Le rein joue un rôle essentiel dans l’élimination des médicaments et de leurs métabolites, de ce fait il assure la défense de l'organisme contre de potentiels xénobiotiques toxiques. Particulièrement, les transporteurs des tubules proximaux rénaux qui ont un rôle dans la sécrétion tubulaire des médicaments. Ainsi, ils sont des déterminants important de la biodisponibilité des xénobiotiques dans l’organisme.Dans cette thèse nous nous sommes intéressés à l’implication des transporteurs rénaux humains hOAT1 et hOAT3 dans des interactions médicamenteuses moyennant des modèles in vitro. Après construction et validation des modèles d’études cellulaires HEK-hOAT1 et HEK-hOAT3, nous avons testé l’effet des inhibiteurs de la pompe à protons sur le transport du méthotrexate par les OATs ainsi que l’effet des antiviraux sur l’influx du tenofovir par ces mêmes transporteurs. Grâce à nos modèles cellulaires nous avons tenté d’expliquer in vitro de probables interactions médicamenteuses décrites en clinique. / The kidney plays an essential role in the elimination of drugs and their metabolites, thus it ensures the defense of the body against potential toxic xenobiotic. Particularly, the secretory transporters in the proximal tubule are major determinants of the disposition of xenobiotic in the body.In this thesis we investigated the involvement of human organic anions transporters hOAT1 and hOAT3 in drug drug interactions through study on in vitro cell models. After construction and validation of cells models studies HEK-hOAT1 and HEK-hOAT3, we tested the effect of proton pump inhibitors on methotrexate transport by OATs and the effect of antivirals on the influx of tenofovir by these two transporters. With our models we tried to explain in vitro probable drug interactions described in the clinic.

The organic cation transporters (OCTs) play a critical role in the absorption, distribution and elimination of many drugs, hormones, herbal medicines, and environmental toxins. Given the broad substrate specificity of OCTs, they fall victim to the high susceptibility for contributing to harmful drug-drug interactions. Further defining how human (h)OCTs mechanistically bind to its broad array of substrates will provide significant insight to the understanding and prediction of drug-drug interactions in polypharmacy patients and the advancement of future rational drug design for therapeutics targeting OCTs. The goal of the current study was to elucidate the critical amino acid residues for transporter-substrate binding interactions on human (h)OCT1 and 2 utilizing in silico molecular modeling techniques (homology modeling and automated docking), as well as in vitro mutagenesis and kinetic transport experiments. Three-dimensional homology models were generated for hOCT1 and 2 using Piriformospora indica phosphate transporter (PiPT) serving as template. A putative binding pocket was identified and used to dock the prototypical substrate MPP+. Docking studies revealed five residues for each transporter (hOCT1 and hOCT2) that may be critical for substrate-transporter interactions. The in silico data was used to guide subsequent in vitro site-directed mutagenesis and kinetic analysis. Four hOCT1 mutants (Gln241Lys, Thr245Lys, Tyr361Ala, and Glu447Lys) and three hOCT2 mutants (Gln242Lys, Tyr362Phe, and Tyr362Ala) showed complete loss of MPP+ transporter activity. Decreased affinity for MPP+ was observed for Phe244Ser and Thr245Ser in hOCT1, and Tyr245Ala in hOCT2. All amino acid residues highlighted in the in vitro experiments may be potentially critical for substrate-transporter interactions particularly Tyr361, Phe244 and Thr245 in hOCT1; and Tyr362 and Tyr245 in hOCT2. Docking of known structurally divergent hOCT1 and hOCT2 substrates revealed similar binding interactions as that identified for MPP+, albeit with some unique residues, suggesting the presence of a large central cavity within both transporters. Through the combination of in silico and in vitro experiments, a putative binding pocket was defined and several residues important for substrate-transporter interaction were identified and verified for hOCT1 and hOCT2. Further defining how OCTs biochemically interact with their broad array of substrates will provide significant insight to the understanding and prediction of drug-drug interactions in polypharmacy patients and the advancement of future rational drug design for therapeutics targeting OCT1 and OCT2.

