The role of protein lactylation in the regulation of CD8+ T cell function in the tumor microenvironment

• Applicant: Dr. Christian Schmidl
• Subject Area: Immunology
• Term: from 2019 to 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 420273313

The tumor microenvironment (TME) is characterized by an accumulation of lactic acid (LA), which has detrimental effects on the anti-tumor response. However, it is unclear how LA impairs tumor infiltrating T cells (TILs) on the molecular level. Recent discoveries established the link between lactylation of lysines (Kla) in histone-tails and gene-expression. Subsequently, the regulation of protein function by non-histone Kla was described. However, the extent of these phenomena is currently completely unknown in human T cells. Re-analysis of human TIL data and new preliminary experiments show cell state-specific regulation of lactate-related metabolic processes and modulation of both histone and non-histone protein lactylation, indicating a direct impact of Kla on human T cell function. With this project, we want to fill the critical knowledge gap on the extent, cell-state-specificity, and functional impact of protein lactylation in human T cells, with a focus on cancer immunology. We will therefore use established proteomic approaches to investigate the extent of Kla globally, and systems biology methods to explore the specific aspect of histone-lactylation and its impact on gene-regulation in detail. By using ex vivo samples and vitro models combined with inhibitors of metabolic pathways, we will reveal how intrinsic factors such as the developmental and activation state shape Kla patterns in human CD8+ T cells. We further hypothesize that exogeneous lactic acid from the TME interplays with intrinsic pathways of Kla with direct impact on the CD8+ T cells’ anti-tumor response. Here, we will investigate Kla patterns in T cells co-cultured with tumor spheroids that have been genetically engineered to secrete high or low levels of LA and confirm results in mouse tumor models mirroring TMEs with high or low LA accumulation. In addition, we will explore the possibility to impact T cell function by manipulating Kla. Using small molecule inhibitors, we will inhibit enzymes that transport lactate or set/erase lactylation marks to modulate lactylation globally. Further, with CRISPR base editing we will also mutate specific lysine residues in proteins that we find differentially lactylated and test the impact of these modifications on the anti-tumor activity on gene-edited T cells. In summary this project’s objective is to prove the hypothesis that protein lactylation in CD8+ T cells impacts their function in a specific and context-dependent manner. Our results will suggest and validate concrete molecular mechanisms of action on how Kla exerts its effects, and to use this knowledge to improve immunotherapeutic applications as a future perspective.

• DFG Programme: Research Grants