Targeting of a Photosensitizer to the Mitochondrion Enhances the Potency of Photodynamic Therapy

Abstract

Photodynamic therapy (PDT) involves use of a photosensitizer, whose activation with light leads to the production of singlet oxygen (SOS), generation of reactive oxygen species (ROS), and initiation of associated cell toxicity. Because a cell's mitochondria constitute sites where oxygen levels are high, ROS can be readily produced, and apoptosis is commonly initiated. Therefore, an ideal PDT agent might be a potent photosensitizer that could naturally accumulate in mitochondria. Although a number of mitochondria-targeting moieties, including triphenylphosphine, guanidinium, and bisguanidium, have been identified, a quantitative comparison of their efficacies in targeting mitochondria has not been performed. In this study, we have prepared triphenylphosphine, guanidinium, and bisguanidium derivatives of the FDA-approved PDT agent verteporfin (Visudyne, benzoporphyrin derivative-monoacid ring A: <b>BPD-MA</b>) and compared their abilities to induce the intracellular perturbations common to potent PDT agents. Cellular parameters examined included subcellular localization of the verteporfin, real-time monitoring of SOS production, quantitation of reactive oxygen species (ROS) generation, analysis of mitochondria and chromatin integrity, characterization of cytoskeletal disruption and evaluation of cytochrome <i>C</i> release as a measure of apoptosis. An analysis of these parameters demonstrates that the triphenylphosphine derivative (<b>0323</b>) has better mitochondria-targeting efficacy, SOS production, and mitochondria membrane toxicity than either unmodified verteporfin or its guanidinium derivatives. Consistent with this potency, <b>0323</b> also induced the most prominent mitochondria swelling, actin depolymerization, pyknosis, and cytochrome <i>C</i> release. We conclude that triphenylphosphine has a better mitochondria-targeting moiety than guanidinium or bis-guanidinium and those PDT photosensitizers with improved cytotoxicities can be prepared by conjugating a mitochondria-targeting moiety to the desired photosensitizer.