Biliary excretion is an important elimination route for many drugs and metabolites. For such compounds, it is important to know the extent of excretion and drug exposure in the bile, e.g., for the risk assessment of drug interactions, liver toxicity and the effects of genetic variants. In this thesis, duodenal aspiration of bile was performed in healthy volunteers and complemented with experiments in an in vivo model in pigs to increase the understanding of the intestinal and hepatobiliary disposition of two direct thrombin inhibitors. The compounds investigated, ximelagatran and AZD0837, are both prodrugs that require bioactivation to exert their pharmacological effect. Upon co-administration with erythromycin and ketoconazole, respectively, altered plasma exposure to ximelagatran and AZD0837 and their respective metabolites has been observed. The main objective of this thesis was to characterize the biliary excretion of the compounds, and investigate whether this elimination route explains the observed drug-drug interactions. High plasma-to-bile AUC ratios were observed, in particular for ximelagatran, its active metabolite melagatran, and AR-H067637, the active metabolite of AZD0837. These high ratios indicate the involvement of active transporters in the biliary excretion of the compounds, which is important since transporters constitute possible sites for drug interactions. The effects of erythromycin and ketoconazole on the plasma exposure of the prodrugs and metabolites were confirmed in both the pig and the clinical studies. The changes seen in plasma for ximelagatran and its metabolites were partly explained by reduced biliary clearance. Inhibited CYP3A4 metabolism likely caused the elevated plasma levels of AZD0837, whereas reduced biliary clearance was seen for AR-H067637 suggesting an effect on its excretion into bile. In summary, the studies led to mechanistic insights in the hepatobiliary disposition of ximelagatran and AZD0837, and demonstrate the value of combined clinical and animal studies for the investigation of the biliary drug excretion.

The general aim of this thesis was to improve the understanding of the in vivo pharmacokinetics and, in particular, the metabolism of finasteride, a 5α-reductase inhibitor used in the treatment of enlarged prostate glands and male pattern baldness. CYP3A4 has been identified as the major enzyme involved in the sequential metabolism of finasteride to ω-OH finasteride (M1) and ω-COOH finasteride (M3). The consequences of induced and inhibited metabolism on the pharmacokinetics of finasteride and its metabolites were investigated in humans and pigs. Both studies included bile collection. The collected human and pig samples were used for the metabolite identification. As expected, induced metabolism led to reduced plasma exposure of finasteride and inhibited metabolism had the opposite effect. The interactions were investigated in detail and included examination of the biliary pharmacokinetics of finasteride and its metabolites. In pigs, the study included monitoring of the hepatic extraction over time, deconvolution and the development of a semi-physiological model for comparison of the effects on the gut wall and liver metabolism. For M3, the concentration ratios of bile to plasma and the renal clearance indicated that carrier-mediated processes are involved in the biliary and urinary excretion. This was not, however, the case for finasteride. The metabolite, M1, could not be quantified either in humans or pigs. Instead, two other OH metabolites, M1 isomers, were identified in humans. These metabolites were found to undergo glucuronide conjugation. In humans, one glucuronide was identified intact and in pigs, both glucuronides were identified intact in bile and in urine. In addition, a glucuronide of M3 was identified in human bile. In conclusion, advances have been made in the understanding of the pharmacokinetics of finasteride, in particular in relation to the metabolism. Hopefully, the findings of this comprehensive investigation can be applied to other drugs and novel chemical entities.

