ERK2 (IC50 = 8.8 nM); ERK1 (IC50 = 23 nM)
MK-8353 is a potent and selective inhibitor of both active and dormant ERK1 and ERK2 kinases (IC50 = 20 and 7 nM, respectively). MK-8353 inhibits CYP 3A4 (pre-incubation) in vitro and exhibits inhibition of CYP 3A4 and 2C8 (IC50 = 1.7 & 3.5 μM), which can cause drug-drug interactions when co-administered with drugs that are primarily metabolized by CYP 2C8 or 3A4. It does not inhibit human CYPs 1A2, 2C9, 2C19, or 2D6. At 0.6 μM, MK-8353 inhibits hERG current by 16%. In A2058, HT-29, and Colo-205 cells, the IC50 values for inhibiting cell prolifertion are 371, 51, and 23 nM, respectively. MK-8353 also stops MEK from phosphorylating ERK, in addition to inhibiting ERK's kinase activity[1]. MK-8353 exhibits kinase selectivity over a 227-human kinase panel; only 3 kinases (CLK2, FLT4, and Aurora B) are inhibited >50% at the 1.0 μM concentration.No other kinase in the panel is inhibited by more than 35% at the 0.1 μM concentration.

MK-8353 is a potent and selective inhibitor of both active and dormant ERK1 and ERK2 kinases (IC50 = 20 and 7 nM, respectively). MK-8353 inhibits CYP 3A4 (pre-incubation) in vitro and exhibits inhibition of CYP 3A4 and 2C8 (IC50 = 1.7 & 3.5 μM), which can cause drug-drug interactions when co-administered with drugs that are primarily metabolized by CYP 2C8 or 3A4. It does not inhibit human CYPs 1A2, 2C9, 2C19, or 2D6. At 0.6 μM, MK-8353 inhibits hERG current by 16%. In A2058, HT-29, and Colo-205 cells, the IC50 values for inhibiting cell prolifertion are 371, 51, and 23 nM, respectively. MK-8353 also stops MEK from phosphorylating ERK, in addition to inhibiting ERK's kinase activity[1]. MK-8353 exhibits kinase selectivity over a 227-human kinase panel; only 3 kinases (CLK2, FLT4, and Aurora B) are inhibited >50% at the 1.0 μM concentration.No other kinase in the panel is inhibited by more than 35% at the 0.1 μM concentration.

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MK-8353 caused a dose-proportional decrease in the phosphorylated/activated forms of ERK1 (pERK1), ERK2 (pERK2), and ribosomal S6 kinase (pRSK) proteins in A2058 cells. Complete suppression of pERK1 and pERK2 was observed at 30 nM MK-8353 [2].
MK-8353 inhibited the in vitro proliferation of a panel of BRAF^V600E-mutant and RAS-mutant cancer cell lines. The half-maximal effective concentration (EC₅₀) values were determined for these cell lines in 3-day viability assays [2].
In patient-derived melanoma cell lines M428 and M435, MK-8353 (0-10 µM) was tested alone or in combination with trametinib for its effects on cell viability after 120 hours of incubation [2].

In male CD1 mice, Sprague Dawley (SD) rats, guinea pigs, beagle dogs, and cynomologus monkeys, the in vivo pharmacokinetics and metabolism of MK-8353 are examined. A half-life range of 1.3-2.8 hours and a mean residence time range of 1.5-4.0 hours are displayed by MK-8353 in all species, with the exception of monkeys, following IV administration. Dogs, mice, and rats all have a respectable oral bioavailability of between 23 and 80 percent, but monkeys have a low oral bioavailability of only 2%. Given the high (135 nm/sec) intestinal permeability and absorption seen in Caco-2 cells, it is likely that these parameters are also high in human beings. Rats have a steady-state volume of distribution that is 0.1 L/kg, whereas it ranges from 0.9 to 3.3 L/kg in mice, dogs, and monkeys. In numerous BRAF-mutant models, MK-8353 exhibits anti-tumor efficacy[1].

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In female athymic nude mice bearing Colo-205 (BRAF^V600E mutant colon cancer) xenografts, oral administration of MK-8353 at 30-60 mpk twice daily inhibited tumor growth in a dose-dependent manner, with the 60 mpk dose leading to tumor regression [2].
In SCID mice bearing SK-MEL-28 (BRAF^V600E mutant melanoma) xenografts, oral administration of MK-8353 at 30-60 mpk twice daily similarly induced dose-dependent tumor growth inhibition and regression [2].
Treatment of Colo-205 xenografts with a single dose of MK-8353 (30-60 mpk) resulted in significant inhibition of pERK in tumor tissue lysates as early as 1 hour post-dose, although pERK levels rebounded after 10-12 hours. Suppression of pERK was also observed in matched normal skin samples from tumor-bearing mice [2].

