Bemcentinib

Bemcentinib (R428) belongs to small molecule inhibitors and is a highly selective oral Axl inhibitor (IC50 = 14 nM) with oral bioavailability and potent cell permeability. This compound effectively inhibits cancer cell migration and invasion, blocks tumor dissemination, and prolongs survival in various tumor models.

cells:14 nM (cell free)

Methods: In HCC827 parental and erlotinib-resistant ER3 and ER10 cells, Bemcentinib (0.1–2.0 μM) was administered for 120 hours, and real-time growth curves were measured using IncuCyte.
Results: Bemcentinib dose-dependently inhibited cell proliferation and induced growth arrest in resistant cells. [1]
Methods: In SET-2 and BaF3-EpoR-JAK2V617F cells, Bemcentinib (0.5–5 μM) was administered for 48 hours; cells were also treated for 24 hours (SET-2) or 12 hours (BaF3).
Results: After 48 hours of treatment, WST-1 assay demonstrated dose-dependent inhibition of cell viability; BrdU incorporation and Annexin V staining confirmed inhibition of cell proliferation and induction of apoptosis. [2]
Methods: In LX2 human hepatic stellate cells, Bemcentinib (0.25 μM) was administered as a 1-hour pretreatment followed by GAS6 stimulation; in primary mouse Kupffer cells, Bemcentinib (0.25 μM) pretreatment was followed by LPS stimulation for 2 hours.
Results: ELISA demonstrated inhibition of MCP-1 release and p-AKT levels; RT-qPCR confirmed reduced expression of IL-1β and IL-6 mRNA. [3]

Methods: In an HCC827 cell xenograft nude mouse model, Bemcentinib (50 or 100 mg/kg, twice daily, oral gavage) was combined with erlotinib (50 mg/kg, once daily) for 138 days of treatment.
Results: The combination therapy significantly delayed the emergence of tumor resistance, with maximum tumor suppression maintained until the experimental endpoint. [1]
Methods: In a SET-2 cell xenograft NSG mouse model, Bemcentinib (50 mg/kg, twice daily, oral gavage) was administered until tumors reached 1500 mm³, achieving 60% tumor growth inhibition.
Results: In a BaF3-EpoR-JAK2V617F systemic model, Bemcentinib (50 mg/kg, twice daily) significantly prolonged survival, reduced splenomegaly, and improved anemia. [2]
Methods: In C57BL/6 mice with MCD- or HFD-induced NASH models, Bemcentinib (50 mg/kg, twice daily, oral gavage) was administered during the final 2 weeks of dietary induction.
Results: Bemcentinib significantly reduced hepatic collagen deposition (confirmed by hydroxyproline assay and Sirius Red staining) and decreased expression of pro-inflammatory and pro-fibrotic genes. [3]

A five-point R428 dose titration was performed in radiometric in vitro kinase assays on 133 kinases at the Km(ATP) for each kinase. Axl, Mer, and Tyro3 assays were also performed using a fluorescence polarization protocol. HER2 activity was determined by Z'-LYTE assay [1].

MDA-MB-231 or 4T1 cells (1 × 10^5) were allowed to migrate through Matrigel toward 20% FCS in an 8-μm pore 24-well Transwell plate at 37°C for 16 to 24 h. Noninvaded cells and Matrigel were removed by swabbing. Invaded cells were fixed in 4% formaldehyde, stained with 1% crystal violet, and quantified as for Axl cell-based assay. Cells were preincubated with R428 for 3 h. R428 was added to both upper and lower Transwell chambers [1].

Female BALB/c mice were inoculated in the mammary fat pad with 0.5 × 10^6 4T1 cells. Forty-eight hours after inoculation, mice were randomized into treatment groups (n = 10). Oral dosing with R428 (7–75 mg/kg twice daily) or vehicle continued until days 19 to 21. Cisplatin (1.2 or 4 mg/kg) was administered i.v. once weekly. Body weight and tumor size were measured thrice per week. Lungs were exposed postmortem. Total number and size of surface lung macrometastases were measured (small, <2 mm; medium, ≥2 mm and <3 mm; large, ≥3 mm). Half of each primary tumor was snap frozen in liquid nitrogen. The other half, and the livers were fixed in paraformaldehyde/lysine/periodate solution, paraffin embedded and sectioned (5 μm thick). Two H&E-stained liver sections per animal were examined microscopically for micrometastases in three view fields. Synergism was determined using Clark's synergy calculation [1].

H2O: < 1 mg/mL (insoluble or slightly soluble)

Ethanol: < 1 mg/mL (insoluble or slightly soluble)

DMSO: 23.4 mg/mL (46.19 mM), Sonication and heating are recommended.

10% DMSO+40% PEG300+5% Tween-80+45% Saline: 1 mg/mL (1.97 mM), Sonication is recommended.