RG Bosch

Medicine 5

Head of Department:
Prof. Dr. med. Andreas Mackensen

T cell-based immunotherapy of uveal melanoma

Team AG Bosch

Barbara Bock, Dr. Jacobus Bosch, Dr. Margarethe Karg


Uveal melanoma

Uveal melanoma is the most common cancer of the eye in adults. Primary uveal melanoma can be treated with a variety of therapies that may limit the growth of the primary tumor in the eye and partially preserve vision. However, none of these treatment modalities prevents the development of metastases, which predominantly arise in the liver and universally remain fatal. Metastatic patients have been treated with conventional chemotherapies, targeted therapies and immunotherapy by checkpoint blockade. Unfortunately, none of these modalities, including checkpoint blockade, have improved overall survival. The need for novel therapies is evident, particularly since significant progress in the treatment of metastatic cutaneous melanoma has been made in recent years. Novel therapies are being explored for their effectiveness against uveal melanoma metastases and immunotherapy may be a potential option.
 

T cell-based immunotherapy and uveal melanoma

Uveal melanoma may be particularly responsive to T-cell-based immunotherapy because it originates in the immune-privileged eye. The localization of the primary tumor in the immune-privileged eye excuses the tumor cells from continuous immunological pressure. This may render primary uveal melanoma more immunogenic than tumor cells from non-privileged sites and allow expression of novel tumor antigens to which the patient's endogenous T cell repertoire is not tolerized (concept reviewed in Lab Refs. 1-3).
 

MHC II uveal melanoma cell-based vaccines

MHC II uveal melanoma cell-based vaccines were generated that target the activation of tumor-reactive CD4+ and CD8+ T cells. These cell-based vaccines consist of primary uveal melanoma cells genetically modified to express major histocompatibility complex class II (MHC II) alleles syngeneic to the recipient and the costimulatory molecule CD80 (B7.1). We showed that these uveal melanoma cell-based vaccines activate CD4+ T cells within total peripheral blood lymphocytes (PBMC) to react with primary uveal melanoma cells and cross-react with metastatic uveal melanoma cells (Lab Ref. 4). In addition, uveal melanoma cell-based vaccines directly activate a diverse repertoire of purified, naïve CD4+ T cells (Lab Ref. 5). Furthermore, cytotoxic CD8+ T cells (CTL) were activated to become cytolytic towards primary and metastatic uveal melanoma cells (Lab Ref. 6). The expression of CD80 blocked the interferon gamma (IFNγ)-mediated upregulation of programmed-death-ligand 1 (PD-L1) and thereby prevented T cell suppression during vaccine priming and boosting of responding T cells (Lab Ref. 7). Adhesion via ICAM-1/LFA-1 interactions facilitated the cell-to-cell contact required for direct CD4+ cell activation by the vaccines (Lab Ref. 5).
 

Redirection of T cells to target uveal melanoma cells

T cell activation to uveal melanoma cells is attempted by redirecting T cells to target human melanoma-associated chondroitin sulphate proteoglycan (MCSP). Our data indicate that primary and metastatic uveal melanoma cell lines express MCSP. Redirection of T cells is studied using three different immunotherapeutic approaches; (i) MCSP/CD3-bispecific single-chain antibody constructs (BITE; collaboration Micromet/Amgen), (ii) MCSP/CD3/Fc-trifunctional antibodies (Triomab; collaboration Trion Research, Martinsried, Germany) and (iii) MCSP-specific chimeric antigen receptor (CAR) modified T cells (in house collaboration with Dept. of Dermatology). Collectively, these studies may indicate that MCSP could serve as a target for T cell-based immunotherapy of uveal melanoma.
 

The role of midkine in tumor-associated lymphangiogenesis, immune cell infiltration and metastasis of ocular malignant melanoma

Collaboration with Prof. Ludwig M. Heindl and Prof. C. Cursiefen, Dept. of Ophthalmology, University Hospital Cologne.

In this collaborative project’s first phase (2013 – 2015), we identified that outgrowth of new lymphatic vessels from preexisting ones (lymphangiogenesis) is a decisive risk factor for metastatic spread and an indicator of poor prognosis in ocular melanoma (Lab Refs. 8,9).

In the second project phase (2015 – 2018), we investigated the underlying cellular and molecular mechanisms both in vitro and in vivo in a novel mouse model of conjunctival melanoma (Lab Refs. 10-12). This led us to identify midkine as a key cytokine in tumor-associated lymphangiogenesis, immune cell infiltration and metastasis in uveal and conjunctival melanoma.

