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The brain of Alzheimer???s disease (AD) is characterized by accumulations of ??-amyloid peptide aggregates which promote neurodegentartive dysfunction. Comprehensive understanding of the interaction between ??-amyloid aggregates and acetylcholine (ACh) neurocycle is required to uncover the physiological processes related to AD and might result in improving therapeutic approaches for AD. Pharmacokinetics (PK) and pharmacodynamics (PD) techniques were applied to allow predicting the extent of the interaction of certain doses of AD drugs and ??-amyloid inhibitors and levels of ACh as well. Although many researchers focused on the ??-amyloid interactions, the mechanisms by which ??-amyloid affects cholinergic neurons and reduction of ACh are still unclear. The prediction of ACh and drug concentrations in the tissues and body needs an understanding of the physiology and mechanisms of ??-amyloid aggregates processes and their compilation into a mechanistic model In this work, two hypotheses are proposed to investigate the dynamic behavior of the interaction between ??-amyloid peptide aggregates and cholinergic neurocycle and the possible therapeutic approaches through proposing pharmacokinetic/pharmacodynamics (PK/PD) models to represent the impact of ??-amyloid aggregates in AD. The effect of ??-amyloid peptide aggregates is formulated through incorporating ??- amyloid aggregates into non-linear model for the neurocycle of ACh where the presynaptic neuron is considered as compartment 1 and both synaptic cleft and postsynaptic neurons are considered as compartment 2. In the first hypothesis which is choline leakage hypothesis, ??-amyloid peptide aggregates are considered to be located in the membrane of the presynaptic neuron and create pathways inside the membrane to allow for the intracellular choline to leak outside the cholinergic system. It is observed that ??-amyloid aggregates via the choline leakage hypothesis could cause significant reductions of ACh and choline levels in both compartments. Furthermore, the process rates of ACh synthesis and hydrolysis have been affected negatively by a wide range of ??-amyloid aggregate concentrations. It is found that as the input rate of ??-amyloid aggregates to compartment 1 increases, the loss of choline from compartment 1 increases leading to an increase in the intracellular concentration of ??-amyloid. In the second hypothesis, ??-amyloid peptide aggregates are proposed to interact with the enzyme ChAT which is responsible for the synthesis of ACh in compartment 1; three different kinetic mechanisms are suggested to account for the interaction between ??-amyloid aggregates and ChAT activity. In the first and second kinetic mechanisms, ??-amyloid aggregate is supposed to attack different species in the enzyme. It is found that there is a significant decrease in the rate of ACh synthesis in compartment 1 and ACh concentrations in both compartments. However, it is observed that there is no effect on choline levels in both compartments, the rate of ACh hydrolysis in compartment 2, pH, and ACh levels in compartment 2. In the third kinetic mechanism, all species in ChAT are attacked by ??-amyloid aggregates; it is observed that at very high input rates of ??-amyloid aggregates, the oscillatory behavior dominates all components of the neurocycle of ACh. The disturbance observed in ACh levels in both compartments explains the harmful effect of the full attack of ??-amyloid aggregates to all species of ChAT. It is found that to contribute significantly in ACh neurocycle, choline leakage hypothesis needs concentration of ??-amyloid aggregates lower than that needed in ChAT activity hypothesis which is in agreement with experimental observations. The significant decrease in ACh levels observed in both choline leakage and loss of ChAT activity hypotheses leads to cognitive loss and memory impairment which were observed in individuals with AD. A one-compartment drug PK/PD model is proposed to investigate a therapeutic approach for inhibiting ??-amyloid aggregation via choline leakage hypothesis where the maximum feed rate of ??-amyloid (KL2 = 1) is considered. The drug is assumed to interact with the tissues of the presynaptic neurons where ??-amyloid aggregates are located. The PK/PD model is built based on the effect of ??-amyloid aggregates via choline leakage hypothesis where the maximum feed rate of ??-amyloid aggregates is considered. The dynamic behavior of all concentrations of ??-amyloid aggregates, choline, ACh, acetate, and pH in both compartments in addition to the rate of ACh synthesis in compartment 1 and ACh hydrolysis are investigated by monitoring the impacts of the drug on ??-amyloid aggregates and cholinergic neurocycle over a wide range of the input drug dosage. The PK/PD model is able to predict the reduction in levels of ??-amyloid aggregates and the increase in choline and ACh, in both compartments as well as both rates of ACh synthesis and hydrolysis catalyzed. The parameters of the PK/PD model such as maximum concentration (Cmax), maximum time (Tmax), area under the curve (AUC), and maximum effect (Emax) were investigated. It was found that it takes a longer time (Tmax) (3-5 h) to reach Emax as the drug dose increases. Furthermore, AUC was found to increase with increasing drug dosage. The results of the current work show that drugs / therapeutic agents inhibiting ??- amyloid aggregation in the brain represent a likely successful therapeutic approach to give systematic highlights to develop future trials, new diagnostic techniques, and medications for AD. This study is helpful in designing PK and PD and developing experimental animal models to support AD drug development and therapy in the future.

