MEK1 (IC50 = 2 nM); MEK2 (IC50 = 2 nM)

GSK1120212 has an IC50 range of 0.92 nM to 3.4 nM and inhibits the phosphorylation of MBP regardless of the isotypes of Raf and MEK. c-Raf, B-Raf, ERK1 and ERK2 are not inhibited by GSK1120212's kinase activity. Furthermore, the other 98 kinases are not significantly inhibited by GSK1120212 in a significant way. The human colorectal cancer cell lines are effectively inhibited by GSK1120212. The cells with the highest sensitivity to GSK1120212 have IC50 values of 0.48 nM and 0.52 nM, respectively, and are known to have a constitutively active B-Raf mutant in HT-29 and COLO205. The IC50 range for GSK1120212 in cell lines with a K-Ras mutation is 2.2–174 nM, indicating a wide range of sensitivity. In contrast, GSK1120212 is found to be resistant to COLO320 DM cells, which carry the wild-type gene in both B-Raf and K-Ras. This is true even at 10 μM. All sensitive cell lines undergo a 24-hour GSK1120212 treatment that results in cell-cycle arrest at the G1 phase. In most colorectal cancer cell lines, treatment with GSK1120212 causes an upregulation of p15INK4b and/or p27KIP1. In all vulnerable cell lines, GSK1120212 prevents constitutive ERK phosphorylation. Both HT-29 and COLO205 cells experience apoptosis induction from GSK1120212; however, COLO205 cells are more vulnerable to this induction than HT-29 cells.[1] GSK1120212 blocks tumor necrosis factor-α and interleukin-6 production from peripheral blood mononuclear cells (PBMCs).[2]

GSK1120212 can effectively stop the growth of the HT-29 xenograft when given orally at doses of 0.3 mg/kg or 1 mg/kg once daily for 14 days. At doses of 1 mg/kg, the tumor growth is almost entirely stopped. A single oral dose of 1 mg/kg GSK1120212 completely inhibits the phosphorylation of ERK1/2 in the tissues of established tumors, and after 14 days of treatment, the levels of the proteins p15INK4b and p27KIP1 are both increased. Tumor regression is seen in the COLO205 xenograft model even at a dose of 0.3 mg/kg. In 4 out of 6 mice receiving a dose of 1 mg/kg, the tumor completely regresses, resulting in tumor volume that cannot be measured.[1] Adjuvant-induced arthritis (AIA) and type II collagen-induced arthritis (CIA) in Lewis rats or DBA1/J mice, respectively, are almost completely suppressed by the administration of GSK1120212 at 0.1 mg/kg.[2]

Non-phosphorylated myelin basic protein (MBP) is coated onto an ELISA plate, and the active form of B-Raf/c-Raf is combined with unphosphorylated MEK1/MEK2, and ERERK2, in the presence of varying concentrations of GSK1120212. Using an anti-phospho-MBP antibody, MBP phosphorylation can be seen.

Exposure to GSK1120212 is followed by a 24-hour preculture of exponentially growing cells in 96-well tissue culture plates. An in vitro toxicology assay kit with sulforhodamine B as its main component measures cell growth. Both adherent and floating cells are gathered and fixed with 70% ethanol for use in the apoptosis assay. The cells are washed in PBS before being suspended in 100 g/mL RNase and 25 μg/mL propidium iodide (PI) and heated to 37 °C for 30 minutes in the dark. Using the flow cytometer Cytomics FC500 or Guava EasyCyte plus, the DNA content of each individual cell is identified.

Mice: To create the A549 (human non-small cell lung carcinoma) model, tissue cultured cells are harvested aseptically and then digested with trypsin. Between 5×106 and 107 cells in 50% martigel are subcutaneously injected into female athymic mice (strain nu/nu). Before treatment, tumors are given one to four weeks to establish. The recommended doses of trametinib are given orally in amounts of 0.2 mL/20 g by weight. Vernier calipers are used every two weeks to measure tumors. When vehicle tumors reached a volume greater than 1000 mm3, antitumor activity was considered to be present. Tumor growth inhibition is defined as the percentage volume difference in tumor growth between the treated and control tumors at that time.

