Dabrafenib (GSK2118436) is a selective, orally bioavailable inhibitor of Mutant B-raf (BRAF) protein kinase with potential antineoplastic activity. B-raf belongs to the the raf/mil family of serine/threonine protein kinases and plays a role in regulating the MAP kinase/ERKs signaling pathway, which may be constitutively activated due to BRAF gene mutations. Mutations in BRAF are associated with increased growth and proliferation of cancer cells. Dabrafenib (GSK2118436) selectively binds to and inhibits the activity of B-raf, which may inhibit the proliferation of tumor cells which contain a mutated BRAF gene. Administration of the mutant BRAFi dabrafenib is associated with induction of keratinocytic proliferation, which in some cases develops features of low-grade malignancy.
|Cell lines||A375P, SKMEL28 and Colo205 cell lines|
|Preparation method||A375P-F11 assay: A375P cells were plated in 96-well plates by limiting dilution and single cell-derived clones were harvested and tested for sensitivity to B-Raf inhibitors. The F11 clone was selected for future studies and was named A375P-F11. Cellular pSmad Assay to Measure Anti-TGF-β Activity: Activity of compounds was tested in a mechanistic assay in HepG2 cells. Cells were seeded in 12-well plates at a density of 500,000 cells/well and allowed to adhere overnight at 37℃/5% CO2. Media (BME+10% serum) was removed and compound in serum free media was added to the cells for 45 minutes at 37oC/5% CO2. Cells were stimulated with 1 ng/ml TGF-β (R&D systems) for 60 minutes. Cells were lysed in buffer (25 mM Tris-HCl ph: 7.5, 2 mM EDTA, 2 mM EGTA,1% Triton X-100, 0.1 % SDS, 50 mM sodium-β-glycerophosphate, 2 mM sodium orthovanadate, 12.5 mM sodium pyrophosphate, protease and phosphatase inhibitor cocktails) for 30 minutes, scraped, collected, clarified by centrifugation and prepared for western blots in LDS/reducing reagent (Invitogen). Samples were resolved on 4-12% Bis-Tris gels, transferred to PVDF, and probed for total and phospho-Smad2 using antibodies from Cell Signaling. Gels were imaged using the odyssey blot scanner (Licor) and quantified using Licor software. Phospho:total Smad2 ratios were determined and the IC50 was defined as the concentration of compound which decreased the phospho:total ratio by 50%.|
|Animal models||A375P F11 Melanoma Xenograft in nude mice|
|Formulation||0.5% hydroxypropylmethylcellulose (Sigma) and 0.2% Tween-80 in distilled water pH 8.0|
|Dosages||0.2 mL/20g daily|
|Body Surface Area (m2)||0.007||0.025||0.15||0.05||0.02||0.5|
|Animal A (mg/kg) = Animal B (mg/kg) multiplied by||Animal B Km|
|Animal A Km|
For example, to modify the dose of resveratrol used for a mouse (22.4 mg/kg) to a dose based on the BSA for a rat, multiply 22.4 mg/kg by the Km factor for a mouse and then divide by the Km factor for a rat. This calculation results in a rat equivalent dose for resveratrol of 11.2 mg/kg.
Discovery of Dabrafenib: A Selective Inhibitor of Raf Kinases with Antitumor Activity against B-Raf-Driven Tumors.
Rheault TR, et al. ACS Med Chem Lett. 2013 Feb 7;4(3):358-62. PMID: 24900673.
Cutaneous Manifestations of Dabrafenib (GSK2118436): A Selective Inhibitor of Mutant BRAF in patients with Metastatic Melanoma.
Anforth et al. Br J Dermatol. 2012 Jul 16. PMID: 22804352.
Dabrafenib in BRAF-mutated metastatic melanoma: a multicentre, open-label, phase 3 randomised controlled trial.
Hauschild et al. Lancet. 2012 Jul 28;380(9839):358-65. PMID: 22735384.
Dabrafenib in patients with melanoma, untreated brain metastases, and other solid tumours: a phase 1 dose-escalation trial.
Falchook et al. Lancet. 2012 May 19;379(9829):1893-901. PMID: 22608338.
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RAF709 is a novel Raf kinase inhibitor with IC50 values of 0.4 and 1.5 nM for c-Raf and b-Raf, respectively.
L-779450 is a potent, ATP-competitive Raf kinase inhibitor (IC50 =10 nM) that displays > 7, > 30 and > 70-fold selectivity over p38α, GSK3β and Lck respectively.
Sorafenib is a multikinase inhibitor of Raf-1, B-Raf and VEGFR-2 with IC50 of 6 nM, 22 nM and 90 nM, respectively.
PF-04880594 is a RAF inhibitor for BRAF/BRAFV599E and c-RAF with IC50 of 0.19 nM/0.13 nM and 0.39 nM, >100-fold selectivity over CSNK1, JNK2/3 and p38.
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