UvA-DARE ( Digital Academic Repository ) Evaluation of fluorescence measurement techniques for tumour detection in vivo

Abstract The work described in this thesis focuses on measurement techniques to detect tumour-localising fluorescent dyes, and specifically the Double Ratio (DR) and Normalised Fluorescence Ratio (NFR) techniques developed at the Laser Centre, AMC, Amsterdam. The aim was to investigate how suitable these techniques are for tumour detection in vivo. This was done experimentally as well as theoretically with an analytical model. In chapter 1 the general aspects of tumour detection in vivo using fluorescent dyes are discussed.The work described in this thesis focuses on measurement techniques to detect tumour-localising fluorescent dyes, and specifically the Double Ratio (DR) and Normalised Fluorescence Ratio (NFR) techniques developed at the Laser Centre, AMC, Amsterdam. The aim was to investigate how suitable these techniques are for tumour detection in vivo. This was done experimentally as well as theoretically with an analytical model. In chapter 1 the general aspects of tumour detection in vivo using fluorescent dyes are discussed. In chapter 2 measurements are described which confirm that the DR value is independent of variations in tissue colour. Fluorescence intensity measured on normal moles and normal skin of volunteers after application of a fluorophore was significantly lower in the moles due to absorption. The DR value was equal in all measured tissues which shows that it is independent of the colour of the tissue. This is an advantage for tumour diagnosis on pigmented tissues. Chapter 3 describes pharmacokinetics measurements on normal piglet skin after oral administration of a fluorophore. The pharmacokinetic behaviour of the fluorophore, measured with the DR technique, varied between the piglets. These variations were thought to be caused by anaesthesia. Chapter 4 describes measurements with the DR and NFR techniques on patients with basal cell carcinoma. Both techniques showed increased values on basal cell carcinoma compared to normal skin, which proves that the techniques can be used for tumour detection in vivo. It was concluded that the NFR technique is more suitable for pharmacokinetics measurements than the DR technique. Chapter 5 gives the results of a photodynamic therapy study on basal cell carcinoma related to the measurements presented in chapter 4. Good treatment results were achieved with treatment 1-2 days after low dose fluorophore application (0.1 mg/kg mTHPC). This agreed well with time for maximum tumour to normal fluorophore concentration ratios, found with the NFR technique. In chapter 6 influence of varying tissue parameters, such as tumour depth, optical properties and autofluorescence, on the NFR value is described. This was investigated experimentally in mice and rats with intradermal tumours, and mathematically with an analytical model. The NFR value on uncovered tumours showed a threeto tenfold increase compared to a tumour covered by skin. It was found that even with high fluorophore tumour-selectivity deeper lying tumours cannot be detected if the blood concentration is high or autofluorescence intensity is low. Chapter 7 presents results of a photodynamic therapy study using mTHPC in a rat liver tumour model. Complete tumour remission was found in 87% of the treated animals. The NFR value and fluorophore concentration was measured to show that mTHPC was retained in the tumours. Chapter 8 describes a study on the relation between the NFR value and the fluorophore concentration using the same tumour model as applied in chapter 7. Pharmacokinetics behaviour measured with the two methods agreed reasonably well. Chapter 9 presents a theoretical evaluation of several fluorescence detection techniques Different tissue parameters were varied and their influence on the techniques was studied and summarised. In chapter 10 the presented work is discussed and conclusions are drawn. One important conclusion is that the DR and NFR techniques, in combination with a tumour selective fluorescent dye, were found to be suitable for tumour detection in vivo.