Formulation and characterisation of an effective particulate delivery vehicle for the novel sub-unit vaccine antigen, Ag85B-ESAT-6

This research focused on the formation of particulate delivery systems for the sub-unit fusion protein, Ag85B-ESAT-6, a promising tuberculosis (TB) vaccine candidate. Initial work concentrated on formulating and characterising, both physico-chemically and immunologically, cationic liposomes based on the potent adjuvant dimethyl dioctadecyl ammonium (DDA). These studies demonstrated that addition of the immunomodulatory trehalose dibehenate (TDB) enhanced the physical stability of the system whilst also adding further adjuvanticity. Indeed, this formulation was effective in stimulating both a cell mediated and humoural immune response. In order to investigate an alternative to the DDA-TDB system, microspheres based on poly(DL-lactide-co-glycolide) (PLGA) incorporating the adjuvants DDA and TDB, either alone or in combination, were first optimised in terms of physico-chemical characteristics, followed by immunological analysis. The formulation incorporating PLGA and DDA emerged as the lead candidate, with promising protection data against TB. Subsequent optimisation of the lead microsphere formulation investigated the effect of several variables involved in the formulation process on physico-chemical and immunological characteristics of the particles produced. Further, freeze-drying studies were carried out with both sugar-based and amino acid-based cryoprotectants, in order to formulate a stable freexe-dried product. Finally, environmental scanning electron microscopy (ESEM) was investigated as a potential alternative to conventional SEM for the morphological investigation of microsphere formulations. Results revealed that the DDA-TDB liposome system proved to be the most immunologically efficient delivery vehicle studied, with high levels of antibody and cytokine production, particularly gamma-interferon (IFN-ϒ), considered the key cytokine marker for anti-mycobacterial immunity. Of the microsphere systems investigated, PLGA in combination with DDA showed the most promise, with an ability to initiate a broad spectrum of cytokine production, as well as antigen specific spleen cell proliferation comparable to that of the DDA-TDB formulation.

Cellular delivery of hammerhead ribozymes

Hammerhead ribozymes are potent RNA molecules which have the potential to specifically inhibit gene expression by catalysing the trans-cleavage of mRNAs. However, they are unstable in biological fluids and cellular delivery poses a problem. Site-specific chemical modification of hammerhead ribozymes was evaluated as a means of enhancing biological stability. Chimeric, 2'-O-methylated ribozymes, containing only five unmodified ribonucleotides, were catalytically active in vitro (kcat = 1.46 min-1) and were significantly more stable in serum and lysosomal enzymes than unmodified (all-RNA) counterparts. Furthermore, they remained undegraded in cell-containing media for up to 8 hours. Stability enhancement allowed cellular uptake properties of radiolabelled ribozymes to be assessed following exogenous delivery. Studies in vulval and glial cell lines indicated that chimeric ribozymes became cell-associated via an inefficient process, which was energy and concentration dependant. A considerable proportion of ribozymes remained bound to cell-surface components, however, a small proportion (<1%) were internalised via mechanisms of adsorptive and / or receptor mediated endocytosis. Fluorescent microscopy indicated that ribozymes were localised within endosomal / lysosomal vesicles following cell entry. This was confirmed by immuno-electron microscopy, which allowed the detection of biotin-labelled ribozymes within the cell ultrastructure. Despite the predominant localisation within endocytic vesicles, a small proportion of internalised ribozymes appeared able to exit these compartments and penetrate target sites within the nucleus and cytoplasm. The ribozymes designed in this report were directed against the epidermal growth factor receptor mRNA, which is over-expressed in a malignant brain disease called glioblastoma multiforme. In order to examine the fate of ribozymes in the brain, the distribution of FITC-labelled ribozymes was examined following intra-cerebro ventricular injection to mice. FITC-ribozymes demonstrated high punctate pattern of distribution within the striatum and cortex, which appeared to represent localisation within cell bodies and dendritic processes. This suggested that delivery to glial cells in vivo may be possible. Finally, strategies were investigated to enhance the cellular delivery of ribozymes. Conjugation of ribozymes to anti~transferrin receptor antibodies improved cellular uptake 3-fold as a result of a specific interaction with transferrin receptors. Complexation with cationic liposomes also significantly improved cell association, however, some toxiclty was observed and this could be a limitation to their use. Overall, it would appear that hammerhead ribozymes can be chemically stabilised to allow direct exogenous administration in vivo. However, additional delivery strategies are probably required to improve cellular uptake, and thus, allow ribozymes to achieve their full potential as pharmaceutical agents. KEYWORDS: Catalytic