Absorption, Distribution and Excretion
Following oral administration, trametinib is rapidly and readily absorbed. The absorption was examined in patients with solid tumours and BRAF V600 mutation-positive metastatic melanoma. Following the administration of trametinib tablets 0.125 mg (0.0625 times the approved recommended adult dosage) to 4 mg (2 times the approved recommended adult dosage) daily, both Cmax and AUC increased dose-proportionally. Intersubject variability in AUC and Cmax at steady state is 22% and 28%, respectively. Trametinib accumulates with daily repeat dosing with a mean accumulation ratio of 6.0 at 2 mg once daily dose. Steady-state was achieved by Day 15. The mean absolute bioavailability of trametinib is 72% for oral tablets and 81% for oral solution. The Tmax is 1.5 hours. A high-fat, high-calorie meal (approximately 1000 calories) decreased trametinib AUC by 24% and Cmax by 70%, and delayed Tmax by approximately four hours as compared with fasted conditions.
Following oral administration of [¹⁴C]-trametinib, greater than 80% of excreted radioactivity was recovered in the feces while less than 20% of excreted radioactivity was recovered in the urine with less than 0.1% of the excreted dose as the parent molecule.
The apparent volume of distribution (V_c/F) is 214 L.
The apparent clearance is 4.9 L/h.

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Metabolism / Metabolites
Trametinib predominantly undergoes deacetylation mediated by carboxylesterases (i.e., carboxylesterase 1b/c and 2) and other hydrolytic enzymes. The deacetylated metabolite may further be glucuronidated. _In vitro_ findings suggest that deacetylation may also be accompanied by mono-oxygenation, hydroxylation, and glucuronidation. CYP3A4-mediated oxidation is a minor pathway. Four metabolites (M1/2/3/4) have been characterized in patients with advanced cancers. _In vitro_, the M1 and M3 metabolites demonstrated approximately equal or 10-fold less potent phospho-MEK1-inhibiting activity than the parent compound. Following a single dose of [¹⁴C]-trametinib, approximately 50% of circulating radioactivity represented the parent compound. According to findings from metabolite profiling after repeat dosing of trametinib, unchanged parent drug accounted for greater than or equal to 75% of drug-related material in plasma.

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Biological Half-Life
The estimated elimination half-life is 3.9 to 4.8 days.

Hepatotoxicity
In large clinical trials, abnormalities in routine liver tests were common with serum aminotransferase elevations occurring in 39% to 60% and alkaline phosphatase in 24% to 67% of patients treated with trametinib. However, elevations in ALT above 5 times the ULN were uncommon, occurring in 0% to 5% of patients and generally resolving rapidly with temporary discontinuation or dose adjustment. In the prelicensure controlled trials of trametinib with or without dabrafenib, no cases of clinically apparent acute liver injury or hepatic failure were reported. There have yet to be published cases of clinically apparent hepatotoxicity attributed to trametinib. However, it has been used for a short time only.
Likelihood score: E* (unproven but suspected cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
No information is available on the clinical use of trametinib during breastfeeding. Because trametinib is 97% bound to plasma proteins, the amount in milk is likely to be low. However, its half-life is 3.9 to 4.8 days and it might accumulate in the infant. The manufacturer recommends that breastfeeding be discontinued during trametinib therapy and for 4 months after the last dose.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
Trametinib is 97.4% bound to human plasma proteins.

[1]. Int J Oncol . 2011 Jul;39(1):23-31.

[2]. Inflamm Res . 2012 May;61(5):445-54.

Trametinib dimethyl sulfoxide is an addition compound obtained by combining equimolar amounts of trametinib and dimethyl sulfoxide. Used for the treatment of patients with unresectable or metastatic melanoma with BRAF V600E or V600K mutations, and who have not received prior BRAF inhibitor treatment. It has a role as an antineoplastic agent and an EC 2.7.11.24 (mitogen-activated protein kinase) inhibitor. It contains a dimethyl sulfoxide and a trametinib.
Trametinib Dimethyl Sulfoxide is a dimethyl sulfoxide (DMSO) solvated form of trametinib, an orally bioavailable inhibitor of mitogen-activated protein kinase kinase (MAP2K; MAPK/ERK kinase; MEK) 1 and 2, with potential antineoplastic activity. Upon oral administration, trametinib specifically binds to and inhibits MEK 1 and 2, resulting in an inhibition of growth factor-mediated cell signaling and cellular proliferation in various cancers. MEK 1 and 2, dual specificity serine/threonine and tyrosine kinases often upregulated in various cancer cell types, play a key role in the activation of the RAS/RAF/MEK/ERK signaling pathway that regulates cell growth.
See also: Trametinib (has active moiety).

