CSCs positive for Lgr5, CD44, and CD133 are regulated by the mTORC2-hedgehog axis in cancer cells and are potentially responsible for the high recurrence rates of CRC. HCT-116 (A-I) and SW480 (J-L) colon cancer cells were transfected with DDK-tag empty vector or DDK-tag UBXN2A. After 48 hours, cells were subjected to flow cytometry analysis using three CSC markers (Lgr5, CD44, and CD133). As previously described for HCT-116 cells, overexpression of UBXN2A has no significant effect on CD133 (I). UBXN2A overexpression led to a significant reduction of CD44 (J) and Lgr5 (K) as well as CD133 (L), which exhibits increased tumor sphere-forming efficiency and increased tumorigenic potential in the SW480 cell line (N=4, **** p< 0.0001, mean +/− SD).