Dual apototic/necrotic response induced by photodynamic treatment with a Zn(II) cationic phthalocyanine in melanoma cells

FEDERICO VALLI; GARCÍA VIOR M. CECILIA; LEONOR P. ROGUIN; JULIETA MARINO

Buenos Aires

Congreso; Reunión Conjunta de Sociedades Biomédicas.; 2017

Reunión Conjunta de Sociedades Biomédicas

Melanoma is an aggressive form of skin carcinoma, highly resistant to traditional therapies. Photodynamic therapy (PDT) is an alternative form of treatment, which combines a photosensitizer, visible light and molecular oxygen to produce reactive oxygen species (ROS) that selectively destroy target cells. In order to find an efficient photosensitizer to be used in melanoma treatment, we evaluated the effect of a sulfur-linked cationic zinc( II) phthalocyanine (Pc13) on a panel of melanoma cells (B16F0, B16F10, WM35, M1/15, A375). Incubation with Pc13 and irradiation, diminished cell viability in a concentration and light dose-dependent manner in all the cell lines studied, with IC50 values ranging from 0.20±0.03 mM to 3.60±0.23 mM for B16F0 and A375 cells, respectively. The most sensitive melanoma cells B16F0 were further employed for studying the mechanisms of cell death triggered by Pc13. Acridine orange/ethidium bromide dual staining showed morphological changes characteristic of both necrosis (42±4%) and apoptosis (21±2%) 3h post irradiation of cells treated with 0.2 μM Pc13. Under these experimental conditions, a significant decrease in the levels of Bcl-2, Bcl-xL and Bid, and a reduction of pro-caspase-3 were observed by Western Blot, evidencing an apoptotic response. Furthermore, a time dependent increase of hypodiploid cell population and cell cycle arrest in G0/G1 were assessed by propidium iodide staining and flow cytometry analysis. In addition, permeabilization of plasma membrane, as sign of necrosis, was evaluated by measuring the release of lactate dehydrogenase (LDH). A light dose and photosensitizer concentration dependent increase of LDH activity was detected in Pc13 treated-cells culture mediums. Taken together, these results indicate that a dual apoptotic and necrotic response is triggered by Pc13 photoactivation in melanoma cells, suggesting that combined mechanisms of cell death could result in a promising alternative for melanoma treatment.