MLi-2

Catalog No.S9694 Batch:S969401

Technical Data

Formula	C₂₁H₂₅N₅O₂
Molecular Weight	379.46	CAS No.	1627091-47-7
Solubility (25°C)*	In vitro	DMSO	76 mg/mL (200.28 mM)
		Water	76 mg/mL (200.28 mM)
		Ethanol	8 mg/mL (21.08 mM)
* <1 mg/ml means slightly soluble or insoluble. * Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations. * Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

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Biological Activity

Description

MLi-2 is an orally active and highly selective inhibitor of LRRK2. MLi-2 exhibits exceptional potency in a purified LRRK2 kinase assay in vitro with IC50 of 0.76 nM, a cellular assay monitoring dephosphorylation of LRRK2 pSer935 LRRK2 with IC50 of 1.4 nM, and a radioligand competition binding assay with IC50 of 3.4 nM.

Targets

LRRK2 ^[1] (purified LRRK2 kinase assay in vitro)	LRRK2 ^[1] (cellular assay monitoring dephosphorylation of LRRK2 pSer935 LRRK2)	LRRK2 ^[1] (radioligand competition binding assay)
0.76 nM	1.4 nM	3.4 nM

In vitro

MLi-2 is a structurally novel, highly potent, and selective LRRK2 kinase inhibitor with central nervous system activity. MLi-2 exhibits exceptional potency in a purified LRRK2 kinase assay in vitro (IC50 = 0.76 nM), a cellular assay monitoring dephosphorylation of LRRK2 pSer935 LRRK2 (IC50 = 1.4 nM), and a radioligand competition binding assay (IC50 = 3.4 nM). MLi-2 has greater than 295-fold selectivity for over 300 kinases in addition to a diverse panel of receptors and ion channels.^[1]

In vivo

Acute oral and subchronic dosing in MLi-2 mice resulted in dosedependent central and peripheral target inhibition over a 24-hour period as measured by dephosphorylation of pSer935 LRRK2. Treatment of MitoPark mice with MLi-2 is well tolerated over a 15-week period at brain and plasma exposures >100× the in vivo plasma IC50 for LRRK2 kinase inhibition as measured by pSer935 dephosphorylation. Morphologic changes in the lung, consistent with enlarged type II pneumocytes, are observed in MLi-2-treated MitoPark mice.^[1]

Protocol (from reference)

Cell Assay:^{^[1]}	Cell lines SH-SY5Y cells Concentrations 10 serial 3-fold dilutions, top concentration 5 μM Incubation Time 90 min Method A human neuroblastoma cell line, SH-SY5Y, is used for the LRRK2 stable cell line construction. SH-SY5Y cells are cultured in Dulbecco’s modified Eagle’s medium/F-12 supplemented with GlutaMax, 10% tetracycline (Tet)-free fetal bovine serum, nonessential amino acids, pen-strep at 37°C and 5% carbon dioxide. Parental cells are transfected with plasmid constructs that overexpress full-length human LRRK2 wildtype or mutant (G2019S) under the control of a Tet-inducible promoter. Transfected cells are selected and maintained with hygromycin (2 mg/ml) and zeocin (100 mg/ml). Cells are seeded into six-well plates and induced with Tet (2 mg/ml) for 72 hours prior to treatment. After 90 minutes of MLi-2 incubation, cells were mechanically lifted, pelleted, and lysed with lysis buffer supplemented with protease, and phosphatase inhibitors. Pellets are further bead homogenized and then spun at 13,200 rpm for 20 minutes at 4°C. Supernatants are then removed for subsequent Western blot analyses.
Animal Study:^{^[1]}	Animal Models male C57Bl/6 mice Dosages 1 mg/kg, 2 mg/kg, 3 mg/kg, 10 mg/kg, 30 mg/kg, 100 mg/kg Administration IV, Oral gavage

Cell Assay:^{^[1]}

Cell lines
SH-SY5Y cells
Concentrations
10 serial 3-fold dilutions, top concentration 5 μM
Incubation Time
90 min
Method
A human neuroblastoma cell line, SH-SY5Y, is used for the LRRK2 stable cell line construction. SH-SY5Y cells are cultured in Dulbecco’s modified Eagle’s medium/F-12 supplemented with GlutaMax, 10% tetracycline (Tet)-free fetal bovine serum, nonessential amino acids, pen-strep at 37°C and 5% carbon dioxide. Parental cells are transfected with plasmid constructs that overexpress full-length human LRRK2 wildtype or mutant (G2019S) under the control of a Tet-inducible promoter. Transfected cells are selected and maintained with hygromycin (2 mg/ml) and zeocin (100 mg/ml). Cells are seeded into six-well plates and induced with Tet (2 mg/ml) for 72 hours prior to treatment. After 90 minutes of MLi-2 incubation, cells were mechanically lifted, pelleted, and lysed with lysis buffer supplemented with protease, and phosphatase inhibitors. Pellets are further bead homogenized and then spun at 13,200 rpm for 20 minutes at 4°C. Supernatants are then removed for subsequent Western blot analyses.

Animal Study:^{^[1]}

Animal Models
male C57Bl/6 mice
Dosages
1 mg/kg, 2 mg/kg, 3 mg/kg, 10 mg/kg, 30 mg/kg, 100 mg/kg
Administration
IV, Oral gavage

References

[1] Matthew J Fell, et al. J Pharmacol Exp Ther. 2015 Dec;355(3):397-409.

Selleck's MLi-2 has been cited by 1 publication

Pharmacological Inhibition of LRRK2 Exhibits Neuroprotective Activity in Mouse Photothrombotic Stroke Model [ Exp Neurobiol, 2024, 33(1):36-45]	PubMed: 38471803

Pharmacological Inhibition of LRRK2 Exhibits Neuroprotective Activity in Mouse Photothrombotic Stroke Model [ Exp Neurobiol, 2024, 33(1):36-45]

PubMed: 38471803

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SHIPPING AND STORAGE
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