BF211,a bufalin (BF) derivative,exhibits stronger anti-cancer activity than BF but with potential cardiotoxicity.Fibroblast activation protein-α (FAPα) is a membrane-bound protease specifically expressed by carcinoma-associated fibroblasts,thus has been used for the selective delivery of anticancer agents.In this study,we used a FAPα-based prodrug strategy to synthesize a dipeptide (Z-GlyPro)-conjugated BF211 prodrug named BF211-03.BF211-03 was hydrolyzed by recombinant human FAPα (rhFAPα) and cleaved by homogenates of human colon cancer HCT-116 or human gastric cancer MGC-803 xenografts.In contrast,BF211-03 showed good stability in plasma and in the homogenates of FAPα-negative normal tissues,such as heart and kidney.In HCT-116 and MGC-803 cells with low levels of FAPα expression,BF211-03 displayed a lower in vitro cytotoxicity than BF211 with approximately 30 to 40-fold larger IC50 values,whereas in human breast cancer MDA-MB-435 cells with high levels of FAPα expression,the IC50 value difference between BF211-03 and BF211 was small (approximately 4-fold).Although the cytotoxicity of BF211-03 against tumor cells was dramatically decreased by the chemical decoration,it was restored after cleavage of BF211-03 by rhFAPα or tumor homogenate.In HCT-116 tumorbearing nude mice,doubling the dose of BF211-03,compared with BF211,caused less weight loss,but showing similar inhibitive effects on tumor growth.Our results suggest that BF211-03 is converted to active BF211 in tumor tissues and exhibits anti-tumor activities in tumor-bearing nude mice.FAPα-targeted BF211-03 displays tumor selectivity and may be useful as a targeting agent to improve the safety profile of cytotoxic natural products for use in cancer therapy.