Home Angiogenesis Syk inhibitor - FLT3 inhibitor - Apoptosis related activator R406 (free base) For research use only.

R406 (free base)

R406 (free base) is a potent Syk inhibitor with IC50 of 41 nM in a cell-free assay, strongly inhibits Syk but not Lyn, 5-fold less potent to Flt3. R406 (free base) triggers apoptosis. Phase 1.

R406 (free base) Chemical Structure

R406 (free base) Chemical Structure

CAS No. 841290-80-0

Purity & Quality Control

R406 (free base) Related Products

Signaling Pathway

Syk Signaling Pathways

Syk Signaling Pathway

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
MV411 Function assay 72 hrs Inhibition of Flt3 in human MV411 cells assessed as assessed as proliferation after 72 hrs incubation by spectrophotometry, EC50=0.01μM 24779514
TF1 Function assay 1 hr Inhibition of Jak2 in erythropoietin-stimulated human TF1 cells assessed as assessed as phospho-Stat5 after 1 hr incubation, EC50=0.013μM 24779514
neutrophils cells Function assay Inhibition of SYK in human neutrophils cells assessed as reduction in FcepsilonR1/FcgammaR-mediated signaling responses, EC50=0.033μM 22257213
SK-M-MC Function assay 1 hr Inhibition of Ret in human SK-M-MC cells assessed as assessed as phosphorylation after 1 hr incubation, EC50=0.036μM 24779514
mast cells Function assay 1 hr Inhibition of cKit in stem cell factor-stimulated bone marrow derived mouse mast cells assessed as phosphorylation after 1 hr incubation, EC50=0.046μM 24779514
B-cells Function assay Inhibition of SYK in human B-cells cells assessed as reduction in FcepsilonR1/FcgammaR-mediated signaling responses, EC50=0.048μM 22257213
Ramos cells Function assay Inhibition of Syk in antihuman IgM-stimulated human Ramos cells assessed as decrease in BCR-mediated BLNK phosphorylation by cellular assay, EC50=0.053μM 24779514
mesangial cells Function assay Inhibition of SYK in cultured human mesangial cells assessed as reduction in FcepsilonR1/FcgammaR-mediated signaling responses, EC50=0.056μM 22257213
SK-N-SH Function assay Inhibition of Ret in human SK-N-SH cells, EC50=0.08μM 22257213
bone marrow cells Function assay Inhibition of IL3 dependent proliferation in C57/B16 mouse bone marrow cells using [3H]thymidine by liquid scintillation counting, IC50=0.147μM 24726806
B-cells Function assay 1 hr Inhibition of Syk in alphaIgM-stimulated human B cells assessed as cell proliferation after 1 hr incubation by flow cytometry, EC50=0.151μM 24779514
THP1 Function assay Inhibition of SYK in human THP1 cells assessed as reduction in FcepsilonR1/FcgammaR-mediated signaling responses, EC50=0.171μM 22257213
B-cells Function assay 1 hr Inhibition of Syk in alphaIgM-stimulated human B cells assessed as CD86 expression after 1 hr incubation by flow cytometry, EC50=0.335μM 24779514
Ramos Function assay Inhibition of Syk in anti IgM-stimulated human Ramos cells assessed as BLNK phosphorylation by cellular assay, IC50=0.457μM 24726806
Rec1 Function assay 2.5 uM 6 hrs Inhibition of Syk phosphorylation in human Rec1 cells at 2.5 uM incubated for 6 hrs by Western blotting method 25222877
Rec1 Function assay 2.5 uM 6 hrs Inhibition of Lyn phosphorylation in human Rec1 cells at 2.5 uM incubated for 6 hrs by Western blotting method 25222877
Rec1 Function assay 2.5 uM 6 hrs Inhibition of BTK phosphorylation in human Rec1 cells at 2.5 uM incubated for 6 hrs by Western blotting method 25222877
SJ-GBM2 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SJ-GBM2 cells 29435139
A673 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for A673 cells 29435139
SK-N-MC qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for SK-N-MC cells 29435139
NB-EBc1 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for NB-EBc1 cells 29435139
Saos-2 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Saos-2 cells 29435139
OHS-50 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for OHS-50 cells 29435139
Rh41 qHTS assay qHTS of pediatric cancer cell lines to identify multiple opportunities for drug repurposing: Primary screen for Rh41 cells 29435139
Click to View More Cell Line Experimental Data

