Tuning and targeting semiconducting polymer nanoparticles to enhance in vivo photoacoustic imaging

Photoacoustic imaging (PAI) is a promising imaging modality which combines high spatial resolution with excellent contrast generation. To fulfil the potential of using PAI in clinical settings for cancer detection, the development of novel contrast agents with strong absorptions in the near infrared and high tumour specificity in vivo is required. Semiconducting polymer nanoparticles (SPNs) encapsulating organic semiconducting polymers in lipid nanoparticles have emerged as excellent candidates for photoacoustic (PA) contrast generation: they retain the polymer’s ability to generate high PA contrast, and the lipid formulation grants SPNs excellent physiological properties. However, these SPNs rely so far on the enhanced retention and permeability (EPR) effect for tumour accumulation. The reliability of this passive mode of accumulation in clinical settings has been recently called into question. To address this, the formulation of targeted SPNs using EGFR-targeting peptides was explored. SPNs based on novel semiconducting polymers as well as commercially available polymers were formulated via the mini-emulsion and nanoprecipitation methods. Lipid formulations included PEGylated lipids as well as functional PEG lipids on which N-terminal Cysteine-modified EGFR-targeting peptides were conjugated via the thiol-maleimide Michael addition. The synthetic accessibility of both the pre- and post-formulation functionalisation strategies was assessed in Chapter 3. To quantify surface functionalisation, a novel NMR characterisation strategy for the routine quantification of maleimide moieties tethered to the surface of nanoparticles was proposed. To compare the targeting efficiency of EGFR-targeting peptides A-R, D4 and GE11, a library of peptide-dye conjugates was constructed using fluorescein-5-maleimide. This library included scrambled controls and short PEG spacers introduced between the targeting sequence and the cysteine residue. The synthesis of these conjugates is described in Chapter 4. While synthesising these peptides, preliminary data supporting the intramolecular transcyclisation of the thiol-maleimide adducts was obtained. The intramolecular transcyclisation of thiol-maleimide adducts is underreported in the literature and clinically relevant. The thiazine rearrangement products are protected from thiol-exchange which, in physiological conditions, can lead to the partial loss of functionality of targeting ligands synthesised using the thiol-maleimide reaction