Genome-wide association study of major recurrent depression in the U.K. population.

OBJECTIVE: Studies of major depression in twins and families have shown moderate to high heritability, but extensive molecular studies have failed to identify susceptibility genes convincingly. To detect genetic variants contributing to major depression, the authors performed a genome-wide association study using 1,636 cases of depression ascertained in the U.K. and 1,594 comparison subjects screened negative for psychiatric disorders. METHOD: Cases were collected from 1) a case-control study of recurrent depression (the Depression Case Control [DeCC] study; N=1346), 2) an affected sibling pair linkage study of recurrent depression (probands from the Depression Network [DeNT] study; N=332), and 3) a pharmacogenetic study (the Genome-Based Therapeutic Drugs for Depression [GENDEP] study; N=88). Depression cases and comparison subjects were genotyped at Centre National de Génotypage on the Illumina Human610-Quad BeadChip. After applying stringent quality control criteria for missing genotypes, departure from Hardy-Weinberg equilibrium, and low minor allele frequency, the authors tested for association to depression using logistic regression, correcting for population ancestry. RESULTS: Single nucleotide polymorphisms (SNPs) in BICC1 achieved suggestive evidence for association, which strengthened after imputation of ungenotyped markers, and in analysis of female depression cases. A meta-analysis of U.K. data with previously published results from studies in Munich and Lausanne showed some evidence for association near neuroligin 1 (NLGN1) on chromosome 3, but did not support findings at BICC1. CONCLUSIONS: This study identifies several signals for association worthy of further investigation but, as in previous genome-wide studies, suggests that individual gene contributions to depression are likely to have only minor effects, and very large pooled analyses will be required to identify them.

Antiretroviral drug toxicity in relation to pharmacokinetics, metabolic profile and pharmacogenetics.

Introduction: Besides therapeutic effectiveness, drug tolerability is a key issue for treatments that must be taken indefinitely. Given the high prevalence of toxicity in HIV therapy, the factors implicated in drug-induced morbidities should be identified in order to improve the safety, tolerability and adherence to the treatments. Current approaches have focused almost exclusively on parent drug concentrations; whereas recent evidence suggests that drug metabolites resulting from complex genetic and environmental influences can also contribute to treatment outcome. Pharmacogenetic variations have shown to play a relevant role in the variability observed in antiretroviral drug exposure, clinical response and sometimes toxicity. The integration of pharmacokinetic, pharmacogenetic and metabolic determinants will more probably address current therapeutic needs in patients. Areas covered: This review offers a concise description of three classes of antiretroviral drugs. The review looks at the metabolic profile of these drugs and gives a comprehensive summary of the existing literature on the influence of pharmacogenetics on their pharmacokinetics and metabolic pathways, and the associated drug or metabolite toxicity. Expert opinion: Due to the high prevalence of toxicity and the related risk of low adherence to the treatments, association of kinetic, genetic and metabolic markers predictive of therapeutic or toxicity outcomes could represent a more complete approach for optimizing antiretroviral therapy.

