Chinese‐German Cooperation Group Tumor Immunology: Another inspiring Meeting in Deidesheim

Abstract

Immunology is a truly international science which greatly benefits from the exchange and collaboration of scientists from different part of the world. The German Society for Immunology (Deutsche Gesellschaft für Immunologie, DGfI) maintains close connections with several national immunological societies to foster bilateral scientific interactions. Next to the initiation of joint meetings with the Japanese Society for Immunology (JSI), regular bilateral meetings with the Chinese Society for Immunology (CSI) is the oldest of those DGfI activities. Fortunately, the bilateral collaboration with the CSI has been generously supported from the beginning by the Sino-German Center for Research Promotion/Chinesisch-Deutsches Zentrum für Wissenschaftsförderung (SGC/CDZ) which is jointly funded and organized by the Deutsche Forschungsgemeinschaft (DFG) and the National Natural Science Foundation of Chinas (NSFC). The SGC is jointly led by a German and a Chinese director. As summarized by the new German director Dr. Karin Zach, the SGC offers various programs to support bilateral Chinese-German interactions in all fields of science. This includes sponsoring of Workshops and Mobility Programs, preparatory visits to initiate cooperations, Summer Schools, and others (see http://www.sinogermanscience.org.cn/). Several programs specifically support young scientists (advanced PhD students, post-doctoral researchers). Every year, about 30 excellent doctoral students nominated by Chinese universities can participate in the annual Lindau Nobel Laureate Meetings and spend additional time in German top research institutions and universities. Over the years (the program was initiated in 2004), a substantial number of them received a Lindau-follow up fellowship to conduct research in Germany. The series of bilateral DGfI-CSI workshops started in 2009 with the first meeting held at Burg Wanzleben near Magdeburg/Germany, organized by S. Meuer (Heidelberg). In fact, the first three meetings including the follow-up meetings 2011 in Beijing and 2013 at Kloster Eberbach/Germany were made possible on the basis of successful joint applications in the “Workshop” program of the SGC. Here, D. Kabelitz (Kiel) was the applicant from the German side, while X. Cao, W. He and B. Huang were the Chinese applicants in 2010, 2011 and 2013, respectively. Each of these workshops included a total of 30 immunologists, half of them from Germany and half from China. This initial workshop series served to get to know each other and to explore areas of common scientific interest. The unique ambience of the meeting venues and enjoyable social activities were added values which cannot be experienced at meetings with several hundreds or even thousands of participants. Inspired by the success of these bilateral workshops, D. Kabelitz (Germany) and X. Cao (China) submitted an application for a “Collaboration Group Tumor Immunology” to the SGC. The idea of this funding instrument was to intensify the bilateral interactions in a specific area of research. Following the discussions at the previous workshops, tumor immunology was identified as a suitable topic of great interest to basic and clinical immunologists in both countries, offering additional perspectives for clinical translation. The proposal was approved by the SGC, and the first meeting of the Cooperation Group took place in November 2015 in Chengdu/China. In addition to the applicants D. Kabelitz and X. Cao, the Cooperation Group comprised six members from the German side (S. Meuer, J. Wienands, C. Watzl, T. Kamradt, A. Radbruch, H.-M. Jäck) and four members from the Chinese side (Z. Tian, B. Huang, X. Quin, T. Chen). This notwithstanding, the SGC funds allowed to finance again the participation of a total of 30 delegates at four Meetings of the Collaboration Group. In addition, some funds were available to sponsor short laboratory visits. We continued to have the meetings alternating in Germany and in China. The second meeting took place in Dresden/Germany in December 2016 1, the third meeting in Hangzhou/China in November 2017, and the final meeting of the Cooperation Group in March 2019 in Deidesheim/Germany. S. Meuer (Heidelberg) as the main organizer of the meetings in Germany selected wonderful places which offered unique opportunities for additional cultural and social activities, and the same was certainly true for the meetings in China. In the following we briefly highlight the presentations at the recent Deidesheim meeting which was attended by 18 immunologists from Germany and 16 from China (Figure 1). The first speaker was Xuetao Cao (Tianjin) who highlighted the crosstalk of epigenetic modifiers and innate molecules in inflammation and cancer. They observed that the translocation of the cytoplasmic β subunit of the IFN-γ receptor to the plasma membrane required for functional