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Background: Metastatic melanoma is one of the most aggressive tumours and is also very resistant to current therapeutic approaches. The aim of this investigation was the in vitro study of the anti-proliferative effects of fotemustine (FM; 100 and 250 mu M), bevacizumab (5 mu g/ml) and proton irradiation (12 and 16 Gy) on resistant HTB140 human melanoma cells. Methods: Viability was estimated by sulphorhodamine B assay, while cell proliferation was analyzed by 5-bromo-2-deoxyuridine assay. Cell cycle distribution and apoptosis were examined using flow cytometry. Results: Cell viability and proliferation were reduced after all applied treatments. The level of apoptosis significantly increased after treatment with FM, protons or a combination of all agents, while the apoptotic index ranged from 1.2 to 9.2. Proton irradiation, as well as combined treatment with bevacizumab and protons or 100 mu M FM, bevacizumab and protons, have reduced melanoma cell proliferation through the induction of G1 phase arrest. Single FM (250 mu M) or bevacizumab treatment and their combination, as well as the joint application of these 2 agents with protons, reduced cell proliferation and provoked G2 phase accumulation. Conclusion: The analyzed treatments reduced cell viability and proliferation, triggered G1 or G2 cell cycle phase accumulation and stimulated apoptotic cell death. Copyright (C) 2010 S. Karger AG, Basel