MiR-199a-5p represses the stemness of cutaneous squamous cell carcinoma stem cells by targeting Sirt1 and CD44ICD cleavage signaling.

ABSTRACT

Tumorigenic cancer stem cells (CSCs) exist in various tumors including the cutaneous squamous cell carcinoma (cSCC) as a minor subpopulation and are tightly associated with metastasis and therapeutic resistance. Better understanding of CSCs properties is essential for the novel therapeutic strategy targeted toward these cancers. The cSCC stem cells (cSCCSCs) were enriched from a cSCC cell line A431 by repeated sphere culture, and identified via the expression analysis of stemness marker genes and CD44 proteolysis. MiR-199a-5p was previously reported to be related with the proteolysis modulation of CD44, so the specific regulation mechanisms were verified by overexpression in vitro and in vivo. MiR-199a-5p is under-expressed in cSCCSCs and functions as a tumor suppressive molecule. Overexpression of miR-199a-5p reduced the stemness of cSCCSCs and inhibited cell proliferation. By targeting the deacetylase Sirt1, miR-199a-5p inhibited cellular proteolysis of CD44 and reduced the CD44 intracellular domain (CD44ICD) release and nuclear translocation. Overexpression of CD44ICD reversed the effects of miR-199a-5p overexpression or Sirt1 silencing, and increased the transcriptional expression of stemness genes. Our results revealed that the miR-199a-5p/Sirt1/CD44ICD signaling pathway regulates cSCCSCs progression by affecting its migration ability and tumorigenicity, therefore can be utilized to develop a curative approach for cSCC.Abbreviations CSCs cancer stem cells; cSCC cutaneous squamous cell carcinoma; cSCCSCs cSCC stem cells; CD44ICD CD44 intracellular domain; HA hyaluronic acid; HNSCC hand and neck squamous cell carcinoma; ESCC esophageal squamous cell carcinoma;MMPs matrix metalloproteinases; SFM sphere formation medium; EGF epidermal growth factor; bFGF basic fibroblast growth factor; BSA bovine serum albumin; CCK-8 cell counting kit-8.