IFNγ signaling in macrophages upon Listeria monocytogenes infection depends on E-cadherin. The assembly of a functional cytokine receptor is thus an important part of the innate immune response against intracellular bacterial infection 2. Xuetao Cao also discussed recent findings how humoral immunity can contribute to tumor progression. In a mouse model of breast cancer, the primary tumor induced HSPA4 antibody-producing B-cell accumulation in lymph nodes which selectively promoted lymph node metastasis in a CXCR4-dependent manner. Importantly, high serum levels of anti-HSPA4 IgG were observed in breast cancer patients with poor prognosis, underscoring the role of tumor-educated B cells for metastasis formation. These finding open possible avenues for new therapeutic strategies, e.g. by diminishing tumor HSPA4 expression by small molecule inhibitors 3. Zhigang Tian (Hefei) discussed the important role of the T-cell Ig and ITIM domain (TIGIT) co-inhibitory receptor in controlling the exhaustion of NK cells. TIGIT-dependent NK cell exhaustion is also observed in Rag2−/− mice but is actually diminished in TIGIT-deficient mice. Antibody-mediated TIGIT blockade inhibited tumor growth in various in vivo models of melanoma, breast and colon cancer 4. Therefore, TIGIT might be a promising additional target for checkpoint inhibitor immunotherapy of cancer. Reinhold Schmidt (Hannover) highlighted recent advances in our understanding of the relationship of primary immunodeficiencies and autoimmunity. As an example, the CTLA-4 haploinsufficiency with autoimmune infiltration (CHAI) syndrome can present with autoantibody-mediated cytopenia, organ-specific autoimmunity and lymphocytic infiltration of non-lymphoid organs. He also discussed the activated PI3-kinase-δ syndrome (APDS) which is a gain-of-function mutation leading to lymphoproliferative senescence and immunodeficiency 5. APDS can be efficiently targeted by the specific PI3K-δ inhibitor Leniolisib 6. Bo Huang (Beijing) has been studying tumor-repopulating cells (TRC) for some time. He recently reported that TRC induce PD1 on CD8 T cells thereby favoring CD8 T-cell exhaustion 7. Further characterization of the features of TRC identified the stiffness of tumor cells as a decisive factor for sensitivity towards perforin-dependent killing. Pore imaging by atomic force microscopy revealed lower pore-forming capacity of perforin in the “softer” TRC when compared to other tumor cells. Strategies to enhance the stiffness of tumor cells (notably TRC) may thus help to enhance cancer immunotherapy. Sven Brandau (Essen) closed the first session with a quantitative analysis of the neutrophil-T cell interaction in human head-and-neck cancer. There are multiple subsets of myeloid-derived suppressor cells 8, and Arginase Ihigh CD11b+CD16+ mature neutrophils are the most suppressive in the peripheral blood of cancer patients. Novel approaches to study in situ conjugate formation between T cells and neutrophils/MDSC identified intratumoral hotspots where T cells engaged with neutrophil/MDSC were granzyme-negative whereas other intratumoral T cells were 40–50% granzyme-positive, supporting the in vivo relevance of tumor-associated suppressive neutrophils. The second session was focused on B cells. Michael Reth (Freiburg) discussed recent studies where CRISPR/Cas9 was used to inactivate individual or several genes of the BCR complex, i.e. heavy chain, light chain, Igα (CD79a), or Igβ (CD79b) in the Ramos Burkitt lymphoma cell line. The results showed that Igβ can be expressed on the cell surface in the absence of other BCR components, and Igβ together with the BCR co-receptor CD19 is required for competitive growth in cell culture 9. Jürgen Wienands (Göttingen) has a long-standing interest in the molecular pathways of BCR signaling in normal and malignant B cells. Previously, his group has characterized several regulators of BCR signaling such as SLP65 and NTAL. More recently, they identified the ubiquitously expressed adaptor protein Cbl-interacting protein of 85 kDa (CIN85) as a novel regulator of NK-κB activation in B cells. Patients with a germline mutation in the X-chromosomal CIN85 gene suffer from a primary antibody deficiency due to intrinsic defects in BCR signaling 10. Katharina Pracht (Erlangen) presented her results on the role of miR-148a for the maintenance of long-lived plasma cells. miR-148a targets the transcription factors Bach2, MiTF, PTEN and Bim. In a tamoxifen-inducible KO mouse model, the loss of miR-148a was associated with a reduction of long-lived plasma cells resulting from both apoptosis and modulated chemokine receptor expression. The final talk in this session was given by Andreas Radbruch (Berlin) who discussed the characteristics of CD69+ bone marrow-resident memory T cells 11 and the role of stroma cells for the maintenance of long-lived plasma cells. Under hypoxic in vitro conditions, cell contact with ST2 stroma cells together with