Therapeutic drug monitoring and metabolites profil analysis of antiviral drugs

The widespread use of combination antiretroviral therapy (ARVs) has considerably improved the prognosis of patients infected with HIV. Conversely, considerable advances have been recently realized for the therapy of hepatitis C infection with the recent advent of potent new anti-HCV drugs that allow an increasing rate HCV infection cure. Despite their overall efficacy, a significant number of patients do not achieve or maintain adequate clinical response, defined as an undetectable viral load for HIV, and a sustained virological response (or cure) in HCV infection. Treatment failure therefore still remains an important issue besides drugs toxicities and viral resistance which is not uncommon in a significant percentage of patients who do not reach adequate virological suppression. The reasons of variability in drug response are multifactorial and apart from viral genetics, other factors such as environmental factors, drug- drug interactions, and imperfect compliance may have profound impact on antiviral drugs' clinical response. The possibility of measuring plasma concentration of antiviral drugs enables to guide antiviral drug therapy and ensure optimal drug exposure. The overall objective of this research was to widen up the current knowledge on pharmacokinetic and pharmacogenetic factors that influence the clinical response and toxicity of current and newly approved antiretroviral and anti-HCV drugs. To that endeavour, analytical methods using liquid chromatography coupled with tandem mass spectrometry have been developed and validated for the precise and accurate measurement of new antiretroviral and anti-HCV drugs . These assays have been applied for the TDM of ARVs and anti-HCV in patients infected with either HIV or HCV respectively, and co-infected with HIV- HCV. A pharmacokinetic population model was developed to characterize inter and intra-patient variability of rilpivirine, the latest marketed Non Nucleoside Reverse transcriptase (NNRTI) Inhibitor of HIVand to identify genetic and non genetic covariates influencing rilpivirine exposure. None of the factors investigated so far showed however any influence of RPV clearance. Importantly, we have found that the standard daily dosage regimen (25 mg QD) proposed for rilpivirine results in concentrations below the proposed therapeutic target in about 40% of patients. In these conditions, virologie escape is a potential risk that remains to be further investigated, notably via the TDM approach that can be a useful tool to identify patients who are at risk for being exposed to less than optimal levels of rilpivirine in plasma. Besides the last generation NNRTI rilpivirine, we have studied efavirenz, the major NNRTI clinically used so far. Namely for efavirenz, we aimed at identifying a potential new marker of toxicity that may be incriminated for the neuropsychological sides effects and hence discontinuation of efavirenz therapy. To that endeavour, a comprehensive analysis of phase I and phase II metabolites profiles has been performed in plasma, CSF and in urine from patients under efavirenz therapy. We have found that phase II metabolites of EFV constitute the major species circulating in blood, sometimes exceeding the levels of the parent drug efavirenz. Moreover we have identified a new metabolite of efavirenz in humans, namely the 8-OH-EFV- sulfate which is present at high concentrations in all body compartments from patients under efavirenz therapy. These investigations may open the way to possible alternate phenotypic markers of efavirenz toxicity. Finally, the specific influence of P-glycoprotein on the cellular disposition of a series ARVs (NNRTIs and Pis] has been studies in in vitro cell systems using the siRNA silencing approach. -- Depuis l'introduction de la thérapie antirétrovirale (ARVs) la morbidité et la mortalité liées au VIH ont considérablement diminué. En parallèle le traitement contre le virus de l'hépatite C (VHC) a connu récemment d'énormes progrès avec l'arrivée de nouveaux médicaments puissants, ce qui a permis une augmentation considérable de la guérison de l'infection par le VHC. En dépit de l'efficacité de ces traitements antiviraux, les échecs thérapeutiques ainsi que les effets secondaires des traitements restent un problème important. Une réponse imparfaite ou la toxicité du traitement est certainement multifactorielle. Le suivi thérapeutique des médicaments [Therapeutic Drug Monitoring TDM) à travers la mesure des concentrations plasmatiques constitue une approche importante pour guider le traitement médicamenteux et de s'assurer que les patients sont exposés à des concentrations optimales des médicaments dans le sang, et puissent tirer tout le bénéfice potentiel du traitement. L'objectif global de cette thèse était d'étudier les facteurs pharmacocinétiques et pharmacogénétiques qui influencent l'exposition des médicaments antiviraux (ARVs et anti- VHC) récemment approuvés. A cet effet, des méthodes de quantification des concentrations plasmatiques des médicaments antirétroviraux, anti-VHC ainsi que pour certains métabolites ont été développées et validées en utilisant la Chromatographie liquide couplée à la spectrométrie de masse tandem. Ces méthodes ont été utilisées pour le TDM des ARVs et pour les agents anti-VHC chez les patients infectés par le VIH, et le VHC, respectivement, mais aussi chez les patients co-infectés par le VIH-VHC. Un modèle de pharmacocinétique de population a été développé pour caractériser la variabilité inter-et intra-patient du médicament rilpivirine, un inhibiteur non nucléosidique de la transcriptase de VIH