Abstract: Objective To study the effect of cell division cycle 42(Cdc42) small interfering RNA(siRNA) on the malignant phenotype of human colon cancer cells. Methods The protein expression levels of Cdc42 in five colon cancer cell lines were examined with Western blot. Three Cdc42 sequence-specific small interfering RNA and negative control RNA were synthesized and transfected into Lovo and SW620 cells which have high expression of Cdc42 by Lipofectamine^TM2000. The expression of Cdc42 mRNA and protein were examined with RT-PCR and Western blot respectively after 48h. Cell growth rate was measured by Cell Counting Kit-8. Wound-healing and invasion assays were used to examine the abilities of migration and invasion of the cells, respectively. Results Both RT-PCR and Western blot revealed that Cdc42-siRNA notably down-regulated Cdc42 expression at mRNA levels, especially the Cdc42-siRNA1. Western blot also revealed notably down-regulated Cdc42 expression at protein levels. The proliferation of colon cells was inhibited after iRNA treatment. The migrating and invasive abilities of the cells were also suppressed. Conclusion Cdc42 siRNA could effectively suppress the proliferation, migration and invasion of colon cancer cells. Cdc42 might be a potential target for molecular targeting therapy.