the cytokine APRIL is sufficient to maintain bone marrow plasma cells alive for extended periods of time. Stroma cell contact induces PI3K/AKT signaling which down-regulates transcription factors FoxO1 and FoxO3a which is critical for survival of memory plasma cells. APRIL appears to prevent the activation of caspase-12, while PI3K signaling prevents the activation of caspase-3. In conclusion, continued contact to bone marrow stromal cells is required for maintenance of memory plasma cells. The topic of the third session was cytotoxic effector cells and -mechanisms, and T cell/tumor interplay. Dieter Kabelitz (Kiel) summarized features of human Vδ2 T cells which make them attractive candidates for cellular immunotherapy. These cells recognize in a HLA-independent manner small pyrophosphates which are overproduced by many tumor cells; hence Vδ2 T cells recognize and kill various cancer cells with no need for HLA-restricted recognition of specific tumor antigens 12. In a recent as yet unpublished collaboration with Zhinan Yin (Guangzhou) it was found that the anti-oxidant and epigenetic modifier Vitamin C enhances the proliferative activity, IFNγ production and cytotoxic activity of human γδ T cells. These results suggest that supplementation of Vδ2 T cell expansion in vitro with Vitamin C might increase the therapeutic efficacy of adoptively transferred γδ T cells in cancer patients. Jaydeep Bhat (Berlin) presented his recent work on the modulation of the γδ T cell/tumor cell interplay in vitro by epigenetic modifiers, specifically the HDAC inhibitor valproic acid (VPA). VPA treatment increased cell surface expression and release of certain NKG2D ligands on pancreatic and prostate cancer cells while it down-modulated NKG2D receptor expression on γδ T cells upon co-culture with tumor cells. A flow cytometric method was established to quantify H3K9 acetylation in co-cultured γδ T cells. This approach allowed to correlate alterations in memory marker expression on γδ T cells co-cultured with tumor cells as a consequence of VPA treatment and thus provided new insights into epigenetic regulation of human γδ T cells 13. NK cells use various pathways to kill their tumor target cells. Having established a sophisticated experimental system which allows to monitor the activity of granzyme B and caspase-8 in individual NK cells upon contact with tumor target cells, Carsten Watzl (Dortmund) studied the serial killing and the respective contribution of granzyme and caspase-8 14. Concomitant with the modulation of death-associated molecules (up-regulation of CD95L, down-regulation of TRAIL, perforin, granzyme B), he observed a transition from the use of granzyme to caspase-8 mediated killing over time. Ottmar Janssen (Kiel) focused on the characterization of various lysosome-related effector vesicles (LREV) in human cytotoxic cells, which can be isolated by differential gradient centrifugation and are characterized by different pre-formed effector proteins like FasL, perforin and granzymes. The two isoforms of the pore-forming protein granulolysin (15 kDa, 9 kDa) were associated with different vesicles and were differentially mobilized by PKC or PKC plus ionomycin activation 15. For the first time, his group could also demonstrate that dipeptidyl peptidase 4 (CD26), an ectopeptidase with multiple functions in the immune and endocrine system, is stored in LREV of cytotoxic effector cells, thereby identifying circulating lymphocytes as a possible source of soluble CD26 in serum (unpublished results). In the final talk of the session, Chen-Yu Zhang (Nanjing) discussed the role of tumor-secreted miR-214 in the induction of Treg and the resulting tumor immune evasion. Underscoring the importance of this pathway, antisense approaches to down-regulate miR-214 were found to block Treg expansion and tumor growth in mice 16. The role of macrophages in tumor immunology was discussed in the fourth session. Tobias Bopp (Mainz) identified a novel mechanism how highly glycolytic tumors like melanoma, via acidification of the tumor microenvironment, trigger the transcriptional repressor ICER in tumor-associated macrophages leading to their non-inflammatory and tumor-promoting polarization 17. In line, ICER-deficient macrophages displayed an inflammatory M1 phenotype and controlled melanoma (but not colon carcinoma) growth. Maode Lai (Hangzhou) used immunohistochemistry and laser capture microdissection to study immune cell composition and gene expression in different areas in a large series of colorectal carcinoma samples. 