The biochemical activity of MK-8353 against ERK1 and ERK2 was evaluated using an IMAP kinase assay to measure inhibition of activated kinases, and a MEK1-ERK2-coupled assay to measure inhibition of non-activated ERK2. IC₅₀ values were derived from these assays [2].
Kinase selectivity profiling was performed against a panel of 227 human kinases to assess the specificity of MK-8353 [2].

A2058 cells (1 × 106 cells per 10-cm dish) were given progressively higher doses of MK-8353 (nM) for 24 hours. Immunoblot analysis was conducted on whole cell lysates.

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For cell proliferation assays, cancer cell lines were plated in 96-well plates. The following day, cells were treated with a 9-point serial dilution of MK-8353 (0.001–10 µM) or DMSO control for 3 days. Cell viability was then assessed using a luminescent cell viability assay kit. EC₅₀ values were calculated from the dose-response curves [2].
For immunoblot analysis of signaling proteins, cells were treated with increasing concentrations of MK-8353 for 24 hours. Cells were then lysed, proteins were separated by SDS-PAGE, transferred to membranes, and probed with specific antibodies against phosphorylated and total forms of ERK1/2 and RSK. Band intensity was detected by chemiluminescence [2].
For experiments with patient-derived M428 and M435 melanoma cells, cells were treated in duplicate with MK-8353 (0–10 µM) and trametinib (0–1 µM) alone or in combination for 120 hours. Cell viability was determined using a luminescent assay [2].

Female athymic nude mice (inoculated with Colo-205 cancer cells) or SCID mice (inoculated with SK-MEL-28 melanoma cells)
30, 45 and 60 mg/kg
p.o.

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For human cancer xenograft models, female athymic nude or SCID mice (4-6 weeks old) were inoculated subcutaneously with cancer cells (e.g., 5 x 10⁶ Colo-205 or SK-MEL-28 cells) suspended in 50% Matrigel. Tumors were allowed to grow to approximately 100 mm³ before treatment initiation [2].
Mice were then randomized into treatment groups (10 mice per group). MK-8353 was administered by oral gavage at doses of 30, 45, or 60 mpk, twice daily. The control group received 20% hydroxypropyl-β-cyclodextrin (HPBCD) vehicle by oral gavage twice daily [2].
Tumor dimensions (length, width, height) were measured with calipers twice weekly to calculate tumor volume. Animal body weight was also monitored twice weekly [2].
For pharmacodynamic analysis, groups of mice (3 per group) bearing Colo-205 xenografts were administered a single dose of MK-8353 or vehicle. Tumors and normal skin from the same animals were harvested at various time points (e.g., 1, 2, 4, 6, 8, 10, 12 hours) post-dose for immunoblot or immunohistochemical analysis of pERK [2].

In female nude mice, plasma exposure [area under the curve (AUC₀₋₂₄)] following oral administration of MK-8353 at doses of 30 to 60 mpk was approximately 15.3 to 55 µM·h [2]. In a study of healthy volunteers (P07652), the median time to peak plasma concentration (tₘₐₓ) of MK-8353 following a single oral dose (10–400 mg) was 1.5–3 hours. The terminal half-life (t₁/₂) ranged from 4.2 to 8.9 hours. Both peak serum concentration (Cₘₐₓ) and AUC₀₋₂₄ increased in a dose-proportional manner [2]. In cancer patients (MK-8353-001 study), after twice-daily oral administration, tₘₐₓ was 2 to 5 hours, and the mean terminal half-life was 5 to 14 hours. Steady-state plasma concentrations were reached after 8 days of continuous administration, with a cumulative fold increase of approximately 2-fold by day 15. Drug exposure (AUC and Cₘₐₓ) in cancer patients was approximately twice that in healthy volunteers at similar doses [2].

In healthy volunteers, single doses of MK-8353 up to 400 mg were generally safe and well tolerated. Associated adverse events included diarrhea, pruritus, and rash. No serious adverse events occurred.[2] In cancer patients, the most common adverse events associated with MK-8353 (of any grade) included diarrhea (44%), fatigue (40%), nausea (32%), maculopapular rash (28%), and vomiting (28%). Grade 3/4 adverse events included diarrhea (16%), asymptomatic indirect elevation of serum bilirubin (16%), and maculopapular rash (8%). No drug-related deaths occurred.[2] Dose-limiting toxicities (DLTs) were observed in the twice-daily 400 mg and 800 mg dose groups. Dose-limiting toxicities (DLTs) included grade 3 diarrhea, hyperbilirubinemia, nausea, vomiting, fatigue, and grade 3 rash leading to missed doses. The twice-daily dose of 400 mg was determined as the initial maximum tolerated dose (MTD).[2]
Reversible unconjugated hyperbilirubinemia was observed in three patients receiving ≥400 mg twice daily. This was attributed to the inhibition of the UGT1A1 enzyme by MK-8353 (in vitro IC₅₀ 0.58 µM), rather than high drug exposure [2].

