It has been proposed that the use of cross-over or dose escalation designs for dose ranging studies in combination with more informative analysis could lead to a better characterisation of the dose response relationship. In example presented, the dose response relationship for the 3-hydroxy-3-methylglutaryl Coenzyme A inhibitors (HMG COA) inhibitor, simvastatin, was estimated from a cross-over study which covered the current recommended dose range (10 to 40 mg). Analysis using nonlinear mixed effects modelling approach demonstrated that the selected doses only covered 20% (70% to 90%) of the upper part of the estimated dose response relationship. It was concluded that a lower dose strength would be required to allow adjustment within the log-linear portion of the dose response relationship. The clinical implications of potential relationships between the pre-treatment cholesterol level and the model parameters were explored through prediction and simulation. On simulating the relationship between dose and the percentage of patients who would achieve reductions to below a recognised target concentration, it was found that a different set of dosages may better optimise clinical response. Where strict experimental design is invalidated by study design or restricted recruitment, the resulting data can be unbalanced and not easily analysed by standard statistical methods. In the example presented, the number and size of doses of dofetilide used to test for PK/PD differences between patients with ischaemic heart disease (ISH) and healthy volunteers were different. A population PK/PD modelling approach was implemented, and no difference between the two groups could be detected. The Cmax and peak QTc ranged were predicted to be narrower following a fixed dose regimen in comparison to a dose per kilogram regimen. However, after incorporating the PK/PD variability, this was not predicted to manifest into an overall increase in the risk of Torsades de Pointes.

Although there is a renewed interest in the therapeutic potential of cannabinoids, pharmacological and physiological characterisation of these promising compounds is currently not well documented in the respiratory system. The aim of this study is to increase our understanding of possible roles of cannabinoids in the airways. Apart from CB1 and CB2 receptor-mediated actions, cannabinoid compounds can also target TRPV1 receptors, ion channels or the orphan GPR55. In isolated guinea-pig bronchi, WIN55212-2 probably exerted its inhibitory effect on sensory nerves through CB2-like receptors. VIR did not act prejunctionally but its excitatory action was mediated through TRPV1 receptors. Δ9-THC activated sensory nerves presumably involving CB1 receptors. It was speculated that GPR55 might be activated by VIR and antagonized by CBD. CBD revealed multiple mechanisms of actions: it antagonized effects mediated by TRPV1 and NK2 receptors, modulated mast cell function and showed anti-allergic activity in an in vitro model of bronchial asthma. In a human bronchial epithelial cell line the functional expression of CB1 receptors could not be confirmed. Cannabinoids examined in this study were ineffective to induce signal transduction which would be linked to ion channel activity or to intracellular Ca2+ changes. Only VIR might trigger a CB1 receptor-independent signalling pathway in these cells. In conclusion, the findings presented in this thesis reflect the diversity of cannabinoid pharmacology in the airways. They show for the first time that CBD has the ability to reduce antigen-induced bronchoconstriction, indicating relevance in bronchial asthma.

The role of NAD(P)H dependent reductase mediated redox cycling in DNA strand breakage and cytotoxicity by doxorubicin (DOX), mitozantrone (MIT), CI941 and a series of alkylaminoanthraquinones (AQ) based on MIT has been investigated in MCF-7 human breast cancer cells. lAQ, 1,8AQ and DOX were found to redox cycle in MCF-7 cells as evidenced by stimulation of NADPH oxidation, superoxide anion formation and hydroxyl radical formation associated with generation of a drug free radical in MCF-7 S9 cell fraction. Furthermore, DOX formed a free radical, stimulated oxygen consumption and produced hydroxyl radicals in intact viable MCF-7 cells. 1,5AQ formed a free radical species and produced hydroxyl radicals but did not stimulate NADPH oxidation or superoxide anion formation in MCF-7 S9 fraction. Reactive oxygen formation correlated with the DNA strand breakage and cytotoxicity produced by IAQ, I,SAQ, 1,8AQ and DOX in MCF-7 cells. DOX also produced plasmid DNA strand breakage in the presence of purified cytochrome P4S0 reductase or xanthine oxidase. lAQ, 1,SAQ and 1,8AQ mediated plasmid DNA strand breakage only in the presence of purified cytochrome P4S0 reductase. MIT, CI941 and 1,4AQ did not redox cycle in MCF-7 cell S9 fraction yet produced equivalent cellular DNA strand breakage to DOX and AQ's at LDSO concentrations. The order of cytotoxicity of these compounds after a 1 hour exposure ~nd at least 6 d~ys further cell growth was;- (LDSO);- CI941(I.SxlO-lUM) >MIT(S.2xlO- YM) >lAQ(6.0xlO- 8M) >1,8AQ(O.5xlO- 6M) >1,4AQ(I.2xIO- 6M) >DOX(3.0xlO- 6M) >1,SAQ(12.3xIO- 6M). Variation in cellular uptake between the agents did not account for differences in cytotoxicity and DNA strand breakage observed. The 1,4 substitution pattern of MIT, CI941 and 1,4AQ appears to prevent metabolic activation. In further studies;- i) MIT and CI941 were more potent inhibitors of 3 MCF-7 topoisomerase I activity than other quinones, ii) MIT and DOX-Fe + complexes mediated plasmid DNA strand breakage in the presence of reduced glutathione iii) Only MIT was oxidatively activated by horseradish peroxidase!H202' producing plasmid DNA damage. The results indicate that future development of quinone anti tumour agents should concentrate on compounds which do not redox cycle but have functional groups that assist iron binding, activation by peroxidases and/or topoisomerase inhibition. In this respect, these agents should possess chromophore hydroxyl groups as these appear to be responsible for the increased cytotoxicity of MIT and CI941 compared to the other AQ's.

