Absorption, Distribution, Metabolism And Excretion Genes Research Articles

Innovation in cancer pharmacotherapy through integrative consideration of germline and tumor genomes.

Precision cancer medicine is widely established, and numerous molecularly targeted drugs for various tumor entities are approved or in development. Personalized pharmacotherapy in oncology has so far been based primarily on tumor characteristics, e.g., somatic mutations. However, the response to drug treatment also depends on pharmacological processes summarized under the term ADME (absorption, distribution, metabolism, and excretion). Variations in ADME genes have been the subject of intensive research for more than five decades, considering individual patients' genetic makeup, referred to as pharmacogenomics (PGx). The combined impact of a patient's tumor and germline genome is only partially understood and often not adequately considered in cancer therapy. This may be attributed, in part, to the lack of methods for combined analysis of both data layers. Optimized personalized cancer therapies should, therefore, aim to integrate molecular information about the tumor and the germline, taking into account existing PGx guidelines for drug therapy. Moreover, such strategies should provide the opportunity to consider genetic variants of previously unknown functional significance. Bioinformatic analysis methods and corresponding algorithms for data interpretation need to be developed to consider PGx data in interdisciplinary molecular tumor boards, where cancer patients are discussed to provide evidence-based recommendations for clinical management based on individual tumor profiles. Significance Statement The era of personalized oncology has seen the emergence of drugs tailored to genetic variants associated with cancer biology. However, full potential of targeted therapy remains untapped due to the predominant focus on acquired tumor-specific alterations. Optimized cancer care must integrate tumor and patient genomes, guided by pharmacogenomic principles. An essential prerequisite for realizing truly personalized drug treatment of cancer patients is the development of bioinformatic tools for comprehensive analysis of all data layers generated in modern precision oncology programs.

Structural variation of the coding and non-coding human pharmacogenome

Genetic variants in drug targets and genes encoding factors involved in drug absorption, distribution, metabolism and excretion (ADME) can have pronounced impacts on drug pharmacokinetics, response, and toxicity. While the landscape of genetic variability at the level of single nucleotide variants (SNVs) has been extensively studied in these pharmacogenetic loci, their structural variation is only poorly understood. Thus, we systematically analyzed the genetic structural variability across 908 pharmacogenes (344 ADME genes and 564 drug targets) based on publicly available whole genome sequencing data from 10,847 unrelated individuals. Overall, we extracted 14,984 distinct structural variants (SVs) ranging in size from 50 bp to 106 Mb. Each individual harbored on average 10.3 and 1.5 SVs with putative functional effects that affected the coding regions of ADME genes and drug targets, respectively. In addition, by cross-referencing pharmacogenomic SVs with experimentally determined binding data of 224 transcription factors across 130 cell types, we identified 1276 non-coding SVs that overlapped with gene regulatory elements. Based on these data, we estimate that non-coding structural variants account for 22% of the genetically encoded pharmacogenomic variability. Combined, these analyses provide the first comprehensive map of structural variability across pharmacogenes, derive estimates for the functional impact of non-coding SVs and incentivize the incorporation of structural genomic data into personalized drug response predictions.

Association between Germline Single-Nucleotide Variants in ADME Genes and Major Molecular Response to Imatinib in Chronic Myeloid Leukemia Patients.

Imatinib is the most common first-line tyrosine kinase inhibitor (TKI) used to treat chronic-phase chronic myeloid leukemia (CP-CML). However, only a proportion of patients achieve major molecular response (MMR), so there is a need to find biological factors that aid the selection of the optimal therapeutic strategy (imatinib vs. more potent second-generation TKIs). The aim of this retrospective study was to understand the contribution of germline single-nucleotide variants (gSNVs) in the achievement of MMR with imatinib. In particular, a discovery cohort including 45 CP-CML patients was analyzed through the DMET array, which interrogates 1936 variants in 231 genes related to the absorption, distribution, metabolism and excretion (ADME) process. Variants statistically significant in the discovery cohort were then tested in an extended and independent cohort of 137 CP-CML patients. Finally, a total of 7 gSNVs (ABCG1-rs492338, ABCB11-rs496550, ABCB11-rs497692, CYP2D6-rs1135840, CYP11B1-rs7003319, MAT1A-rs4934027 and SLC22A1-rs628031) and one haplotype in the ABCB11 gene were significantly associated with the achievement of MMR with first-line imatinibtreatment. In conclusion, we identified a genetic signature of response to imatinib in CP-CML patients that could be useful in selecting those patients that may benefit from starting imatinib as first-line therapy, therefore avoiding the toxicity related to second-generation TKIs.