[1]. ACS Med Chem Lett . 2018 Jun 14;9(7):761-767.

[2]. JCI Insight . 2018 Feb 22;3(4):e92352.

MK-8353 is an indazole compound with the structure 1H-indazole, substituted at position 3 with a 6-(propan-2-oxy)pyridin-3-yl group, and at position 5 with a {[(3S)-3-(methylthio)-1-(2-{4-[4-(1-methyl-1H-1,2,4-triazol-3-yl)phenyl]-3,6-dihydropyridin-1(2H)-yl}-2-oxoethyl)pyrrolidine-3-yl]carbonyl}amino group. In vitro studies have shown that MK-8353 is a potent and selective inhibitor of ERK1 and ERK2 (IC50 values of 23.0 nM and 8.8 nM, respectively). Developed by Merck, this drug is currently in clinical development for the treatment of advanced/metastatic solid tumors. It is an EC 2.7.11.24 (mitogen-activated protein kinase) inhibitor, an antitumor drug, and an inducer of apoptosis. It belongs to the class of indazoles, triazoles, dihydropyridines, pyridines, aromatic ethers, secondary amides, pyrrolidine amides, N-alkylpyrrolidines, methyl sulfides and tertiary amides.
ERK inhibitor MK-8353 is an orally effective extracellular signal-regulated kinase (ERK) inhibitor with potential antitumor activity. After oral administration, MK-8353 can inhibit ERK phosphorylation and activation of ERK-mediated signal transduction pathways, thereby preventing the proliferation and survival of ERK-dependent tumor cells. The mitogen-activated protein kinase (MAPK)/ERK pathway is often upregulated in various tumor cell types and plays a role in the proliferation, differentiation and survival of tumor cells.

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MK-8353 (SCH900353) is a dual-mechanism ERK1/2 inhibitor. It can not only bind to and inhibit the intrinsic kinase activity of ERK1/2, but also induce conformational changes, thereby preventing phosphorylation by upstream kinases (such as MEK)[2].
In a phase I clinical trial (MK-8353-001) in patients with advanced solid tumors, MK-8353 showed antitumor activity. Of the 15 evaluable patients, 3 achieved partial response (PR), all of whom carried BRAF V600E mutant melanoma, resulting in an objective response rate of 20% in evaluable patients[2].
The phase II recommended dose (RP2D) was not formally established because the study was terminated early by the sponsor for strategic reasons rather than toxicity. However, in the expanded cohort, doses of up to 400 mg orally twice daily were considered safe and well-tolerated[2].
Pharmacodynamic assessments of consecutive normal skin biopsies using pERK immunohistochemistry showed no consistent and significant dose-dependent inhibition in the patient population, highlighting the variability of this potential biomarker[2].

C37H41N9O3S

691.8449

691.31

C, 64.23; H, 5.97; N, 18.22; O, 6.94; S, 4.63

1184173-73-6

1184173-73-6;1951448-73-9 (HCl);1951448-74-0 (HCl hydrate);1951428-60-6 (hydrate); 1951428-61-7 (x HCl);

58282870

Off-white to light brown solid powder

4.3

2

9

10

50

1220

1

CC(C)OC1=NC=C(C=C1)C2=NNC3=C2C=C(C=C3)NC(=O)[C@@]4(CCN(C4)CC(=O)N5CCC(=CC5)C6=CC=C(C=C6)C7=NN(C=N7)C)SC

KPQQGHGDBBJGFA-QNGWXLTQSA-N

InChI=1S/C37H41N9O3S/c1-24(2)49-32-12-9-28(20-38-32)34-30-19-29(10-11-31(30)41-42-34)40-36(48)37(50-4)15-18-45(22-37)21-33(47)46-16-13-26(14-17-46)25-5-7-27(8-6-25)35-39-23-44(3)43-35/h5-13,19-20,23-24H,14-18,21-22H2,1-4H3,(H,40,48)(H,41,42)/t37-/m0/s1

(3S)-3-methylsulfanyl-1-[2-[4-[4-(1-methyl-1,2,4-triazol-3-yl)phenyl]-3,6-dihydro-2H-pyridin-1-yl]-2-oxoethyl]-N-[3-(6-propan-2-yloxypyridin-3-yl)-1H-indazol-5-yl]pyrrolidine-3-carboxamide

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: ≥ 2.08 mg/mL (3.01 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 20.8 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: ≥ 2.08 mg/mL (3.01 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 20.8 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.)