This study was conducted to formulate a nonviral delivery system for the delivery of small interfering RNA (siRNA) to B16 melanoma cells in vitro. For this purpose, oleic and stearic acid modified derivatives of branched polyethylenimine (PEI) were prepared and evaluated. The hydrophobically modified polymers increased siRNA condensation up to 3 folds as compared to the parent PEI. The modified PEIs exhibited up to 3-fold higher siRNA protection from degradation in fetal bovine serum as compared to the parent PEI. The formulated complexes were shown to enter B16 cells in a time-dependent fashion, reaching over 90% of the cells after 24 h, as compared to only 5% of the cells displaying siRNA uptake in the absence of any carrier. A proportional reduction in siRNA cell uptake was observed with reduced polymeric content in the formulations. When used to deliver various doses of siRNA to B16 cells, the modified PEIs were superior or comparable to some of the commercially available transfection agents; the hydrophobically modified polymers gave 3-fold increased siRNA delivery than the parent PEI, approximately 5-fold higher delivery than jetPEI and Metafectene, a comparable delivery to Lipofectamine 2000, but a 1.6-fold decreased delivery compared to INTERFERin, which was the most efficient reagent in our hands. Using an siRNA specific for integrin alpha(v), a dose-dependent decrease in integrin alpha(v) levels was demonstrated in B16 cells by flow cytometry, revealing a more pronounced reduction of integrin alpha(v) levels for oleic- and stearic-acid modified PEIs. The overall results suggested that the hydrophobically modified PEIs provide a promising delivery strategy for siRNA therapeutic applications.