Colorectal cancer (CRC) has a very high mortality rate worldwide. Although various therapies have been developed to treat CRC, the need for novel therapeutic approaches has been increasing due to severe side effects and limited efficacy of current treatments. Recently, although research on the gut microbiome and its association with colon cancer has been growing, the mechanisms of gut microbiome inhibition in CRC remain insufficiently understood. Thus, in this study, we investigated the growth-inhibitory effects of the culture supernatant of DS3316, isolated from infant feces, on CRC cell lines (HCT116 and SNUC5). And RNA-seq analysis revealed an increase in apoptosis-related terms induced by DS3316 treatment. Also, we found the non-toxicity of DS3316 in human iPSC-derived intenstine organoid. Thus, we suggested that DS3316 inhibits the growth of colorectal cancer cell lines without affecting normal cells. And DS3316 is expected to be a promising candidate for the development of microbiome-based colorectal cancer therapies. Furthermore, its combined use with various colorectal cancer treatment methods could lead to the proposal of more effective therapeutic approaches.