A strategy formulation process for the delivery of technology enabled service delivery systems

The Product Service Systems, servitization, and Service Science literature continues to grow as organisations seek to protect and improve their competitive position. The potential of technology applications to deliver service delivery systems facilitated by the ability to make real time decisions based upon ‘in the field’ performance is also significant. Research identifies four key questions to be addressed. Namely: how far along the servitization continuum should the organisation go in a single strategic step? Does the organisation have the structure and infrastructure to support this transition? What level of condition monitoring should it employ? Is the product positioned correctly in the value chain to adopt condition monitoring technology? Strategy consists of three dimensions, namely content, context, and process. The literature relating to PSS, servitization, and strategy all discuss the concepts relative to content and context but none offer a process to deliver an aligned strategy to deliver a service delivery system enabled by condition based management. This paper presents a tested iterative strategy formulation methodology which is the result of a structured development programme.

On the Poisson Process of Order k Space

2010 Mathematics Subject Classification: 60E05, 62P05.

Methodology to create cost estimation models for engineer-to-order operations

This research addresses the problem of cost estimation for product development in engineer-to-order (ETO) operations. An ETO operation starts the product development process with a product specification and ends with delivery of a rather complicated, highly customized product. ETO operations are practiced in various industries such as engineering tooling, factory plants, industrial boilers, pressure vessels, shipbuilding, bridges and buildings. ETO views each product as a delivery item in an industrial project and needs to make an accurate estimation of its development cost at the bidding and/or planning stage before any design or manufacturing activity starts. ^ Many ETO practitioners rely on an ad hoc approach to cost estimation, with use of past projects as reference, adapting them to the new requirements. This process is often carried out on a case-by-case basis and in a non-procedural fashion, thus limiting its applicability to other industry domains and transferability to other estimators. In addition to being time consuming, this approach usually does not lead to an accurate cost estimate, which varies from 30% to 50%. ^ This research proposes a generic cost modeling methodology for application in ETO operations across various industry domains. Using the proposed methodology, a cost estimator will be able to develop a cost estimation model for use in a chosen ETO industry in a more expeditious, systematic and accurate manner. ^ The development of the proposed methodology was carried out by following the meta-methodology as outlined by Thomann. Deploying the methodology, cost estimation models were created in two industry domains (building construction and the steel milling equipment manufacturing). The models are then applied to real cases; the cost estimates are significantly more accurate than the actual estimates, with mean absolute error rate of 17.3%. ^ This research fills an important need of quick and accurate cost estimation across various ETO industries. It differs from existing approaches to the problem in that a methodology is developed for use to quickly customize a cost estimation model for a chosen application domain. In addition to more accurate estimation, the major contributions are in its transferability to other users and applicability to different ETO operations. ^

Creating Memories: The Interactive Process of Service Delivery

A model is presented that captures the complex nature of the service experience in an attempt to aid in the design, development and evaluation of service delivery personnel and systems.

Evaluating the Appropriateness of Project Delivery Systems for Different Trenchless Methods

Trenchless methods have been considered to be a viable solution for pipeline projects in urban areas. Their applicability in pipeline projects is expected to increase with the rapid advancements in technology and emerging concerns regarding social costs related to trenching methods. Selecting appropriate project delivery system (PDS) is a key to the success of trenchless projects. To ensure success of the project, the selected project delivery should be tailored to trenchless project specific characteristics and owner needs, since the effectiveness of project delivery systems differs based on different project characteristics and owners requirements. Since different trenchless methods have specific characteristics such rate of installation, lengths of installation, and accuracy, the same project delivery systems may not be equally effective for different methods. The intent of this paper is to evaluate the appropriateness of different PDS for different trenchless methods. PDS are examined through a structured decision-making process called Fuzzy Delivery System Selection Model (FDSSM). The process of incorporating the impacts of: (a) the characteristics of trenchless projects and (b) owners’ needs in the FDSSM is performed by collecting data using questionnaires deployed to professionals involved in the trenchless industry in order to determine the importance of delivery systems selection attributes for different trenchless methods, and then analyzing this data. The sensitivity of PDS rankings with respect to trenchless methods is considered in order to evaluate whether similar project delivery systems are equally effective in different trenchless methods. The effectiveness of PDS with respect to attributes is defined as follows: a project delivery system is most effective with respect to an attribute (e.g., ability to control growth in costs ) if there is no project delivery system that is more effective than that PDS. The results of this study may assist trenchless project owners to select the appropriate PDS for the trenchless method selected.

