Journal of Psychopharmacology http://jop.sagepub.com/
Genetic differences in cytochrome P450 enzymes and antidepressant treatment response Karen Hodgson, Katherine Tansey, Mojca Zvezdana Dernovsek, Joanna Hauser, Neven Henigsberg, Wolfgang Maier, Ole Mors, Anna Placentino, Marcella Rietschel, Daniel Souery, Rebecca Smith, Ian W Craig, Anne E Farmer, Katherine J Aitchison, Sarah Belsy, Oliver SP Davis, Rudolf Uher and Peter McGuffin J Psychopharmacol published online 20 November 2013 DOI: 10.1177/0269881113512041 The online version of this article can be found at: http://jop.sagepub.com/content/early/2013/11/18/0269881113512041
Published by: http://www.sagepublications.com
On behalf of:
British Association for Psychopharmacology
Additional services and information for Journal of Psychopharmacology can be found at: Email Alerts: http://jop.sagepub.com/cgi/alerts Subscriptions: http://jop.sagepub.com/subscriptions Reprints: http://www.sagepub.com/journalsReprints.nav Permissions: http://www.sagepub.com/journalsPermissions.nav
>> OnlineFirst Version of Record - Nov 20, 2013 What is This?
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
512041 2013
JOP0010.1177/0269881113512041Journal of PsychopharmacologyHodgson et al.
Original Paper
Genetic differences in cytochrome P450 enzymes and antidepressant treatment response Karen Hodgson1, Katherine Tansey1, Mojca Zvezdana Dernovšek2, Joanna Hauser3, Neven Henigsberg4, Wolfgang Maier5, Ole Mors6, Anna Placentino7, Marcella Rietschel8, Daniel Souery9, Rebecca Smith1, Ian W Craig1, Anne E Farmer1, Katherine J Aitchison1,10, Sarah Belsy11, Oliver SP Davis1,12, Rudolf Uher1,13 and Peter McGuffin1
Journal of Psychopharmacology 0(0) 1–9 © The Author(s) 2013 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/0269881113512041 jop.sagepub.com
Abstract Aims: Antidepressant response varies between patients, possibly due to differences in the rate cytochrome P450 enzymes metabolise antidepressants into inactive compounds. Drug metabolism rates are influenced by common variants in the genes encoding these enzymes. However, it remains unclear whether treatment outcomes can be predicted by either CYP450 genotype or antidepressant serum concentration. Methods: In GENDEP (a pharmacogenetic study of depressed individuals treated with either escitalopram or nortriptyline), serum concentrations of antidepressants and their primary metabolite were measured after eight weeks treatment and variants in CYP2D6 and CYP2C19 were genotyped. Results: Amongst patients taking escitalopram (n=223), the genotype CYP2C19 was significantly associated with escitalopram serum concentrations and desmethylescitalopram:escitalopram ratio. For those taking nortriptyline (n=161), the CYP2D6 genotype was significantly associated with nortriptyline and 10-hydroxynortriptyline serum concentrations and 10-hydroxynortriptyline:nortriptyline ratio. CYP450 genotypes conferring greater enzyme activity were linked to lower drug serum concentrations and higher metabolite:drug ratios. Nonetheless, no significant association was found between either CYP450 genotype or antidepressant serum concentration and treatment response. Conclusions: While there is a significant relationship between the CYP450 genotype and serum concentrations of escitalopram and nortriptyline, the genotypes are not predictive of differences in treatment response for either drug. Furthermore, differences in antidepressant serum concentrations are not associated with variability in treatment response. Keywords Antidepressant, pharmacogenetics, depression, drug metabolism, cytochrome P450 enzymes
1MRC
Social, Genetic and Developmental Psychiatry Centre, Institute of Psychiatry, King’s College London, London, UK 2University Psychiatric Clinic, Ljubljana, Slovenia 3Laboratory of Psychiatric Genetics, Department of Psychiatry, Poznan University of Medical Sciences, Poznan, Poland 4Croatian Institute for Brain Research, Medical School, University of Zagreb, Zagreb, Croatia 5Department of Psychiatry, University of Bonn, Bonn, Germany 6Centre for Psychiatric Research, Aarhus University Hospital, Risskov, Denmark 7Psychiatric Unit (UOP 23), Department of Mental Health, Spedali Civili Hospital of Brescia, Biological Psychiatry Unit, IRCCS-FBF, Brescia, Italy 8Central Institute of Mental Health, Medical Faculty Mannheim/ Heidelberg University, Mannheim, Germany 9Laboratoire de Psychologie Médicale, Université Libre de Bruxelles; Psy Pluriel – Centre Européen de Psychologie Médicale, Brussels, Belgium
10Department
of Psychiatry, University of Alberta, Edmonton, Canada 11Toxicology Unit, Department of Clinical Biochemistry, King’s College Hospital, London, UK 12The Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK; European Bioinformatics Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, UK 13Department of Psychiatry, Dalhousie University, Halifax, Canada Corresponding author: Karen Hodgson, MRC Social, Genetic and Developmental Psychiatry Centre, Institute of Psychiatry, King’s College London, De Crespigny Park, London, SE5 8AF, UK. Email: [email protected]
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
2
Journal of Psychopharmacology 0(0)
Introduction Cytochrome P450 (CYP450) enzymes are involved in the metabolism of 75% of marketed drugs (Guengerich, 2007). A number of common polymorphisms in the genes encoding the CYP450 enzymes are associated with differences in enzyme activity (see for example, van der Weide and Steijns, 1999). Given that variability in the rate of drug metabolism could alter the concentrations of active compound, it has been suggested that genotyping of CYP450 enzymes may improve the efficacy of many drug therapies by matching drug doses to an individual’s genetically predicted drug metabolism rate (Ingelman-Sundberg, 2004). Antidepressants are amongst those drugs that are metabolised by CYP450 enzymes. The tricyclic antidepressant nortriptyline is metabolised to both cis- and trans- forms of 10-hydroxynortriptyline via the CYP2D6 enzyme (Olesen and Linnet, 1997). The selective serotonin reuptake inhibitor (SSRI) citalopram is a racemic compound and exists as either the R- or S-entantiomer, with S-citalopram being responsible for the desired antidepressant pharmacological effect. Escitalopram, which comprises solely S-citalopram, is also available as an antidepressant. Three CYP enzymes are involved in the metabolism of escitalopram to its primary metabolite desmethylescitalopram; CYP3A4, CYP2C19 and, to a lesser extent, CYP2D6 (Olesen and Linnet, 1999). Genetic variation in the CYP3A4 gene has been reported to be rare and have little impact on the enzymatic activity of CYP3A4 (Lamba et al., 2002). However, genetic differences in CYP2C19 and CYP2D6 have been linked to variability in the