No description available.

No description available.

No description available.

Mitochondria are important to the pharmaceutical industry. Mitochondrial perturbation is central to the aetiology of many poorly treated human diseases (e.g. neurodegenerative diseases, diabetes and cancer) and hence mitochondria are increasingly becoming therapeutic targets. However, drug-induced mitochondrial toxicity (DIMT) has also gained recognition as a significant cause of safety related attrition (including multiple post-market withdrawals). The present work ultimately aimed to discern the mechanisms downstream of DIMT in vitro and in vivo with two tool compounds which act via inhibition of the mitochondrial electron transport chain (ETC) (GSK932121A) or via uncoupling of mitochondrial oxidative phosphorylation (GSK2617971A). The first objective was to compare mitochondrial functional responses between in vitro (treatment of mitochondria isolated from control animals or in vitro cell lines) and in vivo scenarios using oxygen consumption rate (OCR) and calcium retention capacity (CRC) assays. The direction of change in these assays was often different between the in vitro and in vivo situation. Whilst seemingly contradictory, these differences reflect an adapted state in vivo which results from the initial insult in combination with compensatory changes made by the tissue to maintain energy production. In contrast, only the initial, unconfounded, response to treatment is observable in vitro. Integration of both is key to the future risk assessment of mitochondrially-active drug candidates. The second objective was to use a systems approach to build candidate adverse outcome pathways (AOP)/modes of action (MOA) for both tool compounds. AOP/MOAs were constructed based on key events observed in vivo. With both compounds, effect on body temperature, increased lactate/pyruvate ratio, gene expression changes indicative of a fasted state (induction of gluconeogenic genes) and mitochondrial ultrastructure effects were key events observed in the adverse outcome. Future work using additional compounds with mitochondrial activities would help validate the proposed AOP/MOAs. The third objective was to highlight markers of underlying mitochondrial perturbation with utility for future pre-clinical monitoring in the absence of overt DIMT. A number of mitochondrial functional changes (e.g. effect on CRC) were observed prior to the onset of overt toxicity in vivo, with both compounds and these endpoints have potential use in tailored future studies with mitochondrially-active drug candidates. These findings add to the knowledgebase on DIMT, particularly with regard to the candidate AOP/MOAs which have been postulated. This will facilitate the selection of more sensitive and functional endpoints for use in future toxicology studies with drugs where mitochondrial impairment is suspected.

No description available.

No description available.

The diagnosis and treatment of pathologies of the gastrointestinal (GI) tract are performed routinely by gastroenterologists using endoscopes and colonoscopes, however the small intestinal tract is beyond the reach of these conventional systems. Attempts have been made to access the small intestines with wireless capsule endoscopes (WCE). These pill-sized cameras take pictures of the intestinal wall and then relay them back for evaluation. This practice enables the detection and diagnosis of pathologies of the GI tract such as Crohn's disease, small intestinal tumours such as lymphoma and small intestinal cancer. The problems with these systems are that they have limited diagnostic capabilities and they do not offer the ability to perform therapy to the affected areas leaving only the options of administering large quantities of drugs or surgical intervention. To address the issue of administering therapy in the small intestinal tract this thesis presents an active swallowable microrobotic platform which has novel functionality enabling the microrobot to treat pathologies through a targeted drug delivery system. This thesis first reviews the state-of-the-art in WCE through the evaluation of current and past literature. A review of current practises such as flexible sigmoidoscopy, virtual colonoscopy and wireless capsule endoscopy are presented. The following sections review the state-of-the-art in methods of resisting peristalsis, drug targeting systems and drug delivery. A review of actuators is presented, in the context of WCE, with a view to evaluate their acceptability in adding functionality to current WCEs. The thesis presents a novel biologically-inspired holding mechanism which overcomes the issue of resisting natural peristalsis in the GI tract. An analysis of the two components of peristaltic force, circumferential and longitudinal peristaltic contractions, are presented to ensure correct functionality of the holding mechanism. A detailed analysis of the motorised method employed to deploy the expanding mechanism is described and a 5:1 scale prototype is presented which characterises the gearbox and validates the holding mechanism. The functionality of WCE is further extended by the inclusion of a novel targeting mechanism capable of delivering a metered dose of medication to a target site of interest in the GI tract. A solution to the problem of positioning a needle within a 360 degree envelope, operating the needle and safely retracting the needle in the GI tract is discussed. A comprehensive analysis of the mechanism to manoeuvre the needle is presented and validation of the mechanism is demonstrated through the evaluation of scale prototypes. Finally a drug delivery system is presented which can expel a 1 ml dose of medication, stored onboard the capsule, into the subcutaneous tissue of the GI tract wall. An analysis of the force required to expel the medication in a set period of time is presented and the design and analysis of a variable pitch conical compression spring which will be used to deliver the medication is discussed. A thermo mechanical trigger mechanism is presented which will be employed to release the compressed conical spring. Experimental results using 1:1 scale prototype parts validate the performance of the mechanisms.