et d'identifier les variables génétiques et non génétiques influençant l'exposition au médicament. Aucun des facteurs étudiés n'a montré d'influence notable sur la clairance de la rilpivirine. Toutefois, la concentration résiduelle extrapolée selon le modèle de pharmacocinétique de population qui a été développé, a montré qu'une grande proportion des patients présente des concentrations minimales inférieures à la cible thérapeutique proposée. Dans ce contexte, la relation entre les concentrations minimales et l'échappement virologique nécessite une surveillance étroite des taux sanguins des patients recevant de la rilpivirine. A cet effet, le suivi thérapeutique est un outil important pour l'identification des patients à risque soient sous-exposés à lai rilpivirine. Pour identifier de nouveaux marqueurs de la toxicité qui pourraient induire l'arrêt du traitement, le profil des métabolites de phase I et de phase II a été étudié dans différentes matrices [plasma, LCR et urine) provenant de patients recevant de l'efavirenz. Les métabolites de phase II, qui n'avaient à ce jour jamais été investigués, constituent les principales espèces présentes dans les matrices étudiées. Au cours de ces investigations, un nouveau métabolite 8- OH-EFV-sulfate a été identifié chez l'homme, et ce dernier est. présent à des concentrations importantes. L'influence de certains facteurs pharmacogénétique des patients sur le profil des métabolites a été étudiée et ouvre la voie à de possibles nouveaux marqueurs phénotypiques alternatifs qui pourraient possiblement mieux prédire la toxicité associée au traitement par l'efavirenz. Finalement, nous nous sommes intéressés à étudier dans un modèle in vitro certains facteurs, comme la P-glycoprotéine, qui influencent la disposition cellulaire de certains médicaments antirétroviraux, en utilisant l'approche par la technologie du siRNA permettant de bloquer sélectivement l'expression du gène de cette protéine d'efflux des médicaments. -- Depuis l'introduction de la thérapie antiretrovirale (ARVs] la morbidité et la mortalité liées au VIH ont considérablement diminué. En parallèle le traitement contre le virus de l'hépatite C (VHC) a connu récemment d'énormes progrès avec l'arrivée de nouveaux médicaments puissants, ce qui a permis une augmentation considérable de la guérison de l'infection par le VHC. En dépit de l'efficacité de ces traitements antiviraux, les échecs thérapeutiques ainsi que les effets secondaires des traitements restent un problème important. Il a pu être démontré que la concentration de médicament présente dans l'organisme est corrélée avec l'efficacité clinique pour la plupart des médicaments agissant contre le VIH et contre le VHC. Les médicaments antiviraux sont généralement donnés à une posologie fixe et standardisée, à tous les patients, il existe cependant une importante variabilité entre les concentrations sanguines mesurées chez les individus. Cette variabilité peut être expliquée par plusieurs facteurs démographiques, environnementaux ou génétiques. Dans ce contexte, le suivi des concentrations sanguines (ou Therapeutic Drug Monitoring, TDM) permet de contrôler que les patients soient exposés à des concentrations suffisantes (pour bloquer la réplication du virus dans l'organisme) et éviter des concentrations excessives, ce qui peut entraîner l'apparition d'intolérence au traitement. Le but de ce travail de thèse est d'améliorer la compréhension des facteurs pharmacologiques et génétiques qui peuvent influencer l'efficacité et/ou la toxicité des médicaments antiviraux, dans le but d'améliorer le suivi des patients. A cet effet, des méthodes de dosage très sensibles et ont été mises au point pour permettre de quantifier les médicaments antiviraux dans le sang et dans d'autres liquides biologiques. Ces méthodes de dosage sont maintenant utilisées d'une part dans le cadre de la prise en charge des patients en routine et d'autre part pour diverses études cliniques chez les patients infectés soit par le HIV, le HCV ou bien coinfectés par les deux virus. Une partie de ce travail a été consacrée à l'investigation des différents facteurs démographiques, génétiques et environnementaux qui pourraient l'influencer la réponse clinique à la rilpivirine, un nouveau médicament contre le VIH. Toutefois, parmi tous les facteurs étudiés à ce jour, aucun n'a permis d'expliquer la variabilité de l'exposition à la rilpivirine chez les patients. On a pu cependant observer qu'à la posologie standard recommandée, un pourcentage relativement élevé de patients pourrait présenter des concentrations inférieures à la concentration sanguine minimale actuellement proposée. Il est donc utile de surveiller étroitement les concentrations de rilpivirine chez les patients pour identifier sans délai ceux qui risquent d'être sous-exposés. Dans l'organisme, le médicament subit diverses transformations (métabolisme) par des enzymes, notamment dans le foie, il est transporté dans les cellules et tissus par des protéines qui modulent sa concentration au site de son action pharmacologique. A cet effet, différents composés (métabolites) produits dans l'organisme après l'administration d'efavirenz, un autre médicament anti-VIH, ont été étudiés. En conclusion, nous nous sommes intéressés à la fois aux facteurs pharmacologiques et génétiques des traitements antiviraux, une approche qui s'inscrit dans l'optique d'une stratégie globale de prise en charge du patient. Dans ce contexte, le suivi des concentrations sanguines de médicaments constitue une des facettes du domaine émergent de la Médecine Personnalisée qui vise à maximiser le bénéfice thérapeutique et le profil de tolérance des médicaments antiviraux