serum concentrations of active antidepressant between patients. Previous reports have found genetic variation in CYP2C19 to be related to serum concentrations of escitalopram, although not of its primary metabolite desmethylescitalopram (Huezo-Diaz et al., 2012; Rudberg et al., 2008). Similarly nortriptyline concentrations have been associated with polymorphisms in CYP2D6, albeit in smaller sample sizes (Dahl et al., 1996; Murphy et al., 2001). While there does appear to be a relationship between CYP450 genotypes and antidepressant serum concentrations, this has not been translated robustly into clinically important differences in treatment response. Kirchheiner and colleagues reviewed the relationship between CYP450 genotypes CYP2D6 and CYP2C19 and serum concentrations of drug, attempting to establish genotypespecific dose adjustments for a range of antidepressants, although they did not examine the effect on treatment outcome (Kirchheiner et al., 2001; Kirchheiner et al., 2004). More recent studies into the role of CYP450 genetic variation on antidepressant treatment response report discrepant findings (Mrazek et al., 2011; Peters et al., 2008; Tsai et al., 2010). The results of a pilot study (HallFlavin et al., 2012) suggested that outcomes could be improved in depressed patients when treatment was directed by a pharmacogenomics algorithm which included information on cytochrome P450 genotypes. However, the sample size in this pilot study was small, and the information contained within the algorithm was not limited to drug-metabolising genes, but also encompassed genes involved in the serotonergic system. To date, there remains a lack of direct evidence for the relationship between CYP450 genotype and differences in antidepressant treatment outcomes. The literature looking at the direct relationship between serum concentrations of antidepressants and treatment response is conflicting. The large inter-individual variability observed in serum concentrations (Reis et al., 2009) is a result of not only the genetic
variability in CYP450 enzymes but also environmental factors such as diet, comedication and comorbidities. Therefore, measuring serum levels of antidepressant allows us to consider all of these elements together when investigating the role that drug metabolism differences may have on treatment response. Some findings have indicated that treatment response is linked to serum concentrations of antidepressant. A complex curvilinear relationship has been reported between peripheral concentrations of circulating nortriptyline and treatment response (Asberg et al., 1971; Perry et al., 1994). In a study with citalopram, however, there is discrepant evidence linking serum concentrations of antidepressant to the likelihood of a treatment response (Nikisch et al., 2004). However, a recent study observed that patients with citalopram levels above 50µg/L had improved depression severity scores than patients with lower levels (Ostad Haji et al., 2011). The Arbeitsgemeinschaft für Neuropsychopharmakologie und Pharmakopsychiatrie (AGNP) Consensus Guidelines for 2011 (Hiemke et al., 2011) recommend therapeutic dose monitoring for both nortriptyline and citalopram. However, particularly in the case of citalopram, this is based on evidence predominantly related to the observed variability in the pharmacokinetics of the drug, rather than direct tests of the relationship with serum levels of the drug and treatment outcomes. Using data from the large European pharmacogenetic GenomeBased Therapeutic Drugs for Depression (GENDEP) study, we explored the roles of CYP450 genotypes and serum levels of drug metabolites in antidepressant response to either an SSRI (escitalopram) or a tricyclic antidepressant (nortriptyline). We had three main objectives: (1) to replicate previous work showing that CYP450 genotype influences serum concentrations of antidepressant; (2) to examine whether CYP450 genotype predicted antidepressant treatment response; and (3) to explore the relationship between serum concentrations of antidepressant and treatment response.
Methods Gendep The GENDEP study was a 12-week open-label part-randomized multicentre, pharmacogenetic study of antidepressant treatment response with two active pharmacological treatment arms, escitalopram and nortriptyline. A full description of GENDEP can be found elsewhere (Uher et al., 2009). The complete GENDEP cohort (Tansey et al., 2012) included 868 treatment-seeking adults (19–72 years, 321 men and 547 women) of white European origin diagnosed with moderate to severe unipolar depression according to ICD-10/DSM-IV established using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN) interview (Wing et al., 1998). Exclusion criteria were personal or family history of bipolar disorder or schizophrenia and/or active substance dependence. Patients with no contraindications were randomly allocated to protocol-driven flexible-dosage nortriptyline (50–150 mg daily) or escitalopram (10–30 mg daily). We offered patients with contraindications for one drug the other antidepressant. The severity of depressive symptoms were measured weekly, using the Montgomery–Åsberg Depression Rating Scale (MADRS) (Montgomery and Asberg, 1979). Prior to treatment, mean scores on the MADRS were 28.76 (SD=6.78). GENDEP was approved by ethics boards in all participating centres and all participants
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 3 provided written consent after the procedures were explained. GENDEP is registered at EudraCT (No.20040–017233–8, http:// eudract.emea.europa.eu) and ISRCTN (No. 03693000, http:// www.controlled-trials.com).
Genotypes Blood samples were buffered in an EDTA solution, and DNA was extracted using a standard protocol (Freeman et al., 2003). We genotyped all patients using the Roche AmpliChip CYP450 (Roche Molecular Diagnostics, Alameda, CA, USA), a microarray that measures 33 variants in CYP2D6 and two variants in CYP2C19. We also genotyped an additional common variant in CYP2C19, the *17 allele (Sim et al., 2006), using a TaqMan SNP genotyping assay on the 7900HT sequence detection system (Applied Biosystems, CA, USA). Genotypes were determined using SDS software (Applied Biosystems). CYP2D6: We categorized individuals into four categories based on the 33 genotypes of CYP2D6; poor (PM), intermediate (IM), extensive (EM) or ultrarapid (UM) metabolizers (Rebsamen et al., 2009). CYP2C19: We categorized individuals into one of six categories; poor (PM), intermediate (IM), intermediate plus (IM+), extensive (EM), extensive plus (EM+) or ultrarapid (UM) metabolizers (Mrazek et al., 2011). Drug-relevant genotypic information was available for 89.6% of the whole sample. Further information can be found in Supplementary Materials.
