Chemoresistance in cancer is linked to a subset of cancer cells termed “cancer stem cells” (CSCs), and in particular, those expressing the CD44 variant appear to represent a more aggressive disease phenotype. Herein, we demonstrate that CD44v6 represents a CSC population with increased resistance to chemotherapeutic agents, and its high expression is frequently associated with poor overall survival (OS) (HR:3.04; CI:1.35–6.85; p<0.001) and disease-free survival (DFS) (HR:5.30, CI:1.64–17.12, p<0.01) in CRC patients. CD44v6+ cells showed elevated resistance to chemotherapeutic drugs and significantly high tumor initiation capacity. The inhibition of CD44v6 resulted in the attenuation of self-renewal capacity and re-sensitization to chemotherapeutic agents. Of note, miRNA profiling of CD44v6+ SDCSCs identified a unique panel of miRNAs indicative of high self-renewal capacity. In particular, miR-1246 was overexpressed in CD44v6+ cells, and associated with poor OS (HR:2.44, CI:1.15–5.18, p<0.05) and DFS (HR:2.37, CI:1.03–5.44, p<0.05) in CRC patients. We demonstrate that CD44v6+ CSCs induced chemoresistance and enhance tumorigenicity in CRC cells, which is in part orchestrated by dysregulated profiles of a distinct panel of miRNAs. These findings provide a rationale for the therapeutic targeting of specific miRNAs, as well as serving as promising prognostic biomarkers in patients with colorectal neoplasia. Overall design: 8 total samples, 4 CD44v6− HCT116 replicates and 4 CD44v6+ HCT116 replicates