Enantiomers' potential in psychopharmacology--a critical analysis with special emphasis on the antidepressant escitalopram

Stereochemistry is now influencing most areas of pharmacotherapy, with a growing awareness in the field of psychiatry and, more specifically, depression. This is due to the fact that the enantiomers of many chiral drugs may have distinct pharmacological, pharmacokinetic and/or pharmacogenetic profiles. Consequently, in some instances there may be an advantage in using a single enantiomer over the racemic form-thus providing a basis for the development of new therapeutic agents, as well as the potential to improve current treatments. This review highlights some of the potential advantages and disadvantages that using single enantiomers might offer. The principles are exemplified through reference to the stereoselective properties of several established chiral psychotropic drugs, including thioridazine, methadone, trimipramine, mianserin, mirtazapine, fluoxetine and citalopram. Emphasis is given to the treatment of depression and how the potential of one pure enantiomer-escitalopram, the S-enantiomer of the selective serotonin reuptake inhibitor citalopram-appears to be fulfilling its preclinical promise in the clinic.

Multicenter Study on the Clinical Effectiveness, Pharmacokinetics, and Pharmacogenetics of Mirtazapine in Depression.

ABSTRACT: Pharmacogenetic tests and therapeutic drug monitoring may considerably improve the pharmacotherapy of depression. The aim of this study was to evaluate the relationship between the efficacy of mirtazapine (MIR) and the steady-state plasma concentrations of its enantiomers and metabolites in moderately to severely depressed patients, taking their pharmacogenetic status into account. Inpatients and outpatients (n = 45; mean age, 51 years; range, 19-79 years) with major depressive episode received MIR for 8 weeks (30 mg/d on days 1-14 and 30-45 mg/d on days 15-56). Mirtazapine treatment resulted in a significant improvement in mean Hamilton Depression Rating Scale total score at the end of the study (P < 0.0001). There was no evidence for a significant plasma concentration-clinical effectiveness relationship regarding any pharmacokinetic parameter. The enantiomers of MIR and its hydroxylated (OH-MIR) and demethylated (DMIR) metabolites in plasma samples on days 14 and 56 were influenced by sex and age. Nonsmokers (n = 28) had higher mean MIR plasma levels than smokers (n = 17): S(+)-enantiomer of MIR, 9.4 (SD, 3.9) versus 6.2 (SD, 5.5) ng/mL (P = 0.005); R(-)-enantiomer of MIR, 24.4 (SD, 6.5) versus 18.5 (SD, 4.1) ng/mL (P = 0.003). Only in nonsmokers, plasma levels of S(+)-enantiomer of MIR and metabolites depended on the CYP2D6 genotype. Therefore, high CYP1A2 activity seen in smokers seems to mask the influence of the CYP2D6 genotype. In patients presenting the CYP2B6 *6/*6 genotype (n = 8), S-OH-MIR concentrations were higher those in the other patients (n = 37). Although it is not known if S-OH-MIR is associated with the therapeutic effect of MIR, the reduction of the Hamilton scores was significantly (P = 0.016) more pronounced in the CYP2B6 *6/*6-genotyped patients at the end of the study. The role of CYP2B6 in the metabolism and effectiveness of MIR should be further investigated.

Therapeutic drug monitoring in cancer - Are we missing a trick?

Therapeutic drug monitoring (TDM) can be defined as the measurement of drug in biological samples to individualise treatment by adapting drug dose to improve efficacy and/or reduce toxicity. The cytotoxic drugs are characterised by steep dose-response relationships and narrow therapeutic windows. Inter-individual pharmacokinetic (PK) variability is often substantial. There are, however, a multitude of reasons why TDM has never been fully implemented in daily oncology practice. These include difficulties in establishing appropriate concentration target, common use of combination chemotherapies and the paucity of published data from pharmacological trials. The situation is different with targeted therapies. The large interindividual PK variability is influenced by the pharmacogenetic background of the patient (e.g. cytochrome P450 and ABC transporters polymorphisms), patient characteristics such as adherence to treatment and environmental factors (drug-drug interactions). Retrospective studies have shown that targeted drug exposure correlates with treatment response in various cancers. Evidence for imatinib currently exists, others are emerging for compounds including nilotinib, dasatinib, erlotinib, sunitinib, sorafenib and mammalian target of rapamycin (mTOR) inhibitors. Applications for TDM during oral targeted therapies may best be reserved for particular situations including lack of therapeutic response, severe or unexpected toxicities, anticipated drug-drug interactions and concerns over adherence treatment. There are still few data with monoclonal antibodies (mAbs) in favour of TDM approaches, even if data showed encouraging results with rituximab and cetuximab. TDM of mAbs is not yet supported by scientific evidence. Considerable effort should be made for targeted therapies to better define concentration-effect relationships and to perform comparative randomised trials of classic dosing versus pharmacokinetically-guided adaptive dosing.

