Subcellular localization of Photofrin® determines the death phenotype of human epidermoid carcinoma A431 cells triggered by photodynamic therapy: When plasma membranes are the main targets

Abstract

Photodynamic therapy (PDT) is a kind of photochemo-therapeutic treatment that exerts its effect mainly through the induction of cell death. Distinct types of cell death may be elicited by different PDT regimes. In this study, the mechanisms involved in the death of human epidermoid carcinoma A431 cells triggered by PDT with Photofrin (a clinically approved photosensitizer) were characterized. Photofrin distributes dynamically in A431 cells; the plasma membranes and Golgi complex are the main target sites of Photofrin after a brief (3 h) and prolonged (24 h) incubation, respectively. Cells with differentially localized Photofrin displayed distinct death phenotypes in response to PDT. The effects of PDT on cells with plasma membrane-localized Photofrin were further studied in details. Cells stopped proliferating post PDT at Photofrin dose >7 micro g/ml, and at higher dose (28 micro g/ml) plasma membrane disruption and cell swelling were observed immediately after PDT. Dramatic alterations of several important signaling events were detected in A431 cells post Photofrin-PDT, including (i) immediate formation of reactive oxygen species (ROS), (ii) rapid activation of c-Jun N-terminal kinase, (iii) delayed activation of caspase-3 and cleavage of polyADP-ribose polymerase and p21-activated kinase 2, and (iv) loss of mitochondrial membrane potential. Intriguingly, the characteristics of typical apoptosis such as phosphatidylserine externalization and DNA fragmentation were not detected in the cell death process caused by this PDT regime. In conclusion, our results show that when plasma membranes are the main targets, Photofrin-PDT can lead to instant ROS formation and subsequent activation of downstream signaling events similar to those elicited by many apoptotic stimuli, but the damage of plasma membranes renders the death phenotype more necrosis like.

Chemotherapeutic Approaches for Targeting Cell Death Pathways

“…Upon excitation, the photosensitizers generate ROS (primarily singlet oxygen) that can lead to cell death [146,147]. When photosensitization compounds localize to the plasma membrane, cells die by necrosis resulting from the loss of plasma membrane integrity [148,149]. Activation of photosensitizers on lysosomes may disrupt the lysosomal membrane and result in the release of lysosomal proteases leading to necrosis [150].…”

Photochemically stimulated drug delivery increases the cytotoxicity and specificity of EGF–saporin

“…The susceptibility of TPPS 2a -PDT mediated EGFR damage seems to be cell line Photofrin (Wong et al 2003), 5-ALA induced protoporphyrin IX (PpIX) (Wong et al 2003), Hypericin (de Witte et al 1993) and RB (Zhuang et al 2003, Schieke et al 2004 as PSs may be caused by a direct oxidation of the receptor at light exposure time, as all of these PSs has been shown to localize to the plasma membrane (Thomas & Pardini 1992, Lin et al 2000, Selbo et al 2001a, Hsieh et al 2003b. In addition, the lipophilicity of these PSs indicate that they can freely diffuse through the plasma membrane, and thereby easily target the intracellular domain of EGFR upon light exposure.…”

“…Assefa et al reported that JNK activation rescues cells from hypericin mediated PDT (Assefa et al 1999) while JNK activation after Photofrin-PDT has been shown not to influence on cytotoxicity (Hsieh et al 2003a). P38 is also a stress induced kinase (Zarubin & Han 2005) shown to be activated after PDT with different photosensitizers.…”

Relationship between subcellular localisation of Foscan® and caspase activation in photosensitised MCF-7 cells

“…It has been assumed that photosensitisers primarily localised in mitochondria are able to induce early apoptosis by rapid loss of mitochondrial transmembrane potential and/or release of apoptosis-inducing factors such as cytochrome c (cyt c), itself leading to post-mitochondrial caspase activation (Kessel and Luo, 1999;Chiu and Oleinick, 2001). Several reports investigated the possibility to modulate the mechanism of PDTinduced cell death using protocols that selectively target proapoptotic organelles (Fabris et al, 2001;Hsieh et al, 2003). Prolonged incubation of photosensitisers (24 h) with cells compared with shorter incubation times (2 -3 h) was accompanied by a more efficient apoptosis after photoirradiation.…”

“…Prolonged incubation of photosensitisers (24 h) with cells compared with shorter incubation times (2 -3 h) was accompanied by a more efficient apoptosis after photoirradiation. This was attributed to photosensitiser relocalisation to mitochondria or Golgi apparatus during incubation (Fabris et al, 2001;Hsieh et al, 2003).…”