Abstract by Josefine Fussing Tengberg

Leucine-rich repeat kinase 2 (LRRK2) is genetically associated with Parkinson’s disease (PD) through both rare coding and common noncoding variants. LRRK2 represents one of the most prevalent genetic risk factors for PD, the second most common neurodegenerative disorder worldwide. Extensive experimental and genetic evidence has implicated lysosomal dysfunction in PD pathogenesis, particularly highlighting the convergence of lysosome-associated genes with PD risk. LRRK2 is one of those lysosome-associated genes and has similarly been implicated in autophagy, phagocytosis, vesicular trafficking, and various lysosomal functions, especially in lysosomal damage response. Pathogenic mutations in LRRK2 increase its kinase activity leading numerous pharmaceutical companies to explore LRRK2 inhibitors as potential treatments for PD, especially LRRK2-related PD.

In the first study in this thesis, the research was focused on lysosomal damage response induced by lysosomotropic drugs in the RAW 264.7 mouse macrophage-like cell line. Lysoso-motropic drugs induce lysosomal rupture, releasing lysosomal proteases into the cytosol and initiating cell death. Many of these drugs additionally activates LRRK2. Prolonged treatment of lysosomotropic drugs like L-leucyl-L-leucine O-methyl ester (LLOMe) resulted in significant cell death. The selective LRRK2 kinase inhibitor, MLi-2, attenuated LLOMe-induced cell death in RAW 264.7 macrophages. Similarly, siRNA-mediated knockdown of LRRK2 and its substrate RAB8A reduced cell death in response to LLOMe. Thus, inhibiting LRRK2 and RAB8A signalling could partially rescue cell death specific to lysosomes.

Functionally validated transcriptome profiling revealed that LRRK2 kinase inhibition modulated cholesterol metabolism and mTOR pathways in response to LLOMe. Additionally, shotgun lipidomics indicated that LRRK2 kinase inhibition could regulate specific lipid species within lysosomes during LLOMe treatment. Thus, impaired LRRK2 signalling may confer a protective effect on lysosomes and enhance cell viability in response to lysosomal stress.

In the second study, the investigation focused on the phagocytic handling of α-synuclein (αSyn) preformed fibrils (PFFs) in proinflammatory stimulated microglia derived from human induced pluripotent stem cells (iPSCs) from healthy and LRRK2 PD patient donors. Treatment with inter-feron-γ (IFNγ) led to a substantial increase in expression and activity of LRRK2. Additionally, IFNγ markedly reduced αSyn PFF phagocytosis, which was not observed with other proinflam-matory cytokines, while lipopolysaccharide (LPS) increased phagocytosis. Cotreatment with MLi-2 reversed the effects of IFNγ on phagocytosis.

In conclusion, LRRK2 inhibition reduced cell death in mouse macrophages subjected to lysoso-mal stress and enhanced αSyn phagocytosis in human iPSC microglia under inflammatory con-ditions. Both studies emphasise the link between the endolysosomal system, LRRK2 and PD.