Effects of Diminished NADPH:cytochrome P450 Reductase in Human Hepatocytes on Lipid and Bile Acid Homeostasis.

NADPH:cytochrome P450 oxidoreductase (POR) is the obligate electron donor for microsomal cytochrome P450 (CYP) enzymes involved in the biosynthesis of endogenous substances like bile acids and other steroids as well as in the oxidative metabolism of xenobiotics. P450 oxidoreductase also supports other redox enzymes in fatty acid and cholesterol pathways. Recently, we have established CRISPR/Cas9-mediated POR knockdown in a human hepatic cell model, HepaRG, and demonstrated the differential effects of limited POR expression on CYP activity. The aim of the present work was to systematically investigate the impact of POR knockdown with a focus on the expression of ADME (absorption, distribution, metabolism, and excretion) genes and related regulators. Functional consequences have been assessed using quantitative mass spectrometry for targeted metabolomics covering bile acids, and cholesterol and its precursors, and for untargeted proteomics. In addition to the previously described alteration of RNA expression of CYP genes, we showed significant downregulation of transcriptional regulators of drug metabolism and transport, including NR1I3 (CAR), NR1I2 (PXR), NR1H4 (FXR), and NR1H3 (LXRα) in cells with POR gene disruption. Furthermore, POR knockdown resulted in deregulated bile acid and cholesterol biosynthesis demonstrated by low levels of cholic acid derivates and increased concentrations of chenodeoxycholic acid derivates, respectively. Systemic effects of POR knockdown on global protein expression were indicated by downregulation of several metabolic pathways including lipid metabolism and biological oxidation reactions. The deduced protein network map corroborates CYP enzymes as direct interaction partners, whereas changes in lipid metabolism and homeostasis are the result of indirect effects. In summary, our results emphasize a widespread role of POR in various metabolic pathways and provide the first human data on the effects of diminished POR expression on drug and endogenous metabolism in a genomeedited HepaRG cell model.

Brazilian cohort and genes encoding for drug-metabolizing enzymes and drug transporters.

Background& aim:Genetic variability in drug absorption, distribution, metabolism and excretion (ADME)genes contributes to the high heterogeneity of drug responses. The present study investigated polymorphisms of ADME genes frequencies and compared the findings with populations from other continents, available in the 1000Genome Project (1KGP) and the Exome Aggregation Consortium (ExAC) databases. Methodology& results:We conducted a study of 100patients in Brazil and a total of 2003 SNPs were evaluated by targeted next-generation sequencing in 148genes, including Phase I enzymes (n=50), Phase II enzymes (n=38) and drug transporters (n=60). Overall, the distribution of minor allele frequency (MAF) suggests that the distribution of 2003 SNPs is similar between Brazilian cohort, 1 KGP and ExAC; however, we found moderate SNP allele-frequency divergence between Brazilian cohort and both 1000KGP and ExAC. These differences were observed in several relevant genes including CYP3A4, NAT2 and SLCO1B1. Conclusion:We concluded that the Brazilian population needs clinical assessment of drug treatment based on individual genotype rather than ethnicity.

Copy number variation profiling in pharmacogenetics CYP-450 and GST genes in Colombian population

BackgroundCopy Number variation (CNVs) in genes related to drug absorption, distribution, metabolism and excretion (ADME) are relevant in the interindividual variability of drug response. Studies of the CNVs in ADME genes in Latin America population are lacking. The objective of the study was to identify the genetic variability of CNVs in CYP-450 and GST genes in a subgroup of individuals of Colombian origin.MethodsGenomic DNA was isolated from 123 healthy individuals from a Colombian population. Multiplex Ligation-Dependent Probe Amplification (MLPA) was performed for the identification of CNVs in 40 genomic regions of 11 CYP-450 and 3 GST genes. The genetic variability, allelic and genotypic frequencies were analyzed.ResultsWe found that 13 out of 14 genes had CNVs: 5 (35.7%) exhibited deletions and duplications, while 8 (57.1%) presented either deletions or duplications.. 33.3% of individuals carried deletions and duplications while 49.6% had a unique type of CNV (deletion or duplication). The allelic frequencies of the CYP and GST genes were 0 to 47.6% (allele null), 0 to 17.5% (duplicated alleles) and 37 to 100% (normal alleles).ConclusionsOur results describe, for the first time, the genomic profile of CNVs in a subgroup of Colombian population in GST and CYP-450 genes. GST genes indicated greater genetic variability than CYP-450 genes. The data obtained contributes to the knowledge of genetic profiles in Latin American subgroups. Although the clinical relevance of CNVs has not been fully established, it is a valuable source of pharmacogenetic variability data with potential involvement in the response to medications.

