The simple ganglioside GM3 increases cyclin-dependent kinase (CDK) inhibitor (CKI) p21WAF1 expression through the accumulation of p53 protein by the PTEN-mediated inhibition of the PI-3K/AKT/MDM2 survival signaling in HCT116 colon cancer cells. GM3 induces p53-dependent transcriptional activity of p21WAF1, as evidenced by the p21WAF1 promoter-driven luciferase reporter plasmid (full-length p21WAF1 promoter and a construct lacking the p53 binding sites). Additionally, ganglioside GM3 enhanced expression of CKI p27kip1 through the PTEN-mediated inhibition of the PI-3K/AKT signaling. Furthermore, the down-regulation of the cyclin E and CDK2 was clearly observed in GM3-treated HCT116 cells, but the down-regulation of cyclin D1 and CDK4 was not. Whereas, suppression of PTEN levels by RNA interference restores the enhanced expression of p53-dependent p21WAF1 and p27 kip1 through inactivation of PTEN-mediated PI-3K/AKT signaling by ganglioside GM3. We conclude that GM3 represents a modulator of cancer cell proliferation and may have potential for use in colorectal cancer therapy.