A (-)- grandisina (GRA) é um produto natural da classe das lignanas e é encontrada em muitas espécies de plantas das regiões Norte e Nordeste do Brasil. Por apresentar diversas propriedades biológicas, como atividade tripanocida, anti-inflamatória, antinociceptiva, e principalmente atividade antileucêmica e antitumoral contra tumores de Ehrlich, a GRA pode ser considerada um potencial candidato a fármaco. Porém, para que a GRA se torne um fármaco são necessárias diversas etapas de estudos, incluindo estudos pré-clínicos de interações medicamentosas (DDI). As DDI ocorrem principalmente devido a inibições diretas e tempo-dependentes das enzimas do citocromo P450 (CYP450), uma superfamília de enzimas responsável por metabolizar cerca de 75% dos fármacos em uso. Os estudos pré-clínicos de DDI envolvem o conhecimento do potencial inibitório do candidato a fármaco sobre essas enzimas e esses estudos podem ser realizados empregando diversos modelos in vitro, como, por exemplo, microssomas hepáticos de humanos (HLM). Assim, nesse estudo foi avaliado o efeito inibitório da GRA sobre a atividade das principais isoformas do CYP450 e também foram determinadas as isoformas que contribuem para a formação dos metabólitos da GRA. Os resultados demonstraram que múltiplas isoformas participam da formação dos metabólitos da GRA, com destaque para a CYP2C9, que participa da formação de todos os metabólitos. Em relação aos estudos de inibição, foi possível concluir que a GRA é um inibidor fraco da CYP1A2 e CYP2D6, com valores de IC50 maiores do que 200 µM e 100 µM, respectivamente, e um inibidor moderado e competitivo da CYP2C9, com IC50 igual a 40,85 µM e Ki igual a 50,60 µM. Para a CYP3A4 o potencial inibitório da GRA foi avaliado utilizando dois substratos distintos. A GRA demonstrou ser tanto um inibidor dose-dependente moderado e competitivo dessa isoforma, quanto um inibidor tempo-dependente baseado em mecanismo com potencial de inativação equiparável ao do irinotecano, inibidor baseado em mecanismo clinicamente significativo. Utilizando a nifedipina como substrato os valores de IC50 e Ki foram 78,09 µM e 48,71 µM, respectivamente. Já os valores dos parâmetros cinéticos de inativação foram KI= 6,40 µM, kinact= 0,037 min-1 e Clinact= 5,78 mL min-1 µmol-1. Para os ensaios empregando o midazolam os valores de IC50 e Ki foram 48,87 µM e 31,25 µM, respectivamente, e os valores dos parâmetros cinéticos de inativação foram KI= 31,53 µM, kinact= 0,049 min-1 e Clinact= 1,55 mL min-1 µmol-1. Com relação a CYP2E1, por sua vez, foi possível observar que a GRA tem capacidade de aumentar a atividade dessa isoforma significativamente a partir da concentração de 4 µM. Portanto, conclui-se que não há risco da GRA apresentar interações medicamentosas com fármacos metabolizados pela CYP1A2 e CYP2D6, enquanto que para a CYP2C9, apesar da GRA ser um inibidor moderado dessa isoforma, o risco é baixo. Já para medicamentos metabolizados pela CYP2E1 e CYP3A4 o risco de DDI existe e isso deve ser cuidadosamente monitorado in vivo, principalmente porque a CYP3A4 é a isoforma responsável por catalisar o metabolismo da maioria dos fármacos. / (-)-grandisin (GRA) is a lignanic natural product found in many species of plants from North and Northeast of Brazil. This compound has several biological properties, such as trypanocide, anti-inflammatory, antinociceptive, antileukemia activity and antitumor activity against Ehrlich tumor. Because of these biological properties, GRA is considered a potential drug candidate, however, before becoming a new drug, GRA has to undergo various tests, including preclinical drug-drug interactions (DDI) studies. Most of the times, DDI occur because of direct and time-dependent inhibitions of cytochrome P450 (CYP450) enzymes, an enzyme superfamily responsible for metabolizing the vast majority of drugs administered. Preclinical drug-drug interactions studies involve the evaluation of the potential of a drug candidate to inhibit this superfamily of enzymes and these studies can be conducted using in vitro models, such as human liver microsomes (HLM). Therefore, in this project, the inhibitory effect of GRA on the activity of some CYP450 isoforms was evaluated and the isoforms that catalyze the formation of GRA\'s metabolites were also determined. Results showed that multiple CYP450 isoforms participate in the GRA\'s metabolites formation, highlighting CYP2C9, which catalyzes the formation of all metabolites. The inhibition studies showed that GRA is a weak inhibitor of CYP1A2 and CYP2D6, with IC50 values greater than 200 µM and 100 µM, respectively, and a