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Project P3

Prof. Dr. Andreas Mackensen
Dept. of Internal Medicine 5 – Haematology/Oncology

University of Erlangen 
Andreas.Mackensen@uk-erlangen.de

 Mike_2

Dr. Michael Aigner

Dept. of Internal Medicine 5 – Haematology/Oncology

University of Erlangen 
Michael.Aigner@uk-erlangen.de

 Topic

Suppression of human T cells by 5‘-methythioadenosine

Summary

Malignant cells develop diverse mechanisms to escape T cell-mediated immune recognition, including a dysregulated metabolism. Studies in melanoma and renal cell carcinoma have revealed loss of 5’-methylthioadenosine phosphorylase (MTAP) expression in vitro and in vivo. The enzyme MTAP is responsible for the catalytic degradation of its substrate 5’-methythioadenosine (MTA). As a direct consequence, MTAP-deficient tumors secrete high amounts of MTA. In the first funding period we could demonstrate that MTA suppresses human CD8+ cytotoxic T cells in vitro by inhibiting proliferation and cytokine secretion. To study the role of endogenously secreted MTA on antitumor immune responses, a human MTAP-deficient, high MTA secreting melanoma cell line was stably transfected with MTAP resulting in a significant reduction of MTA secretion. Comparative mixed tumor/lymphocyte cultures revealed a reduced proliferative capacity in T cells stimulated with allogeneic mock-transfected melanoma cells (high MTA secretors)  compared to T cells stimulated with MTAP-transfected melanoma cells (low MTA secretors). In addition, we were able to identify two distinct effects of MTA on human T cell subpopulations. When resting CD8+ T cells were exposed to MTA upon antigen-specific stimulation with peptide-pulsed APCs, MTA strongly inhibited activation, proliferation, and differentiation of antigen-specific T cells. In contrast, exposure of already activated antigen-specific CD8+ T cells with low doses of MTA resulted in apoptotic death of T cells. Next, we analyzed signaling pathways in CD8+ T cells after MTA exposure and found that MTA leads to downregulation of phosphorylated (p)Akt but not ERK or p38. As pAkt is part of the CD28 co-receptor pathway we postulate that MTA inhibits signal II of T cell activation. To reconstitute tumor-specific T cell responses in vitro and in vivo we aim to overcome the inhibitory effects of MTA by overexpression of MTAP in human CD8+ T cells. Antigen specific CD8+ T cells will be stably transfected with MTAP, characterized and adoptively transferred into tumor bearing, immunodeficient mice to study the antitumor efficacy in vivo. Our previous results have shown that MTA can affect Akt-mediated signaling which is crucial for T cell activation. Therefore we aim to overcome this blockade by using a natural Akt activating cytokine (IL-7) or a small molecule (SC-79) which acts as an Akt activator. To further understand the molecular mechanism of MTA induced T cell suppression in more detail, we aim to characterize the effects of MTA on asymmetric protein methylation after T cell activation. MTA could also interfere with the metabolic potential of T cells and tumor cells thereby exerting in part its immunoregulatory effects. To test this hypothesis the aerobic and/or anaerobic metabolic potential of T cells in relation to tumor cells will be analyzed in short-term or long-term cultures. In a first series of experiments we could show that MTA is also able to inhibit the cytolytic activity of NK cells. Of major interest NK cell subpopulations exhibited differential susceptibility towards MTA. We now aim to characterize the mechanism of distinct sensitivity of NK cell subpopulations to MTA.

References

  1. Meidenbauer, N., A. Zippelius, M.J. Pittet, M. Laumer, S. Vogl, J. Heymann, M. Rehli, B. Seliger, S. Schwarz, F.A. Le Gal, P.Y. Dietrich, R. Andreesen, P. Romero, A. Mackensen. 2004. High frequency of functionally active Melan-A-specific T cells in a patient with progressive mmunoproteasome-deficient melanoma. Cancer Res 64: 6319-6326

  2. Fischer, K., P. Hoffmann, S. Voelkl, N. Meidenbauer, J. Ammer, M. Edinger, E. Gottfried, S. Schwarz, G. Rothe, S. Hoves, K. Renner, B. Timischl, A. Mackensen, L. Kunz-Schughart, R. Andreesen, S.W. Krause, M. Kreutz. 2007. Inhibitory effect of tumor cell-derived lactic acid on human T cells. Blood 109:3812-3819

  3. Singer, K., M. Kastenberger, E. Gottfried, C.G. Hammerschmied, M. Büttner, M. Aigner, B. Seliger, B. Walter, H. Schlösser, A. Hartmann, R. Andreesen, A. Mackensen, M. Kreutz. 2010. Warburg phenotype in renal cell carcinoma: High expression of glucose-transporter 1 (GLUT-1) correlates with low CD8+ T cell infiltrate in the tumor. Int J Cancer Jul 6. [Epub ahead of print].

Co-Workers

Frederik Henrich
Frederik.Henrich@uk-erlangen.de
 

 

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