PF-3644022

Catalog No.S8224

For research use only.

PF-3644022 is a potent freely reversible ATP-competitive inhibitor of MAPKAPK2 (MK2) with an Ki of 3 nM. PF-3644022 also potently inhibits TNFα production with an IC50 of 160 nM.

PF-3644022 Chemical Structure

CAS No. 1276121-88-0

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Biological Activity

Description

PF-3644022 is a potent freely reversible ATP-competitive inhibitor of MAPKAPK2 (MK2) with an Ki of 3 nM. PF-3644022 also potently inhibits TNFα production with an IC50 of 160 nM.

Targets
MK2 [1] TNFα [1]
3 nM(Ki) 160 nM
In vitro

PF-3644022 is a potent freely reversible ATP-competitive compound that inhibits Mitogen-activated protein kinase-activated protein kinase 2(MK2) activity with good selectivity when profiled against 200 human kinases. In the human U937 monocytic cell line or peripheral blood mononuclear cells, PF-3644022 potently inhibits TNFα production with similar activity. PF-3644022 blocks TNFα and IL-6 production in LPS-stimulated human whole blood.<sup><a class="sref" href="#s_ref">[1]</a></sup>

In vivo

PF-3644022 displays good pharmacokinetic parameters in rats and is orally efficacious in both the rat acute LPS-induced TNFα model and the chronic streptococcal cell wall-induced arthritis model,with ED50 values of 6.9 and 20 mg/kg, respectively.<sup><a class="sref" href="#s_ref">[1]</a></sup>

Protocol (from reference)

Kinase Assay:

<sup><a class="sref" href="#s_ref">[1]</a></sup>

  • In Vitro Kinase Assays and Inhibition Kinetics.:

    The kinase activity of MK2 is assayed by using fluorescently labeled heat shock protein 27 (HSP27) peptide and a Caliper LabChip 3000. Phosphorylated peptide is separated from substrate peptide electrophoretically and quantified. All kinase reactions are performed at room temperature in 20 mM HEPES, containing 10 mM MgCl2, 1 mM dithiothreitol, 0.01% bovine serum albumin, and 0.0005% Tween 20, pH 7.5. For endpoint experiments, reactions are terminated during the linear phase by the addition of 30 mM EDTA. The kinase selectivity experiments are performed with the MgATP concentration fixed at the Km(app) determined for each enzyme. The initial velocitie is determined in the presence and absence of PF-3644022 with ATP as the varied substrate while the HSP27 peptide concentration is held constant.

Cell Research:

<sup><a class="sref" href="#s_ref">[1]</a></sup>

  • Cell lines: Human U937 monocytic cell line, hPBMCs
  • Concentrations: 0.1 nM-100μM
  • Incubation Time: 1 h
  • Method:

    U937 cells and hPBMCs are pretreated with PF-3644022 at varying concentrations for 1 h before LPS stimulation at 100 ng/ml. TNFα levels in U937 cell are measured at 4 h after LPS stimulation by using an electrochemoluminescence MesoScale Discovery TNF kit. In hPBMCs, TNFα and IL-6, IL-8 are measured at 16 h after LPS stimulation by using a four-plex human cytokine MSD plate.

Animal Research:

<sup><a class="sref" href="#s_ref">[1]</a></sup>

  • Animal Models: LPS-induced acute endotoxemia inflammation model (adult male Lewis rats), chronic streptococcal cell wall-induced arthritis model (female Lewis rats)
  • Dosages: 1 mg/kg
  • Administration: Oral gavage

Solubility (25°C)

In vitro

Chemical Information

Molecular Weight 374.46
Formula

C21H18N4OS

CAS No. 1276121-88-0
Storage 3 years -20°C powder
2 years -80°C in solvent

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Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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