In Vitro Demonstration of the Heavy-Atom Effect for Photodynamic Therapy

Photodynamic therapy (PDT) is an emerging treatment modality for a range of disease classes,
both cancerous and noncancerous. This has brought about an active pursuit of new PDT agents that can
be optimized for the unique set of photophysical characteristics that are required for a successful clinical
agent. We now describe a totally new class of PDT agent, the BF2-chelated 3,5-diaryl-1H-pyrrol-2-yl-3,5-diarylpyrrol-2-ylideneamines (tetraarylazadipyrromethenes). Optimized synthetic procedures have been
developed to facilitate the generation of an array of specifically substituted derivatives to demonstrate how
control of key therapeutic parameters such as wavelength of maximum absorbance and singlet-oxygen
generation can be achieved. Photosensitizer absorption maxima can be varied within the body's therapeutic
window between 650 and 700 nm, with high extinction coefficients ranging from 75 000 to 85 000 M-1
cm-1. Photosensitizer singlet-oxygen generation level was modulated by the exploitation of the heavy-atom effect. An array of photosensitizers with and without bromine atom substituents gave rise to a series
of compounds with varying singlet-oxygen generation profiles. X-ray structural evidence indicates that the
substitution of the bromine atoms has not caused a planarity distortion of the photosensitizer. Comparative
singlet-oxygen production levels of each photosensitizer versus two standards demonstrated a modulating
effect on singlet-oxygen generation depending upon substituent patterns about the photosensitizer. Confocal
laser scanning microscopy imaging of 18a in HeLa cervical carcinoma cells proved that the photosensitizer
was exclusively localized to the cellular cytoplasm. In vitro light-induced toxicity assays in HeLa cervical
carcinoma and MRC5-SV40 transformed fibroblast cancer cell lines confirmed that the heavy-atom effect
is viable in a live cellular system and that it can be exploited to modulate assay efficacy. Direct comparison
of the efficacy of the photosensitizers 18b and 19b, which only differ in molecular structure by the presence
of two bromine atoms, illustrated an increase in efficacy of more than a 1000-fold in both cell lines. All
photosensitizers have very low to nondeterminable dark toxicity in our assay system.