Unravelling the modes of phototoxicity of NIR absorbing chlorophyll derivative in cancer cells under normoxic and hypoxic conditions
摘要
The efficacy of photodynamic treatment (PDT) against deep-seated tumor is hindered by low penetration depth of light as well as hypoxic conditions which prevails in tumor. To overcome this limitation, Near-infrared (NIR) absorbing photosensitizers have been investigated actively. In the present study we evaluated the PDT efficacy of an NIR absorbing chlorophyll derivative ‘Cycloimide Purpurin-18 (CIPp-18)’ in Human Breast carcinoma (MCF-7) and cervical adenocarcinoma (Hela) cells under normoxic and hypoxic conditions. PDT with CIPp-18 (2.0 µM, 3 h) and NIR light (700 ± 25 nm, 0.36–1.4 J /cm2) induced potent phototoxicity in both the cell lines. Under hypoxic conditions, PDT induced ~ 32% and 42% phototoxicity at LD50 and LD70 light dose, respectively, which corresponds to phototoxic dose under normoxia. CIPp-18 in neat buffer (pH 7.4) showed generation of singlet oxygen (1O2) as well as superoxide (O2·–) radicals. Studies on ROS generation in cells using fluorescence probes and the effect of mechanistic probes of 1O2 (Sodium Azide, Histidine, D2O) and free radicals (DMSO, Mannitol, Cyanocobalamin, SOD-PEG) on phototoxicity show that 1O2 plays major role in phototoxicity under normoxia. Whereas, under hypoxic conditions, PDT led to no significant generation of ROS and phototoxicity remained unaffected by cyanocobalamin, a quencher of O2·–. Moreover, CIPp-18 showed localization in cell membrane and PDT led to more pronounced loss of membrane permeability in cells under hypoxia than for normoxia. These results demonstrate that CIPp-18 is suitable for PDT of cancer cells under hypoxia and also suggest that phototoxicity under hypoxia is mediated via ROS-independent contact-dependent mechanism.
Graphical abstract