AHR-SASP: Aryl hydrocarbon receptor ligands as clinically accessible biomarkers of cellular senescence and modulators of the senescence-associated secretory phenotype

• Applicant: Dr. Anne Großkopf
• Subject Area: Biogerontology and Geriatric Medicine
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 570620404

Life expectancy is increasing globally, but the healthy lifespan is not keeping pace. As a result, the period with age-related disease is expanding, making healthy aging a key medical priority. Cellular senescence and chronic inflammation are central hallmarks of aging. Senescent cells secrete pro-inflammatory molecules, termed senescence-associated secretory phenotype (SASP). While SASP proteins have gained attention as senescence biomarkers, their detection is limited by technical challenges, specificity, availability of assays, and costs. In contrast, metabolites are more easily quantified and respond more readily to interventions. Recent studies suggest that ligands of the aryl hydrocarbon receptor (AHR), especially tryptophan-derived metabolites, are important predictors of biological age. Preliminary work has shown that bread crust extract and various AHR ligands activate AHR in cell lines, with ligand- and cell-specific responses. Functional AHR knockouts in A549 and EA.hy 926 cell lines were previously established via CRISPR/Cas9, as is stress-induced senescence, β-galactosidase detection, and qRT-PCR of senescence markers and SASP factors. Furthermore, 13 SASP factors were measured and used to stratify a high- and low-SASP group in serum samples from the CARLA cohort study. This one-year project aims to explore the physiological relevance of AHR and its ligands concerning cellular senescence and the SASP and to assess their potential as clinically accessible biomarkers. First, this project will investigate the influence of AHR presence and ligand-specific activation on the induction of cellular senescence and SASP modulation in the endothelial cell line EA.hy 926. Using established AHR-knockout and control clones, senescence will be induced, followed by the analysis of cell cycle inhibitors (p16, p21), β galactosidase activity, nuclear envelope erosion, and SASP factor expression. Ligand treatments will include selected tryptophan metabolites and environmental AHR agonists. Second, the AHR-activation potential of ~500 serum samples with predefined high or low SASP status from the CARLA cohort will be assessed using HepG2-AHR, NRF2, and NFκB luciferase reporter cell lines. These assays will evaluate whether the cumulative ligand profile in human serum differentially activates signaling pathways and whether they correlate with the SASP status. Third, targeted metabolomics will be performed on 100 selected serum samples (50 SASP-high and 50 SASP-low) to quantify a panel of 15 tryptophan-derived AHR-(pro)ligands using LC-MS/MS. These data will be correlated with reporter cell activation and SASP protein levels. In summary, this project will contribute to understanding how AHR and its ligands influence cellular senescence and the SASP and whether these metabolites may be regulators and biomarkers of senescence.

• DFG Programme: Research Grants