Integrated construction project delivery system in the U.S. public sector: An information modeling framework

Integrated project delivery (IPD) method has recently emerged as an alternative to traditional delivery methods. It has the potential to overcome inefficiencies of traditional delivery methods by enhancing collaboration among project participants. Information and communication technology (ICT) facilitates IPD by effective management, processing and communication of information within and among organizations. While the benefits of IPD, and the role of ICT in realizing them, have been generally acknowledged, the US public construction sector is very slow in adopting IPD. The reasons are - lack of experience and inadequate understanding of IPD in public owner as confirmed by the results of the questionnaire survey conducted under this research study. The public construction sector should be aware of the value of IPD and should know the essentials for effective implementation of IPD principles - especially, they should be cognizant of the opportunities offered by advancements in ICT to realize this.^ In order to address the need an IPD Readiness Assessment Model (IPD-RAM) was developed in this research study. The model was designed with a goal to determine IPD readiness of a public owner organization considering selected IPD principles, and ICT levels, at which project functions were carried out. Subsequent analysis led to identification of possible improvements in ICTs that have the potential to increase IPD readiness scores. Termed as the gap identification, this process was used to formulate improvement strategies. The model had been applied to six Florida International University (FIU) construction projects (case studies). The results showed that the IPD readiness of the organization was considerably low and several project functions can be improved by using higher and/or advanced level ICT tools and methods. Feedbacks from a focus group comprised of FIU officials and an independent group of experts had been received at various stages of this research and had been utilized during development and implementation of the model. Focus group input was also helpful for validation of the model and its results. It was hoped that the model developed would be useful to construction owner organizations in order to assess their IPD readiness and to identify appropriate ICT improvement strategies.^

Integrated Construction Project Delivery System in the U.S. Public Sector: An Information Modeling Framework

Integrated project delivery (IPD) method has recently emerged as an alternative to traditional delivery methods. It has the potential to overcome inefficiencies of traditional delivery methods by enhancing collaboration among project participants. Information and communication technology (ICT) facilitates IPD by effective management, processing and communication of information within and among organizations. While the benefits of IPD, and the role of ICT in realizing them, have been generally acknowledged, the US public construction sector is very slow in adopting IPD. The reasons are - lack of experience and inadequate understanding of IPD in public owner as confirmed by the results of the questionnaire survey conducted under this research study. The public construction sector should be aware of the value of IPD and should know the essentials for effective implementation of IPD principles - especially, they should be cognizant of the opportunities offered by advancements in ICT to realize this. In order to address the need an IPD Readiness Assessment Model (IPD-RAM) was developed in this research study. The model was designed with a goal to determine IPD readiness of a public owner organization considering selected IPD principles, and ICT levels, at which project functions were carried out. Subsequent analysis led to identification of possible improvements in ICTs that have the potential to increase IPD readiness scores. Termed as the gap identification, this process was used to formulate improvement strategies. The model had been applied to six Florida International University (FIU) construction projects (case studies). The results showed that the IPD readiness of the organization was considerably low and several project functions can be improved by using higher and/or advanced level ICT tools and methods. Feedbacks from a focus group comprised of FIU officials and an independent group of experts had been received at various stages of this research and had been utilized during development and implementation of the model. Focus group input was also helpful for validation of the model and its results. It was hoped that the model developed would be useful to construction owner organizations in order to assess their IPD readiness and to identify appropriate ICT improvement strategies.

Reversibly Photo-Crosslinkable Polycarbonate-Based Polymersomes for Drug Encapsulation and Delivery

