An understanding of the criteria governing lymphatic absorption of drugs from the gastrointestinal tract may lead to selective uptake of drugs via this route. Lymphatic absorption may have potential advantages over the portal absorption of drugs; first pass elimination may be avoided as the drug reaches the systemic circulation before it reaches the liver, the absorption of some poorly absorbed drugs may be improved and it may provide a means of targeting anticancer agents directly to the lymphatics. In the present study, the lymphatic uptake of a series of retinoids was investigated after oral administration. The most important factors which were found to affect lymphatic absorption were, the chemical nature of the compound and the nature of the oily vehicle in which the retinoid was administered. It was found that the greater the lipophilicity of the retinoid, (as defined by the logarithm of its octanol : water partition coefficient, log. P), the greater was its lymphatic absorption. Temarotene, Ro 15-0778, (log. P = 8.7) exhibited a maximum dose adjusted lymphatic absorption rate of 4100 ng/h compared with the less lipophilic Ro 04-3780 (log. P = 6.8) which showed a maximum rate of only 150 ng/h. The oily vehicle in which the retinoid was orally administered strongly influenced the rate of absorption via the lymphatic route. Ro 04-3780 demonstrated a 150 fold difference between the selected oils giving the maximum and the minimum lymphatic absorption rate. The best oils in this role appeared to be those in which the retinoid showed the least solubility. Mesenteric lymph flow rate was also shown to vary depending upon the oil. Basal lymph flow rate in the fasted rat, after dosing with saline, was 1.6 m1/h/kg. Cottonseed oil and soyabean oil promoted an increase in this flow rate to greater than 3.0 ml/h/kg (p < 0.01) when orally administered with Ro 04-3780. Conversely, linoleic acid suppressed the mean lymph flow rate to 0.8 ml/h/kg (p < 0.01) after oral administration with Ro 04-3780. Lymph turbidity was evaluated as an indication of chylomicron formation and transport in the lymph. Since the chylomicron is the particle in which dietary fats enter the lymphatic system, it was thought that lipophilic drugs, which are soluble in dietary lipid, may be carried into the lymphatic system via this pathway. The mixed long chain fatty tri-acyl glycerol oils, cotton seed oil, soyabean oil and peanut oil, when orally dosed to rats, produced the most turbid lymph (25 - 48 times greater than the turbidity produced after an oral dose of saline). These type of oil also promoted the highest lymphatic uptake rate for the retinoids. Other oils including, oleic acid, linoleic acid and MTS (a Miglyol S12 based self-emulsifying oil system), demonstrated much wider ranging extents of lymphatic absorption, but produced lymph with similar but lower turbidity (10 - 12 times greater than the turbidity produced after an oral dose of saline). A self-emulsifying oil system was developed for use in the oral administration of a retinoid. This system (MTS), produced a stable emulsion with a particle size of 500 nm after gentle mixing with an aqueous solvent and contained 80% Miglyol 812 and 20 % surfactants. MTS increased both the lymphatic and portal absorption rates for Ro 15-0778 by three fold compared with Miglyol S12 alone, improving the overall bioavailability but without selective promotion of lymphatic uptake. The effect of feeding, prior to orally dosing with an oil (linoleic acid) containing Ro 10-9359, was to suppress greatly the portal absorption rate of the retinoid from 310 ng/h to less than 25 ng/h. A number of factors, which were believed to be important in the lymphatic absorption of the retinoids, have been investigated here, using the rat as an animal model. The data obtained in this work suggest that lymphatic absorption is a very complex process and the factors which govern this absorptive pathway vary depending upon the nature of the drug being studied and the nature of the orally dosed vehicle.

