Dziemidowicz, Karolina; (2021) Development of antibody-functionalised biomaterials for cancer immunotherapy. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Protein-loaded biomaterials are generating considerable interest due to the growing importance of immunotherapy and tissue engineering in modern medicine. The complex and fragile structures of many therapeutic proteins require advanced delivery methods and careful optimisation of formulation and manufacturing conditions. Electrohydrodynamic processes (EHD) are material fabrication techniques in which a polymer solution is drawn by the influence of an electrical field to produce solid micro-and nano scale fibres and particles. Multiple approaches have been proposed for the incorporation of therapeutic proteins in electrospun scaffolds, including surface functionalization and coaxial electrospinning. These concepts are introduced in Chapter 1. This PhD thesis explores the EHD fabrication, functionalisation with therapeutically relevant proteins, and characterization of polycaprolactone (PCL) materials, with the aim of developing a platform protein delivery technology. Several protein loading methodologies were investigated on electrospun nanofibres and electrosprayed microparticles for applications relevant to regenerative medicine. The experimental procedures used are detailed in Chapter 2. Chapter 3 explored the feasibility of surface functionalisation of electrospun PCL nanoscaffolds with proteins using perfluorophenyl azide chemistry. Examples of the conjugated biomolecules explored include bovine serum albumin, catalase and antibodies (infliximab and OKT3). The covalently conjugated catalase released from the fibres at a much slower rate than physically adsorbed catalase, revealing this approach to be suitable for safe and effective attachment of proteins to hydrocarbon-based biomaterials. The next application explored is the fabrication of surface-functionalised anti-CD3 antibody-modified electrosprayed PCL microparticles, with the ultimate goal of achieving targeted T cell activation when the particles are injected intratumorally. Formulations were prepared by electrospraying PCL particles and subsequent surface functionalization using perfluorophenylazide chemistry (Chapter 4) and by using “click” chemistry to conjugate protein to azide-functionalised PCL (Chapter 5). The developed formulations were extensively characterised with in vitro T cell activation models. It was found that T cell activation can be achieved following stimulation with biomimetic electrosprayed microparticles prepared using either of the explored bioconjugation methods. The final results chapter, Chapter 6, discusses the fabrication and characterisation of electrospun PCL patches loaded with a checkpoint inhibitor monoclonal antibody, ipilimumab. The coaxial electrospinning technology offers a simple solution for the fabrication of antibody-loaded biocompatible scaffolds that can be easily implanted at the desired site of action and release the therapeutic cargo in a sustained fashion. It was also found that electrospinning monoclonal antibodies near to their isoelectric point leads to improved process stability and enhanced protein encapsulation.

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