The leucine-rich repeat kinase 2 (LRRK2) is commonly mutated in familial and sporadic Parkinson’s disease. Merck LRRK2 inhibitor-2 (MLi-2) is a potent and selective LRRK2 inhibitor with an IC50 value of 0.76nM in a purified LRRK2 kinase assay in vitro. It also exhibited high potency in a cellular assay monitoring the dephosphorylation of pSer935 LRRK2 and a radioligand competition binding assay with IC50 values of 1.4nM and 3.4nM, respectively. In mice, oral administration of MLi-2 dose-dependently reduced phosphorylation of Ser935 with the maximal suppression (>90% reduction) observed at 10mg/kg and higher. The in-diet administration of MLi-2 (3–120mg/kg/day) for 11 days significantly reduced LRRK2 protein level in the kidneys of mice. Treatment with MLi-2 (30mg/kg/day) markedly inhibited pSer935 LRRK2 in the brains of MitoPark mice at 3, 9, and 15 weeks of treatment compared with the vehicle-treated group, but did not stop the progressive motor phenotype or prevented the decline in striatal dopamine, DOPAC, or tyrosine hydroxylase levels in MitoPark mice[3].
MLi-2 细胞实验
Cell Line
Concentration
Treated Time
Description
References
Human Embryonic Kidney 293 cells (HEK293)
1 μM
4 hours
To investigate the effect of MLi-2 on 14-3-3 binding to LRRK2. Results showed that 1 μM MLi-2 completely ablated 14-3-3 binding and Ser935 phosphorylation.
Biochem J. 2021 Oct 15;478(19):3555-3573
Mouse Embryonic Fibroblasts (MEFs)
100 nM
2 hours
To evaluate the inhibitory effect of MLi-2 on LRRK2-mediated Rab10 phosphorylation. Results showed that 1 μM MLi-2 reduced Rab10 and Rab12 phosphorylation to almost background levels.
Biochem J. 2021 Oct 15;478(19):3555-3573
HEK293FT cells
1μM
90 minutes
To evaluate the inhibitory effect of MLi-2 on LRRK2 kinase activity, results showed that MLi-2 significantly reduced Rab10 phosphorylation levels.
Mol Neurodegener. 2021 Mar 19;16(1):17
primary hippocampal neurons
10 nM or 30 nM
7 days
Reduced LRRK2 kinase activity increases α-synuclein overlap with presynaptic markers in primary neurons
Acta Neuropathol Commun. 2021 Nov 8;9(1):180
Wild-type and G2019S-LRRK2 mouse microglia
1 µM
24 hours
To investigate the inhibitory effect of MLi-2 on LRRK2 kinase activity and its impact on gene expression. Results showed that MLi-2 significantly inhibited zymosan-induced LRRK2 Ser935 phosphorylation.
Cells. 2023 Dec 26;13(1):53
MLi-2 动物实验
Species
Animal Model
Administration
Dosage
Frequency
Description
References
Mice
G2019S LRRK2 knock-in (KI) mice
In-diet administration
60 mg/kg/day
Once daily for 10 weeks
To evaluate the molecular effects of chronic LRRK2 kinase inhibition, results showed that MLi-2 significantly reduced S1292 LRRK2 and S106 Rab12 phosphorylation levels and affected endolysosomal and mitochondrial pathways.
Mol Neurodegener. 2021 Mar 19;16(1):17
Mice
LRRK2G2019S knock-in mice
Dietary administration
0, 10, or 60 mg/kg/day
Continuous for 3-6 months
To assess the impact of LRRK2 kinase inhibition on tau pathology progression. Results showed that LRRK2 kinase inhibition reversed tau pathology progression in LRRK2G2019S mice but had minimal effect in wild-type mice.
Transl Neurodegener. 2024 Mar 4;13(1):13
Mice
Parkinson's disease model
Dietary administration
30 mg/kg/day
Continuous for 3 months
To assess the effect of MLi-2 on α-synuclein pathology and neuron death. Results showed MLi-2 significantly reduced LRRK2 kinase activity but did not improve motor function, α-synuclein pathology accumulation, or neuron loss.
Acta Neuropathol Commun. 2019 Feb 26;7(1):28
Mice
Vps35 p.D620N knock-in (VKI) mice
Intraperitoneal injection
5 mg/kg
Once daily for 7 days
MLi-2 significantly abolished Rab phosphorylation in VKIs with a clear interaction between treatment and genotype, further supporting that hyperphosphorylation of Rab proteins in VKIs are LRRK2 kinase dependent.
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