moderate and competitive inhibitor of CYP2C9, with IC50 value equal to 40.85 µM and Ki value equal to 50.60 µM. The capability of GRA to inhibit CYP3A4 was evaluated using two different substrates. GRA showed to be a moderate and competitive dose- dependent inhibitor of this isoform and also a mechanism-based time-dependent inhibitor with potential of inactivation comparable to irinotecan, a clinically significant mechanism-based inhibitor. IC50 and Ki values obtained using nifedipine as substrate were 78.09 µM and 48.71 µM, respectively, and inactivation kinetics parameters were KI= 6.40 µM, kinact= 0,037 min-1 e Clinact= 5.78 mL min-1 µmol-1. On the other hand, IC50 and Ki values using midazolam as substrate were 48.87 µM and 31.25 µM, respectively, and the values of inactivation kinetics parameters were KI= 31.53 µM, kinact= 0,049 min-1 and Clinact= 1.55 mL min-1 µmol-1. With respect to CYP2E1, it was observed that GRA increases its activity significantly from a concentration of 4 µM. Therefore, it is possible to conclude that there is no risk of DDI between GRA and drugs metabolized by CYP1A2 and CYP2D6, while for CYP2C9, although GRA is a moderate inhibitor of this isoform, the risk is low. Finally, for drugs metabolized by CYP3A4 and CYP2E1 there is risk of DDI and this should be carefully monitored in humans, mainly because CYP3A4 is an isoform responsible for catalyzing the metabolism of most drugs in use.

Understanding the processes that govern pre-systemic drug absorption and elimination is of high importance in pharmaceutical research and development, and clinical pharmacotherapy, as the oral route remains the most frequently used route of drug administration. The emergence of systems pharmacology has enabled the utilisation of in silico physiologically-based pharmacokinetic (PBPK) modelling and simulation (M&S) coupled to in vitro-in vivo extrapolation in order to perform extrapolation and exploratory M&S in special populations and scenarios were concerns regarding alterations in oral drug exposure may arise, such as following gastrointestinal (GI) surgery or metabolic drug-drug interactions (DDIs).Due to the multi-factorial physiological implications of bariatric surgery, resulting in the partial resection of the GI tract, the inability to rationalise and predict trends in oral drug bioavailability (Foral) following surgery present considerable pharmacotherapeutical challenges. PBPK M&S is a highly implemented approach for the prediction of DDIs. Reoccurring issues have emerged with regards to predictions of the magnitude of mechanism-based inhibition (MBI) where overestimations of DDIs have repeatedly been reported for drugs exhibiting high intestinal extraction. The aim of this thesis was to explore the interplay between oral drug absorption and metabolism occurring in the GI tract through the exploration of the impact of bariatric surgery on oral drug exposure and by theoretically examining the nesting and hierarchy of enterocyte and enzyme turnover and its impact on MBIs in the small intestine. This would be carried out by utilising a systems pharmacology PBPK M&S approach under a general model development framework of identification and characterisation of critical intrinsic factors and parameters, model implementation and validation. Developed post bariatric surgery PBPK models allow a framework to theoretically explore physiological mechanisms associated with altered oral drug exposure pre to post surgery, which could be assigned to the interplay between dissolution, absorption and gut-wall metabolism, where dissolution and formulation properties emerged as the perhaps most important parameters in predicting the drug disposition following surgery. Model validation identified missing critical factors that are essential for additional model refinement. Developed post bariatric surgery PBPK models have the potential of aiding clinical pharmacotherapy and decision-making following surgery. A mechanistic PBPK model was developed to describe the hierarchical dependency of enzyme and enterocyte turnover in the small intestine. Predicted enzyme recovery using the nested enzyme-within-enterocyte turnover model may potentially account for reported overpredictions of mechanism-based inhibition. Developed models in this thesis showcase the advantage of PBPK M&S in the extrapolation of oral drug exposure to special population and the potential of a PBPK approach in understanding underlying the underlying mechanism governing Foral and additionally highlight the need for generation of interdisciplinary data to support model development.

