| Autor: |
Kluss JH; Cell Biology and Gene Expression Section, Laboratory of Neurogenetics, National Institute on Aging, National Institutes of Health, Bethesda, MD, USA.; School of Pharmacy, University of Reading, Whiteknights Campus, Reading, UK., Mazza MC; Cell Biology and Gene Expression Section, Laboratory of Neurogenetics, National Institute on Aging, National Institutes of Health, Bethesda, MD, USA., Li Y; Proteomic Core Facility, National Institute of Neurological Disorders and Stroke, Bethesda, MD, USA., Manzoni C; School of Pharmacy, University of Reading, Whiteknights Campus, Reading, UK.; UCL School of Pharmacy, Brunswick Square, London, UK., Lewis PA; School of Pharmacy, University of Reading, Whiteknights Campus, Reading, UK.; Royal Veterinary College, Royal College Street, London, UK.; Department of Neurodegenerative Diseases, UCL Queen Square Institute of Neurology, London, UK., Cookson MR; Cell Biology and Gene Expression Section, Laboratory of Neurogenetics, National Institute on Aging, National Institutes of Health, Bethesda, MD, USA. cookson@mail.nih.gov., Mamais A; Department of Neuroscience, Center for Translational Research in Neurodegenerative Disease, Norman Fixel Institute for Neurological Diseases, University of Florida College of Medicine, Gainesville, Florida, USA. |
| Abstrakt: |
The most common mutation in the Leucine-rich repeat kinase 2 gene (LRRK2), G2019S, causes familial Parkinson's Disease (PD) and renders the encoded protein kinase hyperactive. While targeting LRRK2 activity is currently being tested in clinical trials as a therapeutic avenue for PD, to date, the molecular effects of chronic LRRK2 inhibition have not yet been examined in vivo. We evaluated the utility of newly available phospho-antibodies for Rab substrates and LRRK2 autophosphorylation to examine the pharmacodynamic response to treatment with the potent and specific LRRK2 inhibitor, MLi-2, in brain and peripheral tissue in G2019S LRRK2 knock-in mice. We report higher sensitivity of LRRK2 autophosphorylation to MLi-2 treatment and slower recovery in washout conditions compared to Rab GTPases phosphorylation, and we identify pS106 Rab12 as a robust readout of downstream LRRK2 activity across tissues. The downstream effects of long-term chronic LRRK2 inhibition in vivo were evaluated in G2019S LRRK2 knock-in mice by phospho- and total proteomic analyses following an in-diet administration of MLi-2 for 10 weeks. We observed significant alterations in endolysosomal and trafficking pathways in the kidney that were sensitive to MLi-2 treatment and were validated biochemically. Furthermore, a subtle but distinct biochemical signature affecting mitochondrial proteins was observed in brain tissue in the same animals that, again, was reverted by kinase inhibition. Proteomic analysis in the lung did not detect any major pathway of dysregulation that would be indicative of pulmonary impairment. This is the first study to examine the molecular underpinnings of chronic LRRK2 inhibition in a preclinical in vivo PD model and highlights cellular processes that may be influenced by therapeutic strategies aimed at restoring LRRK2 physiological activity in PD patients. |