18 In addition to identifying the frequency of CD68+ macrophages at the tumor invasive front as a decisive factor for immune cell infiltration and overall survival, these studies also revealed opposing roles of intracellular and extracellular S100A8 protein on migration and invasiveness. The differentiation of suppressive tumor-associated macrophages (TAM) is influenced by multiple factors. Rainer Glauben (Berlin) reported that oleate (but not stearate) induces a suppressive phenotype in GM-CSF generated macrophages which acquire a TAM marker signature 19. The identification of enzymes connected to lipid droplet formation in tumor-infiltrating TAM may offer new therapeutic targets to counteract suppressive TAM. Using KO mouse models, Lun-Xiu Qin (Shanghai) identified osteopontin (OPN) as an important factor controlling TAM generation in PD-L1 expression via activation of the colony-stimulating factor-1 (CSF1)/CSF1R pathway in hepatocellular carcinoma (HCC) 20. OPN facilitates M2 polarization, and blocking the OPN/CSF1/CSF1R pathway by small molecules (PLX3397) enhanced the efficacy of checkpoint inhibitor treatment in HCC models. Julia Kzhyshkowska (Mannheim) studies chitinase-like proteins (CLP) which are lectins produced by TAM. The CLP YKL-39 expressed by TAM in human breast cancer is chemotactic for monocytes, induces angiogenesis in vitro, and correlates with poor prognosis in breast cancer patients undergoing neoadjuvant chemotherapy, suggesting that it might be a novel therapeutic target 21. As the first speaker in session five, Xiu-Wu Bian (Chongqing) focused on the role of cancer stem cells (CSC) for heterogeneity of tumors and their interaction with the tumor microenvironment. He identified a NOTCH1-SOX2 positive feedback loop as an essential step of the invasion of white matter in the brain by CD9-expressing glioma stem cells 22. Yiwei Chu (Shanghai) introduced the concept of BiTe-T cells where T cells are transduced with BiTe constructs in a way that the BiTe (e.g., Her2/CD3; ref. [23]) are released extracellulary and bind (and therefore activate) both the transduced and bystander T cells. T cells transduced with a novel BiTe targeting glypican-3 (GPC3) expressed on liver cancer cells were therapeutically effective in mouse models. So far, BiTe-T cells exert high potency and lox toxicity and thus may complement existing CAR-T cells and BiTe therapeutic strategies. No doubts, T cells of distinct differentiation stages are important in immune surveillance and tumor control. Consequently, a precise understanding of the molecular pathways of T cell fate decisions is important. Vigo Heissmeyer (Munich) discussed the mechanisms of how RNA-binding proteins of the Roquin family regulate T cell differentiation. Roquin targets are controlled by the endonuclease Regnase-1, and the generation of mice deficient in T-cell expression of Roquin-1/2 and/or Regnase-1 opened new insights 24. Peter Krammer (Heidelberg) closed this session with a report on the therapeutic efficacy of dimethyl fumarate (DMF) in patients with cutaneous T cell lymphoma (CTC). The mild NF-κB inhibitor DMF induces riptosome formation and inhibits cell migration 25. It is a licensed treatment for patients with psoriasis and multiple sclerosis. Here, exciting results were presented on the efficacy of DMT to reduce clinical symptoms and malignant CD4 T cell numbers in patients with CTC and Sézary's syndrome. Thomas Kamradt (Jena), the current DGfI president, opened the sixth session with a discussion of the complex interaction of IL-33 receptor subunits with various receptor tyrosine kinases (RTK). His recent results indicate that the IL-33 induced, TCR-independent induction of IL-13 production in CD4 T cells depends on the activation of the RTK EGFR by its ligand amphiregulin 26. By contrast, the IL-33R needs to interact with the RTK c-kit in mast cells to induce via IL-6 production FoxP3+RoRγt+ Tregs. Moving from cytokine receptor signaling to maintenance of intestinal immune homeostasis, Qingqing Wang (Hangzhou) highlighted her recent findings which have identified a key role of the Na+/K+ ATPase regulating protein FXYD3 for the regulation of mucin-2 secretion in intestinal goblet cells and thus for immune homeostasis. In line, FXYD3 expression was reduced in experimental colitis models but also in clinical samples from patients with inflammatory bowel disease. The next talk given by Zhihong Zhang (Wuhan) illustrated the potential of high-end optical imaging combined with high spatio-temporal resolution to study the tumor microenvironment. This technology is extremely valuable to evaluate the effect of (immuno)therapeutic strategies on the dynamics of immune cell/tumor cell interaction. The final presentation in this session by Friederike Berberich-Siebelt (Würzburg) highlighted the importance of SUMOylation for the activity of the transcription factor NFATc1 in T cells 27. Using a novel transgenic Nfatc1 transgenic mouse with deficient SUMOylation, it was shown that reduced SUMOylation ameliorated experimental autoimmune encephalitis (EAE) through increased Treg expansion which resulted from elevated IL-2 production. In the seventh and final session of the meeting, important additional aspects of tumor immunology were discussed. Li Wu (Beijing) described the development of a novel TLR2 agonist (SUP3) which is more stable