[show abstract][hide abstract] ABSTRACT: While tamoxifen activity is mainly due to endoxifen and the concentration of this active metabolite is, in part, controlled by CYP2D6 metabolic status, clinical correlative studies have produced mixed results.
In an exploratory study, we determined the CYP2D6 metabolic status and plasma concentrations of endoxifen among 224 Filipino and Vietnamese women participating in a clinical trial of adjuvant hormonal therapy for operable breast cancer. We further conducted a nested-case-control study among 48 women (half with recurrent disease, half without) investigating the relationship of endoxifen concentrations and recurrence of disease. We found a significant association of reduced endoxifen plasma concentrations with functionally important CYP2D6 genotypes. High endoxifen concentrations were associated with higher risk of recurrence; with a quadratic trend fitted to a stratified Cox proportional hazards regression model, the likelihood ratio p-value was 0.002. The trend also showed that in 8 out of 9 pairs with low endoxifen concentrations, the recurrent case had lower endoxifen levels than the matched control.
This exploratory analysis suggests that there is an optimal range for endoxifen concentrations to achieve favorable effects as adjuvant therapy. In particular, at higher concentrations (>70 ng.ml), endoxifen may promote recurrence.
[show abstract][hide abstract] ABSTRACT: Background:Change in breast density may predict outcome of women receiving adjuvant hormone therapy for breast cancer. We performed a prospective clinical trial to evaluate the impact of inherited variants in genes involved in oestrogen metabolism and signalling on change in mammographic percent density (MPD) with aromatase inhibitor (AI) therapy.Methods:Postmenopausal women with breast cancer who were initiating adjuvant AI therapy were enrolled onto a multicentre, randomised clinical trial of exemestane vs letrozole, designed to identify associations between AI-induced change in MPD and single-nucleotide polymorphisms in candidate genes. Subjects underwent unilateral craniocaudal mammography before and following 24 months of treatment.Results:Of the 503 enrolled subjects, 259 had both paired mammograms at baseline and following 24 months of treatment and evaluable DNA. We observed a statistically significant decrease in mean MPD from 17.1 to 15.1% (P<0.001), more pronounced in women with baseline MPD 20%. No AI-specific difference in change in MPD was identified. No significant associations between change in MPD and inherited genetic variants were observed.Conclusion:Subjects with higher baseline MPD had a greater average decrease in MPD with AI therapy. There does not appear to be a substantial effect of inherited variants in biologically selected candidate genes.British Journal of Cancer advance online publication, 1 October 2013; doi:10.1038/bjc.2013.587 www.bjcancer.com.
British Journal of Cancer 10/2013; · 5.08 Impact Factor
[show abstract][hide abstract] ABSTRACT: Background
Alternative splicing increases proteome diversity by expressing multiple gene isoforms that often differ in function. Identifying alternative splicing events from RNA-seq experiments is important for understanding the diversity of transcripts and for investigating the regulation of splicing.
We developed Alt Event Finder, a tool for identifying novel splicing events by using transcript annotation derived from genome-guided construction tools, such as Cufflinks and Scripture. With a proper combination of alignment and transcript reconstruction tools, Alt Event Finder is capable of identifying novel splicing events in the human genome. We further applied Alt Event Finder on a set of RNA-seq data from rat liver tissues, and identified dozens of novel cassette exon events whose splicing patterns changed after extensive alcohol exposure.
Alt Event Finder is capable of identifying de novo splicing events from data-driven transcript annotation, and is a useful tool for studying splicing regulation.
[show abstract][hide abstract] ABSTRACT: Formalin-fixed, paraffin-embedded tumors (FFPETs) are a valuable source of DNA for genotype association studies and are often the only germline DNA resource from cancer clinical trials. The anti-estrogen tamoxifen is metabolized into endoxifen by CYP2D6, leading to the hypothesis that patients with certain CYP2D6 genotypes may not receive benefit because of their inability to activate the drug. Studies testing this hypothesis using FFPETs have provided conflicting results. It has been postulated that CYP2D6 genotype determined using FFPET may not be accurate because of somatic tumor alterations. In this study, we determined the concordance between CYP2D6 genotypes generated using 3 tissue sources (FFPETs; formalin-fixed, paraffin-embedded unaffected lymph nodes [FFPELNs]; and whole blood cells [WBCs]) from 122 breast cancer patients. Compared with WBCs, FFPET and FFPELN genotypes were highly concordant (>94%), as were the predicted CYP2D6 metabolic phenotypes (>97%). We conclude that CYP2D6 genotypes obtained from FFPETs accurately represent the patient's CYP2D6 metabolic phenotype.
