Fluorescence Lifetime of PDT Photosensitizers

Abstract

<p> Photodynamic therapy (PDn is an effective treatment option for various types of invasive tumors, the efficacy of which depends strongly on selective cell uptake and selective excitation of the tumor, which relies on proper dosage. The characterization of the fluorescence lifetimes of photosensitizers localized inside living cells may provide the basis for further investigation of in vitro PDT dosage measurements using time-domain spectroscopy and imaging. In this thesis, the fluorescence lifetimes of localized Photofrin ® and delta-aminolevulinic acid (ALA) induced protoporphyrin IX (PpiX) were investigated in living MAT-LyLu (MLL) rat prostate adenocarcinoma cells. Cells were incubated with the photosensitizers, and then treated with light under well-oxygenated conditions using a two-photon fluorescence lifetime imaging microscope (FLIM). Fluorescence lifetime images of these cells were recorded with average lifetimes of 5.5 ± 1.2 ns for Photofrin and 6.3 ± 1.2 ns for ALA-induced PpiX over 600 to 750 nm. Two channel FLIM revealed lifetimes of7.8 ± 0.5 ns for Photofrin® and 10.8 ± 1.7 ns for PpiX over 620 to 645 nm, while photoproducts observed on the second channel yielded lifetimes of 5.1 ± 0.4 ns over 650 to 670 nm for Photofrin® and 6.3 ± 1.0 ns over 670 to 690 nm for PpiX. Fluorescence lifetimes of both photosensitizers were found to be significantly shorter when localized in cells than when measured in solutions, suggesting that photosensitizers' lifetimes go through significant changes when bonded to intracellular components. These changes in lifetime also provide opportunities to quantitatively measure and monitor the binding states of the photosensitizers and their microenvironment, which may be used in real-time PDT dosimetry, as well as for diagnostic purposes. </p>