Laboratory Medicine & Pathology

Nowadays different strategies are being introduced in order to enhance photodynamic therapy (PDT) effectiveness, such as combination of PDT with chemotherapy or improvement of photosensitizer (PS) features. A new combined PDTchemotherapy treatment comprising two drugs widespread in clinical research - the hydrophobic zinc(II)-phthalocyanine (ZnPc) as PS and the common chemotherapeutic agent doxorubicin (DOX) - were tested. Cytotoxicity assay showed that this combination remarkably increases the effectiveness of the treatment by inducing a synergistic cell death effect (lower than 10%) when compared to DOX or ZnPc monotherapy (cell surviving around 80%). In addition, annexin-V detection by flow cytometry, analysis of active caspase-3 and cytochrome c by immunofluorescence and time-lapse videomicroscopy corroborated a fine-tunable effect depending on light dose, leading to apoptotic or necrotic mechanism of cell death. Using DCFH-DA (dichlorodihydrofluorescein diacetate) probe, we demonstrate that a significant higher reactive oxygen species generation into cells was the main cause of the synergistic effect of this combined treatment. Further, mammosphere formation efficiency assay showed a reduced breast cancer stem cell activity in established cell line and primary cells obtained from patients, even using DOX at much lower concentration than clinical level. Finally, studies in human breast cancer xenografts indicated a high efficiency also in vivo. All these results provide novel and valuable information that contribute to consider chemophototherapy as a promising tool in current antitumoral treatments, potentially overcoming resistance to cancer chemotherapy and targeting cancer stem cells