[show abstract][hide abstract] ABSTRACT: Rifampin causes drug interactions by altering hepatic drug metabolism. Since miRNAs have been shown to regulate genes involved in drug metabolism, we determined the effect of rifampin on the expression of hepatic miRNAs. Primary human hepatocytes from seven subjects were treated with rifampin and the expression of miRNAs and CYP mRNAs were measured by TaqMan assays and RNA-seq, respectively. Rifampin induced the expression of 10 clinically important and 13 additional CYP genes and repressed the expression of 9 other CYP genes (p<0.05). Rifampin induced the expression of 33 miRNAs and repressed the expression of 35 miRNAs (p<0.05). Several of these changes were highly negatively correlated with the rifampin-induced changes in the expression of their predicted target CYP mRNAs, supporting the possibility of miRNA-induced regulation of CYP mRNA expression. In addition, several other miRNA changes were positively correlated with the changes in CYP mRNA expression, suggesting similar regulatory mechanisms. Despite the interindividual variability in the rifampin effects on miRNA expression, principal components analysis clearly separated the rifampin- treated samples from the controls. In conclusion, rifampin treatment alters miRNA expression patterns in human hepatocytes and some of the changes were correlated with the rifampin-induced changes in expression of the CYP mRNAs they are predicted to target.
Drug metabolism and disposition: the biological fate of chemicals 08/2013; · 3.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: Plasma derived miRNAs are being used as biomarkers and have been associated with human liver disease and function including fibrosis, inflammation, and drug induced liver injury. They may also be biomarkers of the drug metabolism function of the liver. In order for plasma miRNA to function as a clinical biomarker, predictable variability is necessary during processing from whole blood to plasma. The current study evaluated miRNAs variability in whole blood stored for 0.5, 1, 2, 4, 8, and 12 hrs following the blood draw under clinic conditions (room temperature) prior to the separation of the plasma. Four healthy volunteer subjects were recruited. All subjects' blood was collected twice. Mir-16 and mir-223 were evaluated because many studies have shown them to be reliably present in the plasma and useful for normalization. MiRNA concentrations were measured by real-time PCR. The coefficient of variability of the CT values for subjects for miR-223 and miR-16 ranged from ~3.6 to 6.8% and ~1.48 to 4.1% respectively over the 12 hour incubation. A second blood collection was performed to determine interday variability. The coefficient of variance from the initial blood draw compared to the final blood draw for each subject ranged from 0.42 to 7.9% for miR-16 and 1.7 to 8.3% for miR-223, indicating that these miRNAs have limited interday variability. We conclude that plasma miR-16 or miR-223 concentrations are stable in whole blood at room temperature for up to at least 12 hours.
Drug metabolism and disposition: the biological fate of chemicals 07/2013; · 3.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: BACKGROUND: The class of acetylcholinesterase inhibitors (ChEI), including donepezil, rivastigmine, and galantamine, have similar efficacy profiles in patients with mild to moderate Alzheimer's disease (AD). However, few studies have evaluated adherence to these agents. We sought to prospectively capture the rates and reasons for nonadherence to ChEI and determine factors influencing tolerability and adherence.Methods/design: We designed a pragmatic randomized clinical trial to evaluate the adherence to ChEIs among older adults with AD. Participants include AD patients receiving care within memory care practices in the greater Indianapolis area. Participants will be followed at 6-week intervals up to 18 weeks to measure the primary outcome of ChEI discontinuation and adherence rates and secondary outcomes of behavioral and psychological symptoms of dementia. The primary outcome will be assessed through two methods, a telephone interview of an informal caregiver and electronic medical record data captured from each healthcare system through a regional health information exchange. The secondary outcome will be measured by the Healthy Aging Brain Care Monitor and the Neuropsychiatric Inventory. In addition, the trial will conduct an exploratory evaluation of the pharmacogenomic signatures for the efficacy and the adverse effect responses to ChEIs. We hypothesized that patient-specific factors, including pharmacogenomics and pharmacokinetic characteristics, may influence the study outcomes. DISCUSSION: This pragmatic trial will engage a diverse population from multiple memory care practices to evaluate the adherence to and tolerability of ChEIs in a real world setting. Engaging participants from multiple healthcare systems connected through a health information exchange will capture valuable clinical and non-clinical influences on the patterns of utilization and tolerability of a class of medications with a high rate of discontinuation.Trial Registration: Clinicaltrials.gov: NCT01362686.
