The cell cycle regulatory pathway responsible for the control of the late-G1 checkpoint is found recurrently altered in human malignant melanoma, often due to lack of functional p16 or pRb (pRb-1) proteins. Here we examined the ability of p16-derived peptides to mimic p16 function in two exemplary human melanoma cell lines: the p16-defective, pRb-positive A375M cells and p16-positive, pRb-defective A2058 cells. The synthetic p16-mimicking peptides strongly induced apoptosis in p16-, pRb+ A375M cells in vitro, while they had significantly less activity on p16+, pRb- A2058 cells. The most active p16-mimicking peptide, p16-AP9, also potently inhibited in vivo growth of the A375M melanoma. Treated tumors showed a threefold smaller volume (P < 0.025) and a significant reduction of the mitotic index and of PCNA expression. Growth of A2058 cells in vivo was not affected by treatment with the p16-mimicking peptide. Our results demonstrate that p16-mimicking peptides can induce apoptosis in vitro and that can inhibit tumor growth in vivo in p16-defective, pRb-expressing human melanoma cells, suggesting that p16-mimicking peptides can represent a promising tool for targeted therapy in selected cancer phenotypes.