Down-Regulation of Glucocorticoid-Induced TNF Receptor (GITR) Ligand on Human Tumors by Proteolytic Shedding Increases Anti-Tumor Reactivity of NK Cells.

Members of the TNF superfamily mediate multiple cellular functions, including cellular proliferation, differentiation and cell death. Many members of this protein family are shed from the cell surface as soluble forms, which affects cell-cell interactions by reduction of ligand densities and distally modulates effector cells bearing the respective receptor. The TNF family member Glucocorticoid-induced TNF Receptor (GITR) costimulates effector T cells, modulates apoptosis and NFkappaB and abrogates suppression of murine but not human regulatory T cells. Its cognate ligand GITRL has been found in various healthy tissues. Recently we reported that NK cells express GITR, while tumor cells express GITR ligand (GITRL), and GITR/GITRL interaction downregulates NK cell-mediated anti-tumor effector mechanisms like cytotoxicity and IFN-gamma production. Here we report that human tumor cells spontaneously release a soluble form of GITRL (sGITRL) detectable in culture supernatants by ELISA. Furthermore, we found elevated levels of sGITRL in sera of patients with various malignancies compared to healthy controls. We demonstrate that the release of GITRL from tumor cells can be blocked by inhibition of metalloproteinases, concomitantly causing accumulation of GITRL on the tumor cell surface as determined by FACS analysis. Upregulated GITRL surface expression further increased inhibition of NK cell anti-tumor effector mechanisms, while, in contrast, presence of sGITRL in cocultures of GITRL-expressing tumor cells and GITR-positive NK cells enhanced NK cell cytotoxicity and IFN-gamma production. Thus, in line with the results obtained with other TNF family members, conversion of membrane bound GITRL to its soluble form is associated with downregulation of its function, potentially due to blocking its cognate receptor. Thus, release of sGITRL substantially influences the interaction of tumor cells with NK cells. In addition, determination of sGITRL levels may be implemented as a diagnostic marker in patients with malignancies. Further prospective studies are currently being conducted addressing the value of GITRL as a tumor marker in different tumor entities.