[show abstract][hide abstract] ABSTRACT: Efavirenz increases CYP2C19- and CYP3A-mediated omeprazole metabolism. We hypothesized that CYP2C19 and CYP2B6 genetic polymorphisms influence the extent of induction of omeprazole metabolism by efavirenz. Healthy subjects (n=57) were administered a single 20 mg oral dose of omeprazole on two occasions: with a single 600 mg efavirenz dose; and after a 17-day treatment with efavirenz (600 mg per day). DNA was genotyped for CYP2C19*2, *3 and *17 alleles and CYP2B6*6, *4 and *9 alleles using Taqman assays. Omeprazole, its enantiomers and metabolites were measured by liquid chromatography/tandem mass spectrometry. Our results showed that efavirenz increased omeprazole clearances in all CYP2C19 genotypes in non-stereoselective manner, but the magnitude of induction was genotype dependent. Metabolic ratios of 5-hydroxylation of omeprazole were reduced in extensive and intermediate metabolizers of CYP2C19 (P<0.05). No significant associations were observed between CYP2B6 genotypes and induction by efavirenz on omeprazole metabolism. Our data indicate how interplays between drug interactions and CYP2C19 genetic variations may influence systemic exposure of CYP2C19 substrates.The Pharmacogenomics Journal advance online publication, 30 April 2013; doi:10.1038/tpj.2013.17.
The Pharmacogenomics Journal 04/2013; · 5.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: Up to 25 % of patients discontinue adjuvant aromatase inhibitor (AI) therapy due to intolerable symptoms. Predictors of which patients will be unable to tolerate these medications have not been defined. We hypothesized that inherited variants in candidate genes are associated with treatment discontinuation because of AI-associated toxicity. We prospectively evaluated reasons for treatment discontinuation in women with hormone receptor-positive breast cancer initiating adjuvant AI through a multicenter, prospective, randomized clinical trial of exemestane versus letrozole. Using multiple genetic models, we evaluated potential associations between discontinuation of AI therapy because of toxicity and 138 variants in 24 candidate genes, selected a priori, primarily with roles in estrogen metabolism and signaling. To account for multiple comparisons, statistical significance was defined as p < 0.00036. Of the 467 enrolled patients with available germline DNA, 152 (33 %) discontinued AI therapy because of toxicity. Using a recessive statistical model, an intronic variant in ESR1 (rs9322336) was associated with increased risk of musculoskeletal toxicity-related exemestane discontinuation [HR 5.0 (95 % CI 2.1-11.8), p < 0.0002]. An inherited variant potentially affecting estrogen signaling may be associated with exemestane-associated toxicity, which could partially account for intra-patient differences in AI tolerability. Validation of this finding is required.
Breast Cancer Research and Treatment 04/2013; · 4.47 Impact Factor
[show abstract][hide abstract] ABSTRACT: Background
Beyond estrogen receptor (ER), there are no validated predictors for tamoxifen (TAM) efficacy and toxicity. We utilized a genome-wide cell-based model to comprehensively evaluate genetic variants for their contribution to cellular sensitivity to TAM.DesignOur discovery model incorporates multidimensional datasets, including genome-wide genotype, gene expression, and endoxifen-induced cellular growth inhibition in the International HapMap lymphoblastoid cell lines (LCLs). Genome-wide findings were further evaluated in NCI60 cancer cell lines. Gene knock-down experiments were performed in four breast cancer cell lines. Genetic variants identified in the cell-based model were examined in 245 Caucasian breast cancer patients who underwent TAM treatment.ResultsWe identified seven novel single-nucleotide polymorphisms (SNPs) associated with endoxifen sensitivity through the expression of 10 genes using the genome-wide integrative analysis. All 10 genes identified in LCLs were associated with TAM sensitivity in NCI60 cancer cell lines, including USP7. USP7 knock-down resulted in increasing resistance to TAM in four breast cancer cell lines tested, which is consistent with the finding in LCLs and in the NCI60 cells. Furthermore, we identified SNPs that were associated with TAM-induced toxicities in breast cancer patients, after adjusting for other clinical factors.Conclusion
Our work demonstrates the utility of a cell-based model in genome-wide identification of pharmacogenomic markers.
[show abstract][hide abstract] ABSTRACT: Background: Beyond estrogen receptor (ER), there are no validated predictors for tamoxifen (TAM) efficacy and toxicity. We utilized a genome-wide cell-based model to comprehensively evaluate genetic variants for their contribution to cellular sensitivity to TAM.