Doctor of Philosophy / Department of Diagnostic Medicine/Pathobiology / Michael D. Apley / The discovery of modern antimicrobials in the early 20th century revolutionized treatment of infectious diseases. Less than 100 years later, antimicrobial resistance has become a global threat to public health. With the rise of antimicrobial resistance, the question that remains to be answered is: Can dosing regimens provide maximal clinical efficacy, yet minimize the development of antimicrobial resistance? A pharmacokinetic / pharmacodynamic approach was utilized to investigate oxytetracycline regimens that would impart efficacy while minimizing the potential for resistance development due to plasmid transfer. An in vitro pharmacodynamic model was used to quantify the response of a Pasteurella multocida isolate to two oxytetracycline dosing regimens. The PK/PD index most closely related to efficacy was the Cmax:MIC. The in vitro pharmacodynamic model was then used to investigate the effects of antimicrobial exposure on plasmid transfer. A mixed population of oxytetracycline-susceptible and resistant bacteria was exposed to two dosing regimens and plasmid transfer was quantified. When oxytetracycline concentrations exceeded the MIC of the recipient, development of resistance was suppressed. The same donor and recipient bacteria were used in an in situ swine model to validate the in vitro findings. Following surgical implantation of porous membrane straws containing the mixed bacterial population, animal subjects in the treatment groups received one of two oxytetracycline treatments. Oxytetracycline concentrations in the plasma and interstitial fluid were quantified. Plasmid transfer within the implant membranes was quantified and correlated to pharmacokinetic measures in the animal. Plasmid transfer rates in the implant membranes did not correlate to the investigated pharmacokinetic parameters. The study methodologies in this dissertation should serve as a foundation for future studies in antimicrobial pharmacokinetic/pharmacodynamic research. The results presented here show that the bacterial response to oxytetracycline can be optimized in a concentration dependent manner and that antimicrobial resistance development through plasmid transfer can be suppressed in vitro when oxytetracycline concentrations exceed the MIC of the recipient bacteria. These results suggest that a proper balance between clinical efficacy and minimizing antimicrobial resistance can be achieved for oxytetracycline through appropriate dosing regimens and drug formulations.

Problems of emerging antibiotic resistance are becoming a serious threat worldwide, and at the same time, the interest to develop new antimicrobials has declined. There is consequently a need for efficient methods to develop new treatments that minimize the risk of resistance development and that are effective on infections caused by resistant strains. Based on in silico mathematical models, describing the time course of exposure (Pharmacokinetics, PK) and effect (Pharmacodynamics, PD) of a drug, information can be collected and the outcome of various exposures may be predicted. A general model structure, that characterizes the most important features of the system, has advantages as it can be used for different situations. The aim of this thesis was to develop Pharmacokinetic-Pharmacodynamic (PKPD) models describing the bacterial growth and killing after mono- and combination exposures to antibiotics and to explore the predictive ability of PKPD-models across preclinical experimental systems. Models were evaluated on data from other experimental settings, including prediction into animals. A PKPD model characterizing the growth and killing for a range of E. coli bacteria strains, with different MICs, as well as emergence of resistance, was developed.  The PKPD model was able to predict results from different experimental conditions including high start inoculum experiments, a range of laboratory and clinical strains as well as experiments where wild-type and mutant bacteria are competing at different drug concentrations. A PKPD model, developed based on in vitro data, was also illustrated to have the capability to replicate the data from an in vivo study. This thesis illustrates the potential of PKPD models to characterize in vitro data and their usage for predictions of different types of experiments. The thesis supports the use of PKPD models to facilitate development of new drugs and to improve the use of existing antibiotics.

