Bemcentinib 是具有选择性的、高效的Axl抑制剂，其IC50=14 nM。

R428 is a selective inhibitor of Axl (IC50: 14 nM) and has been investigated for the treatment of NSCLC.

Bemcentinib （R428） activity was limited to the tyrosine kinase subfamily. Of the 133 kinases, Axl was most potently inhibited by R428. With the exception of Tie-2, Ftl-1, Flt-3, Ret, and Abl, kinase inhibition by R428 was at least 10 times lower than observed for Axl. R428 dose-dependently suppressed invasion of both human MDA-MB-231 and murine 4T1 breast cancer cell lines [1]. Addition of R428 (50 nM-1 μM) resulted in a dose-dependent inhibition of differentiation of 3T3-F442A preadipocytes into mature adipocytes. Oil Red O staining ranged between 84 and 35% of that of DMSO control at R428 concentrations between 50 nM and 1 μM. Inhibition of Axl signaling by R428 in differentiating preadipocytes was confirmed by the Axl cell-based assay, yielding lower values (A450) for phospho-Akt activity upon treatment with 1 μM R428 compared with medium control or DMSO control [2]. The Axl inhibitor R428 showed a mean IC50 dose of ～ 2.0μM for the primary CLL B cells after 24 hours of treatment and normal B-, T- and natural killer (NK) cells showed no significant amount of cell death at this dose of R428 (2.5μM) under similar experimental conditions [3].

R428 treatment reduced lung metastasis. R428 (7 mg/kg twice daily) significantly suppressed both total metastatic burden and the number of larger metastases. R428 suppressed both tumor angiogenesis and vascular endothelial growth factor (VEGF)–induced corneal neovascularization in vivo [1]. At day 35, the last day of HFD feeding, the body weight in both groups treated with R428 (75 mg/kg/day or 25 mg/kg twice daily, p.o.) was significantly lower than in the corresponding vehicle-treated groups. Compared with the start of the experiment, body weights at the end were significantly increased in both vehicle-treated groups, but not in R428-treated groups [2].

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].

1037624-75-1

C30H34N8

506.64

R428;BGB324

DMSO:10 mg/mL(19.7 mM)
H2O:<1 mgml
Ethanol:<1 mgml
Powder: -20°C for 3 years
In solvent: -80°C for 2 years