Serum drug concentrations In week 8 of the study, when steady state serum concentrations have been achieved, we measured serum concentrations of both antidepressants and their primary metabolites (Hiemke and Hartter, 2000; Linder and Keck, 1998). Blood samples were taken in the morning, and patients were asked to postpone taking their morning dose of antidepressant until after the samples were collected. All serum analyses were performed at the Department of Clinical Biochemistry, Kings College Hospital, London (UK). Escitalopram, desmethylcitalopram, nortriptyline and total 10-hydroxynortriptyline were measured using achiral turbulent flow liquid chromatography (Couchman, 2012) using an Aria Transcend TLX-II system (ThermoFisher Scientific, San Jose, USA). Detection was by tandem mass spectrometry (MS/MS) (TSQ Vantage, ThermoFisher Scientific, Hemel Hampstead, UK) in positive ionisation mode using atmospheric pressure chemical ionisation (APCI), with two selected reaction monitoring (SRM) transitions used for each analyte. The cis- and trans-isomers of 10-hydroxynortriptyline were resolved and assay calibration was based on the cis-isomer. Sample preparation in both cases was by protein precipitation. Samples, calibration standards, and internal quality control solutions (50 µL) were vortexed (5 min) with a protein crash solution including internal standard (250 µL), at 4oC, in an eppendorf tube. After centrifugation (5 min, 13 000 rpm), the supernatant was transferred to the well of a 96-well plate and 40 µL injected. The method has been fully validated according to FDA guidelines (FDA, 2001). In all cases, detection was by positive mode electrospray ionisation. SRM was used (two m/z transitions per analyte). Each
assay was calibrated using seven calibration solutions over the following ranges; escitalopram, desmethylescitalopram, and nortriptyline: 10–500 µg/L, cis-10-hydroxynortriptyline: 10–1000 µg/L. Internal standards were: LU-102–020, an escitalopram analogue (for escitalopram and desmethylescitalopram) and nortriptyline-D3 (for nortriptyline and cis-10-hydroxynortriptyline). The response for trans-10-hydroxynortriptyline was assumed to be the same as that of cis-10-hydroxynortriptyline. Ion suppression or enhancement was not observed using standard testing procedures. Serum levels of antidepressant were available for 266 patients taking escitalopram and 191 patients taking nortriptyline. In 23 individuals taking escitalopram, serum measurements were taken but antidepressant levels were found to be below the limit of accurate detection (10 µg/L). It is not known whether this is due to non-compliance or very high rates of drug metabolism, but these cases were excluded from the analysis. Of those with serum measurements, 86.97% also had information available on the dose of antidepressant taken that week (see Figure 1 for further details). For ease of interpretation, standardised serum measurements were calculated, with a mean of 0 and a standard deviation of 1. There were no significant differences between patients with and without serum concentration measurements, regarding the baseline characteristics age, sex, initial depression severity or CYP genotype (Supplementary Materials). However, significant differences were observed in terms of treatment response outcomes. In those without serum measurements, 52% dropped out of the study prior to week 8 of the trial. However, when treatment responses, as measured by adjusted percentage change in MADRS scores (Uher et al., 2010), were compared in those who remained in the study until at least week 8, those with serum measurements available (mean=4.115, SE=1.214) were more likely to have responded to treatment than those without serum measurements (mean= −3.769, SE=2.811), a significant difference (Welch’s t-test: t(200.158) = −2.575, p=0.011).
Comedication Patients reported all medications taken throughout the study, in addition to the prescribed antidepressant. We categorized these comedications using the FDA classification of in vivo inhibitors or inducers (FDA, 2011). In the week that we took the blood sample for the measurement of serum levels, 5.81% of patients with serum concentration data reported taking a CYP2C19 and/or CYP2D6 inhibiting drug, but no patients reporting taking CYP2C19 and/or CYP2D6 inducing drugs. A full list of all cytochrome P450-inhibiting comedications taken can be found in the Supplementary Materials.
Statistical analyses We performed all analyses using STATA 11 (StataCorp., 2009). We fitted linear mixed effects models with maximum likelihood. Each analysis included the following fixed effects; age, sex and cytochrome P450-inhibiting comedications, as well as centre of recruitment as a random effect. Given the different metabolic pathways of nortriptyline and escitalopram, all analyses were performed in a drug-specific manner. For escitalopram, CYP2C19 was examined as the relevant CYP450 genotype and we included CYP2D6 genotype as a fixed
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
4
Journal of Psychopharmacology 0(0)
Figure 1. GENDEP sample characteristics.
effect covariate given the smaller reported role of the CYP2D6 enzyme (Olesen and Linnet, 1999). For nortriptyline, CYP2D6 was used as the relevant CYP450 genotype in the nortriptylinespecific analysis.
Cytochrome P450 genotype and serum level of antidepressant. We examined the relevant CYP450 genotype as a potential predictor of three standardised serum measures: concentrations of the antidepressant, its primary metabolite, and the metaboliteto-drug ratio. Daily dose of drug was entered as a covariate to consider dose-independent effects (CYP450 genotype was found to be unrelated to dose).
Cytochrome P450 genotype and treatment response. We also examined CYP450 genotypes as potential predictors of antidepressant treatment response. Given the availability of weekly data on treatment response, we employed repeated measures of depression severity. The model included fixed effects of baseline depression severity, and both linear and quadratic effects of time. We entered centre of recruitment and individual as random effects.
Serum concentration of antidepressant and treatment response. To further explore the relationship between drug metabolism and treatment response, we assessed serum concentration of antidepressant as a potential predictor of treatment response, again using the repeated measures of depression severity. The model included fixed effects of baseline depression severity, linear
and quadratic effects of time with centre of recruitment and individual as random effects. We completed this analysis in three stages. Firstly, we examined serum concentrations of drug and its primary metabolite as predictors. We then repeated this analysis covarying for dosage, to consider how serum concentration might influence treatment response independently from the effects of the dose of drug taken. This was deemed important, given the use of a flexible dosing protocol, where daily drug dose could be altered over the course of the study if necessary (for example, in response to limited improvement in depressive symptoms). Thirdly, we also considered a curvilinear association between serum concentrations of antidepressant and treatment response, given previous evidence for such an association for nortriptyline (Asberg et al., 1971). To achieve this, serum measurements were centred and then squared, prior to examination as predictors, again within a linear mixed model framework as described above.
Results Genotypic frequencies of CYP2C19 and CYP2D6 can be found in Supplementary Materials. Patterns match those expected in Caucasian populations, with the most common genotypic group being EM (extensive metaboliser) for both CYP450 enzymes (de Leon et al., 2006; Mrazek et al., 2011; Rebsamen et al., 2009) CYP450 genotype was unrelated to recruitment centre, age, dose or sex, in either those taking escitalopram or nortriptyline. Dose prescribed and serum concentrations of antidepressant are shown
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 5 Table 1. Daily dose prescribed and serum concentration of drug. Drug taken Escitalopram
Nortriptyline
Daily dose (mg) Serum escitalopram (µg/L) Serum desmethylescitalopram (µg/L) Daily dose (mg) Serum nortriptyline (µg/L) Serum total 10-hydroxynortriptyline (µg/L)
N
Mean
SD
235 266 212 169 191 188
16.44 30.74 10.80 104.94 92.58 66.56
6.44 18.11 4.32 31.35 57.40 52.23
n = number of individuals; SD = standard deviation.
in Table 1. Serum levels are comparable to those observed in other studies (Foglia et al., 1997; Vandel et al., 1978). There was a highly significant relationship between prescribed dose and the serum concentrations of both drug and metabolite for both drugs (Supplementary Materials). There was also significant variability between recruitment centres not only in terms of daily dose prescribed, but also in serum concentrations of antidepressant and metabolite when dose-adjusted values were calculated (Supplementary Materials). Therefore, all analyses included recruitment centre as a covariate. There was no observed association between age and dose or doseadjusted serum concentrations of drug.
Nortriptyline. Genetic variation in CYP2D6 significantly predicted dose-adjusted levels of both nortriptyline (n=161, ß=−0.550 SE=0.115, p=1.90*10−6) and total 10-hydroxynortriptyline (n=158, ß=0.329, SE=0.109, p=0.0026). Genotypes encoding more active forms of the CYP2D6 enzyme were linked to lower serum concentrations of nortriptyline and higher concentrations of total 10-hydroxynortriptyline. Genetic variation linked to higher levels of CYP2D6 activity were associated with higher 10-hydroxynortriptyline to nortriptyline ratios (n=157, ß=0.478, SE=0.116, p=3.57*10−5; Figure 3).
