<p>The main objective of this thesis was to investigate the in vivo relevance of membrane transporters and cytochrome P450 (CYP) 3A4-mediated metabolism in the intestine and liver for the bioavailability of drugs in humans after oral administration.</p><p>In the first part of the thesis, the main transport mechanisms involved in the intestinal absorption and bioavailability were investigated for fexofenadine, a minimally metabolized drug, which is a substrate for P-glycoprotein (P-gp) and members of organic anion transporting polypeptide (OATP) family. Jejunal perfusion studies revealed that co-perfusion with verapamil increased the bioavailability of fexofenadine by decreasing the first-pass liver extraction as the low intestinal permeability was unchanged by the transport inhibitors studied. The mechanism behind the interaction probably involves inhibition of OATP-mediated sinusoidal uptake and/or P-gp-mediated canalicular secretion of fexofenadine. Results from the Caco-2 model supported that the intestinal absorption of fexofenadine is mainly determined by the low passive permeability of the drug, even though fexofenadine clearly is a P-gp substrate. </p><p>In the second part of the thesis, the effect of repeated oral administration of the P-gp and CYP3A4 inducer St. John’s wort on the in vivo intestinal permeability and presystemic metabolism of the dual P-gp and CYP3A4 substrate verapamil was investigated in a jejunal perfusion study. St. John’s wort decreased the bioavailability of the enantiomers of verapamil by inducing the CYP3A4-mediated presystemic metabolism, probably mainly in the gut. It was also concluded that induction of efflux transporters, such as P-gp, does not affect the intestinal transport or the gut wall extraction of high permeability substrates like verapamil. Data from Caco-2 cells with induced CYP3A4-activity supported these findings. The plasma levels of the enantiomers of norverapamil also decreased despite an increased formation, which was attributed to induction of CYP3A4 and/or other metabolic routes. </p>

<p>The aims of this thesis were to study the in vivo relevance of membrane transporters for intestinal absorption and the hepatic disposition of drugs in humans and preclinical models. Fexofenadine is a substrate for ABCB1 (P-glycoprotein) and members of the organic anion transporting polypeptide (OATP/SLCO) family. It is marginally metabolised in humans. </p><p>The influence of known inhibitors of ABCB1 and OATPs on the membrane transport and pharmacokinetics of fexofenadine was investigated in Caco-2 and porcine models and in humans. The permeability of fexofenadine remained low, even when significantly altered by the addition of an inhibitor. Using the Loc-I-Gut^® technique in vivo in humans, it was possible to see that the jejunal effective permeability of fexofenadine was unchanged when given with verapamil. However, the systemic exposure and apparent absorption rate of fexofenadine increased. This suggests that the first-pass liver extraction of fexofenadine was reduced by verapamil, probably through the inhibition of sinusoidal OATP-mediated and/or canalicular ABCB1-mediated secretion. The unchanged permeability can be explained by simultaneous inhibition of jejunal apical OATP-uptake and ABCB1-efflux, which would leave fexofenadine to be transported by passive trancellular diffusion. A Loc-I-Gut^® perfusion in the porcine model enabling blood sampling in the portal and hepatic veins and bile collection revealed increased jejunal permeability, but no subsequent verapamil-induced elevation in the systemic exposure of fexofenadine. This indicates a species-related difference in the localisation of and/or the substrate specificity of fexofenadine for the transporters involved. The absence of an effect on the first-pass liver extraction in the porcine model might be caused by the observed lower liver exposure of verapamil.</p><p>Finally, a novel intubation technique enabling dosing of fexofenadine in the jejunum, ileum and the colon showed that fexofenadine was absorbed less along the length the intestine in agreement with the properties of a low permeability drug.</p>

