Scavenger-receptor targeted photodynamic therapy

Covalent conjugation of a photosensitizer to a ligand that specifically rec ognized and internalized by a cell-surface receptor may be a way of improvi ng the selectivity of photodynamic therapy (PDT). The class A Type-I scaven ger receptor of macrophages, which among other ligands recognizes maleylate d serum albumin and has a high capacity is a good candidate for testing thi s approach, Chlorin(e6) was covalently attached to bovine serum albumin to give conjugates with molar substitution ratios of 1:1 and 3:1 (dye to prote in), and these conjugates could then be further modified by maleylation, A novel way of purifying the conjugates by acetone precipitation was develope d in order to remove traces of unbound dye that could not be accomplished b y size-exclusion chromatography, Conjugates were characterized by polyacryl amide gel electrophoresis and thin-layer chromatography, Photosensitizer up take was measured by target J774 murine macrophage-like cells and nontarget OVCAR-5 human ovarian cancer cells, and phototoxicity was examined after i llumination by a 660 nm diode laser by a tetrazolium assay, All of the puri fied conjugates were taken up by and after illumination killed J774 cells w hile there was only small uptake and no phototoxicity toward OVCAR-5 cells. The higher dye:protein ratio and maleylation of the conjugates both produc ed higher uptakes and lower survival ratios in J774 cells. The uptake and p hototoxicity by J774 cells were decreased after incubation at 4 degreesC de monstrating internalization, and confocal microscopy with organelle-specifi c green fluorescent probes showed largely lysosomal localization. Uptake an d phototoxicity by J774 cells could both be competed by addition of the sca venger receptor ligand maleylated albumin. These data show that scavenger r eceptor-targeted PDT gives a high degree of specificity toward macrophages and may have applications in the treatment of tumors and atherosclerosis.