Sponsor: DFG (Project 27, SFB854 “Molecular Organization of Cellular Communication within the Immune System”)

 In the past few years immunotherapeutic approaches have revolutionized cancer treatment. Adoptive T cell therapies and monoclonal antibodies directed against the immunoregulatory receptors CTLA4 and PD-L1/PD-1 could induce durable responses in a subset of patients with advanced melanoma and subsequently in a variety of other cancer types. However, many patients show either primary or adaptive resistance to these novel therapies, indicating substantial medical need to better understand the underlying mechanisms and develop more effective treatment protocols. Recently, the role of CD4⁺ T cells has received increasing attention in this highly active research field. While CD4⁺ effector T cell subpopulations can participate in anti-tumor immunity, both natural and induced CD4⁺ Treg subpopulations can orchestrate an immunosuppressive environment and promote tumor growth. Furthermore, evidence is emerging that naïve CD4⁺ T cell possess remarkable functional plasticity allowing them to rapidly adapt to the requirements in local tissue environments and to acquire effector, memory and regulatory phenotypes.

The principal goal of A27 is to better understand the molecular and cellular mechanisms governing the phenotype and function of CD4⁺ T cells in tumor tissues during immunotherapeutic intervention in vivo using experimental mouse melanoma models. We hypothesize that progressively growing melanomas create a T cell-suppressive inflammatory microenvironment that shifts CD4⁺ effector T cells towards anergic, memory and regulatory phenotypes. This shift is likely promoted by tumor cells and by myeloid immune cells which are recruited into tumors in response to tissue injury. In our work we will focus on the role of neutrophils because we discovered in the current funding period that they are reactively recruited from the bone marrow in response to different cancer immunotherapies and limit the induction of anti-tumoral CD8⁺ T cells. We found that neutrophils rapidly acquire an immunosuppressive, protumorigenic phenotype in T cell-inflamed melanoma tissues. To demonstrate interactions between tumor specific CD8⁺ T cells and neutrophils we took advantage of the highly neutrophil-specific and sensitive catchup^IVM fluorescent reporter mouse model established in the CRC854. 

After our move to Magdeburg we optimized a comparable treatment protocol that included the use of melanocyte-specific TCR-transgenic CD4⁺ T cells and heterologous prime boost vaccination. In the 3^rd funding period we will use all these tools to examine how anti-tumoral CD4⁺ T cells are regulated by neutrophils and Tregs in melanoma-bearing mice in vivo. We will use CRISPR/Cas9 genome editing of our Hgf-Cdk4 mouse melanoma lines to generate variants with antigen loss and variants expressing model neo-antigens. Using these cells we will dissect the anti-tumoral effector mechanisms of CD4⁺ T cells through detailed spatial and temporal analyses of tumor tissues focusing on their interaction dynamics with tumor cells, neutrophils and Tregs. In cooperation with CRC854 members we will expand our experimental in vivo model by incorporating novel fluorescent reporter systems for proximal TCR signaling in adoptively transferred T cells and using intravital imaging. These experiments will open up the possibility to not only investigate the interaction dynamics of cells of the immune system, but also to obtain information on the underlying molecular signaling processes. Taken together, these experiments will provide important insights how effector functions of CD4⁺ T cells could be harnessed for the treatment of melanoma.