The main objective of this thesis was to investigate the in vivo relevance of membrane transporters and cytochrome P450 (CYP) 3A4-mediated metabolism in the intestine and liver for the bioavailability of drugs in humans after oral administration. In the first part of the thesis, the main transport mechanisms involved in the intestinal absorption and bioavailability were investigated for fexofenadine, a minimally metabolized drug, which is a substrate for P-glycoprotein (P-gp) and members of organic anion transporting polypeptide (OATP) family. Jejunal perfusion studies revealed that co-perfusion with verapamil increased the bioavailability of fexofenadine by decreasing the first-pass liver extraction as the low intestinal permeability was unchanged by the transport inhibitors studied. The mechanism behind the interaction probably involves inhibition of OATP-mediated sinusoidal uptake and/or P-gp-mediated canalicular secretion of fexofenadine. Results from the Caco-2 model supported that the intestinal absorption of fexofenadine is mainly determined by the low passive permeability of the drug, even though fexofenadine clearly is a P-gp substrate. In the second part of the thesis, the effect of repeated oral administration of the P-gp and CYP3A4 inducer St. John’s wort on the in vivo intestinal permeability and presystemic metabolism of the dual P-gp and CYP3A4 substrate verapamil was investigated in a jejunal perfusion study. St. John’s wort decreased the bioavailability of the enantiomers of verapamil by inducing the CYP3A4-mediated presystemic metabolism, probably mainly in the gut. It was also concluded that induction of efflux transporters, such as P-gp, does not affect the intestinal transport or the gut wall extraction of high permeability substrates like verapamil. Data from Caco-2 cells with induced CYP3A4-activity supported these findings. The plasma levels of the enantiomers of norverapamil also decreased despite an increased formation, which was attributed to induction of CYP3A4 and/or other metabolic routes.

The aims of this thesis were to study the in vivo relevance of membrane transporters for intestinal absorption and the hepatic disposition of drugs in humans and preclinical models. Fexofenadine is a substrate for ABCB1 (P-glycoprotein) and members of the organic anion transporting polypeptide (OATP/SLCO) family. It is marginally metabolised in humans. The influence of known inhibitors of ABCB1 and OATPs on the membrane transport and pharmacokinetics of fexofenadine was investigated in Caco-2 and porcine models and in humans. The permeability of fexofenadine remained low, even when significantly altered by the addition of an inhibitor. Using the Loc-I-Gut® technique in vivo in humans, it was possible to see that the jejunal effective permeability of fexofenadine was unchanged when given with verapamil. However, the systemic exposure and apparent absorption rate of fexofenadine increased. This suggests that the first-pass liver extraction of fexofenadine was reduced by verapamil, probably through the inhibition of sinusoidal OATP-mediated and/or canalicular ABCB1-mediated secretion. The unchanged permeability can be explained by simultaneous inhibition of jejunal apical OATP-uptake and ABCB1-efflux, which would leave fexofenadine to be transported by passive trancellular diffusion. A Loc-I-Gut® perfusion in the porcine model enabling blood sampling in the portal and hepatic veins and bile collection revealed increased jejunal permeability, but no subsequent verapamil-induced elevation in the systemic exposure of fexofenadine. This indicates a species-related difference in the localisation of and/or the substrate specificity of fexofenadine for the transporters involved. The absence of an effect on the first-pass liver extraction in the porcine model might be caused by the observed lower liver exposure of verapamil. Finally, a novel intubation technique enabling dosing of fexofenadine in the jejunum, ileum and the colon showed that fexofenadine was absorbed less along the length the intestine in agreement with the properties of a low permeability drug.

<p><strong>Bakgrund: </strong>I dagens vårdarbete är behandling av venösa bensår en vanlig omvårdnadsåtgärd. Smärta, immobilitet samt social isolering relaterat till venösa bensår, är några av de faktorer som påverkar patientens livskvalitet. <strong>Syfte: </strong>Syftet med litteraturstudien var att beskriva hur sjuksköterskan på bästa sätt kan främja läkningen av venösa bensår. <strong>Metod: </strong>Beskrivande litteraturstudie sammanställd utifrån 14 kvantitativa samt sju kvalitativa studier publicerade mellan åren 2001 till 2009. Databaserna Cinahl och PubMed användes i sökningen av vetenskapliga artiklar. Sökorden som användes var Leg Ulcer, Nursing, Activity, Pain, Venous leg ulcer, Treatment, Management, Exercise, Bandages samt Psychological. <strong>Resultat: </strong>De omvårdnadsåtgärder som visade sig ha stor betydelse för sårläkning var kompression/sårvård, fysisk aktivitet, psykologiskt stöd samt eftervård. Sjuksköterskans kunskap och förmåga att kunna se patienten som en helhet var av stor vikt för god omvårdnad och förbättrad sårläkning. <strong>Slutsats: S</strong>juksköterskor behöver förbättra sina kunskaper angående sårvårdsbehandling. Sjuksköterskan bör ha en holistisk syn på patienten.</p><p> </p>

