Three mesostructured silica-type carriers, MCM-41 and MCM-41 functionalized by a postsynthesis grafting procedure with hydrophilic aminopropyl groups (MCM-APTES) and hydrophobic vinyl moieties (MCM-VTES), respectively, were investigated in order to elaborate drug delivery systems (DDS) for irinotecan molecules. All studied drug delivery systems exhibited higher cytotoxicity on murine embrionary fibroblastic (MEF) cells than free irinotecan at the same content of the cytostatic agent, whereas no toxicity was observed for the three unloaded carriers. The cytotoxic effect of irinotecan loaded on MCM-41-type carriers continued to increase even 24 h after ceasing the cell exposure to the drug and remained significantly higher than that of free irinotecan. The cellular uptake of silica-type hybrids was investigated by labelling MCM-APTES with Rhodamine B. In the case of the studied DDS, an endocytotic mechanism was found to be involved in the cell uptake process, and it was used to explain the cytotoxicity differences between free irinotecan and drug loaded on MCM-41-type supports.