Silencing NKG2D Ligand-targeting MiRNAs Enhances Natural Killer Cell-mediated Cytotoxicity In Breast Cancer

Breast cancer(BC)accounted for 25%of all cancer cases worldwide and 15%of all cancer deaths among females in 2012.Late-stage BC is often associated with an immunosuppressive microenvironment and lacks of antitumor immune responses.Natural killer(NK)cells mediate direct cytotoxic activity against tumor cells and provide an early host defense against the transformed cells.NKG2D is one of the major activating receptors of natural killer(NK)cells and binds to several ligands(NKG2DLs).NKG2DLs are expressed on malignant cells and sensitize them to early elimination by cytotoxic lymphocytes.We investigated the clinical importance of NKG2DLs and the mechanism of NKG2DL regulation in breast cancer(BC).Among the NKG2DLS MICA/B and ULBP1/2/3,the expression levels of MICA/B in BC tissues were inversely associated with the Tumor Node Metastasis(TNM)stage.We first found that the high expression of MICB,but not MICA,was an independent prognostic factor for overall survival in patients with BC.Investigation into the mechanism revealed that a group of microRNAs(miRNAs)belonging to the miR-17-92 cluster,especially miR-20a,decreased the expression of ULBP2 and MICA/B.These miRNAs down-regulated the expression of MICA/B by targeting the MICA/B 3'-untranslated region(3'-UTR)and down-regulated ULBP2 by inhibiting the MAPK/ERK signaling pathway.Functional analysis showed that the silencing of NKG2DL-targeting miRNAs in BC cells increased NK cell-mediated cytotoxicity in vitro and inhibited immune escape in vivo.Additionally,histone deacetylase inhibitors(HDACis)increased NKG2DL expression in BC cells by inhibiting members of the miR-17-92 cluster.Thus,targeting miRNAs with antisense inhibitors or HDACis may represent a novel approach for increasing the immunogenicity of BC.