Overcoming immunosenescence: strategies to modulate DNA damage and senescence in immune cells and fibroblasts

Thesis

Growing evidence suggests that DNA damage and cellular senescence govern age-related immune dysfunction, termed immunosenescence. Interventions that prevent the harmful consequences of DNA damage (genoprotectors) are therefore an attractive target to improve older people’s immunity. Low-dose mTOR inhibition is a potent anti-ageing strategy and improves adaptive immune responses in older people, though how this intervention affects DNA damage and senescence in stromal and immune cells is unclear. In this thesis, I first optimised and validated DNA damage-induced senescence (DDIS) in primary human fibroblasts and observed that these express high levels of WRN, a DNA repair factor. Using phage display, I identified several WRN-interacting sequences that resemble human proteins involved in mTOR signalling and protein synthesis. Accordingly, both DDIS fibroblasts and age-associated peripheral immune subsets displayed high levels of mTOR activity and p53/p21-dependent DDR signalling. Continuous treatment with a pan- mTOR inhibitor, AZD8055, rescued a wide range of senescence phenotypes in DDIS fibroblasts, including the presence of DNA damage foci, suggesting that mTOR inhibition is genoprotective. To test whether this also occurred in immune cells, I developed a DNA damage assay in primary human T cells and found that both rapamycin and AZD8055 prevented the accumulation of DNA lesions, reduced DDR signalling, and increased cell survival after genotoxic treatment. Using an optimised autophagy and DNA damage assay, I observed that mTOR inhibition may elicit these genoprotective effects by inducing autophagy. Finally, I report the results from the UK’s first single-blind, placebo-controlled trial of the impact of low-dose rapamycin in older adults on features of immunosenescence, using 27-colour spectral flow cytometry. Here, rapamycin treatment decreased the expression of immune exhaustion markers and p21 across immune subsets. These data support the notion that mTOR inhibition is a genoprotective strategy and provide a mechanism for its gero- and immunoprotective effects in older people.

Authors: Kell, L

• DOI: 10.5287/ora-dpx4g99a5
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: ageing; senescence; immunosenescence; mTOR inhibition; immune system ageing; DNA damage; mTOR activity; cellular senescence; rapamycin; DNA repair
• Subjects: Old age; Immunosenescence; Immunity; T cells; DNA damage; Aging; Immunology; DNA repair