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Purpose: Control of cancer metastasis is one of the most important issues in cancer treatment. We previously demonstrated that carbon particle irradiation suppresses the metastatic potential of cancer cells, and many studies have reported that photon irradiation promotes it. The purpose of this study was to investigate the effect of carbon beam on non–small-cell lung cancer (NSCLC) cell aggressiveness and gene expression. Methods and Materials: A549 (lung adenocarcinoma) and EBC-1 (lung squamous cell carcinoma) cells were treated with 290 MeV/nucleon carbon ion beam at the Heavy Ion Medical Accelerator in Chiba or with 4-MV X-ray at Osaka University. We tested proliferative, migratory, and invasive activities by cell proliferation assay, Boyden chamber assay, and Matrigel chemoinvasion assay, respectively. cDNA microarray and reverse transcription polymerase chain reaction were also performed to assess mRNA expression alteration. Results: X-irradiation increased cell proliferation of A549 cells at 0.5 Gy, whereas high-dose X-ray reduced migration and invasion of A549 cells. By contrast, carbon beam irradiation did not enhance proliferation, and it reduced the migration and invasion capabilities of both A549 and EBC-1 cells more effectively than did X-irradiation. Carbon beam irradiation induced alteration of various gene expression profiles differently from X-ray irradiation. mRNA expression of ANLN, a homologue of anillin, was suppressed to 60% levels of basal expression in carbon beam–irradiated A549 cells after 12 h. Conclusion: Carbon beam effectively suppresses the metastatic potential of A549 and EBC-1 cells. Carbon beam also has different effects on gene expressions, and downregulation of ANLN was induced only by carbon beam irradiation. [Copyright &y& Elsevier]