LX2343

Kinase experiment:

Inhibition of BACE1 enzyme by LX2343 is assayed using BACE1 activity kits in vitro. Briefly, BACE1 substrate (250 nM), BACE1 enzyme (0.35 U/mL), and varied concentrations of LX2343 (5, 10, and 20 μM) are sequentially incubated for 1 h at 37°C in the dark. Fluorescence intensity is measured with excitation and emission wavelengths at 545 and 585 nm, respectively[1].

Cell experiment:

SH-SY5Y cells are transfected with mRFP-GFP-LC3 plasmids via an adenovirus. The cells are treated without or with Streptozotocin (0.8 mM) in combination with 5 or 20 μM LX2343 for 4 h and then fixed with 4% paraformaldehyde and observed using an Olympus Fluoview FV1000 confocal microscope[1].

Animal experiment:

Mice[1]APP/PS1 [B6C3-Tg(APPswe, PS1dE9)] transgenic mice are used. Genotyping to confirm APP/PS1 DNA sequences in their offspring is performed by assaying the DNA from tail biopsies, with Tg-negative mice as a negative control. Twenty male APP/PS1 mice are divided into two groups with ten non-transgenic mice in one group to serve as a negative control. The two 6-month transgenic groups are administered 10 mg/kg per day of LX2343 or vehicle, and the 6-month non-transgenic group is administered the vehicle for 100 d via intraperitoneal injection. After 100 d of administration, MWM assays are applied to evaluate the cognitive abilities of the mice for 8 d under continuous LX2343 treatment. Upon completion of the MWM test, the mice are euthanized, and the brains are removed and bisected at the mid-sagittal plane. The right hemispheres are frozen and stored at -80°C, and the left hemispheres are fixed in 4% paraformaldehyde[1].

References:

[1]. Guo XD, et al. Small molecule LX2343 ameliorates cognitive deficits in AD model mice by targeting both amyloid β production and clearance. Acta Pharmacol Sin. 2016 Sep;37(10):1281-1297.