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The facility to controlled triggered release from a “cage” system remains a key requirement for novel drug delivery. Earlier studies have shown that Bis-Azo PC based photosensitive liposomes are beneficial for drug delivery. Thus, the aim of this project was to develop photosensitive liposomes that can be used for the controlled release of drugs through UV irradiation, particularly therapeutic agents for the treatment of psoriasis. Bis-Azo PC was successfully synthesized and incorporated into a range of liposomal formulations, and these liposomes were applied for the controlled release of BSA-FITC. Bis-Azo PC sensitized liposomes were prepared via interdigitation fusion method. IFV containing optimum cholesterol amount in terms of protein loading, stability and photo-trigger release of protein was investigated. Further studies investigated the stability and triggered release of the HMT from IFV. Finally, permeation behavior of HMT and HMT-entrapped IFV through rat skin was examined using Franz cell. Results from protein study indicated that the stable entrapment of the model protein was feasible as shown through fluorescence spectroscopy and maximum of 84% protein release from IFV after 12 min of UV irradiation. Moreover, stability studies indicated that IFV were more stable at 4 0C as compared to 25 0C. Hence, DPPC:Chol:Bis-Azo PC (16:2:1) based IFV was chosen for the controlled release of HMT and these studies exhibited that photo-trigger release and stability data of HMT-entrapped IFV are in line with the protein results. Franz cell work inferred that HMT-entrapped IFV attributed to slower skin permeation as compared to HMT. CLSM also demonstrated that HMT can be used as a fluorescent label for the in vitro skin study. Overall, the work highlighted in this thesis has given useful insight into the potentials of Bis-Azo PC based IFV as a promising carrier for the treatment of psoriasis.

Nanoparticles offer an ideal platform for the delivery of small molecule drugs, subunit vaccines and genetic constructs. Besides the necessity of a homogenous size distribution, defined loading efficiencies and reasonable production and development costs, one of the major bottlenecks in translating nanoparticles into clinical application is the need for rapid, robust and reproducible development techniques. Within this thesis, microfluidic methods were investigated for the manufacturing, drug or protein loading and purification of pharmaceutically relevant nanoparticles. Initially, methods to prepare small liposomes were evaluated and compared to a microfluidics-directed nanoprecipitation method. To support the implementation of statistical process control, design of experiment models aided the process robustness and validation for the methods investigated and gave an initial overview of the size ranges obtainable in each method whilst evaluating advantages and disadvantages of each method. The lab-on-a-chip system resulted in a high-throughput vesicle manufacturing, enabling a rapid process and a high degree of process control. To further investigate this method, cationic low transition temperature lipids, cationic bola-amphiphiles with delocalized charge centers, neutral lipids and polymers were used in the microfluidics-directed nanoprecipitation method to formulate vesicles. Whereas the total flow rate (TFR) and the ratio of solvent to aqueous stream (flow rate ratio, FRR) was shown to be influential for controlling the vesicle size in high transition temperature lipids, the factor FRR was found the most influential factor controlling the size of vesicles consisting of low transition temperature lipids and polymer-based nanoparticles. The biological activity of the resulting constructs was confirmed by an invitro transfection of pDNA constructs using cationic nanoprecipitated vesicles. Design of experiments and multivariate data analysis revealed the mathematical relationship and significance of the factors TFR and FRR in the microfluidics process to the liposome size, polydispersity and transfection efficiency. Multivariate tools were used to cluster and predict specific in-vivo immune responses dependent on key liposome adjuvant characteristics upon delivery a tuberculosis antigen in a vaccine candidate. The addition of a low solubility model drug (propofol) in the nanoprecipitation method resulted in a significantly higher solubilisation of the drug within the liposomal bilayer, compared to the control method. The microfluidics method underwent scale-up work by increasing the channel diameter and parallelisation of the mixers in a planar way, resulting in an overall 40-fold increase in throughput. Furthermore, microfluidic tools were developed based on a microfluidics-directed tangential flow filtration, which allowed for a continuous manufacturing, purification and concentration of liposomal drug products.