A New Panel-Based Next-Generation Sequencing Method for ADME Genes Reveals Novel Associations of Common and Rare Variants With Expression in a Human Liver Cohort.

We developed a panel-based NGS pipeline for comprehensive analysis of 340 genes involved in absorption, distribution, metabolism and excretion (ADME) of drugs, other xenobiotics, and endogenous substances. The 340 genes comprised phase I and II enzymes, drug transporters and regulator/modifier genes within their entire coding regions, adjacent intron regions and 5′ and 3′UTR regions, resulting in a total panel size of 1,382 kbp. We applied the ADME NGS panel to sequence genomic DNA from 150 Caucasian liver donors with available comprehensive gene expression data. This revealed an average read-depth of 343 (range 27–811), while 99% of the 340 genes were covered on average at least 100-fold. Direct comparison of variant annotation with 363 available genotypes determined independently by other methods revealed an overall accuracy of >99%. Of 15,727 SNV and small INDEL variants, 12,022 had a minor allele frequency (MAF) below 2%, including 8,937 singletons. In total we found 7,273 novel variants. Functional predictions were computed for coding variants (n = 4,017) by three algorithms (Polyphen 2, Provean, and SIFT), resulting in 1,466 variants (36.5%) concordantly predicted to be damaging, while 1,019 variants (25.4%) were predicted to be tolerable. In agreement with other studies we found that less common variants were enriched for deleterious variants. Cis-eQTL analysis of variants with (MAF ≥ 2%) revealed significant associations for 90 variants in 31 genes after Bonferroni correction, most of which were located in non-coding regions. For less common variants (MAF < 2%), we applied the SKAT-O test and identified significant associations to gene expression for ADH1C and GSTO1. Moreover, our data allow comparison of functional predictions with additional phenotypic data to prioritize variants for further analysis.

Building a Targeted Approach to Assess Risk of Anthracycline Induced Cardiomyopathy Amongst Paediatric Cancer Patients

