Bioassay-guided fractionation of a CHCl3 extract of the leaves of Xanthium italicum Moretti led to the isolation of four xanthanolides: xanthatin (1), 4-epixanthanol (2), 4-epi-isoxanthanol (3), and 2-hydroxyxanthinosin (4). Their structures were determined by means of 1D and 2D NMR spectroscopy, including 1H-1H COSY, NOESY, HSQC and HMBC experiments, which resulted in complete and unambiguous 1H and 13C NMR chemical shift assignments. The isolated compounds 1-4 were evaluated for their antiproliferative activities, and were demonstrated to exert significant cell growth inhibitory activity against human cervix adenocarcinoma (HeLa), skin carcinoma (A431), and breast adenocarcinoma (MCF7) cells.