CYP450-inhibiting comedication. All the CYP450-inhibiting
CYP450 genotype. There was no significant relationship
comedications reported were classified by the FDA as ‘weak’ inhibitors of CYP2D6 or CYP2C19. Amongst patients on escitalopram, those taking CYP2C19-inhibiting comedications had significantly higher concentrations of escitalopram (mean=41.5, SD=16.97, n=12) than those not taking these medications (mean=30.23, SD=18.03, n=254; t(264)=−2.12, p=0.035). These comedications did not affect desmethylescitalopram concentrations (t(210)=1.44, p=0.15). Patients taking nortriptyline along with CYP2D6-inhibiting comedications had higher concentrations of nortriptyline (mean=145, SD=45.14, n=9) than those without comedications (mean=89.99, SD=56.79, n=182; t(189)=−2.86, p=0.0047). They also had significantly higher levels of total 10-hydroxnortriptyline (mean=102.33, SD=58.82, n=9) than those without comedications (mean=66.86, SD=51.48, n=179; t(186)=−2.00, p=0.047). All analyses included CYP450-inhibiting comedications as a covariate. Additionally, when those individuals taking CYP450-inhibiting comedications are excluded from the analysis, the pattern of findings remains the same.
Predicting circulating serum levels of antidepressants Escitalopram. Genetic variation in CYP2C19 significantly predicted serum concentration of escitalopram (n=223, ß= −0.242 SE=0.042, p=9.35*10−9); genotypes that encode more active forms of the CYP2C19 enzyme were associated with lower levels of escitalopram. CYP2C19 genotype was not associated with concentration of desmethylescitalopram (n=183, ß=0.085, SE=0.046, p=0.065), but was significantly associated with the ratio of desmethylescitalopram to escitalopram (n=182, ß=0.288, SE=0.057, p=4.93*10−7); higher levels of enzyme activity were associated with higher metabolite to drug ratios (Figure 2).
Predicting antidepressant treatment response between genetic variation in CYP450 enzymes and response to treatment with antidepressants in either of the two medications examined in this study (escitalopram, n=443, ß =0.165, SE= 0.233, p=0.478; nortriptyline, n=334, ß =0.127, SE=0.524, p=0.807).
Serum concentrations of antidepressant drug. In the first stage of the analysis, there was a significant association between serum concentrations of escitalopram and treatment response (n=266, ß=0.870, SE=0.345, p=0.012). However, this association was not in the expected direction, with higher serum concentrations of escitalopram predicting poorer treatment response. Neither desmethylescitalopram concentrations nor the ratio of desmethylescitalopram to escitalopram significantly predicted response (Supplementary Materials). In patients taking nortriptyline, higher concentrations of total 10-hydroxynortriptyline were significantly linked to poorer treatment response (n=188, ß=1.403, SE=0.446, p=0.002). There was no significant association between nortriptyline concentrations and treatment response or metabolite to drug ratios (Supplementary Materials). Nonetheless, in the second stage of the analysis when drug dose was added as a covariate, no significant relationship was observed between antidepressant treatment response and serum concentrations of drug, primary metabolite or metabolite:drug ratio, for either antidepressant studied (Table 2). There was no curvilinear relationship between serum concentrations of drug, primary metabolite or metabolite to drug ratio and treatment response, for either drug, either with or without controlling for the effect of dose (Supplementary Materials).
Discussion Despite hopes that pharmacogenetic examination of pharmacokinetic processes would prove clinically useful (Ingelman-Sundberg,
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
6
Journal of Psychopharmacology 0(0)
Figure 2. Relationship between CYP2C19 genotype and concentration of the antidepressant escitalopram.
Figure 3. Relationship between CYP2D6 genotype and concentrations of the antidepressant nortriptyline.
2004), in the GENDEP sample the observed association between CYP450 genotype and serum concentrations of antidepressant does not translate into differences in treatment response for either escitalopram or nortriptyline. Furthermore, additional investigation indicates that differences in serum concentrations of antidepressant do not drive the observed variability in treatment response in this dataset, even when curvilinear relationships are considered. While significant associations were initially observed between higher serum concentrations of both escitalopram and total 10-hydroxynortriptyline and poorer treatment response, controlling for the effects of drug dosage removes these effects. Thus, it seems likely that the relationship is due to the dosing protocol used in
GENDEP. Serum samples were taken after eight weeks treatment, during which time clinicians were able to follow a protocol-driven flexible dosing schedule, informed by depressive symptoms and side-effects. Thus, higher drug doses were prescribed to patients who were failing to adequately respond to treatment. Therefore we would suggest that, given the lack of association between serum concentrations of antidepressant and treatment response when dosage is controlled for, together with the absence of any significant relationship between CYP450 genotype and drug dosage, the evidence indicates that differences in serum concentrations of antidepressants do not predict treatment response. We also found no evidence of a curvilinear relationship between
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 7 Table 2. Association between serum concentrations of antidepressant drug and treatment response, controlling for the effects of dose. Dose-adjusted levels
n
ß (SE)
p
Escitalopram Desmethylescitalopram Desmethylescitalopram: escitalopram Nortriptyline Total 10-hydroxynortriptyline Total 10-hydroxynortriptyline : nortriptyline
235 189 188 169 167 165
0.345 (0.385) −0.460 (0.463) −0.212 (0.356) −0.073 (0.466) 0.716 (0.511) 0.086 (0.466)
0.370 0.320 0.553 0.876 0.161 0.854
n = number of individuals; ß = regression coefficient; SE = standard error.
serum concentrations of antidepressant and treatment response, in contrast with previous reports on treatment response to nortriptyline (Asberg et al., 1971). Uher et al. (Uher et al., 2012) calculated that, for studies addressing predictors of antidepressant treatment outcomes, continuous biomarkers (such as serum levels in this study) should explain at least 6.3% of the variance in treatment response in order to be clinically significant. Using G*Power (http://www.psycho. uni-duesseldorf.de/abteilungen/aap/gpower3/download-and-register), it was calculated that a sample size of n=120 would be needed to detect an effect size of this magnitude with p=0.05, and power of 80%. This study exceeds this sample size, and thus is adequately powered to detect clinically significant associations between serum levels of antidepressant and treatment response. No formal multiple-hypothesis testing corrections were applied to control for false-positive associations. Nonetheless, the key significant findings of CYP450 genotype being associated with antidepressant levels are highly significant (escitalopram, p=9.35*10−9; nortriptyline, p=1.90*10−6), and are thus unlikely to be false positives. Neither CYP450 genotype nor dose-adjusted serum levels predicted treatment response, even at the uncorrected cut-off for statistical significance of p
Genetic differences in cytochrome P450 enzymes and antidepressant treatment response Karen Hodgson, Katherine Tansey, Mojca Zvezdana Dernovsek, Joanna Hauser, Neven Henigsberg, Wolfgang Maier, Ole Mors, Anna Placentino, Marcella Rietschel, Daniel Souery, Rebecca Smith, Ian W Craig, Anne E Farmer, Katherine J Aitchison, Sarah Belsy, Oliver SP Davis, Rudolf Uher and Peter McGuffin J Psychopharmacol published online 20 November 2013 DOI: 10.1177/0269881113512041 The online version of this article can be found at: http://jop.sagepub.com/content/early/2013/11/18/0269881113512041
Published by: http://www.sagepublications.com
On behalf of:
British Association for Psychopharmacology
Additional services and information for Journal of Psychopharmacology can be found at: Email Alerts: http://jop.sagepub.com/cgi/alerts Subscriptions: http://jop.sagepub.com/subscriptions Reprints: http://www.sagepub.com/journalsReprints.nav Permissions: http://www.sagepub.com/journalsPermissions.nav
>> OnlineFirst Version of Record - Nov 20, 2013 What is This?