and more potent than standard TLR2 ligands like Pam3CSK4. SUP3 upregulated CD80/CD86 expression and cross-presentation in dendritic cells and induced potent CTL activity but only low levels of inflammatory cytokines in vivo, making it a promising new adjuvant 28. Xiao-Feng Qin (Suzhou) presented exciting new data on the non-redundant function of CD38 as a novel checkpoint inhibitor. Tumor-expressed CD38 (cyclic ADP ribose hydrolase) affects intratumoral adenosine levels which favors tumor growth 29. CD38-negative tumor cells were more susceptible to killing by cytotoxic lymphocytes. In consequence, targeting CD38 by antibodies or small molecule inhibitors of its enzymatic activity offers fascinating perspectives for overriding adaptive resistance to anti-PD-L1 treatment. Yuting Ma (Suzhou) discussed the important aspect of how mental and psychological stress may affect the outcome of chemo- and immunotherapy in cancer patients. To this end, she has established experimental systems to study how social defeat-triggered mental stress impacts on chemo- and immunotherapy in transplantable and primary tumor models. Mental stress exerted multiple negative effects which could at least partially be reverted by glucocorticoid receptor antagonists. On-going studies are devoted to identifying stress-associated gene patterns which might pave the way for novel therapeutic targets to assist chemo- immunotherapy of cancer patients. Finally, Limin Zheng (Guangzhou) summarized his studies on the myeloid cell compartment in cancer patients. He found increased numbers of myeloid-based hematopietic stem and progenitor cells in the blood of cancer patients, correlating with clinical stage and survival rates 30. More recently, his group identified the spleen as a site of tumor-induced extramedullary hematopoiesis and a niche to prime myeloid progenitor cells to promote tumor progression 31. Interestingly, splenectomy was found to increase the efficacy of anti-PD-L1 checkpoint therapy. The meeting was rounded up by two key note talks delivered by prominent scientists from nearby Heidelberg. Michail (Mischa) Sitkovsky (Boston; currently on sabbatical leave in Heidelberg) elaborated on the important role of oxygen and adenosine levels in the tumor microenvironment for anti-tumor immune responses. High intracellular cAMP levels are immunosuppressive, as is the hypoxic tumor microenvironment. Therefore, approaches to block adenosine receptor 2A 32 and to increase oxygenation 33 may have great potential to augment existing immunotherapies. Dirk Jäger, Head of the National Center for Tumor Diseases in Heidelberg, summarized the importance of T cell numbers at the invasive margin of solid tumors as analyzed by T cell density scores across tumor sections. T cell high tumors displayed a chemotactic gene signature at the invasive margin and responded to chemotherapy, while T cell low tumors showed a macrophage gene signature and did not response to chemotherapy. Strategies to enhance T cell migration into tumors were evaluated in a sophisticated human tumor explant model, and CCR5 was identified as a therapeutically important target with CCR5 inhibitors being tested in treatment-resistant cancer patients 34. Importantly, the established tumor explant model can also be used for evaluation of cellular therapeutics like CAR-T cells. Overall, this was an exciting meeting which highlighted major areas of current developments in tumor immunology. Naturally, excellent science is the core of such bilateral meetings. However, social activities outside the meeting room are important adds-on which help to develop lasting friendship. A sightseeing tour to Heidelberg Castle and Old City was followed by a reception at the impressive Tower of the Sino-German Hi Tech Park which serves to promote cooperation in science, economy and industry. The 17th floor terrace offered a spectacular view over the Rhine valley. With Deidesheim being located in the heart of the “Pfalz” wine region, it was a must to visit one of the most famous wineries and enjoying tasting of local wine. While funding by the SGC was crucial to organize the series of bilateral meetings, other lasting interactions have been established over the years. Every year, two young Chinese scientists are invited to attend the DGfI Spring School in Ettal/Germany and an additional week in a German research institution. Vice versa, two to three German advanced PhD students or post-docs are invited by the CSI to attend their annual meeting and to present their work in a young investigator's symposium and to visit Chinese laboratories. Overall, we can proudly conclude that bilateral interactions between CSI and DGfI are now an important asset of both societies. In the future, these interactions will be developed further by including more excellent young researcher from both sides. The organization of the next bilateral meeting to be held in China in 2020 is already being conceptualized.