MLi-2 is a potent and selective inhibitor of leucine-rich repeat kinase 2 (LRRK2), with an IC₅₀ value of 0.76nM [1]. MLi-2 exhibits excellent central nervous system penetration and serves as an important tool compound for studying LRRK2 function and the pathological mechanisms of Parkinson’s disease (PD)[2-3]. Additionally, MLi-2 has been shown to alleviate DOPA-responsive dystonia[4].
In vitro, treatment with MLi-2 (100nM) for 2 hours in SH-SY5Y and MEF cells expressing either wild-type or 4xSA (S910A/S935A/S955A/S973A) mutant LRRK2 significantly inhibited LRRK2 kinase activity, as evidenced by a marked reduction in the phosphorylation levels of Rab8a (T72) and Rab10 (T73), as well as decreased autophosphorylation of LRRK2 at residues S910, S935, S955, and S973[5]. MLi-2 (1μM) treatment for 24 hours in primary microglia derived from wild-type and G2019S-LRRK2 mice significantly reduced LRRK2 phosphorylation at Ser935, indicating effective inhibition of LRRK2 kinase activity. RNA-Seq analysis revealed that MLi-2 treatment primarily upregulated genes associated with the regulation of ion transmembrane transport and downregulated genes involved in the positive regulation of apoptosis[6].
In vivo, MLi-2 (8 or 45mg/kg/day) was administered via diet to wild-type and LRRK2 G2019S knock-in mice injected with Alzheimer’s disease-derived tau fibrils. Treatment began one week prior to tau injection and continued for 6 months. MLi-2 significantly reduced tau pathology progression in cortical regions of G2019S mutant mice, with the effect being more pronounced at the 45mg/kg/day dose. MLi-2 also reversed the altered direction of pathology spread caused by the G2019S mutation. Additionally, long-term MLi-2 treatment led to enlargement of pneumocytes in the lungs[7]. MLi-2 (10mg/kg) was administered via intraperitoneal injection to 12-month-old LRRK2 G2019S knock-in (KI) mice to assess its effects on dopamine transporter (DAT) function. Although MLi-2 significantly inhibited LRRK2 kinase activity (as shown by a 75% reduction in pSer1292 LRRK2 levels), MLi-2 failed to reverse the abnormally elevated DAT uptake activity (Vmax) observed in G2019S KI mice[8].
References:
[1] Fell MJ, Mirescu C, Basu K, et al. MLi-2, a Potent, Selective, and Centrally Active Compound for Exploring the Therapeutic Potential and Safety of LRRK2 Kinase Inhibition. J Pharmacol Exp Ther. 2015 Dec;355(3):397-409.
[2] Hatcher JM, Zwirek M, Sarhan AR, et al. Development of a highly potent and selective degrader of LRRK2. Bioorg Med Chem Lett. 2023 Oct 1;94:129449.
[3] Tengberg JF, Russo F, Benned-Jensen T, et al. LRRK2 and RAB8A regulate cell death after lysosomal damage in macrophages through cholesterol-related pathways. Neurobiol Dis. 2024 Nov;202:106728.
[4] Roman KM, Dinasarapu AR, Cherian S, et al. Striatal cell-type-specific molecular signatures reveal potential therapeutic targets in a model of dystonia. Neurobiol Dis. 2025 Aug;212:106981.
[5] Kania E, Long JS, McEwan DG, et al. LRRK2 phosphorylation status and kinase activity regulate (macro)autophagy in a Rab8a/Rab10-dependent manner. Cell Death Dis. 2023 Jul 15;14(7):436.
[6] Nazish I, Mamais A, Mallach A, et al. Differential LRRK2 Signalling and Gene Expression in WT-LRRK2 and G2019S-LRRK2 Mouse Microglia Treated with Zymosan and MLi2. Cells. 2023 Dec 26;13(1):53.
[7] Lubben N, Brynildsen JK, Webb CM, et al. LRRK2 kinase inhibition reverses G2019S mutation-dependent effects on tau pathology progression. Transl Neurodegener. 2024 Mar 4;13(1):13.
[8] Domenicale C, Mercatelli D, Albanese F, et al. Dopamine Transporter, PhosphoSerine129 α-Synuclein and α-Synuclein Levels in Aged LRRK2 G2019S Knock-In and Knock-Out Mice. Biomedicines. 2022 Apr 12;10(4):881.
MLi-2是一种高效、选择性的富含亮氨酸重复激酶2（LRRK2）抑制剂，其IC₅₀值为0.76nM[1]，MLi-2具有良好的中枢神经系统穿透能力，是研究LRRK2功能和帕金森病（PD）病理机制的重要工具化合物[2-3]。此外，MLi-2还被证明具有缓解DOPA反应性肌张力障碍的功能[3-4]。
在体外，MLi-2（100nM）处理表达野生型或4xSA（S910A/S935A/S955A/S973A）突变型LRRK2的SH-SY5Y细胞和MEF细胞2小时，显著抑制了LRRK2激酶活性，表现为Rab8a（T72）和Rab10（T73）磷酸化水平显著下降，同时降低了LRRK2自身在S910/S935/S955/S973位点的磷酸化水平[5]。MLi-2（1μM）处理野生型和G2019S-LRRK2小鼠原代小胶质细胞24小时，显著抑制了LRRK2在Ser935位点的磷酸化水平，提示其有效抑制LRRK2激酶活性。RNA-Seq分析显示，MLi-2处理主要上调了与离子跨膜转运调控相关的基因表达，并下调了与细胞凋亡正向调控相关的基因[6]。
在体内，MLi-2（8或45mg/kg/天）通过饮食给药，用于处理注射了阿尔茨海默病来源tau纤维的野生型和LRRK2 G2019S敲入小鼠，给药时间为tau注射前1周开始，并持续6个月。MLi-2在G2019S突变小鼠中显著减少了皮质区域的tau病理进展，尤其在45mg/kg/天的剂量下效果更为明显，同时逆转了由G2019S突变引起的病理扩散方向改变。此外，长期给予MLi-2还导致小鼠肺泡细胞（pneumocytes）体积增大[7]。MLi-2（10mg/kg）通过腹腔注射给药，用于处理12月龄的LRRK2 G2019S敲入（KI）小鼠，以评估其对多巴胺转运体（DAT）功能的影响。尽管MLi-2显著抑制了LRRK2激酶活性（pSer1292 LRRK2水平下降75%），但未能逆转G2019S KI小鼠中异常升高的DAT摄取活性（Vmax）[8]。