Molecule

Description

Malignant melanoma, Leukaemia, Malignant melanoma, Pancreatic cancer, Solid tumours

Description

GSK1120212 (JTP-74057, trametinib) is a highly specific and potent MEK1 and MEK2 inhibitor with IC50 of 0.92 nM and 1.8 nM, respectively.

Targets

IC₅₀

In Vitro

GSK1120212 inhibits the phosphorylation of MBP regardless of the isotype of Raf and MEK, with IC50 ranging from 0.92 nM to 3.4 nM. GSK1120212 demonstrates no inhibition of the kinase activities of c-Raf, B-Raf, ERK1 and ERK2. In addition, GSK1120212 does not show drastic inhibitory activity against the other 98 kinases. GSK1120212 displays potent inhibitory activity against human colorectal cancer cell lines. HT-29 and COLO205 cells, which are known to have a constitutively active B-Raf mutant, are most sensitive to GSK1120212 with IC50 0.48 nM and 0.52 nM, respectively. The cell lines bearing a K-Ras mutation show a wide range of sensitivity to GSK1120212 with IC50 of 2.2-174 nM. In contrast, COLO320 DM cells, bearing the wild-type gene in both B-Raf and K-Ras, are found to be resistant to GSK1120212 even at 10 μM. GSK1120212 treatment for 24 hours induces cell-cycle arrest at the G1 phase in all sensitive cell lines. Consistently, GSK1120212 treatment leads to upregulation of p15^INK4b and/or p27^KIP1 in most of the colorectal cancer cell lines. GSK1120212 inhibits constitutive ERK phosphorylation in all sensitive cell lines. GSK1120212 induces apoptosis both in HT-29 and COLO205 cells, but that COLO205 cells are more sensitive to GSK1120212 than HT-29 cells in terms of apoptosis induction. ^[1] GSK1120212 blocks tumor necrosis factor-α and interleukin-6 production from peripheral blood mononuclear cells (PBMCs). ^[2]

In Vivo

Oral administration of GSK1120212 at 0.3 mg/kg or 1 mg/kg once daily for 14 days is effective in inhibiting the HT-29 xenograft growth, and 1 mg/kg of GSK1120212 almost completely blocks the tumor increase. The phosphorylation of ERK1/2 is completely inhibited in the established tumor tissues by single oral dose of 1 mg/kg GSK1120212, and both p15^INK4b and p27^KIP1 protein levels are upregulated after 14 days of treatment with GSK1120212. In the COLO205 xenograft model, tumor regression is observed even at a dose of 0.3 mg/kg. At a dose of 1 mg/kg, a complete regression is obtained in 4 out of 6 mice in which the tumor degenerates to the point that tumor volume is not measurable. ^[1] Administration of GSK1120212 at 0.1 mg/kg almost completely suppresses adjuvant-induced arthritis (AIA) and type II collagen-induced arthritis (CIA) in Lewis rats or DBA1/J mice, respectively. ^[2]

Clinical Trials

A Phase I study to evaluate the effect of repeat oral dosing of GSK1120212 on cardiac repolarization in subjects with solid tumors is currently ongoing.

Features

More potent than PD0325901 or AZD6244

Description

GSK1120212 exhibits an additive or a synergistic effect in combination with the standard-of-care agents, 5-fluorouracil, oxaliplatin or SN-38. ^[1] The combination of GSK2118436 and GSK1120212 effectively inhibits cell growth, decreases ERK phosphorylation, decreases cyclin D1 protein, and increases p27^KIP1 protein in the resistant melanoma cell clones. Moreover, the combination of GSK1120212 with GSK2126458 enhances cell growth inhibition and decreases S6 ribosomal protein phosphorylation in these clones. ^[3] GSK1120212 in combination with GSK2126458 enhances uveal melanoma cell death in a mutant GNAQ- and GNA11-dependent manner. ^[4] A Phase I study of GSK1120212 and GSK1120212 in combination with Gemcitabine in Japanese subjects with solid tumors is currently ongoing.

Raf-MEK-ERK cascade kinase assay

Non-phosphorylated myelin basic protein (MBP) is coated onto an ELISA plate, and the active form of B-Raf/c-Raf is mixed with unphosphorylated MEK1/MEK2 and ERERK2 in 10 μM ATP and 12.5 mM MgCl₂ containing MOPS buffer in the presence of various concentrations of GSK1120212. The phosphorylation of MBP is detected by the anti-phospho-MBP antibody.

Cell Lines

HT-29, HCT-15, HCT116, COLO205, LS-174T, SW480, SW620, T84, LoVo and COLO320

Concentrations

Dissolved in DMSO, final concentrations ~10 μM

Incubation Time

3 or 4 days

Methods

Exponentially growing cells are precultured in 96-well tissue culture plates for 24 hours and then exposed to GSK1120212. Cell growth is determined by an in vitro toxicology assay kit, sulforhodamine B based. For apoptosis assay, both floating and adherent cells are collected and fixed with 70% ethanol. After washing with PBS, the cells are suspended in 100 μg/mL RNase and 25 μg/mL propidium iodide (PI) and incubated at 37 °C for 30 minutes in the dark. The DNA content of each single cell is determined using the flow cytometer Cytomics FC500 or Guava EasyCyte plus.

Animal Models

Female BALB/c-nu/nu mice inoculated subcutaneously with HT-29 or COLO205 cells

Formulation

Dissolved in 10% Cremophor EL-10% PEG400

Doses

~1 mg/kg/day

Administration

Orally