Design: Our discovery model incorporates multidimensional datasets, including genome-wide genotype, gene expression, and endoxifen-induced cellular growth inhibition in the International HapMap lymphoblastoid cell lines (LCLs). Genome-wide findings were further evaluated in NCI60 cancer cell lines. Gene knock-down experiments were performed in four breast cancer cell lines. Genetic variants identified in the cell-based model were examined in 245 Caucasian breast cancer patients who underwent TAM treatment.
Results: We identified seven novel single-nucleotide polymorphisms (SNPs) associated with endoxifen sensitivity through the expression of 10 genes using the genome-wide integrative analysis. All 10 genes identified in LCLs were associated with TAM sensitivity in NCI60 cancer cell lines, including USP7. USP7 knock-down resulted in increasing resistance to TAM in four breast cancer cell lines tested, which is consistent with the finding in LCLs and in the NCI60 cells. Furthermore, we identified SNPs that were associated with TAM-induced toxicities in breast cancer patients, after adjusting for other clinical factors.
Conclusion: Our work demonstrates the utility of a cell-based model in genome-wide identification of pharmacogenomic markers.
[show abstract][hide abstract] ABSTRACT: PURPOSE: Many breast cancer survivors (BCS) take multiple medications for health problems associated with the treated cancer and other noncancer comorbidities. However, there is no published, large-scale descriptive evaluation of medication use in BCS compared to midlife women. The purpose of this study was (1) to compare the number and types of prescription medications and over-the-counter medications between BCS and midlife women without cancer and (2) to assess possible drug-drug interactions by evaluating the cytochrome P450 isoform properties of medications (inductors and inhibitors) in both groups. METHODS: A cross-sectional, descriptive, comparative design was used. Baseline data from 98 BCS and 138 midlife women without cancer was analyzed from a behavioral intervention trial for menopausal symptoms. RESULTS: BCS were taking significantly more prescription medications and a larger variety of different types of medication classifications (p < 0.05) after controlling for group differences (race, noncancer comorbid conditions, marital status, income, and smoking) in demographics. Twenty-four women were taking at least one medication considered to be a cytochrome P450 isoforms (CYP) inhibitor or inducer capable of clinical drug-drug interactions with no differences in CYP inhibitors or inducers found between groups. CONCLUSION: BCS are taking a vast array of medications during survivorship. It is unclear if prescription medications are managed by a single healthcare provider or several providers. Clinical implications are to monitor for possible interactions among the various prescription medications, over-the-counter medications, and supplements. Implications for behavioral and biomedical research are that clinical studies need to carefully assess and account for multiple medication uses. RELEVANCE OF THE STUDY: The findings of this study are relevant to research and practice for both oncology and general practitioners. The importance of assessing medication information provides information about symptom management in individuals surviving cancer. In addition, the potential interaction of drugs impacts efficacy of various treatments and impacts compliance by patients.
Supportive Care in Cancer 02/2013; · 2.09 Impact Factor
[show abstract][hide abstract] ABSTRACT: Polymorphisms in CYP2D6 and CYP2C19 affect the efficacy and safety of tricyclics, with some drugs being affected by CYP2D6 only, and others by both polymorphic enzymes. Amitriptyline, clomipramine, doxepin, imipramine, and trimipramine are demethylated by CYP2C19 to pharmacologically active metabolites. These drugs and their metabolites, along with desipramine and nortriptyline, undergo hydroxylation by CYP2D6 to less active metabolites. Evidence from published literature is presented for CYP2D6 and CYP2C19 genotype-directed dosing of tricyclic antidepressants.Clinical Pharmacology & Therapeutics (2013); advance online publication 13 March 2013. doi:10.1038/clpt.2013.2.
