Tumor microenvironment-directed epigenome remodeling of tumor infiltrating T cells

• Applicant: Dr. Christian Schmidl
• Subject Area: Immunology
• Term: from 2019 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 420273313

Tumor infiltrating lymphocytes (TILs) are often dysfunctional in the tumor microenvironment (TME), which prevents them from killing cancer cells. This dysfunctional state is thought to be molecularly fixed by a distinct epigenetic program, which hinders durable T cell reprogramming by immunotherapy, and could explain why currently only a minority of patients benefits from so-called checkpoint inhibitors. A chronic T cell receptor (TCR) stimulation was suggested as the main driver of epigenetic fixation of T cell dysfunction irrespective of signals from the TME. However, these studies are confounded by very strong TCR stimulation that likely overrides any other signal the T cells receive. Together with the fact that many studies document a significant impact of costimulatory and coinhibitory signals from the TME on T cell function, I want to challenge the current working model, and prove the hypothesis of tumor microenvironment-directed epigenome remodeling in T cells.I propose to employ a murine transplanted tumor model that allows us to provide low, intermediate and high TCR stimulation in combination with controlled, inducible expression of coinhibitory and costimulatory signals to T cells. By using single-cell chromatin mapping methods, we will identify signal-dependent epigenetic signatures in heterogeneous immune responses, and identify the underlying gene-regulatory programs. We will then delete candidate transcription factors and epigenetic modifiers to dissect their function in establishing and maintaining signal-specific T cell (dys-)functional states in vivo by using a novel single-cell CRISPR method that allows the investigation of hundreds to thousands of knockouts in parallel. With this we will identify concrete targets to enhance immunotherapy and prevent T cell dysfunction, but also provide basic mechanistic insights into the specific molecular pathways that regulate gene expression in T cells. Finally, we will develop high-throughput paired T cell receptor sequence and chromatin accessibility profiling from the same single cells, which allows investigating tumor-directed epigenome remodeling of T cells in primary tumor tissues in relation to their T cell receptor sequence. With the proposed work plan I will establish the concept of tumor microenvironment-directed epigenome remodeling in tumor infiltrating lymphocytes. The acquired data will significantly improve our basic understanding of T cell (dys-)function in tumors, and reveal pathways that establish, maintain, and overcome epigenetically “fixed” T cell states. With the newly developed methods from this proposal for paired T cell receptor and epigenome sequencing, we can leverage our findings from mouse models to primary patient samples. As an outlook, the proposal will set the base for developing molecular markers that predict response to immunotherapy, and reveal novel targets to enhance T cell function for clinical applications.

• DFG Programme: Research Grants