Proton pump inhibitors (PPIs) are commonly coadministered with clopidogrel, an antiplatelet agent, to patients with acute coronary syndrome (ACS). Mechanistic studies suggest that PPIs have the potential to competitively inhibit the bioactivation of clopidogrel and may attenuate its antiplatelet action in the body. The clinical implications of this drug-drug interaction have been extensively studied; however reported findings are inconsistent. More recently, several studies have questioned whether PPIs are associated with adverse cardiovascular events independent of clopidogrel. Given that PPIs and clopidogrel are widely used, it is critical to better understand the clinical impact of the concomitant treatment with both drugs. This thesis includes four studies that investigate the clinical effects of the drug-drug interaction between clopidogrel and PPIs. Chapter 2, a systematic review and meta-analysis, summarizes findings from 118 studies. Findings do not provide strong evidence for an association between adverse cardiovascular events and the use of PPIs when used alone, in combination with clopidogrel, or in combination with other antiplatelets. Chapters 3, 4, and 5 present analyses of real-world data comprised of electronic medical records. Results of these analyses demonstrate 1) that the concomitant use of clopidogrel and PPIs among inpatients was consistent with clinical guidelines suggested by the FDA (Chapter 3); 2) a lack of association between PPI use vs nonuse and four adverse cardiovascular outcomes among clopidogrel users (Chapter 4); and 3) a lack of association between PPI use vs nonuse and adverse cardiovascular outcomes among prasugrel users or ticagrelor users (Chapter 5). Collectively, our findings do not provide evidence of an elevated risk of adverse cardiovascular outcomes with the combined use of PPIs and clopidogrel. Although pharmacodynamic and pharmacokinetic studies have demonstrated an interaction between these two drugs, our findings support the opinion that the biological interaction does not translate into adverse clinical events among patients with acute coronary syndrome.

Background: Warfarin is an anticoagulant that is used for the prophylactic and therapeutic treatment for a wide range of thrombo-embolic disorders. The prescribing and monitoring of warfarin therapy is challenging due to the fact that warfarin exhibits numerous interactions with other drugs and a variety of factors that influence the dosing of warfarin. Objective: The general objective of this study was to investigate the prevalence of drugs prescribed with warfarin that may have a potential drug-drug interaction (DDI) with warfarin. Methods: This was a cross-sectional, observational or qualitative study that was conducted on medicine claims data of a pharmaceutical benefit management company for patients receiving warfarin therapy for a six year period, ranging from 1 January 2005 to 31 December 2010. Drug products that were co-prescribed with warfarin were also identified from the medicine claims database. The total number of prescriptions for all drug products during the study period were analysed and compared to the warfarin dataset. This was done by means of the SAS 9.1® computer package (SAS Institute, 2004). The total number of prescriptions and medicine items claimed from the database during the study period were respectively 49 523 818 and 118 305 941. Potential DDls between warfarin and coprescribed drugs were identified and classified according to a clinically significant rating. The clinically significance ratings of potential DDls are described in three degrees of severity, identified as major, moderate and minor (Tatro, 2011 :xiv). Results: The database consisted of 427 238 warfarin prescriptions and 427 744 warfarin medicine items, which represented 0.9% of the total number of prescriptions and 0.4% of total number of medicine items. The total number of patients who claimed warfarin prescriptions through the database represented 0.9% (n=68 575) of the total number of patients who claimed prescriptions in the total database (2005-2010). General practitioners prescribed the highest frequency of warfarin medicine items, representing 58.3% (n=249 202) of the total number prescribed. The age group that claimed the highest