Biosynthesis of Poly-3-Hydroxybutyrate and its Application in Controlled Drug Delivery

Puthussery, H. 2019. Biosynthesis of Poly-3-Hydroxybutyrate and its Application in Controlled Drug Delivery. PhD thesis University of Westminster School of Life Sciences

TitleBiosynthesis of Poly-3-Hydroxybutyrate and its Application in Controlled Drug Delivery
TypePhD thesis
AuthorsPuthussery, H.

Controlled drug delivery has become a research focus in view of many of the drawbacks of conventional drug delivery including increased biodistribution and lack of target specificity, resulting side effects, and thus decreased patient compliance. Controlled delivery is the sustained administration of therapeutic agents, keeping dosage fluctuations within the therapeutic window. Controlled drug delivery relies on the design and modification of specialised drug delivery systems (DDS) for encapsulating and delivering therapeutic agents. Biodegradable polymers have been exploited extensively for this purpose, since post-administrative procedures for removal of these systems from the body is not a requisite, because their degradation can be catalysed by physiological enzymes and autocatalytic hydrolysis. Particulate DDS such as microspheres and nanospheres have gained prominence in this context, and the optimisation of these to cater to the type of target, therapeutic agent to be encapsulated and the mode of delivery is a growing need.
Polyhydroxyalkanoates (PHAs) are a class of natural polyesters that are receiving great attention in many biomedical applications such as tissue regeneration, synthesis of medical devices and drug delivery. Poly(3-hydroxybutyrate), P(3HB), is a short chain length variety of this polymer which has specifically been recognised for its potential to form DDS. In the first part of this study the solid-in-oil-in-water technique was used to optimise production of P(3HB) microspheres. The optimization process used a response surface methodology based on the Box Behnken design. The effect of process parameters such as polymer concentration, surfactant concentration and stirrer speed on microsphere size was analysed. The results of these optimisation experiments were used to tailor micro and nanospheres of different size ranges and these were characterised for their size distribution, porosity, amount of surface residual surfactant and hydrophobicity.
The second part of this study focused on developing micro and nanospheres with encapsulated therapeutic agents relevant in cardiovascular diseases. Encapsulation of two antiproliferative drugs namely rapamycin and tacrolimus were studied in the context of their application in drug eluting stents. Rapamycin encapsulated P(3HB) and PLLA microspheres were compared for their physical properties and drug release kinetics and their release kinetics were found to be comparable. However, P(3HB) microspheres were more suitable for applications as their degradation products were found to be less acidic than that of PLLA microspheres. Tacrolimus encapsulated micro and nanospheres of P(3HB) were compared, to analyse drug encapsulation and release kinetics with respect to size of the spheres. Microspheres were found to have increased encapsulation efficiency, with a slower drug release rate, in comparison with nanospheres. Vascular Endothelial Growth Factor (VEGF), commonly used in the context of cardiovascular regeneration, was encapsulated within P(3HB) micro and nanospheres and were embedded in collagen scaffolds. The differences in release of VEGF from spheres embedded and not embedded in the collagen scaffold were quantified and it was established that embedding in the scaffolds reduced burst release.
In the next stage of this study, P(3HB) microspheres were developed as novel target specific photodynamic drug delivery systems, with potential anticancer therapy applications. Three derivatives of a commonly used photosensitiser, namely porphyrin, were encapsulated within P(3HB) microspheres. The reduction in cell viability of these with respect to drug loading was quantified. Encapsulation efficiency was found to increase with drug loading and reduction in cell viability was found to be a direct reflection of this. These microspheres were made target specific by adsorbing an antibody ‘Anti- HER-2’, which specifically binds to HER2, a cancer marker overexpressed on breast cancer cells. The incorporation of the antibody was found to result in increased cytotoxicity of the formulations, for all three derivatives.
P(3HB) is a crystalline, hydrophobic polymer and degrades through surface erosion. There are applications that necessitate drug delivery systems that can retain moisture (especially in the context of wound healing) and a quicker release of drug moieties. Bearing this requirement in mind, a conjugate of P(3HB) with hyaluronic acid (HA) was synthesised by Contipro a.s. and the final part of this study examined the optimisation of microsphere synthesis using these conjugates. As this conjugate was amphiphilic, a water in oil emulsion, coupled with crosslinking was employed for the synthesis process. Microspheres of ~20 m diameter were synthesised, and hydrophilic and hydrophobic drugs were encapsulated within them. The microspheres were found to be fast degrading, maintaining their structure up to a maximum of 11 days. The drug release kinetics from these showed an increased burst release compared to P(3HB) microspheres.

PublisherUniversity of Westminster
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Ahmed, D., Puthussery, H., Basnett, P., Knowles, J., Lange, S. and Roy, I. 2021. Controlled Delivery of Pan-PAD-Inhibitor Cl-Amidine Using Poly(3-Hydroxybutyrate) Microspheres. International Journal of Molecular Sciences. 22 (23) e12852.

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