Sotrastaurin (AEB071)

Cell lines

SUDHL-4 and OCI-LY8 DLBCL cells

Preparation Method

The OCI-LY8 cell line was cultured in Iscove’s modified Dulbecco’s medium supplemented with 10% fetal bovine serum and 1% penicillin–streptomycin, and the SUDHL-4 cell line was cultured in RPMI 1640 containing 10% FBS and 1% penicillin–streptomycin. Cells were incubated in a humidified atmosphere at 37°C in 5% CO₂. Sotrastaurin (40mM) was dissolved in dimethyl sulfoxide (DMSO) and stored at -20°C. SUDHL-4 and OCI-LY8 cells (0.1ml) were seeded into wells of 96- well plates at a density of 2×10⁵cells/ml, treated with various concentrations of Sotrastaurin (5, 10, 15, 20, 25, 30, or 40μM) for 24, 48 or 72h in a 37°C incubator. At the end of the incubation, CCK-8 assay was used for cell proliferation detection. Cell apoptosis was measured using the Annexin-V/PI apoptosis detection kit. Cell cycle was assessed by flow cytometry. Protein was extracted for western blot analysis.

Reaction Conditions

5-40µM; 24-72h

Applications

Treatment of SUDHL-4 and OCI-LY8 DLBCL cells with Sotrastaurin blocked cell proliferation, induced apoptosis, and triggered G1 arrest accompanied by down-regulation of MCT-1, cyclin D1, p-ERK and p-AKT and up-regulation of p53 and cleaved caspases-3/8/9.

Animal models

C57BL/6 mice

Preparation Method

A total of seventy-two 8-week-old C57BL/6 mice, 36 females and 36 males, were raised to the age of three months. Mice were then randomly assigned to one of six groups according to sex (n=6 mice each), including two groups of Sham-operated (intragastric administration of PBS only), two groups of medial meniscus (DMM) surgery and two groups of DMM+Sotrastaurin treatment (intragastric administration of 10mg/kg Sotrastaurin). For DMM surgery, mice were anaesthetized with chloral hydrate and then subjected to surgical transection of the medial meniscotibial ligament of the right knee. Sham mice received the same surgical procedure without the transection of the medial meniscotibial ligament. Two days post-surgery, mice were intragastrically administered with Sotrastaurin (treatment groups) or equivalent volume of PBS (Sham and DMM groups only) every other day for 4 or 8 weeks. No adverse reactions or animal fatalaties were recorded during the treatment period, and all mice exhibited normal behavioural activity. Mice were killed at the respective endpoints and the knee joints were excised, cleaned of soft tissues, fixed in 4% PFA, and then processed for micro-computed tomography (micro-CT) and histological assessments.

Dosage form

10mg/kg; i.g.; every other day for 4 or 8 weeks

Applications

Sotrastaurin significantly reduced subchondral bone loss and articular cartilage degeneration, decreased TRAP⁺ osteoclast numbers, and improved OARSI scores in DMM-induced OA mice.

References:
[1] Chang G, Zheng J, Xiao W, et al. PKC inhibition of sotrastaurin has antitumor activity in diffuse large B-cell lymphoma via regulating the expression of MCT-1. Acta Biochim Biophys Sin (Shanghai). 2018;50(4):399-407.
[2] Pang C, Wen L, Qin H, Zhu B, Lu X, Luo S. Sotrastaurin, a PKC inhibitor, attenuates RANKL-induced bone resorption and attenuates osteochondral pathologies associated with the development of OA. J Cell Mol Med. 2020;24(15):8452-8465.