To examine possible miRNAs involved in mediating CSC properties in ESCC, we first collected spheres of three ESCC cell lines (Eca109, Kyse150, and TE‐1) by serially replating in ultralow attachment plates (Figure 1A); the stemness of these spheres was identified by measuring several CSC markers (ALDH1, SOX2, NANOG, OCT4, BMI‐1, ABCG2, CD90 and CD271) (Figure S1A–C). This evidence concerns the gene POU5F1 and esophageal squamous cell carcinoma.