IntroductionAnthracyclines are chemotherapeutic agents used in the treatment of over 70% of childhood and adolescent malignancies and their use has significantly improved cancer survival rates (Lipshultz, Alvarez et al. 2008). However, a major side effect of this drug class is the potential to develop subclinical, irreversible and fatal cardiomyopathy with the risk of sudden cardiac death. Despite the high incidence of cardiotoxicity in paediatric and adolescent cancer survivors as well as the emerging evidence that alteration in the genes that control drug absorption, distribution, metabolism and excretion (ADME) may be predictive of a patient's sensitivity to this complication, there are no current standard approaches to identify the 'at risk' group (Visscher, Amstutz et al. 2011). Consequently, there is no way to tailor therapy according to a patient's individual risk.This study aims to create a functional assay that will assess patient sensitivity to anthracyclines using a cohort of paediatric cancer survivors, with and without, evidence of anthracycline cardiotoxicity (ACT). A novel, exciting approach will be used through the creation of patient specific cardiomyocytes from induced pluripotent stem cells (iPSC). Through the exposure of iPSC derived cardiomyocytes to anthracyclines, a series of functional assays will be performed to help determine the patient's sensitivity to this drug toxicity. The cardiomyocytes in both cases and controls will be assessed for variations in the genes involved in drug ADME. The genetic analysis will help to validate the results of our collaborators at the British Columbia (BC) Children's Hospital (Vancouver) and the Canada Pharmacogenomic Network for Drug Sensitivity (CPNDS).Methods: An audit of paediatric patients diagnosed across two tertiary institutes between 2000-2015 was performed. From this patients were deemed to be either cases (no evidence of cardiac damage) or controls (evidence of cardiac damage). The degree of cardiac damage was defined as severe (ejection fraction < 24 %), intermediate (ejection fraction 24 - 27 % or a decline of > 10 % ejection fraction from baseline). Patients were then enrolled to the study upon routine presentation to clinic. Upon enrolment peripheral blood mononuclear cells (PBMC) were collected for DNA/RNA storage and a cell pellet was cryopreserved for iPSC development. Following storage, the aims of the project are to 1) Establish a biobank from anthracycline treated childhood cancer survivors, including both sensitive and resistant patients (cases and controls). 2) To confirm the capacity of patient iPSC derived cardiomyocytes to predict ACT and to explore the mechanistic basis and functional responses of human cardiomyocytes to anthracyclines. 3) To perform detailed genomic profiling of the cohort and assess if genetic variation in ADME genes is predictive of ACT in the Australian paediatric population. 4) To document the diastolic changes evident on echocardiography and correlate these findings with long-term anthracycline cardiotoxicity in all enrolled patients.Results: To date 56 cases and 199 controls have enrolled onto the study and had their blood processed accordingly. The median age of the cases is 11.5 years and controls 8.0 years. Of the 56 cases, 52 % (n = 29) are female and 48 % (n = 27) are male. The predominant anthracycline exposure in the cases was doxorubicin alone (47.3 %), daunorubicin and doxorubicin (29.1 %) and daunorubicin alone (14.1 %). Overall 63 % of cases and controls had a diagnosis of Acute Lymphoblastic leukaemia. The median anthracycline dose exposure in the cases was 172 mg/m2 (range 22 - 438) versus 96 mg/m2 (range 18 - 389) in the controls. To date we have successfully been able to derive cardiomyocytes from 8 cases and 8 controls and have begun a range of viability assays to assess susceptibility to anthracycline exposure.Conclusions: We hope over the next 12 months to complete whole exome sequencing and microarray SNP analysis to further characterise the genetic basis of anthracycline cardiomyopathy. Furthermore, using human induced pluripotent stem cell derived cardiomyocytes as a functional platform we hope to further characterise the molecular basis of the disease and create a novel functional assay to stratify patients according to their risk of anthracycline induced cardiomyopathy. DisclosuresNo relevant conflicts of interest to declare.

CML Patients with Primary Resistance or Suboptimal Response to TKI Therapy Have Variants in Genes Affecting Drug Absorption and Metabolism

BACKGROUNDChronic myeloid leukemia in chronic phase (CML CP) is successfully treated with tyrosine kinase inhibitors (TKIs) but a proportion of patients show primary or secondary resistance to TKI therapy. BCR-ABL1 mutations are the primary cause for secondary resistance to TKI therapy, but the causes for primary resistance are poorly known. Putative variants in genes affecting drug absorption, distribution, metabolism and excretion (ADME) have not been comprehensively analyzed. We set out to screen CML CP patients who either had primary resistance to TKI therapy or suboptimal response with large ADME gene panel to examine if these genes play a role in poor TKI responses.METHODSWe selected samples from 21 CML CP patients treated in our institute who either had primary resistance (N=6) or responded suboptimallyto TKI treatment (n=15) but did not have BCR-ABL1 mutations. DNA samples from the time of diagnosis were sequenced with massively parallel sequencing using a comprehensive custom ADME gene panel covering exons and introns of 382 genes. Over 200 healthy volunteers who had been sequenced with the same gene panel served as population controls. Loss-of-function and novel variants were validated using capillary sequencing.We tested the differences of variant allele frequencies between CML patients and healthy controls by performing the Cochran-Armitage trend test using additive coding (0,1,2) for genotypes homozygous for the major allele, heterozygous, and homozygous for the minor allele, respectively. CML patients with optimal response (n=24) were used as a control group.RESULTSSequencing of 21 poor responding CML CP patients revealed a total of 658 nonsynonymousgene variants. Majority of the variants (94%) were missense variants whose effects on gene function are uncertain. Eigthvariants were previously unreported. 18 variants were predicted to lead to a complete loss-of-function of the genes affected. Principal component analysis did not reveal clear global differences between the CML patients and healthy controls in the frequencies of ADME gene variants. However, the Cochran-Armitage trend test showed a number of variants whose frequencies in CML samples were significantly different from the healthy controls (table 1). Two of the loss-of-function variants were in the top 10 variants, whose frequencies differed between CML patients and healthy controls. ACAD11 rs41272317 variant was observed in 14.2% of patients whereas its frequency in healthy controls was 0.5% (p=0.003). Similarly DPYD rs3918290 was observed in 14.2% of patients and 3.3% of healthy controls (p=0.006). ACAD11 encodes an acyl-CoA dehydrogenase enzyme with a preference for carbon chain lengths between 20 and 26, and has been shown to be an important metabolic target for p53 pro-survival function. DPYD codesdihydropyrimidine dehydrogenase, which is involved in the metabolism ofpyrimidines and the anticancer agent 5-fluorouracil. Sanger sequencing confirmed both variants in patient samples. From the top10 missense variants, SLC15A1 rs144553892 variant was significantly more often observed in poor responding CML patients than in healthy controls (14.2 and 0.9%, respectively, p=0.003). SLC15A1 is a solute carrier family 15 member 1 which has a known function as a co-transporter foroligopeptides. Recently solute carriers have been identified to have significant impact on cancer therapy. Screening of additional CML patients is ongoing to confirm the frequency of these variants in optimally responding CML patients.CONCLUSIONSCML patients with primary resistance or suboptimal response to TKI therapy have mutations in genes affecting drug intake and metabolism, which can be causally associated with poor therapy responses. Validation in larger patient cohorts is warranted. [Display omitted] DisclosuresPorkka:Novartis: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding. Mustjoki:Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Ariad: Research Funding.