Therapeutic monitoring of antidepressants in the era of pharmacogenetics studies.

As for other drugs, there is a large interindividual variability of the plasma concentrations of antidepressants for a given dose. Within the last 2 decades, a very large number of pharmacogenetic studies have made it possible to understand the importance of genetic factors on the disposition of drugs in the organism, many of them at the levels of drug metabolism. Polymorphism of CYP2D6 and of other drug-metabolizing enzymes may thus lead to very large differences in drug exposure between patients and possibly also to toxicity or ineffective drug concentrations in some subjects. In consequence, dose recommendations of antidepressants based on genotypes, justified by the principle of administering bioequivalent individualized drug doses, are now proposed. However, blood (and thus possibly brain) concentrations also depend on other factors than the genetic makeup of the patients. Therapeutic drug monitoring of antidepressants allows us to take into account the influence of factors such as comedications, diet, smoking habit, impaired organ function, and compliance. Therapeutic drug monitoring and genotyping are thus complementary, and their combined use contributes to improve pharmacotherapy with antidepressants and other drugs.

Satisfaction survey with DNA cards method to collect genetic samples for pharmacogenetics studies

Background: Pharmacogenetic studies are essential in understanding the interindividual variability of drug responses. DNA sample collection for genotyping is a critical step in genetic studies. A method using dried blood samples from finger-puncture, collected on DNA-cards, has been described as an alternative to the usual venepuncture technique. The purpose of this study is to evaluate the implementation of the DNA cards method in a multicentre clinical trial, and to assess the degree of investigators' satisfaction and the acceptance of the patients perceived by the investigators.Methods: Blood samples were collected on DNA-cards. The quality and quantity of DNA recovered were analyzed. Investigators were questioned regarding their general interest, previous experience, safety issues, preferences and perceived patient satisfaction. Results: 151 patients' blood samples were collected. Genotyping of GST polymorphisms was achieved in all samples (100%). 28 investigators completed the survey. Investigators perceived patient satisfaction as very good (60.7%) or good (39.3%), without reluctance to finger puncture. Investigators preferred this method, which was considered safer and better than the usual methods. All investigators would recommend using it in future genetic studies. Conclusion: Within the clinical trial setting, the DNA-cards method was very well accepted by investigators and patients (in perception of investigators), and was preferred to conventional methods due to its ease of use and safety.

AGNP consensus guidelines for therapeutic drug monitoring in psychiatry: update 2011.

Therapeutic drug monitoring (TDM), i. e., the quantification of serum or plasma concentrations of medications for dose optimization, has proven a valuable tool for the patient-matched psychopharmacotherapy. Uncertain drug adherence, suboptimal tolerability, non-response at therapeutic doses, or pharmacokinetic drug-drug interactions are typical situations when measurement of medication concentrations is helpful. Patient populations that may predominantly benefit from TDM in psychiatry are children, pregnant women, elderly patients, individuals with intelligence disabilities, forensic patients, patients with known or suspected genetically determined pharmacokinetic abnormalities or individuals with pharmacokinetically relevant comorbidities. However, the potential benefits of TDM for optimization of pharmacotherapy can only be obtained if the method is adequately integrated into the clinical treatment process. To promote an appropriate use of TDM, the TDM expert group of the Arbeitsgemeinschaft für Neuropsychopharmakologie und Pharmakopsychiatrie (AGNP) issued guidelines for TDM in psychiatry in 2004. Since then, knowledge has advanced significantly, and new psychopharmacologic agents have been introduced that are also candidates for TDM. Therefore the TDM consensus guidelines were updated and extended to 128 neuropsychiatric drugs. 4 levels of recommendation for using TDM were defined ranging from "strongly recommended" to "potentially useful". Evidence-based "therapeutic reference ranges" and "dose related reference ranges" were elaborated after an extensive literature search and a structured internal review process. A "laboratory alert level" was introduced, i. e., a plasma level at or above which the laboratory should immediately inform the treating physician. Supportive information such as cytochrome P450 substrate and inhibitor properties of medications, normal ranges of ratios of concentrations of drug metabolite to parent drug and recommendations for the interpretative services are given. Recommendations when to combine TDM with pharmacogenetic tests are also provided. Following the guidelines will help to improve the outcomes of psychopharmacotherapy of many patients especially in case of pharmacokinetic problems. Thereby, one should never forget that TDM is an interdisciplinary task that sometimes requires the respectful discussion of apparently discrepant data so that, ultimately, the patient can profit from such a joint eff ort.