This research programme investigates the influence of genetic variability on the clinical pharmacology of carbamazepine (CBZ) and lamotrigine (LTG). Common polymorphisms in genes that may influence the response to CBZ and LTG include CYP3A4 g.-392A>G, CYP3A5 g.6986A>G, CYP1A2 g.5734C>A, EPHX1 c.337T>C, EPHX1 c.416A>G, UGT2B7 c.802C>T, ABCB1 c.1236C>T, ABCB1 c.2677G>T/A, ABCB1 c.3435C>T and SCN2A c.56G>A. The prevalence of these common polymorphisms was evaluated in a 400-strong study population from a single research unit. Minor allele frequency ranged between 3.5% (CYP3A4 -392G) and 48.0% (ABCB1 1236T). Allele and genotype distributions were comparable to published data for other Caucasian populations. The influence of common polymorphisms in drug metabolising enzyme (DME) and sodium channel genes on the optimal dose of CBZ was assessed in a cohort of 70 patients. This study revealed that age and common polymorphisms in the EPHX1 gene (c.337T>C and c.416A>G) are potential predictors for optimal dose of CBZ. The significant effects of EPHX1 variants may be explained by their significant contribution to the inactivation of CBZ. These potential predictors explain approximately 15% of the inter-individual variation in CBZ dose requirements. Preliminary findings suggest that common polymorphisms in DME genes do not form a unique profile which increases the risk of developing intolerable CBZ adverse effects. It is unlikely that common polymorphisms in ABCB1 and SCN2A genes influence the expression and/or activity of their respective proteins to the level at which they can dictate response to LTG therapy. The influence of common polymorphisms in ABCB1 and SCN2A genes on the optimal dose of LTG was assessed in a cohort of 94 patients. Optimal dose in this study was defined as the final dose given to the patients that successfully maintained seizure freedom for at least 1 year with LTG monotherapy. Several basic clinical factors such as age and gender were also examined as potential predictors. The effect of predictors on the optimal dose of LTG was investigated using linear regression analysis. This study revealed that gender and common polymorphisms in the ABCB1 gene (c.1236C>T and c.3435C>T) are potential predictors for optimal dose of LTG. These predictors explain approximately 17% of the inter-individual variation in LTG dose requirement. These findings further highlight the potential role of P-gp in the management of epilepsy. The final study investigated the influence of ABCB1 c.1236C>T and ABCB1 c.3435C>T polymorphisms on the pharmacokinetics of LTG. A total of 156 blood samples from 50 patients receiving LTG monotherapy were available for analysis. The influence of ABCB1 variants was evaluated by population pharmacokinetics. This approach successfully estimated the oral clearance of LTG monotherapy at steady-state. However, the absorption rate constant (Ka) and volume of distribution (Vd) of LTG were poorly estimated. The inclusion of common polymorphisms in the ABCB1 gene in the pharmacokinetic model did not improve the estimation of oral clearance. This may indicate that common variants of ABCB1 do not influence clearance of LTG.

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