In-depth mass-spectrometry reveals phospho-RAB12 as a blood biomarker of G2019S LRRK2-driven Parkinson

Leucine-rich repeat kinase 2 (LRRK2) inhibition is a promising disease-modifying therapy for LRRK2-associated Parkinsons disease (L2PD) and idiopathic PD (iPD). Yet, pharmaco-dynamic readouts and progression biomarkers for disease modification clinical trials are insufficient. Employing phospho-/proteomic analyses we assessed the impact that LRRK2 activating mutations had in peripheral blood mononuclear cells (PBMCs) from a LRRK2 clinical cohort from Spain (n=174) encompassing G2019S L2PD patients (n=37), non-manifesting LRRK2 mutation carriers of G2019S, here, G2019S L2NMCs (n=27), R1441G L2PD patients (n=14), R1441G L2NMCs (n=11), iPD (n=40), and controls (n=45). We identified 207 differential proteins in G2019S L2PD compared to controls (39 up/ 168 down) and 67 in G2019S L2NMCs (10 up/ 57 down). G2019S down-regulated proteins affected the endolysosomal pathway, proteostasis and mitochondria, e.g., ATIC, RAB9A, or LAMP1. At the phospho-proteome level, we observed increases in endogenous phosphorylation levels of pSer106 RAB12 in G2019S carriers, which were validated by immunoblotting after 1 year of follow-up (n=48). Freshly collected PBMCs from 3 G2019S L2PD, 1 R1441G L2PD, 1 iPD, and 5 controls (n=10) showed strong diminishment of pSer106 RAB12 phosphorylation levels after in-vitro administration of the MLi-2 LRRK2 inhibitor. Using machine learning, we identified an 18-feature G2019S phospho-/protein signature capable of discriminating G2019S L2PD, L2NMCs, and controls with 96% accuracy that correlated with disease severity, i.e., UPDRS-III motor scoring. Our study identified pSer106 RAB12 as an endogenous biomarker in easily accessible PBMCs from G2019S carriers and suggests that phospho-/proteomic findings in human PBMCs such as pSer106 RAB12 can be deployed as a universal pharmaco-dynamic readout for L2PD, L2NMCs, and iPD. Future work may determine whether pSer106 RAB12 could help with patient enrichment and monitoring drug efficacy in LRRK2 clinical trials.