GSK2578215A is a potent and selective LRRK2 kinase inhibitor.

LRRK2 (WT):10.9 nM, LRRK2 (G2019S):8.9 nM

Administering 100 mg/kg intraperitoneally (i.p.) of GSK2578215A inhibits the phosphorylation of Ser910 and Ser935 in the spleen and kidneys of mice, but has no effect in the brain.

In SH-SY5Y cells, GSK2578215A impairs autophagic flux by altering autophagosome-lysosome fusion and induces mitochondrial autophagy through Drp-1-mediated mitochondrial fission and mitochondrial-derived ROS signaling. Furthermore, GSK2578215A induces dose-dependent inhibition of Ser910 and Ser935 phosphorylation in HEK293 cells stably transfected with wild-type LRRK2 and LRRK2 [G2019S], and similarly promotes dose-dependent dephosphorylation of Ser910 and Ser935 in endogenous LRRK2 in mouse Swiss 3T3 cells.

PFV integration assay: For quantitative strand transfer assays, donor DNA substrate is formed by annealing HPLC grade oligonucleotides 5′-GACTCACTATAGGGCACGCGTCAAAATTCCATGACA and 5′-ATTGTCATG GAATTTTGACGCGTGCCCTATAGTGAGTC. Reactions (40 μL) contains 0.75 μM PFV IN, 0.75 μM donor DNA, 4 nM (300 ng) supercoiled pGEM9-Zf(?) target DNA, 125 mM NaCl, 5 mM MgSO4, 4 μM ZnCl2, 10 mM DTT, 0.8% (vol/vol) DMSO, and 25 mM BisTris propane–HCl, pH 7.45. Raltegravir is added at indicated concentrations. Reactions are initiated by addition of 2 μL PFV IN diluted in 150 mM NaCl, 2 mM DTT, and 10 mM Tris-HCl, pH 7.4, and stopped after 1 hour at 37 °C by addition of 25 mM EDTA and 0.5% (wt/vol) SDS. Reaction products, deproteinized by digestion with 20 μg proteinase K for 30 minutes at 37 °C followed by ethanol precipitation, are separated in 1.5% agarose gels and visualized by staining with ethidium bromide. Integration products are quantified by quantitative real-time PCR, using Platinum SYBR Green qPCR SuperMix and three primers: 5′-CTACTTACTCTAGCTTCCCGGCAAC, 5′-TTCGCCAGTTAATAGTTTGCGCAAC, and 5′-GACTCACTATAGGGCACGCGT. PCR reactions (20 μL) contained 0.5 μM of each primer and 1 μL diluted integration reaction product. Following a 5-min denaturation step at 95 °C, 35 cycles are carried out in a CFX96 PCR instrument, using 10 seconds denaturation at 95 °C, 30 seconds annealing at 56 °C and 1 minutes extension at 68 °C. Standard curves are generated using serial dilutions of WT PFV IN reaction in the absence of INSTI.