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
512041 2013
JOP0010.1177/0269881113512041Journal of PsychopharmacologyHodgson et al.
Original Paper
Genetic differences in cytochrome P450 enzymes and antidepressant treatment response Karen Hodgson1, Katherine Tansey1, Mojca Zvezdana Dernovšek2, Joanna Hauser3, Neven Henigsberg4, Wolfgang Maier5, Ole Mors6, Anna Placentino7, Marcella Rietschel8, Daniel Souery9, Rebecca Smith1, Ian W Craig1, Anne E Farmer1, Katherine J Aitchison1,10, Sarah Belsy11, Oliver SP Davis1,12, Rudolf Uher1,13 and Peter McGuffin1
Journal of Psychopharmacology 0(0) 1–9 © The Author(s) 2013 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav DOI: 10.1177/0269881113512041 jop.sagepub.com
Abstract Aims: Antidepressant response varies between patients, possibly due to differences in the rate cytochrome P450 enzymes metabolise antidepressants into inactive compounds. Drug metabolism rates are influenced by common variants in the genes encoding these enzymes. However, it remains unclear whether treatment outcomes can be predicted by either CYP450 genotype or antidepressant serum concentration. Methods: In GENDEP (a pharmacogenetic study of depressed individuals treated with either escitalopram or nortriptyline), serum concentrations of antidepressants and their primary metabolite were measured after eight weeks treatment and variants in CYP2D6 and CYP2C19 were genotyped. Results: Amongst patients taking escitalopram (n=223), the genotype CYP2C19 was significantly associated with escitalopram serum concentrations and desmethylescitalopram:escitalopram ratio. For those taking nortriptyline (n=161), the CYP2D6 genotype was significantly associated with nortriptyline and 10-hydroxynortriptyline serum concentrations and 10-hydroxynortriptyline:nortriptyline ratio. CYP450 genotypes conferring greater enzyme activity were linked to lower drug serum concentrations and higher metabolite:drug ratios. Nonetheless, no significant association was found between either CYP450 genotype or antidepressant serum concentration and treatment response. Conclusions: While there is a significant relationship between the CYP450 genotype and serum concentrations of escitalopram and nortriptyline, the genotypes are not predictive of differences in treatment response for either drug. Furthermore, differences in antidepressant serum concentrations are not associated with variability in treatment response. Keywords Antidepressant, pharmacogenetics, depression, drug metabolism, cytochrome P450 enzymes
1MRC
Social, Genetic and Developmental Psychiatry Centre, Institute of Psychiatry, King’s College London, London, UK 2University Psychiatric Clinic, Ljubljana, Slovenia 3Laboratory of Psychiatric Genetics, Department of Psychiatry, Poznan University of Medical Sciences, Poznan, Poland 4Croatian Institute for Brain Research, Medical School, University of Zagreb, Zagreb, Croatia 5Department of Psychiatry, University of Bonn, Bonn, Germany 6Centre for Psychiatric Research, Aarhus University Hospital, Risskov, Denmark 7Psychiatric Unit (UOP 23), Department of Mental Health, Spedali Civili Hospital of Brescia, Biological Psychiatry Unit, IRCCS-FBF, Brescia, Italy 8Central Institute of Mental Health, Medical Faculty Mannheim/ Heidelberg University, Mannheim, Germany 9Laboratoire de Psychologie Médicale, Université Libre de Bruxelles; Psy Pluriel – Centre Européen de Psychologie Médicale, Brussels, Belgium
10Department
of Psychiatry, University of Alberta, Edmonton, Canada 11Toxicology Unit, Department of Clinical Biochemistry, King’s College Hospital, London, UK 12The Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, UK; European Bioinformatics Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, UK 13Department of Psychiatry, Dalhousie University, Halifax, Canada Corresponding author: Karen Hodgson, MRC Social, Genetic and Developmental Psychiatry Centre, Institute of Psychiatry, King’s College London, De Crespigny Park, London, SE5 8AF, UK. Email: [email protected]
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
2
Journal of Psychopharmacology 0(0)
Introduction Cytochrome P450 (CYP450) enzymes are involved in the metabolism of 75% of marketed drugs (Guengerich, 2007). A number of common polymorphisms in the genes encoding the CYP450 enzymes are associated with differences in enzyme activity (see for example, van der Weide and Steijns, 1999). Given that variability in the rate of drug metabolism could alter the concentrations of active compound, it has been suggested that genotyping of CYP450 enzymes may improve the efficacy of many drug therapies by matching drug doses to an individual’s genetically predicted drug metabolism rate (Ingelman-Sundberg, 2004). Antidepressants are amongst those drugs that are metabolised by CYP450 enzymes. The tricyclic antidepressant nortriptyline is metabolised to both cis- and trans- forms of 10-hydroxynortriptyline via the CYP2D6 enzyme (Olesen and Linnet, 1997). The selective serotonin reuptake inhibitor (SSRI) citalopram is a racemic compound and exists as either the R- or S-entantiomer, with S-citalopram being responsible for the desired antidepressant pharmacological effect. Escitalopram, which comprises solely S-citalopram, is also available as an antidepressant. Three CYP enzymes are involved in the metabolism of escitalopram to its primary metabolite desmethylescitalopram; CYP3A4, CYP2C19 and, to a lesser extent, CYP2D6 (Olesen and Linnet, 1999). Genetic variation in the CYP3A4 gene has been reported to be rare and have little impact on the enzymatic activity of CYP3A4 (Lamba et al., 2002). However, genetic differences in CYP2C19 and CYP2D6 have been linked to variability in the serum concentrations of active antidepressant between patients. Previous reports have found genetic variation in CYP2C19 to be related to serum concentrations of escitalopram, although not of its primary metabolite desmethylescitalopram (Huezo-Diaz et al., 2012; Rudberg et al., 2008). Similarly nortriptyline concentrations have been associated with polymorphisms in CYP2D6, albeit in smaller sample sizes (Dahl et al., 1996; Murphy et al., 2001). While there does appear to be a relationship between CYP450 genotypes and antidepressant serum