Gemcitabine-loaded pegylated unilamellar liposomes (200 nm) were proposed for the treatment of multiple myeloma cancer disease. Physicochemical and technological parameters of liposomes were evaluated by using laser light scattering and gel permeation chromatography. The growth-inhibitory activity of gemcitabine-loaded liposomes compared to the free drug was assayed in vitro on U266 (autocrine, interleukin-6-independent) and INA-6 (IL-6-dependent) multiple myeloma cell lines. Liposomes noticeably improved the growth-inhibitory activity of gemcitabine in terms of both dose-dependent and incubation-time effects. Liposomal delivery of gemcitabine consistently and significantly increased induction of apoptosis and caused a complete inhibition of proliferation. Liposomes were able to interact with multiple myeloma cells as demonstrated by confocal laser scanning microscopy and hence to improve the intracellular gemcitabine delivery. Gemcitabine-loaded liposomes were much more effective in vitro than the free drug. This formulation may offer even more in vivo advantages both in terms of drug pharmacokinetic and biodistribution.