindividual variation in therapeutic response. The goal of this study was to systematically evaluate high-throughput genotyping technologies for their ability to assay variation in pharmacogenetically important genes (pharmacogenes). These platforms are either being proposed for or are already widely used for clinical implementation; therefore, knowledge of coverage of pharmacogenes on these platforms would serve to better evaluate current or proposed pharmacogenetic association studies.
Among the genes included in our study are drug metabolizing enzymes, transporters, receptors, and drug targets, of interest to the entire pharmacogenetic community. We considered absolute and LD-informed coverage, minor allele frequency spectrum and functional annotation for a Caucasian population. We also examined the effect of linkage disequilibrium, effect size and cohort size on power to detect SNP associations.
In our analysis of 253 pharmacogenes, we found that no platform showed more than 85% coverage of these genes (after accounting for LD). Furthermore the lack of coverage showed a dramatic increase at minor allele frequencies of less than 20%. Even after accounting for LD, only 30% of missense polymorphisms (which are enriched for low frequency alleles) were covered by HapMap with still lower coverage on the other platforms.
We have conducted the first systematic evaluation of the Axiom Genomic Database, Omni 2.5M and the DMET chip. This study is the first to utilize the 1000 Genomes Project to present a comprehensive evaluative framework. Our results provide a much-needed assessment of microarray-based genotyping and next-generation sequencing technologies’ ability to survey fully the variation in genes of particular interest to the pharmacogenetics community. Our findings demonstrate the limitations of genome-wide methods and the challenges of implementing pharmacogenomic tests into the clinical context.
Pharmacogenomics; microarray; genotyping; sequencing; coverage; LD; minor allele frequency; SNP function
To assess the feasibility of developing a Combined Clinical and Pharmacogenetic Predictive Test, comprised of multiple single nucleotide polymorphisms (SNPs) that is associated with poor bronchodilator response (BDR).
We genotyped SNPs that tagged the whole genome of the parents and children in the Childhood Asthma Management Program (CAMP) and implemented an algorithm using a family-based association test that ranked SNPs by statistical power. The top eight SNPs that were associated with BDR comprised the Pharmacogenetic Predictive Test. The Clinical Predictive Test was comprised of baseline forced expiratory volume in 1 s (FEV1). We evaluated these predictive tests and a Combined Clinical and Pharmacogenetic Predictive Test in three distinct populations: the children of the CAMP trial and two additional clinical trial populations of asthma. Our outcome measure was poor BDR, defined as BDR of less than 20th percentile in each population. BDR was calculated as the percent difference between the prebronchodilator and postbronchodilator (two puffs of albuterol at 180 μg/puff) FEV1 value. To assess the predictive ability of the test, the corresponding area under the receiver operating characteristic curves (AUROCs) were calculated for each population.
The AUROC values for the Clinical Predictive Test alone were not significantly different from 0.50, the AUROC of a random classifier. Our Combined Clinical and Pharmacogenetic Predictive Test comprised of genetic polymorphisms in addition to FEV1 predicted poor BDR with an AUROC of 0.65 in the CAMP children (n= 422) and 0.60 (n= 475) and 0.63 (n= 235) in the two independent populations. Both the Combined Clinical and Pharmacogenetic Predictive Test and the Pharmacogenetic Predictive Test were significantly more accurate than the Clinical Predictive Test (AUROC between 0.44 and 0.55) in each of the populations.
Our finding that genetic polymorphisms with a clinical trait are associated with BDR suggests that there is promise in using multiple genetic polymorphisms simultaneously to predict which asthmatics are likely to respond poorly to bronchodilators.
asthma; bronchodilator response; personalized medicine; pharmacogenetic test; predictive medicine
Fenofibrate, a peroxisome proliferator-activated receptor-alpha (PPARα) agonist, reduces triglyceride (TG) concentrations by 25–60%. Given significant inter-individual variation in TG response, we investigated the association of PPARA rare variants with treatment response in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) study.
We calculated change in TG concentration (ΔTG) among 861 GOLDN participants treated with fenofibrate (160 mg/day) for 3 weeks. From the distribution of ΔTG adjusted for age and sex, the 150 highest and 150 lowest fenofibrate responders were selected from the tails of the distribution for PPARA resequencing. The resequencing strategy was based on VariantSEQrtm technology for the amplification of exons and regulatory regions.
We identified 73 variants with an average minor allele frequency (MAF) of 4.8% (range 0.2%–16%). We tested the association of rare variants located in a coding or regulatory region (MAF<1%, 13 variants) with treatment response group via an indicator variable (presence/absence of ≥1 rare variant) using general linear mixed models to allow for adjustment for family relationship. After adjusting for baseline fasting TG concentration carrying at least one rare variant was associated with low fenofibrate response (odds ratio=6.46; 95% CI 1.4–30.8). Carrier status was also associated with relative change in total cholesterol concentration (P=0.02), but not high density lipoprotein or low density lipoprotein concentration.
Rare, potentially functional variants in PPARA may play a role in TG response to fenofibrate, but future experimental studies will be necessary to replicate the findings and confirm functional effects.
clinical trials; lipids; DNA sequence analysis; PPARalpha
To identify panels of genetic variants that predict treatment-related coronary heart disease (CHD) outcomes in hypertensive patients on one of four different classes of initial antihypertensive treatment. The goal was to identify subgroups of people based on their genetic profile who benefit most from a particular treatment.
Candidate genetic variants (n=78) were genotyped in 39,114 participants from GenHAT, ancillary to ALLHAT. ALLHAT randomized hypertensive participants (>=55 years) to one of four treatments (amlodipine, chlorthalidone, doxazosin, lisinopril). The primary outcome was fatal CHD or non-fatal MI (mean follow-up=4.9 years). A pharmacogenetic panel was derived within each of the four treatment groups. ROC curves estimated the discrimination rate between those with and without a CHD event, based on the addition of the genetic panel risk score.
For each treatment group, we identified a panel of genetic variants that collectively improved prediction of CHD to a small but statistically significant extent. Chlorthalidone (A): NOS3, rs3918226; SELE, rs5361; ICAM1, rs1799969; AGT, rs5051; GNAS, rs7121; ROC comparison p=.004; Amlodipine (B): MMP1, rs1799750; F5, rs6025; NPPA, rs5065; PDE4D, rs6450512; MMP9, rs2274756; ROC comparison p=.006; Lisinopril (C): AGT, rs5051; PON1, rs705379; MMP12, rs652438; F12, rs1801020; GP1BA, rs6065; PDE4D, rs27653; ROC comparison p=.01; Doxazosin (D): F2, rs1799963; PAI1, rs1799768; MMP7, rs11568818; AGT, rs5051; ACE, rs4343; MMP2, rs243865; ROC comparison p=.007. Each panel was tested for a pharmacogenetic effect; panels A, B and D showed such evidence (p=.009, .006, and .001 respectively), panel C did not (p=.09).
Because each panel was associated with CHD in a specific treatment group but not the others, this research provides evidence that it may be possible to use gene panel scores as a tool to better assess antihypertensive treatment choices to reduce CHD risk in hypertensive individuals.
pharmacogenetics; antihypertensive pharmacogenetics; CVD; gene panels
ADORA2A; caffeine; CYP1A2; pathway; pharmacogenomics