frequency of warfarin prescriptions (n=327 592, 76.6%) and the highest frequency of warfarin medicine items (n=327 984, 76.7%) was age group 4 (consisting of patients 59 years and older). The distribution between females and males regarding warfarin prescriptions claimed (n=205 999, 48.2%; n=221 117, 51.8%) and warfarin medicine items claimed (n=206 232, 48.2%; n=221 390, 51.8%) were almost equal. General practitioners prescribed the highest average PDD (7.01 mg ± 9.86 mg) of warfarin medicine items. Paediatric cardiologists prescribed the lowest average PDD (4.61 mg ± 1.29 mg) of warfarin medicine items. A d-value of 0.1 indicates that there is no practical difference of the average PDD between general practitioners and paediatric cardiologists. The average PDD of warfarin medicine items between females (6.60 mg ± 9.06 mg) and males (6.74 mg± 8.41 mg) was almost equal. The age group who was prescribed the highest average PDD was age group 2 (consisting of patients 20 years to 39 years old) (7.42 mg± 7.42 mg). Age group 4 (consisting of patients 59 years and older) (6.50 mg± 8.90 mg) was prescribed the lowest average PDD of warfarin medicine items. A d-value of 0.1 indicates that there is no practical difference of the average PDDs of warfarin medicine items between these two age groups. The results revealed that drugs with a significance rating (SR) of 1 (n=155 066, 43.3%), 2 (n=30128, 8.4%), 4 (n=137144, 38.3%), and 5 (n=36144, 10.1%) were co-prescribed with warfarin in the six year study period. The five drugs that was co-prescribed with warfarin most frequently was aspirin (n=48 903, 13.6%), thyroxine (n=33 954, 9.5%), amiodarone (n=25 056, 7.0%), simvastatin (n=19 070, 5.3%) and celecoxib (n=10 794, 3.0%). These five drugs have a SR of 1. Conclusions: This study showed that the top five drugs most frequently prescribed with warfarin are aspirin, thyroxine, amiodarone, simvastatin and celecoxib. These drugs can potentially interact with warfarin. The potential interactions of these drugs are rated with a significance rating of 1. This concludes that drugs that can potentially cause life threatening effects and permanent damage are commonly co-prescribed with warfarin. Clinical data concerning the INR or PT must be obtained in order to evaluate whether or not warfarin therapy is changed when a potentially interacting drug is co-prescribed. The age of the patients as well as the duration of warfarin treatment should also be obtained in order to assess whether warfarin treatment is changed with the progression of age. / MPharm (Pharmacy Practice), North-West University, Potchefstroom Campus, 2013

Background: Warfarin is an anticoagulant that is used for the prophylactic and therapeutic treatment for a wide range of thrombo-embolic disorders. The prescribing and monitoring of warfarin therapy is challenging due to the fact that warfarin exhibits numerous interactions with other drugs and a variety of factors that influence the dosing of warfarin. Objective: The general objective of this study was to investigate the prevalence of drugs prescribed with warfarin that may have a potential drug-drug interaction (DDI) with warfarin. Methods: This was a cross-sectional, observational or qualitative study that was conducted on medicine claims data of a pharmaceutical benefit management company for patients receiving warfarin therapy for a six year period, ranging from 1 January 2005 to 31 December 2010. Drug products that were co-prescribed with warfarin were also identified from the medicine claims database. The total number of prescriptions for all drug products during the study period were analysed and compared to the warfarin dataset. This was done by means of the SAS 9.1® computer package (SAS Institute, 2004). The total number of prescriptions and medicine items claimed from the database during the study period were respectively 49 523 818 and 118 305 941. Potential DDls between warfarin and coprescribed drugs were identified and classified according to a clinically significant rating. The clinically significance ratings of potential DDls are described in three degrees of severity, identified as major, moderate and minor (Tatro, 2011 :xiv). Results: The database consisted of 427 238 warfarin prescriptions and 427 744 warfarin medicine