Background: Isosorbide dinitrate has been broadly used in the treatment of various ischemic heart diseases. Isosorbide is a nitric oxide donor which increases blood flow to tumors through vasodilatation and consequently accelerates the access of chemo-drugs to them. Furthermore, this drug has inhibitory effects on angiogenesis, tumor growth and metastasis in vivo. Moreover, its ant-inflammatory effects have also been reported. In the present study we evaluated the effects of isosorbide on the proliferative activity of fibrosarcoma WEHI-164 cell line and peripheral blood mononuclear cells (PBMCs). Methods: WEHI-164 fibrosarcoma cells and human PBMCs were cultured in complete Roswell Park Memorial Institute (RPMI) 1640 medium with 10% fetal bovine serum and 2×104 cells/mL for WEHI-164 and 2×105 cells/mL for PBMCs. The cells were then incubated at the exponential growth phase with different concentrations of isosorbide (4×10-6-1.6×10-3 M) for 24, 48 and 72 hours. Subsequently, isosorbide effects on proliferation of the cells were evaluated by trypan blue dye exclusion (TB) test and MTT assay. Statistical comparisons between groups were made by analysis of variance. Results: The proliferative activity of WEHI-164 fibrosarcoma cells and human PBMCs treated with different concentrations of isosorbide, did not show any significant difference with untreated control cells. Conclusion: The results of this study showed that isosorbide neither had any significant effects on the proliferative activity of fibrosarcoma WEHI-164 cells nor on human PBMCs. Our findings suggest that anti-tumoral effects of isosorbide reported by other investigators may be mediated through non-cytotoxic mechanisms.