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The MAPK pathway is one of the most important pathways for novel anticancer drug development. We performed high-throughput screening for compounds that induce expression of p15INK4b, and identified JTP-74057 (GSK1120212), which is being evaluated in ongoing phase I, II and III clinical trials. We characterized its antitumor activities in vitro and in vivo. JTP-74057 strongly inhibited MEK1/2 kinase activities, but did not inhibit another 98 kinase activities. Treatment by JTP-74057 resulted in growth inhibition accompanied with upregulation of p15INK4b and/or p27KIP1 in most of the colorectal cancer cell lines tested. Daily oral administration of JTP-74057 for 14 days suppressed tumor growth of HT-29 and COLO205 xenografts in nude mice. Notably, tumor regression was observed only in COLO205 xenografts, and COLO205 was much more sensitive to JTP-74057-induced apoptosis than HT-29 in vitro. Treatment with an Akt inhibitor enhanced the JTP-74057-induced apoptosis in HT-29 cells. Finally, JTP-74057 exhibited an additive or a synergistic effect in combination with the standard-of-care agents, 5-fluorouracil, oxaliplatin or SN-38. JTP-74057, a highly specific and potent MEK1/2 inhibitor, exerts favorable antitumor activities in vitro and in vivo. Sensitivity to JTP-74057-induced apoptosis may be an important factor for the estimation of in vivo efficacy, and sensitivity was enhanced by an Akt inhibitor. These results suggest the usefulness of JTP-74057 in therapeutic applications for colorectal cancer patients.[1]

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Objective and design: To examine the effects of a mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1/2-inhibitor, JTP-74057, on inflammatory arthritis development, and compare its anti-arthritic effect with leflunomide. Materials: Human, mouse, and rat peripheral blood mononuclear cells (PBMCs) were used. Lewis rats and DBA/1J mice were used for animal models. Treatment: JTP-74057 was tested between 0.1-100 nM in in-vitro studies. JTP-74057 (0.01-0.3 mg/kg) and leflunomide (2-10 mg/kg) were administered orally in vivo. Methods: PBMCs were stimulated with lipopolysaccharide. Adjuvant-induced arthritis (AIA) and type II collagen-induced arthritis (CIA) was induced in Lewis rats or DBA1/J mice, respectively. Results: 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 or 10 mg/kg of leflunomide. In the CIA, JTP-74057, but not leflunomide, suppressed collagen-reactive T-cell proliferation ex vivo, whereas leflunomide, but not JTP-74057, suppressed anti-collagen antibody production. Conclusions: JTP-74057 exerts potent anti-arthritic effects with a different profile from leflunomide, suggesting that JTP-74057 may be useful as a new therapeutic reagent in the treatment of rheumatoid arthritis.[2]

C28H29FIN5O5S

693.09

693.091

C, 48.49; H, 4.21; F, 2.74; I, 18.30; N, 10.10; O, 11.53; S, 4.62

1187431-43-1

Trametinib;871700-17-3

50992434

White to off-white solid powder

5.523

2

8

5

41

1120

0

IC1C([H])=C([H])C(=C(C=1[H])F)N([H])C1=C2C(=C(C([H])([H])[H])C(N1C([H])([H])[H])=O)N(C1=C([H])C([H])=C([H])C(=C1[H])N([H])C(C([H])([H])[H])=O)C(N(C2=O)C1([H])C([H])([H])C1([H])[H])=O.S(C([H])([H])[H])(C([H])([H])[H])=O

OQUFJVRYDFIQBW-UHFFFAOYSA-N

InChI=1S/C26H23FIN5O4.C2H6OS/c1-13-22-21(23(31(3)24(13)35)30-20-10-7-15(28)11-19(20)27)25(36)33(17-8-9-17)26(37)32(22)18-6-4-5-16(12-18)29-14(2)34;1-4(2)3/h4-7,10-12,17,30H,8-9H2,1-3H3,(H,29,34);1-2H3

N-[3-[3-cyclopropyl-5-(2-fluoro-4-iodoanilino)-6,8-dimethyl-2,4,7-trioxopyrido[4,3-d]pyrimidin-1-yl]phenyl]acetamide;methylsulfinylmethane

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (3.60 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: 2.5 mg/mL (3.60 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (3.60 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)