This thesis describes the preparation of polymersomes from poly(ethylene glycol)-block-polycarbonate (PEG-PC) copolymers functionalized with pendant coumarin groups. Coumarin groups undergo photo-reversible dimerization when irradiated with specific ultraviolet wavelengths, so they can be used to prepare polymers with photo-responsive properties. In this case, the pendant coumarin groups enable stabilization of the polymersome membrane through photo-crosslinking of the hydrophobic block. Initially, several novel cinnamoyl and coumarin functionalized cyclic carbonate monomers were synthesized using ester, ether, or amide linkages. While the homopolymerization of these functionalized monomers proved challenging due to their high melting points, both cinnamoyl and coumarin functionalized monomers were successfully copolymerized with trimethylene carbonate (TMC) at 100 ℃ using a catalyst-free melt polymerization process where the TMC doubled as a solvent for the higher melting point monomer. Using this system, polycarbonate copolymers with up to 33% incorporation of the functionalized monomers were prepared. In addition, an investigation of some anomalous polymerization results identified previously unreported triethylamine-based catalysts for the melt polymerization of carbonate monomers. These studies also demonstrated that the catalyst-free polymerization of TMC occurs faster and at lower temperatures than previously reported. Subsequently, the photo-crosslinking of cinnamoyl and coumarin functionalized polycarbonates was compared and coumarin was identified as the more effective crosslinking agent when using 300-400 nm UV. An investigation of the photo-reversibility of the coumarin dimerization revealed no discernible change in the properties of crosslinked networks, but rapid photo-reversion in dilute solutions. The photo-crosslinking and photo-reversion kinetics of the coumarin functionalized polycarbonates were determined to be second-order in both cases. Finally, the self-assembly of PEG-PC diblock copolymers functionalized with coumarin was examined and both reverse solvent evaporation and solvent displacement were found to induce self-assembly, with hydrophilic mass fractions (f-factors) of 12-28% resulting in the formation of solid microparticles and nanoparticles and f-factors of 33-43% resulting in the formation of polymersomes. The stabilization of these polymersome membranes through photo-initiator-free photo-crosslinking was demonstrated with the crosslinking allowing polymersomes to withstand centrifugation at 12,000 x g. In addition, the encapsulation of calcein, as a model small molecule drug, in the stabilized polymersomes was successfully demonstrated using confocal microscopy.

Design and fabrication of electro-thermal microcantilevers for ultrafast molecular sorting and delivery

Improvements to the current state of the art in microfabricated cantilevers are investigated in order to realize enhanced functionality and increased versatility for use in ultrafast electrophoretic molecular sorting and delivery. Design rationale and fabrication process flow are described for six types of electro-thermal microcantilevers. Devices have been tailored for the process of separating mixtures of heterogeneous molecules into discrete detectable bands based on electrophoretic mobility, and delivering them to a conductive substrate using electric fields. Four device types include integrated heating elements capable of warming samples to catalyze reactions or cleaning the device for reuse. Similar devices have been shown to be capable of targeting temperatures between ambient conditions and the melting point of silicon, to within 0.1˚C precision or better. All microcantilevers types are equipped with a highly doped conductive silicon tip capable of interacting with a conductive substrate to deliver molecules under the presence of an electric field. Devices are equipped with additional electrodes to aid in sorting molecules on the surface of the probe end. Two designs contain two legs and one additional sorting electrode while four designs contain three legs and have two sorting electrodes. Devices having two sorting electrodes are designed to be capable of sorting three or more molecular species, a distinctive advancement in the state of the art. A detailed process flow of the fabrication process for all six electro-thermal cantilever designs are explained in detail.

Mobile marketing process of an international event – Case: Falun2015 FIS Nordic World Ski Championships

Any other technology has never affected daily life at this level and witnessed as speedy adaptation as the mobile phone. At the same time, mobile media has developed to be a serious marketing tool for all kinds of businesses, and the industry has grown explosively in recent years. The objective of this thesis is to inspect the mobile marketing process of an international event. This thesis is a qualitative case study. The chosen case for this thesis is the mobile marketing process of Falun2015 FIS Nordic World Ski Championships due to researcher’s interest on the topic and contacts to the people around the event. The empirical findings were acquired by conducting two interviews with three experts from the case organisation and its partner organisation. The interviews were performed as semi-structured interviews utilising the themes arising from the chosen theoretical framework. The framework distinguished six phases in the process: (i) campaign initiation, (ii) campaign design, (iii) campaign creation, (iv) permission management, (v) delivery, and (vi) evaluation and analysis. Phases one and five were not examined in this thesis because campaign initiation was not purely seen as part of the campaign implementation, and investigating phase five would have required a very technical viewpoint to the study. In addition to the interviews, some pre-established documents were exploited as a supporting data. The empirical findings of this thesis mainly follow the theoretical framework utilised. However, some modifications to the model could be made mainly related to the order of different phases. In the revised model, the actions are categorised depending on the time they should be conducted, i.e. before, during or after the event. Regardless of the categorisation, the phases can be in different order and overlapping. In addition, the business network was highly emphasised by the empirical findings and is thus added to the modified model. Five managerial recommendations can be concluded from the empirical findings of this thesis: (i) the importance of a business network should be highly valued in a mobile marketing process; (ii) clear goals should be defined for mobile marketing actions in order to make sure that everyone involved is aware them; (iii) interactivity should be perceived as part of a mobile marketing communication; (iv) enough time should be allowed for the development of a mobile marketing process in order to exploit all the potential it can offer; and (v) attention should be paid to measuring and analysing matters that are of relevance