Biological Activity

Description R406 (free base) is a potent Syk inhibitor with IC50 of 41 nM in a cell-free assay, strongly inhibits Syk but not Lyn, 5-fold less potent to Flt3. R406 (free base) triggers apoptosis. Phase 1.
Targets
FLT3 [1] Syk [1]
(Cell-free assay)
41 nM
In vitro
In vitro R406 is an ATP-competitive inhibitor of Syk with a Ki value of 30 nM. R406 selectively inhibits Syk-dependent signaling with EC50 values ranging from 33 nM to 171 nM, more potently than Syk-independent pathways in different cells. [1] R406 inhibits cellular proliferation of a large panel of diffuse large B-cell lymphoma (DLBCL) cell lines at EC50 values ranging from 0.8 μM to 8.1 μM. R406 treatment (1 μM or 4 μM) induces the activation of caspases 9 and 3, but not caspase 8, leading to significant apoptosis of the majority of DLBCL cell lines. Pretreatment of R406 completely blocks the phosphorylation of SYK525/526 and the SYK-dependent phosphorylation of BLNK in R406-sensitive DLBCLs following B-cell receptor (BCR) crosslinking. [2] R406 potently decreases MMP-9 mRNA levels by 2.8- and 4.3-fold lower than controls after 24 and 48 hours treatment, respectively, and reduces the invasive capacity of the RL cells. [3]
Kinase Assay In-vitro Fluorescence Polarization Kinase Assays
R406 is serially diluted in DMSO and then diluted to 1% DMSO in kinase buffer (20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated BGG). ATP and substrate in kinase buffer are added at room temperature, resulting in a final DMSO concentration on 0.2%. The kinase reactions are performed in a final volume of 20 μL containing 5 μM HS1 peptide substrate and 4 μM ATP and started by addition of 0.125 ng of Syk in kinase buffer. The reaction is allowed to proceed for 40 minutes at room temperature. The reaction is stopped by the addition of 20 μL of PTK quench mix containing EDTA/anti-phosphotyrosine antibody (1X final)/fluorescent phosphopeptide tracer (0.5X final) diluted in FP Dilution Buffer. The plate is incubated for 30 minutes in the dark at room temperature and then read on a Polarion fluorescence polarization plate reader. Data are converted to amount of phosphopeptide present using a calibration curve generated by competition with the phosphopeptide competitor provided in the Tyrosine Kinase Assay Kit. For IC50 determination, R406 is tested at eleven concentrations in duplicate and curve-fitting is performed by non-linear regression analysis using Prism GraphPad Software.
Cell Research Cell lines DHL4, DHL6, DHL8, DHL10, Wsu-NHL, Karpas422 (K422), OCI LY1, LY3, LY4, LY7, LY10, LY18, LY19, Pfeiffer, and Toledo
Concentrations Dissolved in DMSO at a concentration of 10 mM, final concentrations ~ 5 μM
Incubation Time 72 or 96 hours
Method

DLBCL cell lines are treated with serial dilutions of R406 (0.3, 0.6, 1.25, 2.5, or 5 μM) for 72 or 96 hours. Thereafter, cellular proliferation is determined by MTT assay, and cell apoptosis is assessed by using annexin V–FITC/propidium iodide (PI) staining. For the determination of caspase 9, 8, and 3, cells are lysed, size-fractionated by polyacrylamide gel electrophoresis (PAGE), and immunoblotted.
In Vivo
In vivo R406 has shown efficacy in a number of animal models of immune disorders. Oral administration of R406 in mice with immune complex-mediated inflammation significantly inhibits the cutaneous reverse passive Arthus reaction by approximately 72% and 86% at 1 mg/kg and 5 mg/kg, respectively, compared with the control. R406 treatment at 10 mg/kg significantly reduces inflammation and swelling, decreases the progressive arthritis to a lower level in the passive anticollagen antibody-challenged mice, and delays the onset and reduces paw thickening and clinical arthritis by approximately 50% in the K/BxN serum transfer mice model. [1]
Animal Research Animal Models Female C57BL/6 mice challenged intravenously with 1% ovalbumin (OVA) in saline (10 mg/kg) containing 1% Evans blue dye, female Balb/c mice with the anticollagen antibody-induced arthritis, and female C57BL/6 mice with arthritis induced by intraperitoneal
Dosages ~10 mg/kg/day
Administration Orally (prodrug R788)
NCT Number Recruitment Conditions Sponsor/Collaborators Start Date Phases
NCT01725230 Completed
Rheumatoid Arthritis
AstraZeneca
 November 2012 Phase 1
NCT01598571 Completed
Healthy
AstraZeneca
 May 2012 Phase 1
NCT01387308 Completed
Healthy
AstraZeneca
 August 2011 Phase 1
NCT01355354 Completed
Healthy Volunteers|Rheumatoid Arthritis
AstraZeneca
 June 2011 Phase 1

Chemical Information & Solubility

Molecular Weight 470.45 Formula

C22H23FN6O5
CAS No. 841290-80-0 SDF Download R406 (free base) SDF
Smiles CC1(C(=O)NC2=C(O1)C=CC(=N2)NC3=NC(=NC=C3F)NC4=CC(=C(C(=C4)OC)OC)OC)C
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 29 mg/mL ( (61.64 mM) Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Water : Insoluble

Ethanol : Insoluble


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In vivo
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Tech Support

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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