L’un des défis actuels de la lutte contre la tuberculose est de développer un traitement plus court et plus efficace. La modélisation mathématique constitue une approche qui peut nous aider à comprendre les problèmes actuels et favoriser les innovations thérapeutiques. L’objectif de ce travail est de construire un modèle thérapeutique mathématique de la tuberculose pulmonaire basé sur des éléments pharmacocinétiques, pharmacodynamiques et physiopathologiques. La mise en application du modèle pharmacodynamique a été précédée d’une étude théorique sur l’équation de Hill. Cette synthèse a permis de dégager les bases rationnelles de son utilisation en modélisation pharmacologique. En utilisant une approche de population, un modèle pharmacocinétique de diffusion pulmonaire a permis de décrire les concentrations en rifampicine dans le plasma et le poumon chez 34 sujets. Le modèle a ensuite été utilisé pour analyser la valeur d’indices pharmacodynamiques corrélés à l’effet chez 10 000 sujets fictifs, par simulation de Monte Carlo. Les résultats indiquent que la dose de standard de rifampicine conduit à des concentrations globalement peu efficaces et pouvant favoriser la résistance bactérienne. Un premier modèle mathématique du traitement de la tuberculose par la rifampicine, incluant un modèle physiopathologique formel, a enfin été construit. Il permet de simuler la dynamique bactérienne du premier jour de l’infection au dernier jour de traitement. L’ensemble des résultats conduit à une remise en question de la dose standard de rifampicine et suggère une nouvelle hypothèse sur les causes de la persistance de Mycobacterium tuberculosis au cours du traitement antituberculeux / There is a critical need for a shorter tuberculosis treatment to improve tuberculosis control. Mathematical models may be helpful to understand current problems associated with tuberculosis therapy and to suggest innovation resources. The objective of this study is to set up a full mathematical model of tuberculosis treatment by rifampin, based on pharmacokinetic, pharmacodynamic and physiological submodels. Prior to its application in the pharmacodynamic modeling framework, the Hill equation has been the focus of a theoretical study. The various properties of this equation have been reviewed and the rationale of its use in pharmacological modelling has been clarified. Rifampin pharmacokinetics in plasma and lungs was modelled in a population of 34 volunteers by use of a nonparametric population approach. Then, a 10,000 subject Monte Carlo simulation was performed to explore Mycobacterium tuberculosis killing effect and prevention of resistance by rifampin. The results suggest that rifampin pulmonary concentrations obtained with the standard dose are too low to be highly effective and prevent drug resistance in most subjects. Finally, a full mathematical model of tuberculosis treatment, including a physiological model, has been implemented. The model is able to simulate the time-course of bacterial counts from the first day of infection to the last day of treatment. Overall results of this modelling effort indicate that current dosage regimens of rifampin may be optimized. In addition, this work suggests a new hypothesis regarding the bacterial persistence during tuberculosis treatment.

