Abstract
There is compelling evidence for the role of the leucine-rich repeat kinase 2 (LRRK2) and in particular its kinase function in Parkinson's disease. Orally bioavailable, brain penetrant and potent LRRK2 kinase inhibitors are in the later stages of clinical development. Here, we describe a facile and robust assay to quantify LRRK2 kinase pathway activity by measuring LRRK2-mediated phosphorylation of Rab10 in human peripheral blood neutrophils. We use the selective MJFF-pRab10 monoclonal antibody recognising the Rab10 Thr73 phospho-epitope that is phosphorylated by LRRK2. We highlight the feasibility and practicability of using our assay in the clinical setting by studying a few patients with G2019S LRRK2 associated and sporadic Parkinson's as well as healthy controls. We suggest that peripheral blood neutrophils are a valuable resource for LRRK2 research and should be considered for inclusion in Parkinson's bio-repository collections as they are abundant, homogenous and express relatively high levels of LRRK2 as well as Rab10. In contrast, the widely used peripheral blood mononuclear cells are heterogeneous and only a minority of cells (monocytes and contaminating neutrophils) express LRRK2. While our LRRK2 kinase pathway assay could assist in patient stratification based on LRRK2 kinase activity, we envision that it may find greater utility in pharmacodynamic and target engagement studies in future LRRK2 inhibitor trials.
Highlights
Parkinson’s disease is a common and complex neurodegenerative disorder affecting ∼1% of people over the age of 65 [1]
We argue that neutrophils are better suited than peripheral blood mononuclear cells (PBMCs) to study leucine-rich repeat kinase 2 (LRRK2) regulated Rab10 phosphorylation in human peripheral blood
To gain insights into the expression of the LRRK2 and Rab10 proteins in human peripheral blood cells, we interrogated the IMMPROT proteomic database [34]. This suggests that LRRK2 is highly expressed in neutrophils (∼2 × 105 copies per cells) as well as monocytes (∼1 × 105 copies per cells), but is virtually undetectable in B and T lymphocytes as well as natural killer and dendritic cells that constitute most of the PBMCs (Figure 1)
Summary
Parkinson’s disease is a common and complex neurodegenerative disorder affecting ∼1% of people over the age of 65 [1]. The leucine-rich repeat kinase 2 (LRRK2) gene is one of the main genetic contributors and was first discovered to be associated with Parkinson’s in 2004 [5,6]. It is a large (2527 residues, 286 kDa) multi-domain protein including a ROC/COR GTPase and kinase catalytic domains [5,6]. LRRK2 constitutes a pleomorphic risk factor for developing Parkinson’s, and links familial and sporadic forms of the disease [7]. There are common protein-coding and non-protein-coding variants at the LRRK2 locus that moderately increase the risk for developing Parkinson’s [11]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