Traitement de la dépression résistante par l'association citalopram-lithium. Méthodologie d'une étude multicentrique en double aveugle et résultats préliminaires [Treatment of resistant depression with the citalopram-lithium combination. Methodology of a double-blind multicenter study and preliminary results].

Citalopram, a new bicyclic antidepressant, is the most selective serotonin reuptake inhibitor. In a number of double-blind controlled studies, citalopram was compared to placebo and to known tricyclic antidepressants. These studies have shown their efficacy and good safety. The inefficacy of a psychotropic treatment in at least 20% of depressives has led a number of authors to propose original drug combinations and associations, like antidepressant/lithium (Li), antidepressant/sleep deprivation (agrypnia), antidepressant/ECT, or antidepressant/LT3. The aim of this investigation is to evaluate the clinical effectiveness and safety of a combined citalopram/lithium treatment in therapy-resistant patients, taking account of serotonergic functions, as tested by the fenfluramine/prolactin test, and of drug pharmacokinetics and pharmacogenetics of metabolism. DESIGN OF THE STUDY: A washout period of 3 days before initiating the treatment is included. After an open treatment phase of 28 days (D) with citalopram (20 mg D1-D3; 40 mg D4-D14; 40 or 60 mg D15-D28; concomitant medication allowed: chloral, chlorazepate), the nonresponding patients [less than 50% improvement in the total score on the 21 item-Hamilton Depression Rating Scale (HDRS)] are selected and treated with or without Li (randomized in double-blind conditions: citalopram/Li or citalopram/placebo) during the treatment (D29-D35). Thereafter, all patients included in the double-blind phase subsequently receive an open treatment with citalopram/Li for 7 days (D36-D42). The hypothesis of a relationship between serotoninergic functions in patients using the fenfluramine/prolactin test (D1) and the clinical response to citalopram (and Li) is assessed. Moreover, it is evaluated whether the pharmacogenetic status of the patients, as determined by the mephenytoin/dextromethorphan test (D0-D28), is related to the metabolism of fenfluramine and citalopram, and also to the clinical response. CLINICAL ASSESSMENT: Patients with a diagnosis of major depressive disorders according to DSM III are submitted to a clinical assessment of D1, D7, D14, D28, D35, D42: HDRS, CGI (clinical global impression), VAS (visual analog scales for self-rating of depression), HDRS (Hamilton depression rating scale, 21 items), UKU (side effects scale), and to clinical laboratory examens, as well as ECG, control of weight, pulse, blood pressure at D1, D28, D35. Fenfluramine/prolactin test: A butterfly needle is inserted in a forearm vein at 7 h 45 and is kept patent with liquemine. Samples for plasma prolactin, and d- and l-fenfluramine determinations are drawn at 8 h 15 (base line). Patients are given 60 mg fenfluramine (as a racemate) at 8 h 30. Kinetic points are determined at 9 h 30, 10 h 30, 11 h 30, 12 h 30, 13 h 30. Plasma levels of d- and l-fenfluramine are determined by gas chromatography and prolactin by IRNA. Mephenytoin/dextromethorphan test: Patients empty their bladders before the test; they are then given 25 mg dextropethorphan and 100 mg mephenytoin (as a racemate) at 8 h 00. They collect all urines during the following 8 hours. The metabolic ratio is determined by gas chromatography (metabolic ratio dextromethorphan/dextrorphan greater than 0.3 = PM (poor metabolizer); mephenytoin/4-OH-mephenytoin greater than 5.6, or mephenytoin S/R greater than 0.8 = PM). Citalopram plasma levels: Plasma levels of citalopram, desmethylcitalopram and didesmethylcitalopram are determined by gas chromatography--mass spectrometry. RESULTS OF THE PILOT STUDY. The investigation has been preceded by a pilot study including 14 patients, using the abovementioned protocol, except that all nonresponders were medicated with citalopram/Li on D28 to D42. The mean total score (n = 14) on the 21 item Hamilton scale was significantly reduced after the treatment, ie from 26.93 +/- 5.80 on D1 to 8.57 +/- 6.90 on D35 (p less than 0.001). A similar patCitalopram, a new bicyclic antidepressant, is the most selective serotonin reu

AGNP Consensus Guidelines for Therapeutic Drug Monitoring in Psychiatry: Update 2011.

