[Description]:
Agerafenib hydrochloride is a highly potent and orally efficacious inhibitor of BRAFV600E with a Kd of 14 nM.
[Related Catalog]:
[Target]
BRafV600E:14 nM (Kd)
Braf:36 nM (Kd)
CRAF:39 nM (Kd)
c-Kit:2 nM (Kd)
Ret:2 nM (Kd)
LCK:2 nM (Kd)
Abl-1:3 nM (Kd)
VEGFR-2:8 nM (Kd)
CSF-1R:9 nM (Kd)
EPHA2:14 nM (Kd)
EGFR:22 nM (Kd)
c-Met:513 nM (Kd)
JAK-2:4700 nM (Kd)
MEK-1:7100 nM (Kd)
MEK-2:8300 nM (Kd)
[In Vitro]
Agerafenib (CEP-32496) exhibits high potency against several BRAFV600E-dependent cell lines and selective cytotoxicity for tumor cell lines expressing mutant BRAFV600E versus those containing wild-type BRAF. Agerafenib (CEP-32496) exhibits potent binding (BRAFV600E Kd=14 nM) and cellular activity (pMEK IC50=82 nM and A375 proliferation IC50=78 nM), with activity in the proliferation assay. CEP-32496 also exhibits a favorable CYP450 inhibition profile, with measured IC50 values greater than 10 μM versus the CYP1A2, CYP2C9, CYP2D6, and CYP3A4 isoforms and an IC50=3.4 μM versus CYP2C19[1].
[In Vivo]
Oral administration of Agerafenib (CEP-32496) to Colo-205 tumor xenograft-bearing mice results in significant inhibition of pMEK in tumor cell lysates. For instance, a single 30 mg/kg (po) dose of Agerafenib (CEP-32496) leads to a 50 and 75% inhibition of normalized pMEK in tumor lysates at the 2 and 6 h postdose time point, respectively (p<0.03), while a 55 mg/kg (po) dose resulted in a 75% to 57% (p<0.03) inhibition of pMEK at 2 through 10 h post administration, with normalization to baseline by 24 h. Agerafenib (CEP-32496) exhibits an exceptional PK profile in mouse, dog, and cynomolgus monkey. Administration of Agerafenib (CEP-32496) to beagle dogs (single dose of 1 mg/kg iv and 10 mg/kg po) results in low clearance (CL=5.0 (mL/min)/kg) and excellent bioavailability (%F=100). Similarly, in cynomolgus monkey, the administration of Agerafenib (CEP-32496) (single dose of 1 mg/kg iv and 10 mg/kg po) leads to high oral exposure due to low clearance (CL=6.7 mL/min/kg) and excellent bioavailability (%F=100)[1].
[Cell Assay]
A375 cells are seeded at 10,000 cells per well in DMEM with 10% fetal calf serum and allowed to attach. The cells are washed with PBS and switched to DMEM with 0.5% of serum and incubated overnight. The test compounds (e.g., Agerafenib (CEP-32496); 10 μM) are then added at various concentrations with a final DMSO concentration of 0.5% and incubated for 72 h. At the end of incubation, a Cell Titer Blue is added per instructions, and incubation is continued for 3 h. Remaining viable cells are quantified by measuring the strength of the fluorescence signal using SoftMax Pro (excitation at 560 nm and emission at 590 nm). IC50 values are derived using a 9-point curve and are presented as mean values from experiments performed in duplicate[1].
[Animal admin]
Mice[1] Six to eight week old athymic nu/nu nude mice (20-25 g) are inoculated subcutaneously with Colo-205 tumor cells (1×106/mouse) in the right flank. Upon reaching an average tumor volume of 150-200 mm3 (10-12 days post implantation), animals are randomized into treatment groups (n=10 mice/group). Each group is dosed orally for 14 days with either vehicle only (22% HPβCD) or with Agerafenib (CEP-32496) at 10, 30, or 100 mg/kg twice daily (BID), and each dose of drug is given in a volume of 0.1 mL per 20 g of body weight, adjusted for the body weight of the animal. Tumor volumes are measured three times weekly using vernier calipers, and volumes are calculated[1].
[References]
[Related Small Molecules]