Enhanced apoptotic effect of combined modality of 9-hydroxypheophorbide α-mediated photodynamic therapy and carboplatin on AMC-HN-3 human head and neck cancer cells

Photodynamic therapy (PDT) has been developed as an effective treatment for malignant disease. Carboplatin (CBDCA), a less nephrotoxic analog of cisdiamminedichloroplatinum (cisplatin), has been widely used for the treatment of multiple malignancies. In this study, we investigated the cytotoxic and apoptotic effect of combined modality of 9-hydroxypheophorbide α (9-HPbD)-mediated PDT and CBDCA on AMC-HN-3 human head and neck cancer cell line in vitro. The attached AMC-HN-3 cells were incubated with CBDCA (0.04 mg/ml) for 24 h at 37°C and followed by photosensitization with 9-HPbD for 6 h and laser irradiation with 670 nm diode laser at an intensity of 2.0 J/cm2 for activating 9-HPbD for 15 min. Then MTT reduction assay and Hoechst 33342 and propidium iodide (PI) double staining were used respectively to measure the cytotoxicity and nuclear morphology at 24 h after PDT. Expression of caspase-3, -9 and poly(ADP-ribose) polymerase (PARP) was detected at 0, 3, 6 and 12 h after irradiation through Western blotting techniques. Compared with PDT and CBDCA alone groups, there was more cytotoxicity and enhanced apoptotic cell death in combination groups. The peaked expression of cleaved form of caspase-3, -9 and PARP occurred ≈3 h after PDT. There was stronger expression of cleaved caspase-3, -9 and PARP in combination groups than that in PDT or CBDCA alone groups. This study demonstrates that the combined modality resulted in enhanced apoptotic cell death as well as cytotoxic effect on AMC-HN-3 cells in vitro, which suggests the feasibility of combined modality and the possibility of reducing the effective dosage of 9-HPbD and CBDCA and lowering the side effects on normal cells.