Biomimetic coacervates for drug delivery

• Main Author: Lim, Zhi Wei
• Other Authors: Ali Gilles Tchenguise Miserez
• Format: Thesis-Doctor of Philosophy
• Language: English
• Published: Nanyang Technological University 2020
• Subjects: Engineering::Materials::Biomaterials; Science::Medicine::Pharmacy::Pharmaceutical technology
• Online Access: https://hdl.handle.net/10356/137527
• Institution: Nanyang Technological University

Biomimetics has provided inspirations to design innovative materials and advanced technologies. The squid beak of Dosidicus gigas, commonly known as Humboldt squid is a fascinating instance, being a “biotool” with remarkable gradient stiffness, yet fully organic and non-mineralised. The four main components of the squid beak are chitin, pigments, water, and proteins. There are two families of proteins within the squid beak, chitin binding beak proteins (DgCBPs) and histidine-rich beak proteins (DgHBPs). DgHBPs have been demonstrated to undergo coacervation. Among the tested recombinant DgHBPs and biomimetic peptides, DgHBP-2 peptide was found to be most suitable as an encapsulation material, due to its ability to coacervate under physiological conditions, its pH and temperature responsive coacervation. With the aim of exploring DgHBP-2 peptide coacervates as drug delivery systems, characterisation of the coacervates was first done to understand its properties and limitations. Different crosslinking methods were also developed with the aim of stabilising these coacervates. The next study attempts to incorporate DgHBP-2 peptide coacervates into Glucose Responsive Insulin Delivery System (GRIDS). In this GRIDS, Glucose oxidase (GOx) and insulin were both loaded into DgHBP-2 peptide coacervates. GOx functions as a glucose sensor while the coacervates acted as an insulin reservoir. GOx hydrolyses glucose into gluconic acid, bringing about a localised acidic microenvironment that causes the disassembly of coacervates and release of insulin. The encapsulation efficiency of insulin by DgHBP-2 peptide coacervates was more 99% and was found to be cytocompatible. In vitro release assays have shown that the amount and rate of insulin release were influenced by the surrounding glucose concentration, with higher glucose concentration triggering faster release of insulin. This coacervate-based GRIDS could also respond to varying glucose concentrations that mimics blood glucose fluctuations within human body. In addition, the structure of insulin released from coacervates was near identical to the native insulin, indicating that its bioactivity was not affected. Together, these results have shown that coacervate-based GRIDS can be a potential insulin replacement therapy for diabetes. With the successful GRIDS fabrication, the use of coacervates for thermo-chemotherapy for liver cancer was explored next. The effectiveness of chemotherapies in liver cancer are often limited by its severe toxicity and hence chemotherapeutic drugs e.g. Doxorubicin (Dox) should be targeted towards the tumour site. Furthermore, hyperthermia treatments have been shown to improve the efficacy of chemotherapy, decreasing the need to use high concentrations of chemotherapeutic drugs. Magnetic nanoparticles and Dox were co-encapsulated within DgHBP-2 coacervates, followed by crosslinking with 4-methylcatecol and sodium periodate. These Dox-loaded Magnetic coacervates (DMCs) were efficiently uptaken into HepG2 liver cancer cells through an ATP- and endocytosis-independent pathway. Application of an external Alternating current Magnetic Field (AMF) on DMCs increased the temperature to 45oC within the coacervates, destabilizing the droplets and triggering the release of Dox. In vitro studies on DMCs-treated HepG2 cells have shown that they were cytocompatible and capable of triggering cell death under AMF. Moreover, thermo-chemotherapy treatments by DMCs were found to be more effective than either hyperthermia or chemotherapy treatments alone. This thesis highlights the use of biomimetic peptide coacervates as a novel natural encapsulation material. By performing in vitro studies on DgHBP-2 peptide coacervates, their integration into drug delivery system was examined more comprehensively, from the encapsulation of therapeutics to release assays and cytotoxic studies. The experiments presented in this thesis provide proof-of-concept validation that stimuli-responsive coacervate microdroplets can be used environmental-responsive drug delivery systems.