ABSTRACT
Three new copper(II) complexes containing two different pyrazole bound ligands (1, 2) have been synthesized and characterized by IR, LSI-MS (liquid secondary ion mass spectrometry) and elemental analysis. (1)H NMR spectra of the organic ligands have been recorded. We describe the influence of these complexes on particular cancer cell lines and DNA structure by MTT-assay [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], APA (acid phosphatase activity)-assay or CD-spectroscopy and agarose gel electrophoresis methods, together with their physico-chemical properties such as lipophilicity and stability in aqueous solution. The cytotoxic effect on HUVEC (endothelial cells) for the most active complex 4 has been also investigated. Moreover, the ability of these complexes to induce apoptosis in cancer cells has been assessed by using fluorescence microscopy. Our results indicate that dichloridobis{1-[amino(thioxo)methyl]-5-hydroxy-3-phenyl-1H-pyrazole-κN2}copper(II) is the most potent complex among the tested complexes.
Subject(s)
Antineoplastic Agents/chemical synthesis , Coordination Complexes/chemical synthesis , Copper/chemistry , Pyrazoles/chemistry , Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Coordination Complexes/pharmacology , Drug Screening Assays, Antitumor , Drug Stability , Human Umbilical Vein Endothelial Cells/drug effects , Human Umbilical Vein Endothelial Cells/physiology , Humans , Inhibitory Concentration 50ABSTRACT
INTRODUCTION: The toxic influence of photosensitizers in the dark is poorly investigated. In our study we used the photosensitizers liposomal meso-tetrahydroxyphenyl chlorin derivative (Foslipos(®)) and hypericin as well as their 1:1 combination on two different head and neck squamous cell carcinoma (HNSCC) cell lines (UMB-SCC 745 and UMB-SCC 969). MATERIALS AND METHODS: We examined uptake, efflux and localization of the photosensitizers with confocal microscopy. Fluorescence quantification was measured with a micro-plate spectrometer. Special interest was given to effects on cell proliferation (BrdU proliferation assay), RNA quality (Bioanalyzer measurements) and DNA damage (comet assays) in the dark. RESULTS: Foslipos(®) uptake was linear over time and its efflux was not achieved even after 24 h while uptake of hypericin reached a plateau after 5 h and was almost eliminated after 24 h. Localization of Foslipos(®) was organelle-unspecific. Hypericin was found mainly at membranes and in trans-golgi network. Foslipos(®) treated cells showed cell toxicity for the highest concentration (10 µg/mL). In contrast, hypericin was toxic for all concentrations (10-0.6 µg/mL). The photosensitizer combination was non-toxic for all concentrations (10-0.6 µg/mL). No changes in RNA quality were monitored. Initial DNA damage was found only in hypericin treated UMB-SCC 745, which recovered after 3h. No significant DNA damage was found for UMB-SCC 969. CONCLUSION: Our data shows that the combinatorial application decrease photosensitizer toxicity, which can be advantageous in PDT treatments.
Subject(s)
Darkness , Mesoporphyrins/pharmacology , Neoplasms, Squamous Cell/drug therapy , Perylene/analogs & derivatives , Photochemotherapy/methods , Photosensitizing Agents/pharmacology , Anthracenes , Cell Line, Tumor , DNA Damage/drug effects , Dose-Response Relationship, Drug , Drug Combinations , Humans , Liposomes , Mesoporphyrins/administration & dosage , Perylene/administration & dosage , Perylene/pharmacology , Photosensitizing Agents/administration & dosage , RNA/drug effects , trans-Golgi Network/drug effectsABSTRACT
BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is currently the eighth leading cause of cancer death worldwide. The often severe side effects, functional impairments and unfavorable cosmetic outcome of conventional therapies for HNSCC have prompted the quest for novel treatment strategies, including the evaluation of nanotechnology to improve e.g. drug delivery and cancer imaging. Although silica nanoparticles hold great promise for biomedical applications, they have not yet been investigated in the context of HNSCC. In the present in-vitro study we thus analyzed the cytotoxicity, uptake and intracellular fate of 200-300 nm core-shell silica nanoparticles encapsulating fluorescent dye tris(bipyridine)ruthenium(II) dichloride with hydroxyl-, aminopropyl- or PEGylated surface modifications (Ru@SiO2-OH, Ru@SiO2-NH2, Ru@SiO2-PEG) in the human HNSCC cell line UMB-SCC 745. RESULTS: We found that at concentrations of 0.125 mg/ml, none of the nanoparticles used had a statistically significant effect on proliferation rates of UMB-SCC 745. Confocal and transmission electron microscopy showed an intracellular appearance of Ru@SiO2-OH and Ru@SiO2-NH2 within 30 min. They were internalized both as single nanoparticles (presumably via clathrin-coated pits) or in clusters and always localized to cytoplasmic membrane-bounded vesicles. Immunocytochemical co-localization studies indicated that only a fraction of these nanoparticles were transferred to early endosomes, while the majority accumulated in large organelles. Ru@SiO2-OH and Ru@SiO2-NH2 nanoparticles had never been observed to traffic to the lysosomal compartment and were rather propagated at cell division. Intracellular persistence of Ru@SiO2-OH and Ru@SiO2-NH2 was thus traceable over 5 cell passages, but did not result in apparent changes in cell morphology and vitality. In contrast to Ru@SiO2-OH and Ru@SiO2-NH2 uptake of Ru@SiO2-PEG was minimal even after 24 h. CONCLUSIONS: Our study is the first to provide evidence that silica-based nanoparticles may serve as useful tools for the development of novel treatment options in HNSCC. Their long intracellular persistence could be of advantage for e.g. chronic therapeutic modalities. However, their complex endocytotic pathways require further investigations.