LX2343 is a BACE1 enzyme inhibitor with an IC50 value of 11.43±0.36 μM. LX2343 acts as a non-ATP competitive PI3K inhibitor with an IC50 of 15.99±3.23 μM. LX2343 stimulates autophagy in its promotion of Aβ clearance.

β-Amyloid:, Autophagy:, BACE1:11.43 μM, PI3K:15.99 μM

LX2343（5-20 μM）在HEK293-APPsw和CHO-APP细胞中降低Aβ积累的能力呈剂量依赖性，同时在SH-SY5Y细胞和原代星形胶质细胞中促进Aβ清除。通过抑制Aβ产生和促进其清除，LX2343改善了APP/PS1转基因小鼠的认知功能障碍，突显了其在治疗AD中的潜力。Western blot结果显示，LX2343在HEK293-APPsw细胞和CHO-APP细胞中不影响BACE1蛋白水平，而体外BACE1酶活性测定表明，LX2343剂量依赖性地降低BACE1活性（以TDC作为阳性对照），IC50为11.43±0.36 μM。为了测试LX2343与ATP之间是否存在竞争关系，我们研究了不同浓度ATP对LX2343抑制活性的影响。结果表明，ATP的存在对LX2343抑制PI3K的作用几乎没有影响，说明LX2343是PI3K的一种非ATP竞争性抑制剂。在10 μM ATP存在下，LX2343的IC50为13.11±1.47 μM；在50 μM ATP存在下，IC50为13.86±1.12 μM；在100 μM ATP存在下，IC50为15.99±3.23 μM。

APP/PS1小鼠表达嵌合型人源瑞典突变APP和突变人源presenilin 1蛋白，是研究AD痴呆的有效动物模型。利用MWM测试评估了LX2343对记忆障碍的改善效果。在8天的训练试验中，APP/PS1转基因小鼠寻找平台的路径长度和逃逸潜伏期显著长于非转基因小鼠。然而，10 mg/kg LX2343的给药显著对抗了第7和第8天的延长路径长度和逃逸潜伏期。在探针试验中，与接受车辆处理的转基因小鼠相比，接受LX2343处理的转基因小鼠更频繁地穿过隐藏平台的位置。

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

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

LX2343 is dissolved in 3% DMSO and 5% tween-80.MiceAPP/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