Rheumatoid Arthritis (RA) is the commonest inflammatory polyarthritis in the UK, and is associated with significant symptoms, disability and premature mortality. Treatment options in 1980 were restricted to corticosteroids, non-steroidal anti-inflammatory drugs, and a small number of disease modifying anti-rheumatic drugs (DMARDs). There had been few large, well conducted randomised controlled studies such that our knowledge about the relative efficacy and safety of the drugs available was very limited. Trial design also left much to be desired, with inadequate methods of randomisation and/or concealment of allocation being commonplace. There was no consensus about which outcome measures ought to included in trials, and no composite measures of outcome had yet been developed or validated. The limited evidence base and restricted therapeutic armamentarium was reflected in the poor outcome of the disease for many patients: remission was rare, and patients often experienced increasing disability, orthopaedic intervention, work-related unemployment and premature mortality. Within 20 years, the prognosis for patients with newly diagnosed RA has dramatically improved. Modern management results in the majority of patients achieving low disease activity or even remission. The studies incorporated into this thesis have played an important role in the evolution of these management strategies. Early studies focussed on building the evidence base for the use of DMARD monotherapy, demonstrating that sulfasalazine and methotrexate were both safe and effective treatments. The Sulfasalazine-Auranofin trial contributed to the downfall of auranofin, a drug that is no longer manufactured. Contrary to early concerns, methotrexate was proven to be well tolerated, even in the socially deprived population of Greater Glasgow, and this drug had become the DMARD of first choice in the management of RA. DMARD monotherapy, however, is usually not sufficient to maintain good disease control in the long term. The Gold-Hydroxychloroquine study was one of the first studies to investigate the role of combination DMARD therapy in a well conducted, double blind randomised controlled trial (RCT). The results were negative, but a follow up trial demonstrated the superiority of stepping up to Methotrexate-Sulfasalazine combination therapy when compared to sequential monotherapy in the MASCOT study. The West of Scotland Early RA corticosteroid trial was a double blind RCT investigating the role of low dose oral corticosteroids in addition to sulfasalazine therapy. It failed to demonstrate any benefit of low dose steroids, a finding that is at odds with a growing literature that has established corticosteroids as a proven disease modifying therapy. The strategy trials (Tight Control of RA [TICORA] and Triple Therapy in Early RA [TEAR] studies) have been the most influential studies to have been performed in Glasgow. They did not primarily address the issue of whether a drug is effective or whether one drug is more effective than another. Rather, they sought to test a hypothesis drawn from observations in other biologic models (principally Type I Diabetes Mellitus): namely, that i) using currently available DMARDs ‘tight control’ can be achieved if patients are reviewed frequently, their disease assessed formally (using the disease activity score), and their treatment escalated if their disease remained active; ii) that the achievement of tight control would lead to improved outcomes. The studies provided strong evidence that dramatic improvements in symptom control, disability and radiographic progression can be achieved by pursuing this strategy of ‘Tight Control’. National and international clinical guidelines, and international consensus statements have embraced the results of TICORA (and subsequent confirmatory studies), such that regular, frequent assessment of the patient, use of composite measures of disease activity and the adoption of a ‘treat-to-target’ therapeutic strategy have become accepted as ‘best practice’ throughout the world.

This study investigated the mood states of adolescents who have Duchenne Muscular Dystrophy. The specific purpose was twofold. First, to examine whether there were differences in the mood states of adolescents who have Duchenne Muscular and able-bodied adolescents. And second, to explore and evaluate whether participation in a theatrical performance program had a positive effect on the mood states of the adolescents who have Duchenne Muscular Dystrophy. / The statistical findings indicated that there were significant initial and concluding differences in the mood states of the adolescents who have Duchenne Muscular Dystrophy and their able-bodied cohorts. Analysis of the theatrical performance program indicated a short-term pre-post treatment improvement in the mood states of the adolescents who have Duchenne Muscular Dystrophy. This short-term improvement, however, did not sustain itself over time. (Abstract shortened by UMI.)

The health benefits of the fruit of Punica granatum (pomegranate) have been recognised for many centuries. This thesis tested the hypothesis that a potent novel antimicrobial system could be developed based upon the activity of pomegranate rind extract (PRE), particularly when combined with a potentiating agent, and focussed on studying activity against Herpes simplex virus, types 1 and 2 (HSV-1 and HSV-2). High potentiated virucidal action of PRE against HSV-1 was observed when ferrous sulphate (FeSO4) was added; however, this activity diminished rapidly and also gave rise to an unsightly black byproduct. Data was obtained suggesting this was linked to the rapid oxidation of ferrous to ferric (Fe (II) to Fe (III)); however, no such oxidation was indicated when zinc sulphate (ZnSO4) was added instead, prompting an exploration of the effects of PRE + ZnSO4 combination on HSV. The potentiation of PRE by ZnSO4 in virucidal mode was comparable to that observed with FeSO4 (at the concentration examined for both metal salts). More in-depth investigation of the potentiation of PRE and ZnSO4 achieved over a 9000 fold increase in viral destruction in comparison to either agent alone. This activity was not transient and did not produce a (black) byproduct. Other salts of Zn (II) generally performed similarly to ZnSO4, with the main exception of ZnO which was too insoluble in water to test. When the antiviral properties of PRE were examined, the results were again potent and comparable to Aciclovir, the established treatment for Herpes simplex infections. Furthermore, PRE demonstrated even greater potency against Aciclovir-resistant HSV-2. When applied to ex vivo skin, PRE produced a 66% reduction in the level of cyclooxygenase-2 (COX–2) - an important inflammatory mediator - with the presence of ZnSO4 having no effect. PRE and ZnSO4 were formulated as a stable hydrogel, which was able to effectively deliver the biologically active compounds through the epidermal, buccal and vaginal membranes to the epidermal/dermal interface - the major sites of HSV-1 and HSV-2 vesicular clusters during a clinical infection.