<p>The transport of drugs across the blood-brain barrier (BBB) has been investigated in different species using morphine and morphine-6-glucuronide (M6G) as model compounds. The influence of probenecid on the BBB transport of morphine and M6G was investigated, and the consequences of meningitis and severe brain injury on the concentrations of morphine in the brain were examined. All data were obtained by microdialysis, and data analysis using mathematical models was emphasised.</p><p>Morphine is exposed to active efflux at the BBB in rats, pigs and humans. In addition, the half-life of morphine is longer in the brain than in blood in these species. These interspecies similarities show the predictive potential of the two animal models for the BBB transport of morphine in humans. In the pig the exposure of the brain to morphine was higher in the presence of meningitis than when healthy. This was interpreted as a decrease in the active efflux and an increase in the passive diffusion over the injured BBB. In contrast, there was no significant difference in the concentrations of morphine in the “better” (uninjured) or the “worse” (injured) brain tissue in brain trauma patients. The extent of the transport across the BBB is similar for morphine and M6G. However, co-administration of probenecid only increased the brain concentrations of morphine, demonstrating that morphine and M6G are substrates for different efflux transporters at the BBB. An integrated model for the analysis of data obtained by microdialysis was developed. This model makes fewer assumptions about the recovery, the protein binding and the time of the dialysate observation than a previous model and traditional non-compartmental analysis and should, therefore, yield more reliable parameter estimates.</p><p>Knowledge of the consequences of efflux transporters and disease on the brain concentrations of a drug can be useful for individualising the dosing regimen in patients. </p>

<p>To increase the proportion of patients with successful drug treatment, dose individualisation on the basis of one or several patient characteristics, <i>a priori</i> individualisation, and/or on the basis of feedback observations from the patient following an initial dose, <i>a posteriori</i> individualisation, is an option. Efficient tools in optimising individualised dosing strategies are population models describing pharmacokinetics (PK) and the relation between pharmacokinetics and pharmacodynamics (PK/PD).</p><p>Methods for estimating optimal dosing strategies, with a discrete number of doses, for dose individualisation <i>a priori</i> and <i>a posteriori</i> were developed and explored using simulated data. The methods required definitions of (<i>i</i>) the therapeutic target, <i>i.e. </i>the value of the target variable and a risk function quantifying the seriousness of deviation from the target, (<i>ii</i>) a population PK/PD model relating dose input to the target variable in the patients to be treated, and (<i>iii</i>) distributions of relevant patient factors. Optimal dosing strategies, in terms of dose sizes and individualisation conditions, were estimated by minimising the overall risk. Factors influencing the optimal dosing strategies were identified. Consideration of those will have implications for study design, data collection, population model development and target definition.</p><p>A dosing strategy for <i>a priori</i> individualisation was estimated for NXY-059, a drug under development. Applying the estimated dosing strategy in a clinical study resulted in reasonable agreement between observed and expected outcome, supporting the developed methodology.</p><p>Estimation of a dosing strategy for <i>a posteriori</i> individualisation for oxybutynin, a drug marketed for the treatment of overactive bladder, illustrated the implementation of the method when defining the therapeutic target in terms of utility and responder probability, that is, as a combination of the desired and adverse effects.</p><p>The proposed approach provides an estimate of the maximal benefit expected from individualisation and, if individualisation is considered clinically superior, the optimal conditions for individualisation. The main application for the methods is in drug development where the methods can be generally employed in the establishment of dosing strategies for individualisation with relevant extensions regarding population model complexity and individualisation conditions.</p>

<p>During the last years, drugs that prolong the repolarisation phase of the myocardial action potential, due to inhibition of the rapid component of the delayed-rectifying potassium channel (I_Kr) have been in focus. In addition to arrhythmogenic potential, selective Ikr-blockers have also been shown to be embryotoxic and teratogenic in animal studies. The aim of this thesis was to investigate a theory that these developmental toxic results from pharmacologically induced episodes of embryonic cardiac arrhythmias leading to hypoxia related damage in the embryo. Almokalant (ALM) was used as a model compound for selective Ikr-blockers. ALM induced embryonic cardiac arrhythmia, and in similarity with results obtained by maternal hypoxia, ALM induced embryonic death and growth retardation in both rats, and mice. </p><p>The theory of a hypoxia-related mechanism was strengthened by the results that ALM induce phase specific external and visceral defects (e.g. cleft lip/palate, distal digital, cardiovascular, and urogenital defects), and that the skeletal defects (not shown before) showed a clear trend; the later the treatment the more caudal was the site of the defect, which is in accordance with results from maternal hypoxia induced by e.g. lowering of the O₂ content in the air. The spin trapping agent PBN decreased almokalant induced malformations, suggesting that the defects mainly are caused by reoxygenation damage after episodes of severe embryonic dysrhythmia, rather than "pure hypoxia".</p><p>Sotalol was tested in a third species, the rabbit who expresses functional I_Kr channels both in the embryo and in the adult, where it induced developmental toxicity, and indicating that the embryo is more sensitive than the adult towards arrhythmia caused by I_Kr-blockers. </p>