Introduction. 5-HMF and novel INN-compounds are left-shifting AEH, shown to have anti-sickling action by forming transiently covalent Schiff-base adducts with hemoglobin (Hb), thereby increasing the Hb O2-affinity. They are hypothesized to be substrates for aldehyde dehydrogenase (ALDH) in the liver and red blood cells (RBC). Methods. Biopharmaceutical assessments were made for AEH, using calculated physicochemical properties. Their in-vitro hepatic metabolism (mediated by ALDH) was characterized using hepatic cytosol, and in-vitro-in-vivo extrapolations (IVIVE) were made. Inter-species differences in hepatic cytosolic ALDH activity were investigated using acetaldehyde as a model substrate in different mammalian species. Time- and concentration-dependent in-vitro disposition of 5-HMF in human whole blood was fully characterized and quantitatively modeled. In-vitro time- and concentration-dependent pharmacodynamic (PD) profiling of AEH (0.5 – 5 mM) was carried out in normal whole blood. 5-HMF binding to (normal) HbA and (sickle) HbS was studied in systematic time- and concentration-dependency studies using isolated Hb solutions. Quantitative PK/PD models were developed to fit the experimental data by nonlinear regression (Scientist®). Results. 5-HMF and the two INN-compounds were classified as BCS-I and BCS-II, respectively. All AEH were substrates for hepatic ALDH, with predicted low/intermediate hepatic extraction. Intrinsic ALDH activity varied significantly between mammalian species. In whole blood, 5-HMF plasma concentrations declined rapidly (t1/2 of 0.8 – 4 hrs), with nonlinear kinetics, due to saturable Hb-binding. AEH showed a time-dependent, biphasic PD effect in whole blood, suggesting transiently covalent Hb binding, with slow recovery to the baseline, corresponding to dissociation from Hb and subsequent metabolism by RBC-ALDH. Binding studies with HbA and HbS demonstrated slight differences in binding affinity, but sustained adduct formation - with slow dissociation t1/2. A novel semi-mechanistic target-site drug disposition (TSDD)/PD model was developed, integrating the information, for simultaneous modeling of 5-HMF concentrations in plasma, and its effect in whole blood. Conclusions. This translational research investigated in detail the in-vitro PK/PD of AEH, and systematically compared findings with older generation compounds. A (generic) novel TSDD/PD model was developed for disposition of AEH, identifying k-1 (dissociation constant of AEH from Hb) and kmet (RBC-ALDH metabolism rate constant) as key properties for the time course of PD effect.

A buspirona, ansiolítico da segunda geração usada no tratamento de ansiedade, foi analisada fisico-quimica e quimicamente, com o objetivo de fornecer subsídios para a respectiva monografia. Fizeram-se os seguintes testes de caracterização deste fármaco: faixa de fusão, análise elementar, pH, umidade e solubilidade. Realizaram-se igualmente as provas de identificação constante de: análise cromatográfica, ensaio de cloreto, espectro no infravermelho, espectro no ultravioleta, espectro de ressonância magnética protônica, espectro de massa, espectro de carbono-13, termogravimetria, calorimetria exploratória diferencial e cromatografia líquida de alta resolução. Efetuaram-se, também, determinações quantitativas pelos seguintes métodos: micro-Kjeldahl, gravimétrico, potenciométrico com nitrato de prata, doseamento com ácido perclórico em meio acetico por volumetria e potenciometria e espectrofotométrico no ultravioleta. Concluiu-se que: 1) o teste de caracterização mais indicado e o ponto de fusão; 2) das provas de identificação, as mais simples e rápidas são as seguintes: identificação de cloreto, infravermelho e ultravioleta; 3) dos métodos de determinação quantitativa, mais prático e o doseamento com ácido perclórico em meio acético, usando como indicador o vermelho de quinaldina. / Buspirone, a new non-benzodiazepine anxiolytic agent which is effective in the treatment of anxiety, was analyzed by physicochemical and chemical methods, having in mind a monograph of this drug. The fo1lowing characterization tests of this drug were performed: melting range, elementary analysis, pH, loss in drying, solubility. The following identification tests were also carried out: chromatographic analysis, identification of chloride, infrared absorption, ultraviolet absorption, protonic magnetic resonance spectra, mass spectrum, carbon 13C NMR spectroscopy, thermogravimetry and high performance liquid chromatography. Quantitative determinations were obtained through the following methods: micro-Kjeldahl, gravimetry, potenciometry, non-aqueous titration with perchloric acid and ultraviolet spectrometry. The following conclusions were reached: 1) the best characterization test is the melting range; 2) the simplest and quickest tests of identification are: identification of chloride, infrared and ultraviolet spectrophotometry; 3) the most practical method is the non-aqueous assay with perchloric acid.

