The effect of ZN sulfophthalocyanine and laser irradiation (636 nm) on a lung cancer cell line (A549)

• Main Author: Manoto, Sello Lebohang
• Published: 2009
• Subjects: Cancer - Photochemotherapy; Lungs
• Online Access: http://hdl.handle.net/10210/2998

M. Tech. === Photodynamic therapy (PDT) is a type of phototherapy which is based on the interaction of photosensitizer with visible light in the presence of oxygen resulting in the production of reactive oxygen species (Pfaffel-Schubart et al., 2008). PDT has rapidly matured in the past 6 years and is an accepted standard treatment for various cancerous diseases. The main advantages of PDT include significant drug selectivity in tumour cells, absence of toxicity in the dark, possibility to treat only cancer cells and the ability to retreat a tumour in order to improve the prognosis (Pfaffel-Schubart et al., 2008). Photofrins are the most studied photosensitizers and their disadvantages is the inability of these photosensitizers to localize specifically in tumour cells and are retained in normal cells for prolonged periods (Nowis et al., 2005). This factor has stimulated the development of second generation photosensitizers with improved physical, chemical and spectral properties (Sharman et al., 1999). Phthalocyanine compounds are second generation photosensitizers which have a potential as photosensitizers in PDT treatment of many cancers. Traditional treatments of cancer show limited efficiency and other treatment modalities need to be explored. PDT has shown promising results in the treatment of many cancers using phthalocyanine compounds as photosensitizers but little is known about Zn sulfophthalocyanine on lung cancer cells. This study investigated the effects of Zn sulfophthalocyanine photosensitizer and laser irradiation on a lung cancer cell line (A549). Lung cancer cells were cultured in RPMI medium 1640 supplemented with 10% foetal bovine serum and antibiotics and incubated at 37 °C with 5% CO2 and 85% humidity. Cells were divided into 4 groups. Group 1 was an unirradiated control not treated with a photosensitizer. Group 2 was photosensitized at a concentration of 15.8 μM but not irradiated. Group 3 was irradiated but not photosensitized while Group 4 was irradiated and photosensitized at a concentration of 15.8 μM. Laser irradiations were done using a diode laser emitting 636 nm with an output of 110 mW at 5.3 J/cm2. Cell morphology was evaluated using the light inverted microscope. Cell viability was assessed using adenosine triphosphate (ATP) luminescence assay and Trypan blue exclusion test while cell proliferation was measured using the alamarBlue assay. Cytotoxicity was evaluated by assessing membrane permeability for lactate dehydrogenase and DNA damage was evaluated by comet assay. The mode of cell death was assessed by annexin V-FITC apoptosis detection kit using flow cytometry. In addition, expression of Bcl-2/Bax was monitored by western blot analysis and to determine the levels of induced Heat shock protein 70, Hsp 70 ELISA was used. Cells treated with photosensitizer at 15.8 μM and irradiated with 5.3 J/cm2 showed changes in cell morphology, decrease in cellular viability, proliferation and Hsp 70 expression and an increase in cytotoxicity and DNA damage. This indicated that the metalated phthalocyanine was effective in inducing cell death and the analysis of cell death by annexin FITC-V revealed an apoptotic cell death pattern. Furthermore, loss of the antiapoptotic Bcl-2 protein suggested that there was no inhibition of apoptosis while the absence of proapoptotic Bax suggested that other proapoptotic protein might have played a role in the induction of apoptosis. Untreated cells, irradiated cells and cells treated with photosentitizer alone showed no changes in morphology, increase in cellular viability, proliferation, expression of Bcl2/Bax, and Hsp 70 and a decrease in DNA damage and cell membrane damage. However, treatment of cell with photosensitizer alone caused DNA damage. These results indicate that irradiation or photosensitizer alone has no effect on the A549 cells but photosensitizer alone induces DNA damage.