Doctor of Medicine, Eberhard Karls Universitat Tubingen (2012)
Michael Cleary, Postdoctoral Faculty Sponsor
Natural killer (NK) cells are cytotoxic lymphocytes that largely contribute to the efficacy of therapeutic strategies like allogenic stem cell transplantation in acute myeloid leukemia (AML) and application of Rituximab in chronic lymphocytic leukemia (CLL). The tumor necrosis factor (TNF) family member GITR ligand (GITRL) is frequently expressed on leukemia cells in AML and CLL and impairs the reactivity of NK cells which express GITR and upregulate its expression following activation. We developed a strategy to reinforce NK anti-leukemia reactivity by combining disruption of GITR-GITRL interaction with targeting leukemia cells for NK antibody-dependent cellular cytotoxicity (ADCC) using GITR-Ig fusion proteins with modified Fc moieties. Neutralization of leukemia-expressed GITRL by the GITR domain enhanced cytotoxicity and cytokine production of NK cells depending on activation state with NK reactivity being further largely dependent on the engineered affinity of the fusion proteins to the Fc receptor. Compared with wild-type GITR-Ig, treatment of primary AML and CLL cells with mutants containing a S239D/I332E modification potently increased cytotoxicity, degranulation, and cytokine production of NK cells in a target-antigen-dependent manner with additive effects being observed with CLL cells upon parallel exposure to Rituximab. Fc-optimized GITR-Ig may thus constitute an attractive means for immunotherapy of leukemia that warrants clinical evaluation.
View details for DOI 10.1038/mt.2013.11
View details for Web of Science ID 000317110300020
View details for PubMedID 23380816
Ligands of the prototypical activating NK receptor NKG2D render cancer cells susceptible to NK cell-mediated cytolysis if expressed at sufficiently high levels. However, malignant cells employ mechanisms to evade NKG2D-mediated immunosurveillance, such as NKG2D ligand (NKG2DL) shedding resulting in reduced surface expression levels. In addition, systemic downregulation of NKG2D on NK cells of cancer patients has been observed in many studies and was attributed to soluble NKG2DL (sNKG2DL), although there also are conflicting data. Likewise, relevant expression of NKG2DL in leukemia has been reported by some, but not all studies. Hence, we comprehensively studied expression, release, and function of the NKG2D ligands MHC class I chain-related molecules A and B and UL16-binding proteins 1-3 in 205 leukemia patients. Leukemia cells of most patients (75%) expressed at least one NKG2DL at the surface, and all investigated patient sera contained elevated sNKG2DL levels. Besides correlating NKG2DL levels with clinical data and outcome, we demonstrate that sNKG2DL in patient sera reduce NKG2D expression on NK cells, resulting in impaired antileukemia reactivity, which also critically depends on number and levels of surface-expressed NKG2DL. Together, we provide comprehensive data on the relevance of NKG2D/NKG2DL expression, release, and function for NK reactivity in leukemia, which exemplifies the mechanisms underlying NKG2D-mediated tumor immunosurveillance and escape.
View details for DOI 10.4049/jimmunol.1200796
View details for Web of Science ID 000306599100033
View details for PubMedID 22730533
Natural killer (NK) cells play an important role in the immunosurveillance of hematopoietic malignancies. Their reactivity is influenced by activating and inhibitory signals mediated by tumor-expressed ligands for NK receptors. Many members of the tumor necrosis factor (TNF) family modulate differentiation, proliferation, activation and death of both tumor and immune effector cells. The TNF receptor family member glucocorticoid-induced TNFR-related protein (GITR) stimulates anti-tumor immunity in mice, but available data indicate that GITR may mediate different effects in mice and men and impairs the reactivity of human NK cells. Here, we comprehensively studied the expression and function of GITR ligand (GITRL) in leukemia. Among the different leukemia entities, pronounced expression of GITRL on leukemic cells was observed in chronic lymphocytic leukemia (CLL), and the GITR receptor was expressed at significantly higher levels on NK cells of CLL patients compared with healthy controls. Upon GITR-GITRL interaction, signaling via GITRL into the leukemia cells induced the release of interleukin (IL)-6, IL-8 and TNF, which act as growth and survival factors for CLL cells. In addition, GITRL impaired both direct and Rituximab-induced degranulation, cytotoxicity and interferon-? production of NK cells, which could be restored by GITR blocking antibodies. Thus, GITRL may contribute to disease pathophysiology and resistance to direct and Rituximab-induced NK reactivity in CLL.
View details for DOI 10.1038/leu.2011.313
View details for Web of Science ID 000303883500018
View details for PubMedID 22064350
NK cells play an important role in tumor immunosurveillance and largely contribute to the therapeutic success of anti-tumor antibodies like Rituximab. Here, we studied the role of the TNF family member 4-1BB ligand (4-1BBL) during the interaction of NK cells with chronic lymphocytic leukemia (CLL) cells. 4-1BBL was highly expressed on patient B-CLL cells in all 56 investigated cases. Signaling via 4-1BBL following interaction with 4-1BB, which was detected on NK cells of CLL patients but not healthy individuals, led to the release of immunoregulatory cytokines including TNF by CLL cells. CLL patient sera contained elevated levels of TNF and induced 4-1BB upregulation on NK cells, which in turn impaired direct and Rituximab-induced NK-cell reactivity against 4-1BBL-expressing targets. NK-cell reactivity was not only enhanced by blocking the interaction of NK cell-expressed 4-1BB with 4-1BBL expressed by CLL cells, but also by preventing 4-1BB upregulation on NK cells via neutralization of TNF in patient serum with Infliximab. Our data indicate that 4-1BBL mediates NK-cell immunosubversion in CLL, and thus might contribute to the reportedly compromised efficacy of Rituximab to induce NK-cell reactivity in the disease, and that TNF neutralization may serve to enhance the efficacy of Rituximab treatment in CLL.
View details for DOI 10.1002/eji.201141920
View details for Web of Science ID 000302541300022
View details for PubMedID 22144129