La résistance bactérienne est une menace mondiale clairement établie. Les chercheurs se rassemblent afin de trouver de nouvelles solutions et des alternatives aux antibiotiques. Une des causes principales de cette menace est l’utilisation massive des antibiotiques. Il faut cesser d’avoir un recours déraisonné à leur usage. Au sein d’une population vieillissante, ce sont les personnes les plus vulnérables qui en payent le prix. Nous avons utilisé les outils de modélisation pour mieux comprendre les causes engendrant ce fléau. L'objectif de ce travail est de construire des modèles pharmacocinétique de population chez la personne âgée pour la vancomycine et la teicoplanine. À partir des modèles validés en amont ou bien issus de la littérature, des simulations ont été respectivement réalisées pour les glycopeptides et la ciprofloxacine dans le cas d’infections à Staphylococcus aureus résistant à la méthicilline et à Pseudomonas aeruginosa. Elles ont permis de mettre en évidence que lorsque la concentration minimale inhibitrice (CMI) de la bactérie incriminée était proche de la valeur critique de sensibilité, les doses à administrer pour être efficace doivent être augmentées par rapport aux recommandations actuelles. Quant à la prévention de l’émergence de la résistance bactérienne, dans le cas de la ciprofloxacine et de la teicoplanine, les doses à administrer sont telles que la toxicité est inévitable. Les analyses faites aux moyens des simulations de Monte Carlo sur les antibiotiques étudiés ont permis de mieux comprendre les déterminants de la minimisation de l’émergence de la résistance bactérienne à savoir obtenir dans les plus brefs délais la CMI exacte de la bactérie en cause ainsi qu’une estimation des paramètres individuels du patient / Bacterial resistance is a clearly established global threat. Researchers are coming together to find new solutions and alternatives to antibiotics. One of the main causes is the massive use of antibiotics. The unreasonable recourse of their use must be stopped. The population is aging, so it is the most vulnerable people who pay the price. We have used modeling tools to better understand the causes of this scourge. The goal of this work is to construct population pharmacokinetic models in the elderly for vancomycin and teicoplanin. From models validated upstream or from the literature, simulations have been performed for glycopeptides and ciprofloxacin for methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa infections, respectively. They showed that when the minimal inhibitory concentration (MIC) of the incriminated bacterium was close to the critical value of sensitivity, the doses to be administered in order to be effective have to be increased compared to current recommendations. As for the prevention of the emergence of bacterial resistance, in the case of ciprofloxacin and teicoplanin, the doses to be administered are so high that toxicity is unavoidable. Analyses using Monte Carlo simulations on the antibiotics studied provided a better understanding of the determinants of minimizing the emergence of bacterial resistance to obtain as quickly as possible the exact MIC of the bacteria and an estimation of individual patient parameters

A buspirona, ansiolítico da segunda geração usada no tratamento de ansiedade, foi analisada fisico-quimica e quimicamente, com o objetivo de fornecer subsídios para a respectiva monografia. Fizeram-se os seguintes testes de caracterização deste fármaco: faixa de fusão, análise elementar, pH, umidade e solubilidade. Realizaram-se igualmente as provas de identificação constante de: análise cromatográfica, ensaio de cloreto, espectro no infravermelho, espectro no ultravioleta, espectro de ressonância magnética protônica, espectro de massa, espectro de carbono-13, termogravimetria, calorimetria exploratória diferencial e cromatografia líquida de alta resolução. Efetuaram-se, também, determinações quantitativas pelos seguintes métodos: micro-Kjeldahl, gravimétrico, potenciométrico com nitrato de prata, doseamento com ácido perclórico em meio acetico por volumetria e potenciometria e espectrofotométrico no ultravioleta. Concluiu-se que: 1) o teste de caracterização mais indicado e o ponto de fusão; 2) das provas de identificação, as mais simples e rápidas são as seguintes: identificação de cloreto, infravermelho e ultravioleta; 3) dos métodos de determinação quantitativa, mais prático e o doseamento com ácido perclórico em meio acético, usando como indicador o vermelho de quinaldina. / Buspirone, a new non-benzodiazepine anxiolytic agent which is effective in the treatment of anxiety, was analyzed by physicochemical and chemical methods, having in mind a monograph of this drug. The fo1lowing characterization tests of this drug were performed: melting range, elementary analysis, pH, loss in drying, solubility. The following identification tests were also carried out: chromatographic analysis, identification of chloride, infrared absorption, ultraviolet absorption, protonic magnetic resonance spectra, mass spectrum, carbon 13C NMR spectroscopy, thermogravimetry and high performance liquid chromatography. Quantitative determinations were obtained through the following methods: micro-Kjeldahl, gravimetry, potenciometry, non-aqueous titration with perchloric acid and ultraviolet spectrometry. The following conclusions were reached: 1) the best characterization test is the melting range; 2) the simplest and quickest tests of identification are: identification of chloride, infrared and ultraviolet spectrophotometry; 3) the most practical method is the non-aqueous assay with perchloric acid.