<p>Transposons of the Mu superfamily are widespread and have been shown to play an important role in the dissemination of antibiotic resistance among microorganisms. One of these elements, Tn<i>5090</i>/Tn<i>402</i> is the basal vehicle of the type 1 integrons in which mobile resistance gene cassettes are inserted to form clusters and operons. The transposon was shown to preferentially target recombination sites of the serine family of recombinases that occur in many plasmids and transposons. Mutation analysis revealed that DNA-binding of the targeting factor, a serine recombinase, is essential for efficient transposition, while the recombination activity is not required. Truncated elements were frequently observed and in one instance borne on a composite transposon flanked by IS<i>6100</i>. This new transposon, Tn<i>5089</i>, has allowed the translocation of the integron to small mobilizable IncQ-plasmids that lack the targeting factor and thus are incompetent for insertion of Tn<i>5090</i>/Tn<i>402</i>. Another small replicon, by contrast targeting-positive, was completely sequenced.</p><p>The transposon Tn<i>5090</i>/Tn<i>402 </i>carries arrayed transposase-binding sites at the ends, which are supposed to arrange the transposase TniA in the appropriate geometry in a recombinationally active complex with DNA. Footprinting showed that transposase, TniA, binds to four 19 bp repeats on one end and to two 19 bp repeats on the other end.</p><p>Site-specific resolution of Tn<i>5090</i>/Tn<i>402</i> co-integrates<i> </i>was analysed in an <i>in vitro </i>system. The<i> res</i> site was found to be composed of three unusually organized subsites and expression of TniC was shown to be autoregulated by TniC acting as repressor due to an overlap of the <i>res</i> site with the promoter. </p><p>The data presented show several aspects of cooperation between transposition and site-specific recombination. This cooperation has enriched genes and combinations of genes that mediate resistance to antibiotic drugs and promotes lateral transfer of these genes. The organization of sites and subsites in the DNA is a subtle genetic code for the formation of the molecule complexes controlling these genetic events. </p>

<p>Artemisinin is the parent compound of a novel family of antimalarials. Repetitive administrations of artemisinin to both healthy volunteers and malaria patients have been shown to result in decreased plasma concentrations of the compound, most probably due to an autoinduction of different CYP450 enzymes. The aim of this thesis was to investigate the clinical pharmacokinetics and efficacy of different dosage regimens of the drug, and study the kinetics of the enzyme induction. Moreover, the putative interaction of the compound with blood components was investigated in vitro. </p><p>Artemisinin was found to distribute into red blood cells, competing with oxygen for binding to hemoglobin. The compound was stable in plasma and, in contrast to previous reports, did not bind to red blood cell membranes. </p><p>To circumvent the logistical and ethical problems associated with plasma sampling, suitability of saliva as substitute was investigated. Moreover, due to the large number of collected samples, an HPLC method, enabling a direct injection of saliva and plasma samples, was developed. </p><p>Saliva artemisinin concentrations were found to correlate with its unbound plasma levels, making saliva a suitable body fluid for pharmacokinetic studies of the compound. Based on saliva samples, artemisinin was shown to exhibit a dose-dependent kinetics and efficacy in malaria patients, with a possible sex-effect on the metabolism of the compound during the first treatment day. Moreover, the time-dependent kinetics of the compound was observed in both malaria patients and healthy subjects. A physiological approach was utilized to model the autoinduction in the latter group. A model with a feedback mechanism of enzymes was able to describe the data, with estimations of the half-lives of induction (3.15 hrs) and elimination of enzymes (32.9 hrs), as well as pharmacokinetic parameters of artemisinin. </p><p>In conclusion, artemisinin was found to exhibit a fast induction of enzymes, with time- and dose-dependent drug kinetics and dose-dependent antimalarial efficacy. </p>