[show abstract][hide abstract] ABSTRACT: OBJECTIVE: We previously demonstrated that maternal and fetal genotypes are independently associated with neonatal respiratory distress syndrome (RDS). The objective of the current study is to determine the impact of maternal and fetal single nucleotide polymorphisms (SNPs) in key betamethasone (BMZ) pathways on respiratory outcomes that serve as markers for severity of disease. STUDY DESIGN: DNA was obtained from women given BMZ and their infants. Samples were genotyped for 73 exploratory drug metabolism and glucocorticoid pathway SNPs. Clinical variables and neonatal outcomes were obtained. Logistic regression analysis controlling for relevant clinical variables to determine SNP impact on bronchopulmonary dysplasia (BPD), need for respiratory support, and surfactant therapy use was performed. RESULTS: 109 women delivering 117 infants were analyzed. 14.5% of the infants developed BPD, 70.8% needed some respiratory support after birth, and 27.5% needed surfactant. In a multivariable regression analysis, gestational age at delivery was associated with most neonatal respiratory outcomes (p≤0.01) and chorioamnionitis was associated with BPD (p<0.03). Genotypes associated with respiratory severity outcomes were as follows: BPD- Fetal IPO13 (rs4448553; OR 0.01, 95% CI 0.00-0.92); Surfactant use- Maternal IPO13 (rs2428953 and 2486014; OR 13.8, 95%CI 1.80-105.5 and OR 35.5, 95% CI 1.71-736.6, respectively). CONCLUSIONS: Several discrete maternal and fetal SNPs in the Importin 13 gene (IPO13) may be associated with neonatal respiratory outcomes after maternal antenatal corticosteroid treatment for anticipated preterm birth.
American journal of obstetrics and gynecology 01/2013; · 3.28 Impact Factor
[show abstract][hide abstract] ABSTRACT: Background. The genome-wide association studies (GWAS) have been successful during the last few years. A key challenge is that the interpretation of the results is not straightforward, especially for transacting SNPs. Integration of transcriptome data into GWAS may provide clues elucidating the mechanisms by which a genetic variant leads to a disease. Methods. Here, we developed a novel mediation analysis approach to identify new expression quantitative trait loci (eQTL) driving CYP2D6 activity by combining genotype, gene expression, and enzyme activity data. Results. 389,573 and 1,214,416 SNP-transcript-CYP2D6 activity trios are found strongly associated (P < 10(-5), FDR = 16.6% and 11.7%) for two different genotype platforms, namely, Affymetrix and Illumina, respectively. The majority of eQTLs are trans-SNPs. A single polymorphism leads to widespread downstream changes in the expression of distant genes by affecting major regulators or transcription factors (TFs), which would be visible as an eQTL hotspot and can lead to large and consistent biological effects. Overlapped eQTL hotspots with the mediators lead to the discovery of 64 TFs. Conclusions. Our mediation analysis is a powerful approach in identifying the trans-QTL-phenotype associations. It improves our understanding of the functional genetic variations for the liver metabolism mechanisms.
BioMed research international. 01/2013; 2013:493019.
[show abstract][hide abstract] ABSTRACT: Indoleamine 2,3 dioxygenase (IDO) plays an important role in immunoregulation as it is involved in downregulating immune responses to infections. We sought to characterize IDO activity in histoplasmosis and to do so, C57Bl6 mice were infected intranasally with Histoplasma capsulatum. After infection, lung and spleen IDO activity was assessed by HPLC and IDO expression by qRT-PCR. The distribution of IDO was determined by immunohistochemical staining. Cytokine levels were measured in lung and spleen homogenates using cytokine bead array. Fungal burden was quantified by culture. Subcutaneous pellets containing methyltryptophane (1-MT) were employed to inhibit IDO in vivo. Histoplasma infection strongly induced functional lung IDO, with activity at its highest at weeks 1 and 2 and then decreased thereafter as the mice cleared the infection. Lung IDO activity positively correlated with the fungal burden (Rho = 0.845), interferon-γ (Rho = 0.839) and tumor necrosis factor-α (Rho = 0.791) levels, P < 0.001. In contrast, spleen IDO activity was not induced despite high infection burden and cytokine levels. IDO expressing cells were predominately located at the ring edge of Histoplasma-induced granulomas. IDO inhibition prior to infection reduced fungal burdens and inflammation in lungs and spleen. Histoplasma preferentially induces lung IDO, as early as one week after infection. IDO appears to modulate the immune response to Histoplasma infection.
Medical mycology: official publication of the International Society for Human and Animal Mycology 11/2012; · 2.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: Women with reduced CYP2D6 activity have low endoxifen concentrations and likely worse long term benefits from tamoxifen. We
investigated the association between CYP2D6 genotype and tamoxifen-induced hot flashes in a prospective cohort. We collected hot flash frequency and severity data over
12months from 297 women initiating tamoxifen. We performed CYP2D6 genotyping using the AmpliChip CYP450 test and correlated inherited genetic polymorphisms in CYP2D6 and tamoxifen-induced hot flashes. Intermediate metabolizers had greater mean hot flash scores after 4months of tamoxifen
therapy (44.3) compared to poor metabolizers (20.6, P=0.038) or extensive metabolizers (26.9, P=0.011). At 4months, we observed a trend toward fewer severe hot flashes in poor metabolizers compared to intermediate
plus extensive metabolizers (P=0.062). CYP2D6 activity may be a modest predictive factor for tamoxifen-induced hot flashes. The presence or absence of
hot flashes should not be used to determine tamoxifen’s efficacy.