A pathophysiological study was made of spontaneously breathing dogs anaesthetised by a neurolept-analgesic technique and exposed to 100% oxygen at 2 ATA to demonstrate the time-course and mechanism of response. The animals remained apparently normal for some 18 hours, following which the majority developed a fulminating intra-alveolar oedema and died of hypoxaemia within a few hours. There was no evidence of systemic nor pulmonary capillary hypertension, and electron microscopy demonstrated complete absence of damage to the endothelial and Type 1 epithelial cells of the alveolar septum. Changes were detected in the Type 2 cells, and oedemagenesis was attributed to an oxygen-induced depression of surfactant activity.

The PhD project is about the Regulation of vascular tone by endothelium. It involves studying the effects of methylated arginine analogues such as monomethyl arginine (L-NMMA) and asymmetric dimethylarginine (ADMA) on nitric oxide (NO) activity in rat aorta using organ baths containing Krebs solution at 37 ˚C and gassed with 95% O2 and 5% CO2. These two inhibitors of endothelial nitric oxide synthase enzyme (eNOS) showed an anomalous action, as they inhibited basal NO activity (assessed by enhancement of phenylephrine-induced tone and by blocking relaxations produced by superoxide dismutase or the PDE5 inhibitor; T0156) but not that is stimulated by agonists like acetylcholine or the calcium ionophore A23187. After establishing these findings I will try to find the reason(s) for these paradoxical actions of these two endogenously synthesized inhibitors of eNOS.

The androgen receptor (AR) is the central component in regulation of the androgen signalling within the prostate gland. Deregulation of the AR activity is frequently involved in the development of prostate cancer. Treatment of advanced prostate cancer often involves chemotherapy and most of these drugs exert their function by generating genotoxic stress such as DNA damage. Although many cellular responses to DNA damage have been clarified over the past years, the effects of genotoxic stress on AR function remain to be elucidated. Here, the effects of genotoxic agents used in chemotherapeutic regimes were investigated in relation to endogenously expressed AR function in the hormone responsive prostate cancer cell line LNCaP. This led to the novel finding that the topoisomerase 11 inhibitors, etoposide and doxorubicin, and the DNA crosslinking agent, cisplatin, inhibited the AR activity. It was further discovered that this loss of AR activity could not be explained by changes in cell cycle distribution, altered nuclear translocation of the AR, reduced expression of the receptor or by induction of apoptosis. Activation of the tumour suppressor p53 is a central component in various cellular responses to genotoxic stress, however, the inhibition of AR activity in response to genotoxic stress was found to be mediated by a mechanism independent of p53 function. Etoposide reduced AR ligand binding within the first hour of androgen exposure, a response only observed to a minor degree after cisplatin treatment. In contrast, cisplatin caused a loss of serine 81 phosphorylation on the AR after 8 hours of drug exposure, which was a response not seen in etoposide treated cells. Interestingly, further studies revealed that at early timepoints both agents inhibited the hormone stimulated recruitment of AR to androgen response elements (AREs) in the promoter and enhancer regions of an AR regulated gene. A possible involvement of MAPK, P13K or cell cycle checkpoint signalling pathways was investigated, but none were found to be directly involved, however, preliminary studies suggest that fully functional HSP90 may be involved in this aspect of AR regulation.

