Malignant pleural effusion (MPE) is associated with advanced-stage lung cancer. Vinorelbine (VNR) is a semisynthetic vinca alkaloid with strong antitumor activity in non-small cell lung cancer (NSCLC). The purpose of this study was to evaluate the effect of VNR on the production of MPE associated with lung cancer. To investigate the therapeutic efficacy of VNR in the production of MPE in mice, cells of the murine lung cancer cell line, 3LL, were injected into the pleural space of mice, and VNR was administered once intravenously. Treatment with VNR almost completely inhibited tumor growth and MPE production. In addition, immunohistochemical staining revealed that neovascularization and vascular endothelial growth factor (VEGF) expression were markedly decreased in VNR-treated tumors compared with those of the control tumors. Treatment of 3LL cells with high concentrations of VNR (5 or 10 nM) significantly inhibited cell proliferation in vitro. Interestingly, low concentrations of VNR (1 or 2.5 nM) did not affect 3LL cell proliferation, but markedly reduced VEGF expression in these cells. These results suggest that VNR may be effective for the treatment of MPE associated with lung cancer not only by its cytotoxic effect but also through down-regulation of VEGF expression in lung cancer cells.