concentrations, this has not been translated robustly into clinically important differences in treatment response. Kirchheiner and colleagues reviewed the relationship between CYP450 genotypes CYP2D6 and CYP2C19 and serum concentrations of drug, attempting to establish genotypespecific dose adjustments for a range of antidepressants, although they did not examine the effect on treatment outcome (Kirchheiner et al., 2001; Kirchheiner et al., 2004). More recent studies into the role of CYP450 genetic variation on antidepressant treatment response report discrepant findings (Mrazek et al., 2011; Peters et al., 2008; Tsai et al., 2010). The results of a pilot study (HallFlavin et al., 2012) suggested that outcomes could be improved in depressed patients when treatment was directed by a pharmacogenomics algorithm which included information on cytochrome P450 genotypes. However, the sample size in this pilot study was small, and the information contained within the algorithm was not limited to drug-metabolising genes, but also encompassed genes involved in the serotonergic system. To date, there remains a lack of direct evidence for the relationship between CYP450 genotype and differences in antidepressant treatment outcomes. The literature looking at the direct relationship between serum concentrations of antidepressants and treatment response is conflicting. The large inter-individual variability observed in serum concentrations (Reis et al., 2009) is a result of not only the genetic
variability in CYP450 enzymes but also environmental factors such as diet, comedication and comorbidities. Therefore, measuring serum levels of antidepressant allows us to consider all of these elements together when investigating the role that drug metabolism differences may have on treatment response. Some findings have indicated that treatment response is linked to serum concentrations of antidepressant. A complex curvilinear relationship has been reported between peripheral concentrations of circulating nortriptyline and treatment response (Asberg et al., 1971; Perry et al., 1994). In a study with citalopram, however, there is discrepant evidence linking serum concentrations of antidepressant to the likelihood of a treatment response (Nikisch et al., 2004). However, a recent study observed that patients with citalopram levels above 50µg/L had improved depression severity scores than patients with lower levels (Ostad Haji et al., 2011). The Arbeitsgemeinschaft für Neuropsychopharmakologie und Pharmakopsychiatrie (AGNP) Consensus Guidelines for 2011 (Hiemke et al., 2011) recommend therapeutic dose monitoring for both nortriptyline and citalopram. However, particularly in the case of citalopram, this is based on evidence predominantly related to the observed variability in the pharmacokinetics of the drug, rather than direct tests of the relationship with serum levels of the drug and treatment outcomes. Using data from the large European pharmacogenetic GenomeBased Therapeutic Drugs for Depression (GENDEP) study, we explored the roles of CYP450 genotypes and serum levels of drug metabolites in antidepressant response to either an SSRI (escitalopram) or a tricyclic antidepressant (nortriptyline). We had three main objectives: (1) to replicate previous work showing that CYP450 genotype influences serum concentrations of antidepressant; (2) to examine whether CYP450 genotype predicted antidepressant treatment response; and (3) to explore the relationship between serum concentrations of antidepressant and treatment response.
Methods Gendep The GENDEP study was a 12-week open-label part-randomized multicentre, pharmacogenetic study of antidepressant treatment response with two active pharmacological treatment arms, escitalopram and nortriptyline. A full description of GENDEP can be found elsewhere (Uher et al., 2009). The complete GENDEP cohort (Tansey et al., 2012) included 868 treatment-seeking adults (19–72 years, 321 men and 547 women) of white European origin diagnosed with moderate to severe unipolar depression according to ICD-10/DSM-IV established using the Schedules for Clinical Assessment in Neuropsychiatry (SCAN) interview (Wing et al., 1998). Exclusion criteria were personal or family history of bipolar disorder or schizophrenia and/or active substance dependence. Patients with no contraindications were randomly allocated to protocol-driven flexible-dosage nortriptyline (50–150 mg daily) or escitalopram (10–30 mg daily). We offered patients with contraindications for one drug the other antidepressant. The severity of depressive symptoms were measured weekly, using the Montgomery–Åsberg Depression Rating Scale (MADRS) (Montgomery and Asberg, 1979). Prior to treatment, mean scores on the MADRS were 28.76 (SD=6.78). GENDEP was approved by ethics boards in all participating centres and all participants
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 3 provided written consent after the procedures were explained. GENDEP is registered at EudraCT (No.20040–017233–8, http:// eudract.emea.europa.eu) and ISRCTN (No. 03693000, http:// www.controlled-trials.com).
Genotypes Blood samples were buffered in an EDTA solution, and DNA was extracted using a standard protocol (Freeman et al., 2003). We genotyped all patients using the Roche AmpliChip CYP450 (Roche Molecular Diagnostics, Alameda, CA, USA), a microarray that measures 33 variants in CYP2D6 and two variants in CYP2C19. We also genotyped an additional common variant in CYP2C19, the *17 allele (Sim et al., 2006), using a TaqMan SNP genotyping assay on the 7900HT sequence detection system (Applied Biosystems, CA, USA). Genotypes were determined using SDS software (Applied Biosystems). CYP2D6: We categorized individuals into four categories based on the 33 genotypes of CYP2D6; poor (PM), intermediate (IM), extensive (EM) or ultrarapid (UM) metabolizers (Rebsamen et al., 2009). CYP2C19: We categorized individuals into one of six categories; poor (PM), intermediate (IM), intermediate plus (IM+), extensive (EM), extensive plus (EM+) or ultrarapid (UM) metabolizers (Mrazek et al., 2011). Drug-relevant genotypic information was available for 89.6% of the whole sample. Further information can be found in Supplementary Materials.
