Barbara Bock, Dr. Jacobus Bosch, Dr. Margarethe Karg
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).
Barbara Bock, MTA
Margarete Karg, PhD (2018)
Lukas John, Medical Student
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
Ocular Melanoma Foundation
Jacobus Bosch is a Member of the Medical Advisory Board of the Ocular Melanoma Foundation (http://www.ocularmelanoma.org)