Immune escape of malignant gliomas – regulation of NKG2D ligands by TGF-beta and matrix metalloproteinases

Gliomas are the most frequent primary brain tumors with an incidence of up to 5 per 100.000. For tumorigenesis escape from immunosurveillance is essential. A major mediator of anti-tumor immunity is the lectin-like NKG2D immunoreceptor which is expressed by natural killer (NK) and CD8+ alpha/beta and gamma/delta T cells. NKG2D ligands (NKG2DL) are absent from somatic cells (with the exception of the intestinal epithelium), but are induced upon transformation or infection, leading to NK or T cell-mediated lysis of NKG2DL-expressing cells. Gliomas shape their microenvironment by secreting proteolytically active enzymes like matrix metalloproteinases (MMP) and create an immunosuppressive microenvironment by soluble factors like transforming growth factor-beta (TGF-beta). In this study we show novel pathways for the immune escape of gliomas by selective downregulation of NKG2DL by TGF-beta and MMP. Here we report for the first time, that the NKG2DL MHC class I-chain related molecule A (MICA) and UL16-binding protein 2 (ULBP2) are expressed in gliomas in vivo. With increasing grade of malignancy ULBP2 expression decreases, while for MICA a diffuse background staining most likely originating from soluble MICA increases, suggesting that posttranslational cleavage of MICA is enhanced with tumor grade. In vitro, downregulation of TGF-beta by RNA interference (siRNA) in LNT-229 human malignant glioma cells reveals that TGF-beta selectively suppresses MICA and ULBP2 on transcriptional and protein level while MICB, ULBP1 and ULBP3 were not affected. Surprisingly, the almost complete loss of the TGF-beta-dependent MMP-2 and -9 does not result in reduced NKG2DL cleavage from the surface of TGF-beta1/2 siRNA cells, although MICA cleavage occurs in an MMP-dependent manner. Consequently, NKG2DL-dependent NK cell-mediated lysis was enhanced either by depletion of TGF-beta or by inhibition of MMP with the TGF-beta-antagonism exerting the greater effect. These data define a differential regulation of human NKG2DL expression as part of the immunosuppressive actions of TGF-beta and MMP in human malignant gliomas. In summary, a better understanding of MICA and ULBP2 regulation may open up options for therapeutic intervention.