Therapeutic drug monitoring (TDM), i. e., the quantification of serum or plasma concentrations of medications for dose optimization, has proven a valuable tool for the patient-matched psychopharmacotherapy. Uncertain drug adherence, suboptimal tolerability, non-response at therapeutic doses, or pharmacokinetic drug-drug interactions are typical situations when measurement of medication concentrations is helpful. Patient populations that may predominantly benefit from TDM in psychiatry are children, pregnant women, elderly patients, individuals with intelligence disabilities, forensic patients, patients with known or suspected genetically determined pharmacokinetic abnormalities or individuals with pharmacokinetically relevant comorbidities. However, the potential benefits of TDM for optimization of pharmacotherapy can only be obtained if the method is adequately integrated into the clinical treatment process. To promote an appropriate use of TDM, the TDM expert group of the Arbeitsgemeinschaft für Neuropsychopharmakologie und Pharmakopsychiatrie (AGNP) issued guidelines for TDM in psychiatry in 2004. Since then, knowledge has advanced significantly, and new psychopharmacologic agents have been introduced that are also candidates for TDM. Therefore the TDM consensus guidelines were updated and extended to 128 neuropsychiatric drugs. 4 levels of recommendation for using TDM were defined ranging from "strongly recommended" to "potentially useful". Evidence-based "therapeutic reference ranges" and "dose related reference ranges" were elaborated after an extensive literature search and a structured internal review process. A "laboratory alert level" was introduced, i. e., a plasma level at or above which the laboratory should immediately inform the treating physician. Supportive information such as cytochrome P450 substrate- and inhibitor properties of medications, normal ranges of ratios of concentrations of drug metabolite to parent drug and recommendations for the interpretative services are given. Recommendations when to combine TDM with pharmacogenetic tests are also provided. Following the guidelines will help to improve the outcomes of psychopharmacotherapy of many patients especially in case of pharmacokinetic problems. Thereby, one should never forget that TDM is an interdisciplinary task that sometimes requires the respectful discussion of apparently discrepant data so that, ultimately, the patient can profit from such a joint effort.