The Applicability Of Ordered Mesoporous Sba-15 And Its Hydrophobic Glutaraldehyde-bridge Derivative To Improve Ibuprofen-loading In Releasing System.

The mesoporous SBA-15 silica with uniform hexagonal pore, narrow pore size distribution and tuneable pore diameter was organofunctionalized with glutaraldehyde-bridged silylating agent. The precursor and its derivative silicas were ibuprofen-loaded for controlled delivery in simulated biological fluids. The synthesized silicas were characterized by elemental analysis, infrared spectroscopy, (13)C and (29)Si solid state NMR spectroscopy, nitrogen adsorption, X-ray diffractometry, thermogravimetry and scanning electron microscopy. Surface functionalization with amine containing bridged hydrophobic structure resulted in significantly decreased surface area from 802.4 to 63.0 m(2) g(-1) and pore diameter 8.0-6.0 nm, which ultimately increased the drug-loading capacity from 18.0% up to 28.3% and a very slow release rate of ibuprofen over the period of 72.5h. The in vitro drug release demonstrated that SBA-15 presented the fastest release from 25% to 27% and SBA-15GA gave near 10% of drug release in all fluids during 72.5 h. The Korsmeyer-Peppas model better fits the release data with the Fickian diffusion mechanism and zero order kinetics for synthesized mesoporous silicas. Both pore sizes and hydrophobicity influenced the rate of the release process, indicating that the chemically modified silica can be suggested to design formulation of slow and constant release over a defined period, to avoid repeated administration.

Preparation, Characterization And In Vitro Digestibility Of Gellan And Chitosan-gellan Microgels.

Gellan microgels with potential application in delivery systems were obtained by physically cross-linked gellan gum. The microgels were produced by atomization followed by ionotropic gelation using CaCl2 (gellan/Ca) or KCl (gellan/K) as hardening agent and part of them were coated with chitosan in order to improve their resistance to gastric digestion. Size distribution, morphology and zeta potential of microgels were evaluated before and after in vitro digestion process. The long term stability was also evaluated. Spherical microparticles were obtained at gellan concentration above 0.6% w/w, showing average size among 70-120 μm. Most of the coated and uncoated microgels showed stability in aqueous media, except the uncoated gellan/K microgel. The in vitro digestion evaluation showed that all particles maintained their size and shape after the gastric digestion step. However, the enteric digestion caused disintegration of microgels indicating their potential application for enteric delivery systems. The chitosan-coated microgels showed lower degree of fragmentation when compared to the uncoated microgels, indicating that the coating process enable a better control of microgels releasing properties during the enteric digestion.

Delivery Of A Urology Online Course Using Moodle Versus Didactic Lectures Methods.

To subjectively and objectively compare an accessible interactive electronic library using Moodle with lectures for urology teaching of medical students. Forty consecutive fourth-year medical students and one urology teacher were exposed to two teaching methods (4 weeks each) in the form of problem-based learning: - lectures and - student-centered group discussion based on Moodle (modular object-oriented dynamic learning environment) full time online delivered (24/7) with video surgeries, electronic urology cases and additional basic principles of the disease process. All 40 students completed the study. While 30% were moderately dissatisfied with their current knowledge base, online learning course delivery using Moodle was considered superior to the lectures by 86% of the students. The study found the following observations: (1) the increment in learning grades ranged from 7.0 to 9.7 for students in the online Moodle course compared to 4.0-9.6 to didactic lectures; (2) the self-reported student involvement in the online course was characterized as large by over 60%; (3) the teacher-student interaction was described as very frequent (50%) and moderately frequent (50%); and (4) more inquiries and requisitions by students as well as peer assisting were observed from the students using the Moodle platform. The Moodle platform is feasible and effective, enthusing medical students to learn, improving immersion in the urology clinical rotation and encouraging the spontaneous peer assisted learning. Future studies should expand objective evaluations of knowledge acquisition and retention.