OSAnalyzer: A Bioinformatics Tool for the Analysis of Gene Polymorphisms Enriched with Clinical Outcomes.

Background: The identification of biomarkers for the estimation of cancer patients’ survival is a crucial problem in modern oncology. Recently, the Affymetrix DMET (Drug Metabolizing Enzymes and Transporters) microarray platform has offered the possibility to determine the ADME (absorption, distribution, metabolism, and excretion) gene variants of a patient and to correlate them with drug-dependent adverse events. Therefore, the analysis of survival distribution of patients starting from their profile obtained using DMET data may reveal important information to clinicians about possible correlations among drug response, survival rate, and gene variants. Methods: In order to provide support to this analysis we developed OSAnalyzer, a software tool able to compute the overall survival (OS) and progression-free survival (PFS) of cancer patients and evaluate their association with ADME gene variants. Results: The tool is able to perform an automatic analysis of DMET data enriched with survival events. Moreover, results are ranked according to statistical significance obtained by comparing the area under the curves that is computed by using the log-rank test, allowing a quick and easy analysis and visualization of high-throughput data. Conclusions: Finally, we present a case study to highlight the usefulness of OSAnalyzer when analyzing a large cohort of patients.

Single Nucleotide Polymorphisms in Cellular Drug Transporters Are Associated with Intolerance to Antiretroviral Therapy in Brazilian HIV-1 Positive Individuals.

Adverse reactions are the main cause of treatment discontinuation among HIV+ individuals. Genes related to drug absorption, distribution, metabolism and excretion (ADME) influence drug bioavailability and treatment response. We have investigated the association between single nucleotide polymorphisms (SNPs) in 29 ADME genes and intolerance to therapy in a case-control study including 764 individuals. Results showed that 15 SNPs were associated with intolerance to nucleoside and 11 to non-nucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs), and 8 to protease inhibitors (PIs) containing regimens under alpha = 0.05. After Bonferroni adjustment, two associations remained statistically significant. SNP rs2712816, at SLCO2B1 was associated to intolerance to NRTIs (ORGA/AA = 2.37; p = 0.0001), while rs4148396, at ABCC2, conferred risk of intolerance to PIs containing regimens (ORCT/TT = 2.64; p = 0.00009). Accordingly, haplotypes carrying rs2712816A and rs4148396T alleles were also associated to risk of intolerance to NRTIs and PIs, respectively. Our data reinforce the role of drug transporters in response to HIV therapy and may contribute to a future development of personalized therapies.

Genetic architectures of ADME genes in five Eurasian admixed populations and implications for drug safety and efficacy

BackgroundDrug absorption, distribution, metabolism and excretion (ADME) contribute to the high heterogeneity of drug responses in humans. However, the same standard for drug dosage has been applied to all populations...

