Characterising senescence in iPSC-derived glial cells from amyotrophic lateral sclerosis patients expressing the C9orf72 mutation.

Abstract

ALS is a motor neuron disease characteristic of upper and lower motor neuron degeneration, with limited treatment options and no cure. Senescence, a stress state cells enter in response to DNA damage, has recently been identified as a new opportunity for ALS drug discovery. Under chronic conditions, senescence contributes to neuroinflammation, blood-brain barrier (BBB) degeneration, and neuronal death. Upregulated markers of senescence in astrocytes and microglia have been identified in SOD1 ALS animal models and post-mortem brain tissue analysis of ALS patients. However, the cell autonomous nature of senescence development in astrocytes and microglia, the predominant cell type involved in senescence, and their exact role in neurodegeneration, remains unclear. Lastly, the most common ALS mutation, the C9orf72 HRE mutation, has not been well represented in ALS models. Current senolytics have poor BBB permeability and target proteins in a way that may be detrimental to motor neuron viability in ALS.

This thesis sought to characterise senescence in ALS glial cells and to examine the impact of small molecule compound derivatives based off an existing non-BBB-penetrating senolytic, ABT-737. We successfully characterised senescence in iPSC-derived C9orf72 HRE ALS astrocytes for the first time, determining they exhibited a unique senescence signature and upregulation of a key protein, Bcl-xL. Furthermore, we showed that iPSC-derived C9orf72 HRE ALS microglia-like cells do not display upregulated markers of senescence or inflammation, and identified limitations associated with this modelling approach. Finally, we determined that truncation of ABT-737 completely diminished compound binding to target proteins, yet identified a new potential drug lead that increased ALS cell viability through an unknown mechanism independent of senescence. Overall, this thesis has contributed to an understanding of senescence in ALS and will inform future iterative drug design approaches.