items, which represented 0.9% of the total number of prescriptions and 0.4% of total number of medicine items. The total number of patients who claimed warfarin prescriptions through the database represented 0.9% (n=68 575) of the total number of patients who claimed prescriptions in the total database (2005-2010). General practitioners prescribed the highest frequency of warfarin medicine items, representing 58.3% (n=249 202) of the total number prescribed. The age group that claimed the highest frequency of warfarin prescriptions (n=327 592, 76.6%) and the highest frequency of warfarin medicine items (n=327 984, 76.7%) was age group 4 (consisting of patients 59 years and older). The distribution between females and males regarding warfarin prescriptions claimed (n=205 999, 48.2%; n=221 117, 51.8%) and warfarin medicine items claimed (n=206 232, 48.2%; n=221 390, 51.8%) were almost equal. General practitioners prescribed the highest average PDD (7.01 mg ± 9.86 mg) of warfarin medicine items. Paediatric cardiologists prescribed the lowest average PDD (4.61 mg ± 1.29 mg) of warfarin medicine items. A d-value of 0.1 indicates that there is no practical difference of the average PDD between general practitioners and paediatric cardiologists. The average PDD of warfarin medicine items between females (6.60 mg ± 9.06 mg) and males (6.74 mg± 8.41 mg) was almost equal. The age group who was prescribed the highest average PDD was age group 2 (consisting of patients 20 years to 39 years old) (7.42 mg± 7.42 mg). Age group 4 (consisting of patients 59 years and older) (6.50 mg± 8.90 mg) was prescribed the lowest average PDD of warfarin medicine items. A d-value of 0.1 indicates that there is no practical difference of the average PDDs of warfarin medicine items between these two age groups. The results revealed that drugs with a significance rating (SR) of 1 (n=155 066, 43.3%), 2 (n=30128, 8.4%), 4 (n=137144, 38.3%), and 5 (n=36144, 10.1%) were co-prescribed with warfarin in the six year study period. The five drugs that was co-prescribed with warfarin most frequently was aspirin (n=48 903, 13.6%), thyroxine (n=33 954, 9.5%), amiodarone (n=25 056, 7.0%), simvastatin (n=19 070, 5.3%) and celecoxib (n=10 794, 3.0%). These five drugs have a SR of 1. Conclusions: This study showed that the top five drugs most frequently prescribed with warfarin are aspirin, thyroxine, amiodarone, simvastatin and celecoxib. These drugs can potentially interact with warfarin. The potential interactions of these drugs are rated with a significance rating of 1. This concludes that drugs that can potentially cause life threatening effects and permanent damage are commonly co-prescribed with warfarin. Clinical data concerning the INR or PT must be obtained in order to evaluate whether or not warfarin therapy is changed when a potentially interacting drug is co-prescribed. The age of the patients as well as the duration of warfarin treatment should also be obtained in order to assess whether warfarin treatment is changed with the progression of age. / MPharm (Pharmacy Practice), North-West University, Potchefstroom Campus, 2013

La kétamine est un antagoniste des récepteurs NMDA fréquemment utilisée pour des procédures anesthésiques et moins fréquemment pour traiter la douleur chronique. Des travaux de notre laboratoire ont montré que l’utilisation de la kétamine en combinaison avec la xylazine chez des rats âgés d’environ deux ans pouvait prolonger la durée d’anesthésie. Considérant que les patients gériatriques présentent des caractéristiques physiologiques demandant des adaptations au niveau de la pharmacothérapie, que cette classe de patient requiert fréquemment des anesthésies afin de réaliser des interventions chirurgicales et qu’elle représente la sous-population qui consomme le plus de médicaments pour traiter des problèmes de douleur aiguë et chronique, il convient alors d’investiguer l’impact que l’âge peut avoir sur le métabolisme des médicaments. Le but de ce projet est donc d’évaluer l’impact de l’âge sur le métabolisme de la kétamine à l’aide de fractions cellulaires (in vitro) et d’une technique de chromatographie en phase liquide couplée à la spectrométrie de masse. En premier lieu, nous avons dû confirmer la voie métabolique principale de formation de la