Antimicrobial peptides of the cathelicidin family play a central role in the host defense system. Our group has reported previously that cathelicidin-related or cathelicidin-modified antimicrobial peptides, such as FF/CAP-18, have antiproliferative effects on the squamous cell carcinoma cell line SAS-H1 and colon cancer-derived cell line HCT116. Ceragenin CSA-13, which mimics the hydrophobic and cationic morphology of cathelicidin-related peptides, was developed to reduce synthetic costs and resolve stability issues in the presence of proteases. In this study, we evaluated the antiproliferative effect of CSA-13 on HCT116 cells. We evaluated the effects of CSA-13 in HCT116 cells by measuring cell growth, detecting apoptosis, analyzing the cell cycle, and examining mitochondrial membrane depolarization. Treatment with CSA-13 suppressed HCT116 cell proliferation in a dose-dependent manner, increasing the incidence of apoptosis detected by the binding of Annexin V. Furthermore, cell cycle analysis showed that the cell cycle of CSA-13-treated wild-type and p53 null mutant HCT116 cells was arrested at the G1/S phase, indicating that CSA-13 affects the cell cycle by a p53-independent pathway. Our study showed that CSA-13 exerts an antiproliferative effect in cancer cells similar to that of FF/CAP-18, suggesting that membrane-permeabilizing capability is the common underlying mechanism for anticancer and antimicrobial effects of CSA-13 and anitimicrobial peptides.