Abstract
Cancer is a devastating disease responsible for the destruction of millions of lives worldwide, despite the availability of numerous types of treatments. One example of such a treatment is photodynamic therapy (PDT). PDT is a two-step cancer treatment which involves the administration of a photosensitizer (PS) which accumulates in cancerous tissue. This is followed by selective illumination of the tissue with light of a particular wavelength to activate the PS. The activated PS will result in the production of reactive oxygen species (ROS), responsible for induction of cellular damage and destruction of the cancerous tissue. One type of cancer with the potential to be treated with PDT is oesophageal cancer (OC), a particularly aggressive type of cancer. OC holds a benefit for treatment with PDT as it permits easy access for endoscopic tumour illumination with laser light. This study aimed to investigate the effect of the addition of a metal ion to the macrocycle core of H2THPP and to assess the effectiveness of these two novel derivatives, Ag(I)-THPP and Pd(II)-THPP, as potential PSs for PDT of OC. Uptake and localisation of H2THPP, Ag(I)-THPP and Pd(II)-THPP in SNO cells were determined through fluorescence microscopy, while dark toxicity was assessed through analysis of their effect on cell mitochondrial activity. Following irradiation with visible light of 405 nm, the phototoxic effects of Ag(I)-THPP and Pd(II)-THPP were assessed using light microscopy and various homeostatic and cytotoxic assays for mitochondrial activity, cellular respiration, production of ROS and the presence of membrane damage. PI fluorescence was used to look for the presence of cell cycle arrest, while flow cytometry, Hoechst staining of DNA and Western blotting was used for indications of potential induction of cell death. H2THPP showed a high dark toxicity towards SNO cells when prepared as a naked drug dissolved in organic solvents, while displaying signs of compound aggregation when encapsulated in PEG2000-DSPE vesicles, which made this compound unsuitable for further biological use. However, Ag(I)-THPP and Pd(II)-THPP showed improved dark toxicity and more stable formulations when encapsulated in PEG2000-DSPE vesicles, compared to their parent counterpart. At their respective IC50 and IC80 treatment conditions, both Ag(I)-THPP and Pd(II)-THPP resulted in a loss of membrane integrity, reduction in mitochondrial activity, sharp decline in ATP levels, without a significant increase in extracellular levels of LDH, compared to an untreated control population. Ag(I)-THPP and Pd(II)-THPP photodynamic treatments also resulted in an increase in cells with sub-G1 DNA content, externalisation of phosphatidylserine as well as chromatin condensation and marginilisation, without the activation of caspase 3, 7 or 8. The phototoxic effects of Ag(I)-THPP and Pd(II)-THPP were comparable, though the concentration of Pd(II)-THPP required to induce these effects, was much less than that of Ag(I)-THPP. However, despite a higher phototoxicity displayed by Pd(II)-THPP; both compounds show early promise and would be worthy of further investigation as a potential PS for PDT.
M.Sc. (Biochemistry)