Oil and emulsion vehicles have been shown to alter the oral absorption of many drugs. This may be due to enhanced lymph flow and/or altered gastro-intestinal motility in the presence of the oils. The oral absorption of a model compound (DOT) in the presence of three chemically different oils, arachis oil, Miglyol 812 and liquid paraffin was investigated in rats, the influence of lymphatic absorption and gastro-intestinal motility being determined. The findings were applied to the for.mulation of the'steroid prednisolone, in an attempt to produce elevated more uniform plasma drug levels by enhancing lymphatic absorption. The rank order for total DOT absorption from 1m! Volumes of different vehicles was arachis oil > Miglyol 812= water containing 6% Tween 80 > liquid paraffin. The concentration of DDT in lymph collected via thoracic duct cannblae in anaesthetised rats was greatest in the presence of arachis oil, there being no difference between Miglyol 812 and liquid paraffin. Using a gamma camera the gastric emptying rate and total intestinal transit of 9~c-sulphur colloid, an oil phase marker, was shown to be faster in the presence of 1ml liquid paraffin than the other two oils. The oral absorption of 3H-prednisolone was independent of the nature of the oily vehicle (30pl volumes) and was not selectively absorbed into the lymph. Esterification to 3H-prednisolone-21-palmitate increased the lipophilicity of the drug but did not stimulate selective lymphatic absorption and reduced oral absorption following administration in arachis oil. Lymphatic absorption of the ester was not promoted by administration in 1ml arachis oil. Only the lymphatic absorption of compounds exhibiting selective uptake into the lymph may be enhanced by the presence of a suitable lipid vehicle. Altered gastro-intestinal motility in the presence of lipids may have greater potential for enhancing the absorption of a wider variety of compounds.

This thesis describes synthetic studies directed towards a second generation total synthesis of ulapualide A. Ulapualide A is an extraordinary bioactive tris-oxazole based macrolide which was isolated from the egg masses of the marine sponge Hexabranchus sanguineus and exhibits potent antifungal activity with inhibition of leukaemia cell proliferation. The Introduction to this thesis includes an overview of natural product chemistry and draws attention to the 'ulapualide' family of secondary metabolites including their isolation, biological activity, biosynthesis and structural determination. Also included is a summary of a total synthesis of ulapualide A by our research group in Nottingham, together with a review of oxazole containing natural products. The Results and Discussion section of this thesis details our general strategy for an alternative design for the synthesis of the tris-oxazole based macrolide core of ulapualide A. A synthesis of a model system exemplifying this strategy is then described, together with a detailed discussion of polyoxazole ring formation. This is followed by application of the model study to ulapualide A itself, and includes a total synthesis of the polyol C26-C41 side-chain of ulapualide A. The section concludes by describing our synthetic efforts towards the remaining chiral fragment of this natural product, the bottom-chain. The thesis concludes with an Experimental section containing full details of the preparative work completed and listing spectroscopic and analytical data for all new compounds synthesised during the study. An Appendix contains a description of contemporaneous synthetic studies carried out by Panek et al during the course of my PhD studies. X-ray crystallographic and spectroscopic data, together with reprints of publications resulting from our work are also included.

Poly (butyl 2-cyanoacrylate) nanoparticles have been prepared with a range of particle sizes by varying the nature and concentration of stabiliser added to the polymerisation medium. Particle size analysis was performed by photon correlation spectroscopy. The range of diameters produced using dextran stabilisers was found to be approximately 100 to 800nm. This could be extended to 3ym using j3 -cyclodextrin and to 20nm using polysorbate 20. The results infer that the nanoparticles are sterically stabilised. The molecular weight of the cyanoacrylate polymer formed during nanoparticle production was found to be dependent on the type of stabiliser used together with the polymerisation pH and monomer concentration. The bulk of the polymer had a relatively low molecular weight (<2000) which indicates that nanoparticles are formed by an aggregative mechanism. Dextran was found to copolymerise with the monomer to give an interfacial layer of the polysaccharide attached by covalent linkages. By using dextrans bearing charged functional groups it was possible to alter the electrophoretic behaviour of the resulting nanoparticles. Partial oxidation of the surface dextran introduced aldehyde groups which were capable of covalently binding a simple amine, aniline, thereby enhancing the uptake and decreasing the release rate of this compound. This technique may be applicable to the covalent coupling of antibodies or cytotoxic agents to the nanoparticle surface. Nanoparticles were radiolabelled with a technetium-99m-dextran complex and the biodistribution of this colloid determined in rabbits by gamma scintigraphy following intravenous injection. Most of the nanoparticle suspension (approximately 50%) was cleared by the liver and spleen. Coating the nanoparticles with non-ionic surfactants (poloxamer 338 or Tetronic 908) failed to alter significantly this distribution pattern.