mTHPC-mediated photodynamic therapy (PDT): how do drug uptake and distribution influence tumour and normal tissue response?
09 / 2000 - 09 / 2004
Clinical protocols for PDT are based on the assumption that optimum intervals between photosensitiser administration and illumination are times at which there is a maximum differential between drug uptake in tumour and surrounding normal tissue. PDT mediated tumour destruction can, however, occur both via direct cell killing and via vascular mediated damage. The hypothesis to be tested in this project is that drug exposure of endothelial cells in vessels feeding the tumour, and the resultant vascular damage, are more important determinants of PDT response than the tumour drug levels. Increased understanding of the relationship between pharmacokinetics and PDT efficacy should lead to optimisation, and possibly individualisation, of clinical protocols. The purpose of this study is to investigate the relationship between uptake, distribution and clearance of the sensitiser meta-tetrahydroxyphenylchlorin (mTHPC) and efficacy of PDT in the following systems. 1) Human endothelial cells, fibroblasts and tumour cell lines in vitro. 2) Human tumour xenografts in nude mice and normal mouse skin. Vascular damage after PDT will also be assessed using specific perfusion and hypoxia markers. 3) Patients with multiple basal cell carcinomas (BCC). Additional information on plasma pharmacokinetics and normal tissue damage (skin test patches) will also be obtained from patients undergoing PDT for malignant mesothelioma or oral cavity tumours. RESULTS OBTAINED TO DATE: 1) Cellular uptake of mTHPC in vitro increased linearly with increasing drug concentration (24h incubation, 0.1-5 ?g/ml) in 2 human tumour cell lines (HNXOE, HMESO1) and human endothelial cells (HUVECS). Maximum drug uptake was achieved within 4 h for HUVECS but only 20-40% of maximum was achieved within 4 h in the tumour lines. 2) Plasma clearance of mTHPC in nude mice was bi-exponential, with T1/2 values of 1 and 14 h. Maximum drug uptake was seen in liver, lung, kidney and diaphragm within 3 h, but not until 72 h in skin, tumour, oesophagus and muscle. Tumour and skin response to PDT were maximal for illumination at 3 h after mTHPC and minimal at 48-72 h. There was therefore no correlation between tissue drug levels and phototoxicity. 3) Plasma pharmacokinetic data have been obtained from 4 patients with multiple BCC (2 patients were treated twice) and 6 head and neck cancer patients. BCC tumours have been treated with drug light interval of 12-96h. FUTURE DIRECTIONS: 1) In vitro drug uptake studies will be expanded to include human fibroblast and microvascular endothelial cells, MVECs (suggestion of referee). Changes in intracellular localisation of mTHPC with time will be determined using CLSM. Phototoxicity will be compared for MVECs, fibroblasts and tumour cell lines after incubation times giving 50-100% maximum drug uptake. 2) In vivo tumour and normal tissue response to PDT will be correlated with drug levels in tissue and plasma at the time of illumination (results already available). Changes in drug distribution and localisation with time will be evaluated from fluorescence levels in cryostat sections. Vascular damage for illumination at increasing times after sensitisation will be evaluated using specific perfusion and hypoxic markers and using new non-invasive techniques. 3) Clinical pharmacokinetic studies will continue in BCC and head and neck cancer patients undergoing PDT. Drug levels in plasma and tumour at the time of illumination will be correlated with PDT response (BCC patients). Normal tissue toxicity will be evaluated by illumination of skin patches at 24-96h after mTHPC (head and neck patients) and these results will be correlated with drug levels in skin and plasma at the time of illumination.