Two novel bisindole alkaloids, cyclovinblastine A-B (1-2), together with ten known alkaloids 4-deacetoxycyclovinblastine (3), cycloleurosine (4), leurosine (5), vinblastine (6), leurosidine (7), vinblastine N-b'-oxide (8), isoleurosine (9), 4'-deoxyleurosidine (10), 4'-deoxyleurosidine N-b'-oxide (11) and 4-desacetoxyvinblastine (12) were isolated from the leaves of Catharanthus roseus. The structures of 1 and 2 were established by analysis of their NMR and HR-ESI-MS spectroscopic data. All alkaloids (1-12) were evaluated for their cytotoxic activities against the human hepatocellular carcinoma (HepG2) cell line, human colorectal carcinoma (Lovo) cell line, and human breast carcinoma (MCF-7) cell line by the MTT method in vitro, respectively. The results indicated that cytotoxic activities of alkaloids 9, 10 and 12 were much more potent than those of the positive control vinblastine (6). In addition, the structure-activity relationships (SAR) were conducted on the basis of the cytotoxicity of these isolated alkaloids.