Serum drug concentrations In week 8 of the study, when steady state serum concentrations have been achieved, we measured serum concentrations of both antidepressants and their primary metabolites (Hiemke and Hartter, 2000; Linder and Keck, 1998). Blood samples were taken in the morning, and patients were asked to postpone taking their morning dose of antidepressant until after the samples were collected. All serum analyses were performed at the Department of Clinical Biochemistry, Kings College Hospital, London (UK). Escitalopram, desmethylcitalopram, nortriptyline and total 10-hydroxynortriptyline were measured using achiral turbulent flow liquid chromatography (Couchman, 2012) using an Aria Transcend TLX-II system (ThermoFisher Scientific, San Jose, USA). Detection was by tandem mass spectrometry (MS/MS) (TSQ Vantage, ThermoFisher Scientific, Hemel Hampstead, UK) in positive ionisation mode using atmospheric pressure chemical ionisation (APCI), with two selected reaction monitoring (SRM) transitions used for each analyte. The cis- and trans-isomers of 10-hydroxynortriptyline were resolved and assay calibration was based on the cis-isomer. Sample preparation in both cases was by protein precipitation. Samples, calibration standards, and internal quality control solutions (50 µL) were vortexed (5 min) with a protein crash solution including internal standard (250 µL), at 4oC, in an eppendorf tube. After centrifugation (5 min, 13 000 rpm), the supernatant was transferred to the well of a 96-well plate and 40 µL injected. The method has been fully validated according to FDA guidelines (FDA, 2001). In all cases, detection was by positive mode electrospray ionisation. SRM was used (two m/z transitions per analyte). Each
assay was calibrated using seven calibration solutions over the following ranges; escitalopram, desmethylescitalopram, and nortriptyline: 10–500 µg/L, cis-10-hydroxynortriptyline: 10–1000 µg/L. Internal standards were: LU-102–020, an escitalopram analogue (for escitalopram and desmethylescitalopram) and nortriptyline-D3 (for nortriptyline and cis-10-hydroxynortriptyline). The response for trans-10-hydroxynortriptyline was assumed to be the same as that of cis-10-hydroxynortriptyline. Ion suppression or enhancement was not observed using standard testing procedures. Serum levels of antidepressant were available for 266 patients taking escitalopram and 191 patients taking nortriptyline. In 23 individuals taking escitalopram, serum measurements were taken but antidepressant levels were found to be below the limit of accurate detection (10 µg/L). It is not known whether this is due to non-compliance or very high rates of drug metabolism, but these cases were excluded from the analysis. Of those with serum measurements, 86.97% also had information available on the dose of antidepressant taken that week (see Figure 1 for further details). For ease of interpretation, standardised serum measurements were calculated, with a mean of 0 and a standard deviation of 1. There were no significant differences between patients with and without serum concentration measurements, regarding the baseline characteristics age, sex, initial depression severity or CYP genotype (Supplementary Materials). However, significant differences were observed in terms of treatment response outcomes. In those without serum measurements, 52% dropped out of the study prior to week 8 of the trial. However, when treatment responses, as measured by adjusted percentage change in MADRS scores (Uher et al., 2010), were compared in those who remained in the study until at least week 8, those with serum measurements available (mean=4.115, SE=1.214) were more likely to have responded to treatment than those without serum measurements (mean= −3.769, SE=2.811), a significant difference (Welch’s t-test: t(200.158) = −2.575, p=0.011).
Comedication Patients reported all medications taken throughout the study, in addition to the prescribed antidepressant. We categorized these comedications using the FDA classification of in vivo inhibitors or inducers (FDA, 2011). In the week that we took the blood sample for the measurement of serum levels, 5.81% of patients with serum concentration data reported taking a CYP2C19 and/or CYP2D6 inhibiting drug, but no patients reporting taking CYP2C19 and/or CYP2D6 inducing drugs. A full list of all cytochrome P450-inhibiting comedications taken can be found in the Supplementary Materials.
Statistical analyses We performed all analyses using STATA 11 (StataCorp., 2009). We fitted linear mixed effects models with maximum likelihood. Each analysis included the following fixed effects; age, sex and cytochrome P450-inhibiting comedications, as well as centre of recruitment as a random effect. Given the different metabolic pathways of nortriptyline and escitalopram, all analyses were performed in a drug-specific manner. For escitalopram, CYP2C19 was examined as the relevant CYP450 genotype and we included CYP2D6 genotype as a fixed
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
4
Journal of Psychopharmacology 0(0)
Figure 1. GENDEP sample characteristics.
effect covariate given the smaller reported role of the CYP2D6 enzyme (Olesen and Linnet, 1999). For nortriptyline, CYP2D6 was used as the relevant CYP450 genotype in the nortriptylinespecific analysis.
Cytochrome P450 genotype and serum level of antidepressant. We examined the relevant CYP450 genotype as a potential predictor of three standardised serum measures: concentrations of the antidepressant, its primary metabolite, and the metaboliteto-drug ratio. Daily dose of drug was entered as a covariate to consider dose-independent effects (CYP450 genotype was found to be unrelated to dose).
Cytochrome P450 genotype and treatment response. We also examined CYP450 genotypes as potential predictors of antidepressant treatment response. Given the availability of weekly data on treatment response, we employed repeated measures of depression severity. The model included fixed effects of baseline depression severity, and both linear and quadratic effects of time. We entered centre of recruitment and individual as random effects.
Serum concentration of antidepressant and treatment response. To further explore the relationship between drug metabolism and treatment response, we assessed serum concentration of antidepressant as a potential predictor of treatment response, again using the repeated measures of depression severity. The model included fixed effects of baseline depression severity, linear
and quadratic effects of time with centre of recruitment and individual as random effects. We completed this analysis in three stages. Firstly, we examined serum concentrations of drug and its primary metabolite as predictors. We then repeated this analysis covarying for dosage, to consider how serum concentration might influence treatment response independently from the effects of the dose of drug taken. This was deemed important, given the use of a flexible dosing protocol, where daily drug dose could be altered over the course of the study if necessary (for example, in response to limited improvement in depressive symptoms). Thirdly, we also considered a curvilinear association between serum concentrations of antidepressant and treatment response, given previous evidence for such an association for nortriptyline (Asberg et al., 1971). To achieve this, serum measurements were centred and then squared, prior to examination as predictors, again within a linear mixed model framework as described above.
Results Genotypic frequencies of CYP2C19 and CYP2D6 can be found in Supplementary Materials. Patterns match those expected in Caucasian populations, with the most common genotypic group being EM (extensive metaboliser) for both CYP450 enzymes (de Leon et al., 2006; Mrazek et al., 2011; Rebsamen et al., 2009) CYP450 genotype was unrelated to recruitment centre, age, dose or sex, in either those taking escitalopram or nortriptyline. Dose prescribed and serum concentrations of antidepressant are shown
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 5 Table 1. Daily dose prescribed and serum concentration of drug. Drug taken Escitalopram
Nortriptyline
Daily dose (mg) Serum escitalopram (µg/L) Serum desmethylescitalopram (µg/L) Daily dose (mg) Serum nortriptyline (µg/L) Serum total 10-hydroxynortriptyline (µg/L)
N
Mean
SD
235 266 212 169 191 188
16.44 30.74 10.80 104.94 92.58 66.56
6.44 18.11 4.32 31.35 57.40 52.23
n = number of individuals; SD = standard deviation.
in Table 1. Serum levels are comparable to those observed in other studies (Foglia et al., 1997; Vandel et al., 1978). There was a highly significant relationship between prescribed dose and the serum concentrations of both drug and metabolite for both drugs (Supplementary Materials). There was also significant variability between recruitment centres not only in terms of daily dose prescribed, but also in serum concentrations of antidepressant and metabolite when dose-adjusted values were calculated (Supplementary Materials). Therefore, all analyses included recruitment centre as a covariate. There was no observed association between age and dose or doseadjusted serum concentrations of drug.
Nortriptyline. Genetic variation in CYP2D6 significantly predicted dose-adjusted levels of both nortriptyline (n=161, ß=−0.550 SE=0.115, p=1.90*10−6) and total 10-hydroxynortriptyline (n=158, ß=0.329, SE=0.109, p=0.0026). Genotypes encoding more active forms of the CYP2D6 enzyme were linked to lower serum concentrations of nortriptyline and higher concentrations of total 10-hydroxynortriptyline. Genetic variation linked to higher levels of CYP2D6 activity were associated with higher 10-hydroxynortriptyline to nortriptyline ratios (n=157, ß=0.478, SE=0.116, p=3.57*10−5; Figure 3).