Currently, in the third project phase (2018 – 2021), we plan to study in depth the molecular signaling mechanisms and cellular function of midkine in ocular melanoma. Specifically, we will focus on the role of midkine in ocular melanoma-associated lympangiogenesis and metastasis both in vitro and in vivo. In addition, the role of midkine in the tumor microenvironment, specifically tumor cell – stroma interaction via lymphatic endothelial cells and infiltration of tumor infiltrating lymphocytes and tumor-associated macrophages will be addressed (Lab Ref. 13). Based on these findings we will explore whether targeting midkine could be a novel anti-(lymph)angiogenic therapy for attenuation of tumor metastasis and recurrence in our novel mouse model of metastatic conjunctival melanoma.

Further information: http://www.for2240.de/p4-ocular-melanoma
 

Monitoring T cell and tumor cell dynamic cell-to-cell interactions by organ-on-chip and in vitro cell-technology

Collaboration with Prof. Peter Ertl, TU Vienna, Austria

In this collaborative project we developend a complementary cell analysis method to assess the dynamic interactions of tumor cells with resident tissue and immune cells using optical light scattering and impedance sensing to shed light on tumor cell behavior. The combination of electroanalytical and optical biosensing technologies integrated in a lab-on-a-chip allows for continuous, label-free, and non-invasive probing of dynamic cell-to-cell interactions between adherent and non-adherent cocultures, thus providing real-time insights into tumor cell responses under physiologically relevant conditions. For the first time, the direct cell-to-cell interactions of tumor cells with bead-activated primary T cells were continuously assessed using an effector cell to target a cell ratio of 10:1 (Lab Ref. 14).
 

Magnetic flowcytometry (MRCyte) – Subproject: “Hematology and oncology cell models, cell surface marker identification, (pre)clinical testing”

Collaboration with Prof. Oliver Hayden, TU Munich, Germany.

MRCyte-Consortium: Lead: Siemens AG Corporate Technology; Partners: Sensitec, M2 Automation, Chemicell, Microfluidic Chipshop, Elmos Semiconductor, ICT-IMM Fraunhofer, HNO Universitätsklinikum Mainz, Medizin 5 Universitätsklinikum Erlangen.

Methods for hematological-oncological diagnostics, monitoring of therapy and side effects (e.g. cytopenia) are increasingly needed. Complex and expensive diagnostic methods e.g. optical flowcytometry are available in university hospitals or larger cancer centers only. These methods are usually cost-intensive, time-consuming and require highly trained personnel. The goal of this MRCyte Consortium was therefore to develop a cost-effective, easy-to-operate magnetic flowcytometer on the basis of a microfluidic-cartridge as sensor for detection of blood-platelets, CD4+ T cells, leukemic and tumor cells in whole blood. Specifically, in this MRCyte subproject the goal was to give clinical input in the early development of magnetic flowcytometry, to make model cells and blood probes available for technology testing, as wells as performing a benchmark of the experimental MRCyte magnetic flowcytometry with gold standard technology of fluorescence flowcytometry (Lab Ref. 15).

Picture AG Bosch with Prof. Muramatsu, discoverer of midkine

 

Laboratory Members

Barbara Bock, MTA
Margarete Karg, PhD (2018)
Lukas John, Medical Student

Picture AG Bosch reunion


Laboratory Alumni

Sabine Britting, PhD (2013)
Jonas Sommer, Medical Student
Julia Kittler, Medical Student
Teresa Bösl, Molecular Science Master Student
Mirjam Fass, Medical Student
Agnes Hahn, Medical Student

Funding

  • German Research Council (DFG)
  • German Federal Ministry of Education and Research (BMBF)
  • Interdisciplinary Center for Clinical Research Erlangen (IZKF) and ELAN-Program, Erlangen


External Advisors

  • Prof. Dr. S. Ostrand-Rosenberg, Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, USA
  • Prof. Dr. C.J.M. Melief, ISA Pharmaceuticals, Leiden, The Netherlands
  • Prof. Dr. M.J. Jager, Department of Ophthalmology, Leiden University Medical Center, Leiden, The Netherlands
  • Prof. Dr. B.R. Ksander, The Schepens Eye Research Institute and Massachusetts Eye & Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, MA, USA
  • Prof. Dr. A.F. Cohen, Centre for Human Drug Research, Leiden, The Netherlands


