Trametinib is an ATP-noncompetitive inhibitor of MEK 1/2 (IC50s: 0.7/0.9 nM). It shows low inhibition for more than 180 kinases, including B-Raf, c-Raf, and MEK5.

Trametinib (JTP-74057) inhibited the phosphorylation of MBP regardless of the isotype of Raf and MEK (IC50s: 0.92 to 3.4 nM). JTP-74057 demonstrated no inhibition of the kinase activities of c-Raf, B-Raf, ERK1 and ERK2. HT-29 and COLO205 cells, which are known to have a constitutively active B-Raf mutant, were most sensitive to JTP-74057, showing subnanomolar IC50 values. The cell lines bearing a K-Ras mutation showed a wide range of sensitivity to JTP-74057 [1]. Among KRAS mutant cell lines, cell lines with an expression pattern suggestive of epithelial-to-mesenchymal transition were less sensitive to GSK1120212. A proportion of cell lines from certain tissue types not known to carry frequent RAF/RAS mutations also seemed to be sensitive to GSK1120212 [2].

Both 0.3 mg/kg and 1 mg/kg of JTP-74057 were effective in inhibiting the HT-29 xenograft growth, and 1 mg/kg of JTP-74057 blocked the tumor increase almost completely. Neither significant changes in body weight nor gross lesions at autopsy were observed in the mice treated with 1 mg/kg of JTP-74057. The phosphorylation of ERK1/2 was completely inhibited in the established tumor tissues by a single oral dose of 1 mg/kg JTP-74057, and both p15INK4b and p27KIP1 protein levels were upregulated after 14 days of treatment with JTP-74057 [1]. JTP-74057 blocked tumor necrosis factor-α and interleukin-6 production from PBMCs. AIA and CIA development were suppressed almost completely by 0.1 mg/kg of JTP-74057. In the CIA, JTP-74057 suppressed collagen-reactive T-cell proliferation ex vivo [3].

A Raf-MEK-ERK cascade kinase assay was carried out as previously described. Briefly, nonphosphorylated myelin basic protein (MBP) was coated onto an ELISA plate, and the active form of B-Raf/c-Raf was mixed with unphosphorylated MEK1/MEK2 and ERK2 in 10 μM ATP and 12.5 mM MgCl2 containing MOPS buffer in the presence of various concentrations of JTP-74057. The phosphorylation of MBP was detected by the anti-phosphoMBP antibody. Kinase inhibitory activities against a total of 99 kinases were tested by kinase profiler at 10 μM ATP [1].

These cells were maintained in media recommended by the providers. Exponentially growing cells were precultured in 96-well tissue culture plates for 24 h and then exposed to JTP-74057. Cell growth was determined by an in vitro toxicology assay kit, sulforhodamine B based. For combination studies, two compounds were simultaneously added to the HT-29 cells and incubated for 72 h. In the presence of various concentrations of compound A, the 50% inhibitory concentration (IC50) values of compound B were determined. Then, the fixed concentration of compound A versus the IC50 value of compound B was plotted. Conversely, the IC50 values of compound A were determined in the presence of various concentrations of compound B and plotted [1].

Female BALB/c-nu/nu mice were used. On day 0, HT-29 cells or COLO205 cells suspended in ice-cold HBSS (-) were inoculated subcutaneously into the right flank of the mice at 5x10^6 cells/100 μl/site or 1x10^6 cells/100 μl/site, respectively. The acetic acid-solvated form of JTP-74057 was dissolved in 10% Cremophor EL-10% PEG400 and was administered orally once daily for 14 days from the day when the mean tumor volume reached 100 mm^3. The tumor length [L (mm)] and width [W (mm)] were measured using a micro gauge twice a week after the commencement of dosing, and the tumor volume was calculated using the following formula: tumor volume (mm^3) = L x W x W/2. All procedures relating to the use of animals in this study were reviewed and approved by the Institutional Animal Care and Use Committee of Japan Tobacco [1].

871700-17-3

C26H23FIN5O4

615.39

JTP-74057;曲美替尼;GSK1120212;Trametinib

[1]Yamaguchi T, et al. Suppressive effect of an orally active MEK1/2 inhibitor in two different animal models for rheumatoid arthritis: a comparison with leflunomide. Inflamm Res, 2012, 61(5), 445-454. [2]Yamaguchi T, et al. Antitumor activities of JTP-74057 (GSK1120212), a novel MEK1/2 inhibitor, on colorectal cancer cell lines in vitro and in vivo. Int J Oncol, 2011, 39(1), 23-31. [3]KieΔling M K, Nicolay J P, Schl?r T, et al. NRAS mutations in cutaneous T cell lymphoma (CTCL) sensitize tumors towards treatment with the multikinase inhibitor Sorafenib[J]. Oncotarget. 2017 Jul 11;8(28):45687-45697. [4]Wang X, Wu F, Wang H, et al. PDCD6 cooperates with C-Raf to facilitate colorectal cancer progression via Raf/MEK/ERK activation[J]. Journal of Experimental & Clinical Cancer Research. 2020, 39(1): 1-15. [5]Klein C, Roussel G, Brun S, et al. 5-HIAA induces neprilysin to ameliorate pathophysiology and symptoms in a mouse model for Alzheimer’s disease[J]. Acta neuropathologica communications. 2018 Dec 11;6(1):136. [6]Jing J, et al. Comprehensive predictive biomarker analysis for MEK inhibitor GSK1120212. Mol Cancer Ther. 2012 Mar;11(3):720-9.

[1]Jiang Z, Cheng L, Wu Z, et al. Transforming primary human hepatocytes into hepatocellular carcinoma with genetically defined factors. EMBO reports. 2022: e54275 [2]Wang X, Wu F, Wang H, et al. PDCD6 cooperates with C-Raf to facilitate colorectal cancer progression via Raf/MEK/ERK activation. Journal of Experimental & Clinical Cancer Research. 2020, 39(1): 1-15 [3]Sun C Y, Li Y Z, Cao D, et al. Rapamycin and trametinib: a rational combination for treatment of NSCLC. International Journal of Biological Sciences. 2021, 17(12): 3211-3223. [4]Lü Z, Li X, Li K, et al. Nitazoxanide and related thiazolides induce cell death in cancer cells by targeting the 20S proteasome with novel binding modes. Biochemical Pharmacology. 2022: 114913. [5]Klein C, Roussel G, Brun S, et al. 5-HIAA induces neprilysin to ameliorate pathophysiology and symptoms in a mouse model for Alzheimer’s disease. Acta Neuropathologica Communications. 2018 Dec 11;6(1):136 [6]KieΔling M K, Nicolay J P, Schl?r T, et al. NRAS mutations in cutaneous T cell lymphoma (CTCL) sensitize tumors towards treatment with the multikinase inhibitor Sorafenib. Oncotarget. 2017 Jul 11;8(28):45687-45697.

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