Development of a broad-based ADME panel for use in pharmacogenomic studies.

To optimally address the interindividual variability observed in pharmacokinetic drug response, we have created a custom genotyping panel that interrogates most of the key genetic variations present in a set of 181 prioritized genes responsible for the absorption, distribution, metabolism and excretion (ADME) of many therapeutic agents. This consensus list of genes and variants was based on the ADME core and extended gene lists compiled by a group of pharmaceutical companies as having relevance. Although these pharmacokinetic genes and pathways are well known, tools that can interrogate a large number of these genes simultaneously within a single experiment are not currently available. Using novel design strategies, we have developed an optimized and validated ADME genotyping panel, encompassing approximately 3000 variants, that has broad applicability to any study or clinical trial that would benefit from the evaluation of an extensive list of ADME genes. Over the course of three design iterations, overall assay conversion rates were improved from 83 to 97% resulting in a panel that fills in many of the gaps in coverage present on currently available commercial genotyping assays. The utility of the assay has been demonstrated by the screening of more than 1000 samples resulting in the discovery of novel pharmacogenomic associations. The assay, and the underlying methods, will continue to be a valuable tool for use in future pharmacogenomic studies.

PHC-014 Exploratory Analysis of 1,936 SNPs in 225 ADME Genes For Association with Busulfan Clearance in Adult Hematopoietic Stem Cell Recipients

BackgroundBusulfan is used in preparative regimens prior to stem cell transplantation (SCT). There is significant inter-patient variability in busulfan pharmacokinetics (PK) and outcome is related to exposure.To date, only polymorphisms...

ADME pharmacogenetics: investigation of the pharmacokinetics of the antiretroviral agent lopinavir coformulated with ritonavir

An ADME (absorption, distribution, metabolism and excretion)-pharmacogenetics association study may identify functional variants relevant to the pharmacokinetics of lopinavir co-formulated with ritonavir (LPV/r), a first-line anti-HIV agent. An extensive search of literature and web resources helped select ADME genes and single nucleotide polymorphisms (SNPs, functional and HapMap tagging SNPs) with a proven or potentially relevant role in LPV/r pharmacokinetics. The study followed a two-stage design. Stage 1 (discovery) considered a Caucasian population (n=638) receiving LPV/r, where we selected 117 individuals with low LPV clearance (cases) and 90 individuals with high clearance (controls). Genotyping was performed by a 1536-SNP customized GoldenGate Illumina BeadArray. Stage 2 (confirmation) represented a replication study of candidate SNPs from the stage 1 in 148 individuals receiving LPV/r. The analysis led to formal population pharmacokinetic-pharmacogenetic modeling of demographic, environmental and candidate SNP effects. One thousand three hundred and eighty SNPs were successfully genotyped. Nine SNPs prioritized by the stage 1 analysis were brought to replication. Stage 2 confirmed the contribution of two functional SNPs in SLCO1B1, one functional SNP in ABCC2 and a tag SNP of the CYP3A locus in addition to body weight effect and ritonavir coadministration. According to the population pharmacokinetic-pharmacogenetic model, genetic variants explained 5% of LPV variability. Individuals homozygous rs11045819 (SLCO1B1*4) had a clearance of 12.6 l/h, compared with 5.4 l/h in the reference group, and 3.9 l/h in individuals with two or more variant alleles of rs4149056 (SLCO1B1*5), rs717620 (ABCC2) or rs6945984 (CYP3A). A subanalysis confirmed that although a significant part of the variance in LPV clearance was attributed to fluctuation in ritonavir levels, genetic variants had an additional effect on LPV clearance. The two-stage strategy successfully identified genetic variants affecting LPV/r pharmacokinetics. Such a general approach of ADME pharmacogenetics should be generalized to other drugs.