CYP450-inhibiting comedication. All the CYP450-inhibiting
CYP450 genotype. There was no significant relationship
comedications reported were classified by the FDA as ‘weak’ inhibitors of CYP2D6 or CYP2C19. Amongst patients on escitalopram, those taking CYP2C19-inhibiting comedications had significantly higher concentrations of escitalopram (mean=41.5, SD=16.97, n=12) than those not taking these medications (mean=30.23, SD=18.03, n=254; t(264)=−2.12, p=0.035). These comedications did not affect desmethylescitalopram concentrations (t(210)=1.44, p=0.15). Patients taking nortriptyline along with CYP2D6-inhibiting comedications had higher concentrations of nortriptyline (mean=145, SD=45.14, n=9) than those without comedications (mean=89.99, SD=56.79, n=182; t(189)=−2.86, p=0.0047). They also had significantly higher levels of total 10-hydroxnortriptyline (mean=102.33, SD=58.82, n=9) than those without comedications (mean=66.86, SD=51.48, n=179; t(186)=−2.00, p=0.047). All analyses included CYP450-inhibiting comedications as a covariate. Additionally, when those individuals taking CYP450-inhibiting comedications are excluded from the analysis, the pattern of findings remains the same.
Predicting circulating serum levels of antidepressants Escitalopram. Genetic variation in CYP2C19 significantly predicted serum concentration of escitalopram (n=223, ß= −0.242 SE=0.042, p=9.35*10−9); genotypes that encode more active forms of the CYP2C19 enzyme were associated with lower levels of escitalopram. CYP2C19 genotype was not associated with concentration of desmethylescitalopram (n=183, ß=0.085, SE=0.046, p=0.065), but was significantly associated with the ratio of desmethylescitalopram to escitalopram (n=182, ß=0.288, SE=0.057, p=4.93*10−7); higher levels of enzyme activity were associated with higher metabolite to drug ratios (Figure 2).
Predicting antidepressant treatment response between genetic variation in CYP450 enzymes and response to treatment with antidepressants in either of the two medications examined in this study (escitalopram, n=443, ß =0.165, SE= 0.233, p=0.478; nortriptyline, n=334, ß =0.127, SE=0.524, p=0.807).
Serum concentrations of antidepressant drug. In the first stage of the analysis, there was a significant association between serum concentrations of escitalopram and treatment response (n=266, ß=0.870, SE=0.345, p=0.012). However, this association was not in the expected direction, with higher serum concentrations of escitalopram predicting poorer treatment response. Neither desmethylescitalopram concentrations nor the ratio of desmethylescitalopram to escitalopram significantly predicted response (Supplementary Materials). In patients taking nortriptyline, higher concentrations of total 10-hydroxynortriptyline were significantly linked to poorer treatment response (n=188, ß=1.403, SE=0.446, p=0.002). There was no significant association between nortriptyline concentrations and treatment response or metabolite to drug ratios (Supplementary Materials). Nonetheless, in the second stage of the analysis when drug dose was added as a covariate, no significant relationship was observed between antidepressant treatment response and serum concentrations of drug, primary metabolite or metabolite:drug ratio, for either antidepressant studied (Table 2). There was no curvilinear relationship between serum concentrations of drug, primary metabolite or metabolite to drug ratio and treatment response, for either drug, either with or without controlling for the effect of dose (Supplementary Materials).
Discussion Despite hopes that pharmacogenetic examination of pharmacokinetic processes would prove clinically useful (Ingelman-Sundberg,
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
6
Journal of Psychopharmacology 0(0)
Figure 2. Relationship between CYP2C19 genotype and concentration of the antidepressant escitalopram.
Figure 3. Relationship between CYP2D6 genotype and concentrations of the antidepressant nortriptyline.
2004), in the GENDEP sample the observed association between CYP450 genotype and serum concentrations of antidepressant does not translate into differences in treatment response for either escitalopram or nortriptyline. Furthermore, additional investigation indicates that differences in serum concentrations of antidepressant do not drive the observed variability in treatment response in this dataset, even when curvilinear relationships are considered. While significant associations were initially observed between higher serum concentrations of both escitalopram and total 10-hydroxynortriptyline and poorer treatment response, controlling for the effects of drug dosage removes these effects. Thus, it seems likely that the relationship is due to the dosing protocol used in
GENDEP. Serum samples were taken after eight weeks treatment, during which time clinicians were able to follow a protocol-driven flexible dosing schedule, informed by depressive symptoms and side-effects. Thus, higher drug doses were prescribed to patients who were failing to adequately respond to treatment. Therefore we would suggest that, given the lack of association between serum concentrations of antidepressant and treatment response when dosage is controlled for, together with the absence of any significant relationship between CYP450 genotype and drug dosage, the evidence indicates that differences in serum concentrations of antidepressants do not predict treatment response. We also found no evidence of a curvilinear relationship between
Downloaded from jop.sagepub.com at Scientific library of Moscow State University on November 28, 2013
Hodgson et al. 7 Table 2. Association between serum concentrations of antidepressant drug and treatment response, controlling for the effects of dose. Dose-adjusted levels
n
ß (SE)
p
Escitalopram Desmethylescitalopram Desmethylescitalopram: escitalopram Nortriptyline Total 10-hydroxynortriptyline Total 10-hydroxynortriptyline : nortriptyline
235 189 188 169 167 165
0.345 (0.385) −0.460 (0.463) −0.212 (0.356) −0.073 (0.466) 0.716 (0.511) 0.086 (0.466)
0.370 0.320 0.553 0.876 0.161 0.854
n = number of individuals; ß = regression coefficient; SE = standard error.
serum concentrations of antidepressant and treatment response, in contrast with previous reports on treatment response to nortriptyline (Asberg et al., 1971). Uher et al. (Uher et al., 2012) calculated that, for studies addressing predictors of antidepressant treatment outcomes, continuous biomarkers (such as serum levels in this study) should explain at least 6.3% of the variance in treatment response in order to be clinically significant. Using G*Power (http://www.psycho. uni-duesseldorf.de/abteilungen/aap/gpower3/download-and-register), it was calculated that a sample size of n=120 would be needed to detect an effect size of this magnitude with p=0.05, and power of 80%. This study exceeds this sample size, and thus is adequately powered to detect clinically significant associations between serum levels of antidepressant and treatment response. No formal multiple-hypothesis testing corrections were applied to control for false-positive associations. Nonetheless, the key significant findings of CYP450 genotype being associated with antidepressant levels are highly significant (escitalopram, p=9.35*10−9; nortriptyline, p=1.90*10−6), and are thus unlikely to be false positives. Neither CYP450 genotype nor dose-adjusted serum levels predicted treatment response, even at the uncorrected cut-off for statistical significance of p