Breast Cancer Research and Treatment 04/2012; 117(3):571-575. · 4.47 Impact Factor
[show abstract][hide abstract] ABSTRACT: The ESR2 gene encodes the estrogen receptor beta protein. Several studies have shown that genetic variants in the ESR2 gene are associated with a variety of clinical phenotypes. However, very little is known about the functional significance of ESR2 genetic variants. We used a bioinformatics approach to identify regions of the ESR2 promoter that is evolutionarily conserved across the genomes of several species. We resequenced 1.6 kb of the ESR2 gene which included 0.8 kb of the promoter, 0.3 kb of exon ON, and 0.5 kb of the following intron. We identified five single-nucleotide polymorphisms (SNPs) in the ESR2 promoter and one SNP in the intron. Phase analysis indicated that the SNPs likely exist in 11 different haplotypes. Three of the SNPs (rs8008187, rs3829768, rs35036378) were predicted to alter transcription factor binding sites in the ESR2 promoter. All three were detected only in African American subjects. The rs35036378 SNP was in the TATA box and was highly conserved across species. ESR2 promoter reporter assays in LNCaP and SKBR3 cell lines showed that the variant construct containing the rs35036378 SNP allele had approximately 50% less activity relative to the wild-type construct. We conclude that the rs35036378 SNP appears to cause a reduced promoter activity of the ESR2 gene.
[show abstract][hide abstract] ABSTRACT: To determine the impact of maternal and fetal single nucleotide polymorphisms in key betamethasone pathways on neonatal outcomes.
DNA was obtained from women given betamethasone and their infants. Samples were genotyped for 73 exploratory drug metabolism and glucocorticoid pathway single nucleotide polymorphisms. Clinical variables and neonatal outcomes were obtained. Logistic regression analysis using relevant clinical variables and genotypes to model for associations with neonatal respiratory distress syndrome was performed.
One hundred nine women delivering 117 infants were analyzed. Sixty-four infants (49%) developed respiratory distress syndrome. Multivariable analysis revealed that respiratory distress syndrome was associated with maternal single nucleotide polymorphisms in CYP3A5 (odds ratio [OR], 1.63; 95% confidence interval [CI], 1.16-2.30) and the glucocorticoid resistance (OR, 0.28; 95% CI, 0.08-0.95) and fetal single nucleotide polymorphisms in ADCY9 (OR, 0.17; 95% CI, 0.03-0.80) and CYP3A7*1E (rs28451617; OR, 23.68; 95% CI, 1.33-420.6).
Maternal and fetal genotypes are independently associated with neonatal respiratory distress syndrome after treatment with betamethasone for preterm labor.
American journal of obstetrics and gynecology 02/2012; 206(5):447.e17-24. · 3.28 Impact Factor
[show abstract][hide abstract] ABSTRACT: Hepatic nuclear factor 4α (HNF4A) is a nuclear transcription factor that regulates the expression of many genes involved in drug disposition. To identify additional molecular mechanisms that regulate HNF4A, we identified microRNAs (miRNAs) that target HNF4A expression. In silico analyses suggested that HNF4A is targeted by many miRNAs. We conducted in vitro studies to validate several of these predictions. With use of an HNF4A 3'-untranslated region (UTR) luciferase reporter assay, five of six miRNAs tested significantly down-regulated (∼20-40%) the luciferase activity. In HepG2 cells, miR-34a and miR-449a also down-regulated the expression of both the HNF4A protein and an HNF4A target gene, PXR (∼30-40%). This regulation appeared without reduction in HNF4A mRNA expression, suggesting that they must be blocking HNF4A translation. Using additional bioinformatic algorithms, we identified polymorphisms that are predicted to alter the miRNA targeting of HNF4A. Luciferase assays indicated that miR-34a and miR-449a were less effective in regulating a variant (rs11574744) than the wild-type HNF4A 3'-UTR. In vivo, subjects with the variant HNF4A had lower CYP2D6 enzyme activity, although this result was not statistically significant (p = 0.16). In conclusion, our findings demonstrate strong evidence for a role of miRNAs in the regulation of HNF4A.
Drug metabolism and disposition: the biological fate of chemicals 01/2012; 40(4):726-33. · 3.74 Impact Factor