ADME transcriptome in Hispanic versus White donor livers: Evidence of a globally enhanced NR1I3 (CAR, constitutive androstane receptor) gene signature in Hispanics

Previous studies have found that, compared with Whites, Hispanic donor livers had elevated expression of CYP2 enzymes, gene products regulated by the constitutive androstane receptor (CAR). The objectives of the current study were to determine (1) the CAR activation signature in human liver (2) whether other drug detoxification (absorption, distribution, metabolism and excretion (ADME)) genes were differentially expressed in Hispanic versus White livers, and (3) the extent of overlap in the CAR and Hispanic liver transcriptomes. The CAR transcriptome (ADME genes differentially expressed following phenobarbital versus vehicle treatment of human hepatocytes) and the Hispanic liver transcriptome (ADME genes differentially expressed in Hispanic versus White livers) were identified using Affymetrix oligonucleotide arrays. Quantitative real-time polymerase chain reaction (PCR) was used to verify candidate genes in a larger sample size. Comparison of the CAR and Hispanic liver ADME transcriptomes revealed a significant association between the gene changes. Sixty-four per cent of the ADME genes induced more than twofold by phenobarbital were also induced in Hispanics, and 14% of the ADME genes repressed more than twofold by phenobarbital were repressed in Hispanics. In conclusion, compared with Whites, Hispanic donor livers have increased expression of many genes that are transcriptionally regulated by CAR. This result has practical implications to the drug treatment of Hispanic patients.

Compendium of gene expression profiles comprising a baseline model of the human liver drug metabolism transcriptome

Oligonucleotide microarrays were used to study the variability of pharmacokinetics and drug metabolism (PKDM)-related gene expression in 75 normal human livers. The objective was to define and use absorption, distribution, metabolism and excretion (ADME) gene expression variability to discern co-regulated genes and potential surrogate biomarkers of inducible gene expression. RNA was prepared from donor tissue and hybridized on Agilent microarrays against an RNA mass balanced pool from all donors. Clustering of PKDM gene sets revealed donors with distinct patterns of gene expression that grouped genes known to be regulated by the nuclear receptor, pregnane X-receptor (PXR). Fold range metrics and frequency distributions from the heterogeneous human population were used to define the variability of individual PKDM genes in the 75 human livers and were placed in context by comparing expression data with basal ADME gene expression variability in an inbred and diet/environment controlled population of 27 Rhesus livers. The most variable genes in the hepatic transcriptome were mainly related to drug metabolism, intermediary metabolism, inflammation and cell cycle control. Unique patterns of expression across 75 individuals of inducible ADME gene expression allowed their expression to be correlated with the expression of many other genes. Correlated genes for AhR, CAR and PXR responsive genes (CYP1A2, CYP2B6 and CYP3A4) were identified that may be co-regulated and, therefore, provide clues to the identity of surrogate gene or protein markers for CYP induction. In conclusion, microarrays were used to define the variable expression of hepatic ADME genes in a diverse human population, the expression variability of ADME genes was compared with the expression variability in an inbred population of Rhesus monkeys, and genes were defined that may be co-regulated with important inducible CYP genes.

Microarray-based compendium of hepatic gene expression profiles for prototypical ADME gene-inducing compounds in rats and mice in vivo

To examine species-specific aspects of the induction of absorption, distribution, metabolism and excretion (ADME)-related genes, we used 25 000 gene oligonucleotide microarrays to construct a rodent gene-response compendium that compared hepatic gene expression profiles and developed consensus aryl hydrocarbon receptor (AhR), constitutive androstane receptor (CAR) and pregnane X-receptor (PXR) ligand signatures relevant to drug clearance. Twenty-six inducer compounds were chosen from the literature. Rats and mice received one of six dose levels (log2 dose escalation, 32-fold dose range) of each compound daily for 3 days. Animals were necropsied 6–9 h after the last dose, and tissues were collected for RNA analysis. Hepatic gene expression profiles were obtained using Rosetta Resolver® expression analysis system, and ADME-related genes were extracted. Cross-talk among nuclear receptors or hepatoxicity at high dose levels resulted in large signatures (usually >1000 genes at p < 0.01) for most compounds. After ADME gene transcript enrichment, agglomerative clustering separated AhR ligands from CAR/PXR ligands, but it was difficult to distinguish CAR from PXR ligands. Consensus signatures were derived from groups of AhR, CAR and PXR ligands; and cross-talk among responding genes was determined. Many compounds had distinct log dose–response profiles, and relative potencies for ligands were established. Robust responses by CYP1A1, CYP2B10 (CAR responsive in mice) and CYP2B15 (CAR responsive in rats) and CYP3A1 (PXR responsive in rats) were used to benchmark the relative potency of different ligands and to determine the relative selectivity for AhR, CAR or PXR. By using a compendium of gene expression profiles, we defined species-specific induction patterns across the ADME transcriptome.