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

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

Pharmacogenetics of Antiviral Treatment

RESUME - FRANÇAISRésuméDans ce travail de thèse, l'importance de la pharmacogénétique des traitements antiviraux a été évaluée en déterminant, au moyen de trois différentes approches, l'impact de variations génétiques sur la pharmacocinétique de deux traitements antirétroviraux (à savoir l'efavirenz et le lopinavir) ainsi que sur la capacité de pouvoir éliminer le virus de l'hépatite C de façon naturelle ou suite à un traitement médicamenteux.L'influence des variations génétiques sur les taux plasmatiques de l'efavirenz et de ses métabolites primaires a été évaluée par l'analyse d'un seul gène candidat : le cytochrome P450 (CYP) 2A6, impliqué dans une voie métabolique accessoire de l'efavirenz. Cette étude a permis de démontrer que le génotype du CYP2A6 devient cliniquement déterminant en l'absence de fonction du CYP2B6, impliqué dans la voie métabolique principale, et que la perte simultanée des voies métaboliques principales et accessoires entraine une augmen¬tation du risque d'interruption du traitement, soulignant la valeur prédictive du génotypage.L'influence de la génétique sur la clairance du lopinavir a été évaluée par l'analyse à grande échelle de gènes candidats, à savoir les gènes potentiellement impliqués dans l'absorption, le métabolisme, la distribution et l'élimination d'un médicament. Cette étude a permis l'identification de 4 polymorphismes, dans des transporteurs et des enzymes métaboliques, associés à la clairance du lopinavir et expliquant 5% de la variabilité inter¬individuelle de ce phénotype.L'influence de la génétique sur la capacité d'éliminer le virus de l'hépatite C, de façon naturelle ou à la suite d'un traitement, a été évaluée par l'analyse du génome entier. Cette étude a permis l'identification d'un polymorphisme situé à proximité de l'interféron-X3. Quatre variations génétiques potentiellement causales ont ensuite pu être identifiées par reséquencage. Finalement, la contribution nette de ce gène sur l'élimination du virus a pu être évaluée dans une cohorte infectée par une seule et même source, permettant ainsi de contrôler l'effet de la diversité virale, du genre et de la présence de co-infections.Cette thèse a permis de mettre en évidence les diverses méthodes disponibles pour la recherche en pharmacogénétique, ainsi que l'importance du reséquencage pour l'identification de variations génétiques causales.SUMMARY - ENGLISHSummaryIn this thesis work the relevance of pharmacogenetics of antiviral treatment has been assessed by investigating, through three different approaches, the impact of host genetic variation on antiretroviral drug disposition (namely efavirenz and lopinavir) and on natural or treatment-induced clearance of hepatitis C virus.The influence of host genetic variation on efavirenz and its primary metabolite plasma levels was assessed by single candidate gene approach, through comprehensive analysis of cytochrome P450 (CYP) 2A6 - involved in efavirenz accessory metabolic pathway. The study could demonstrate that CYP2A6 genotype became increasingly relevant in the setting of limited CYP2B6 function - involved in efavirenz main metabolic pathway - and that individuals with both main and accessory metabolic pathways impaired were at higher risk for treatment discontinuation, overall emphasizing the predictive power of genotyping.The influence of host genetic variation on lopinavir clearance was assessed by large scale candidate gene approach, through analysis of genes involved in the absorption, distribution, metabolism and elimination. The study identified four genetic variants in drug transporters and metabolizing enzymes that explained 5% of the interindividual variability in lopinavir clearance.The influence of host genetic variation on hepatitis C virus (HCV) natural or treatment- induced clearance was assessed through genome-wide association study approach. This study identified an intergenic polymorphism, part of a linkage disequilibrium block encompassing the interferon-3 gene, as highly associated with treatment-induced and spontaneous HCV clearance. Resequencing and recombinant mapping lead to the identification of four potentially causal genetic variants. Finally, we could assess the net contribution of genetic variants in interferon-3 to clearance by controlling for viral diversity, gender and co-infection status in a single source infected cohort.This thesis highlights the various genetic tools available to pharmacogenetic discovery (candidate gene, pathway or and genome-wide approaches), and the importance of resequencing for mapping of causal variants.

Buprenorphine and norbuprenorphine quantification in human plasma by simple protein precipitation and ultra-high performance liquid chromatography tandem mass spectrometry.

A highly sensitive ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method was developed for the quantification of buprenorphine and its major metabolite norbuprenorphine in human plasma. In order to speed up the process and decrease costs, sample preparation was performed by simple protein precipitation with acetonitrile. To the best of our knowledge, this is the first application of this extraction technique for the quantification of buprenorphine in plasma. Matrix effects were strongly reduced and selectivity increased by using an efficient chromatographic separation on a sub-2μm column (Acquity UPLC BEH C18 1.7μm, 2.1×50mm) in 5min with a gradient of ammonium formate 20mM pH 3.05 and acetonitrile as mobile phase at a flow rate of 0.4ml/min. Detection was made using a tandem quadrupole mass spectrometer operating in positive electrospray ionization mode, using multiple reaction monitoring. The procedure was fully validated according to the latest Food and Drug Administration guidelines and the Société Française des Sciences et Techniques Pharmaceutiques. Very good results were obtained by using a stable isotope-labeled internal standard for each analyte, to compensate for the variability due to the extraction and ionization steps. The method was very sensitive with lower limits of quantification of 0.1ng/ml for buprenorphine and 0.25ng/ml for norbuprenorphine. The upper limit of quantification was 250ng/ml for both drugs. Trueness (98.4-113.7%), repeatability (1.9-7.7%), intermediate precision (2.6-7.9%) and internal standard-normalized matrix effects (94-101%) were in accordance with international recommendations. The procedure was successfully used to quantify plasma samples from patients included in a clinical pharmacogenetic study and can be transferred for routine therapeutic drug monitoring in clinical laboratories without further development.

New developments in laboratory monitoring of HIV-1 infection.

The laboratory tests currently available to the clinician for day-to-day management of HIV infection are generally limited to the measurement of the viral load and of the CD4 cell count. More recently, analysis of drug resistance and of plasma drug levels have been added to the monitoring armamentarium. There are, however, numerous other techniques currently available to researchers that may in the future be incorporated into clinical routine. These include the analysis of human and viral genetic determinants of disease evolution, detailed analyses of immune recovery and reserve, pharmacogenetic determinants of treatment response, and toxicity. These approaches may in the future provide highly individualized disease management.