norkétamine en utilisant le substrat et l’inhibiteur de référence de la voie métabolique du CYP3A qui sont respectivement le midazolam et le kétoconazole. De plus, nous avons caractérisé l’interaction métabolique de la kétamine fréquemment combinée au midazolam. Ensuite, nous avons comparé les profils de dégradation de la kétamine par des fractions S9 hépatiques de rats âgés de 3, 6, 12 et 18 mois. Les valeurs (±ÉT) de Vmax obtenues étaient respectivement de 2.39 (±0.23), 2.61 (±0.18), 2.07 (±0.07) pour les plus jeunes comparé à 0.68 (±0.02) pour le groupe de 18 mois (p < 0.001). Ces résultats suggèrent donc que le CYP3A est rapidement saturé par la présence de kétamine. De plus, la valeur de Km a diminué de 6 à 7 fois pour le groupe de 18 mois comparé aux autres groupes (p < 0.05). Ainsi, on peut émettre l’hypothèse que l’enzyme subit une modification de sa conformation tridimentionnelle, ce qui résulte en une diminution de formation de norkétamine. Finalement, nous avons tenté d’évaluer la contribution du métabolisme cérébral au métabolisme de la kétamine. Toutefois, nous n’avons pas trouvé de différence significative dans les concentrations de kétamine ou de midazolam entre le début de la réaction et 60 minutes plus tard. Le même constat a été observé pour d’autres substrats du CYP3A et CYP2D (dextromethorphan et codéine). D’autres expériences sont nécessaires afin d’arriver à des conclusions définitives. Les différences observées entre les différents groupes d’âge suggèrent que la voie métabolique hépatique du CYP3A est détériorée de façon importante. Les protocoles anesthésiques pour patients âgés devraient donc tenir compte de cette importante diminution d’activité métabolique étant donné que l’exposition aux concentrations systémiques risque d’être significativement augmentée. / Ketamine is a NMDAR antagonist widely used for anesthetic procedures and less often to treat chronic pain. Work in our laboratory showed that the use of ketamine combined to xylazine is a poor anesthetic choice for aged rats of approximately 2 years. Considering that geriatric patients present physiological characteristics demanding adaptations of the pharmacotherapy, that this class of patients frequently needs anesthesia to perform surgeries and that they represent the subpopulation that uses many drugs to treat acute and chronic pain, it is relevant to investigate the impact of aging on drug metabolism. The goal of this project was to assess the impact of aging on ketamine metabolism using cellular fractions (in vitro) and liquid chromatography tandem mass spectrometry. First, we needed to confirm the main metabolic pathway of norketamine formation using well-established reference substrate and inhibitor of CYP3A metabolic pathway which are respectively midazolam and ketoconazole. Furthermore, we also characterized the metabolic interaction of ketamine frequently combined with midazolam. Second, we compared the degradation profiles of ketamine in liver S9 fractions of rats aged of 3, 6, 12 and 18 months. Vmax values (±SD) were respectively of 2.39 (±0.23), 2.61 (±0.18), 2.07 (±0.07) for the younger groups compared to 0.68 (±0.02) for the 18 month older group (p < 0.001). Those results suggest that CYP3A is quickly saturated by the presence of ketamine. Moreover, Km value showed a 6 to 7 fold decrease for the oldest group compared to the younger one (p < 0.05). Therefore, we hypothesized that the enzyme undergoes conformational changes, which results in a decrease of norketamine formation. Finally, we intend to assess the contribution of brain metabolism to the overall ketamine metabolism. Unfortunately, we did not find any significative difference in ketamine or midazolam concentrations between the beginning of the reaction and 60 minutes later. The same ascertainment has been observed with other susbstrates of CYP3A and CYP2D (dextromethorphan and codeine). Other experiences are needed to definetely conlude that brain metabolism is negligible. The differences observed between age groups suggest that the hepatic metabolic pathway of CYP3A is severely impaired. Anesthetic protocols for aging patients should take into consideration the diminished metabolic activity since it may lead to a significant increase of exposition to systemic drug concentrations.