Laboratory Key Publications

  1. Bosch JJ and Ksander BR (2003). Immunology of uveal melanoma; adaptive immunity and basis for immuno-therapy. Ocular Oncology. 11:231-253.
  2. Bosch JJ (2012). Immunotherapy of uveal melanoma. Dev Ophthalmol. 49:137-49. DOI: 10.1159/000328268
  3. Bosch JJ, Heindl LM. [Novel Adjuvant Therapy for Ocular Melanoma]. Klin Monbl Augenheilkd. 2017 May;234(5):670-673. DOI: 10.1055/s-0043-107807
  4. Bosch JJ, Thompson JA, Srivastava MK, Iheagwara UK, Murray TG, Lotem M, Ksander BR, Ostrand-Rosenberg S (2007). MHC class II-transduced tumor cells originating in the immune-privileged eye prime and boost CD4(+) T lymphocytes that cross-react with primary and metastatic uveal melanoma cells. Cancer Res. 67(9):4499-506. DOI: 10.1158/0008-5472.CAN-06-3770
  5. Kittler JM, Sommer J, Fischer A, Britting S, Karg MM, Bock B, Atreya I, Heindl LM, Mackensen A, Bosch JJ (2019). Characterization of CD4+ T cells primed and boosted by MHCII primary uveal melanoma cell-based vaccines. Oncotarget. 10(19): 1812-1828. DOI: 10.18632/oncotarget.26737
  6. Bosch JJ, Iheagwara UK, Reid S, Srivastava MK, Wolf J, Lotem M, Ksander BR, Ostrand-Rosenberg S (2010). Uveal melanoma cell-based vaccines express MHC II molecules that traffic via the endocytic and secretory pathways and activate CD8+ cytotoxic, tumor-specific T cells. Cancer Immunol Immunother. 59(1):103-12. DOI: 10.1007/s00262-009-0729-0
  7. Haile ST*, Bosch JJ*, Agu NI, Zeender AM, Somasundaram P, Srivastava MK, Britting S, Wolf JB, Ksander BR, Ostrand-Rosenberg S (2011). Tumor cell programmed death ligand 1-mediated T cell suppression is overcome by coexpression of CD80. J Immunol. 186(12): 6822-9. DOI: 10.4049/jimmunol.1003682
  8. Heindl LM, Hofmann-Rummelt C, Adler W, Bosch JJ, Holbach LM, Naumann GO, Kruse FE, Cursiefen C (2011). Tumor-associated lymphangiogenesis in the development of conjunctival melanoma. Invest Ophthalmol Vis Sci. 52(10):7074-83. DOI: 10.1167/iovs.11-7902
  9. Heindl LM, Hofmann-Rummelt C, Adler W, Bosch JJ, Holbach LM, Naumann GO, Kruse FE, Cursiefen C (2011). Prognostic significance of tumor-associated lymphangiogenesis in malignant melanomas of the conjunctiva. Ophthalmology. 118(12):2351-60. DOI: 10.1016/j.ophtha.2011.05.025
  10. Schlereth SL, Iden S, Mescher M, Ksander BR, Bosch JJ, Cursiefen C, Heindl LM (2015). A Novel Model of Metastatic Conjunctival Melanoma in Immune-Competent Mice. Invest Ophthalmol Vis Sci. 56(10):5965-73. DOI: 10.1167/iovs.15-17290
  11. Refaian N, Schlereth SL, Koch KR, Notara M, Hos D, Mescher M, Iden S, Bosch JJ, Jager MJ, Cursiefen C, Heindl LM (2015). Comparing the Hem- and Lymphangiogenic Profile of Conjunctival and Uveal Melanoma Cell Lines. Invest Ophthalmol Vis Sci. 56(9): 5691-7. DOI: 10.1167/iovs.15-16829
  12. Koch KR, Refaian N, Hos D, Schlereth SL, Bosch JJ, Cursiefen C, Heindl LM (2014). Autocrine impact of VEGF-A on uveal melanoma cells. Invest Ophthalmol Vis Sci. 55(4): 2697-704. DOI: 10.1167/iovs.13-13254
  13. Grajewski RS, Bosch JJ, Bruns H, Cursiefen C, Heindl LM (2016). The Trojan Horse Tale Revisited: An Eye on Metastatic Spread of Carcinoma Cells. Cancer Immunol Res.4:92-4. DOI: 10.1158/2326-6066.CIR-15-0127
  14. Charwat V, Rothbauer M, Tedde SF, Hayden O, Bosch JJ, Muellner P, Hainberger R, Ertl P (2013). Monitoring dynamic interactions of tumor cells with tissue and immune cells in a lab-on-a-chip. Anal Chem. 85(23):11471-8. DOI: 10.1021/ac4033406
  15. Helou M, Reisbeck M, Tedde SF, Richter L, Bär L, Bosch JJ, Stauber RH, Quandt E, Hayden O (2013). Time-of-flight magnetic flow cytometry in whole blood with integrated sample preparation. Lab Chip. 13(6):1035-8. DOI: 10.1039/c3lc41310a


Ocular Melanoma Foundation

Jacobus Bosch is a Member of the Medical Advisory Board of the Ocular Melanoma Foundation (http://www.ocularmelanoma.org)

 
Contact
Dr. med. Jacobus Bosch
e-mail: jacobus.bosch@uk-erlangen.de
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Comprehensive Cancer Center
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