Home Angiogenesis Syk inhibitor R406

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R406 Syk/FLT3 Inhibitor

Cat.No.S2194

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

Chemical Structure

Molecular Weight: 628.63

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Quality Control

Batch: Purity: 99.43%
99.43

Cell Culture, Treatment & Working Concentration

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
AMO-1 Function Assay 1 μM 3 h reduces migration  26251761
U266 Function Assay 1 μM 3 h reduces migration  26251761
Jeko-1 Growth Inhibition Assay 48 h IC50=5.06826 μM 25835755
Mino Growth Inhibition Assay 48 h IC50=5.70854 μM 25835755
Jeko-1 Apoptosis Assay 5 μM 24 h induces 25.1 ± 3.2 % apoptosis 25835755
primary MCL Apoptosis Assay 2 µM 24 h increases significantly apoptosis  25388373
PBMCs Cell Viability Assay 0-50 μM 24 h DMSO inhibits cell viability dose dependently 25127862
PBMCs Function Assay 5 μM 1 h DMSO decreases the cell migration 25127862
CFSE-CD4+ T  Growth Inhibition Assay 0.0625-1 μM 4 d blocks proliferation of GVHD-derived CD4+ T cells and CD11b+ cells 24679982
CFSE-CD11b+ Growth Inhibition Assay 0.0625-1 μM 8 d blocks proliferation of GVHD-derived CD4+ T cells and CD11b+ cells 24679982
HMECs Function Assay 0-10 μM 20 min inhibits VEGF-stimulated release of NO 24329544
AB5 Apoptosis Assay 0-2.5 μM 48 h DMSO induces apoptosis 23398911
JB7 Apoptosis Assay 0-2.5 μM 48 h DMSO induces apoptosis 23398911
AB5 Growth Inhibition Assay 0-2.5 μM 48 h DMSO induces cell cycle arrest 23398911
JB7 Growth Inhibition Assay 0-2.5 μM 48 h DMSO induces cell cycle arrest 23398911
RL Function Assay 2.5/5 μM 24/48 h DMSO induces a potent decrease in MMP-9 mRNA expression 21926965
RL Function Assay 1/2.5 μM 24 h DMSO reduces the activation of Akt and p70S6K 21926965
platelet  Function Assay 1 μm 5 min inhibits FcγRIIA-mediated platelet aggregation 21848694
platelet  Function Assay 0.05/1/2.5 μM 5 min inhibits the signaling mechanisms downstream of FcγRIIA 21848694
DoHH2 Apoptosis Assay 0/3/10 μM 48 h induces cell death significantly 20875408
Jeko-1  Apoptosis Assay 0/3/10 μM 48 h induces cell death significantly 20875408
Raji  Apoptosis Assay 0/3/10 μM 48 h induces cell death significantly 20875408
DHL4 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
LY7 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
LY3 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
DHL6 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
LY10 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
DHL10 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
Wsu-NHL Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
LY18 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
LY1 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
DHL8 Apoptosis Assay 0/1/4 μM 96 h induces apoptosis dose dependently 18006696
DHL4 Apoptosis Assay 4 μM 96 h induces cleavage of caspases 9 and 3, but not caspase 8 18006696
DHL6 Apoptosis Assay 4 μM 96 h induces cleavage of caspases 9 and 3, but not caspase 8 18006696
LY3 Apoptosis Assay 4 μM 96 h induces cleavage of caspases 9 and 3, but not caspase 8 18006696
LY7 Apoptosis Assay 4 μM 96 h induces cleavage of caspases 9 and 3, but not caspase 8 18006696
DHL4 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
LY7 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
LY3 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
DHL6 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
LY10 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
Wsu-NHL Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
LY18 Function Assay 4 μM 16 h inhibits tonic BLNK tyrosine phosphorylation 18006696
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 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 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
mouse 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 BTK 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 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
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

Solubility

In vitro
Batch:

DMSO : 100 mg/mL (159.07 mM)
(Moisture-contaminated DMSO may reduce solubility. Use fresh, anhydrous DMSO.)

Water : Insoluble

Ethanol : 0 mg/mL

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Mass Concentration Volume Molecular Weight
Dilution Calculator Molecular Weight Calculator

In vivo
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Working concentration: mg/ml;

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Chemical Information, Storage & Stability

Molecular Weight 628.63 Formula

C22H23FN6O5.C6H6O3S

 Storage (From the date of receipt)
CAS No. 841290-81-1 Download SDF Storage of Stock Solutions

Synonyms R406 besylate Smiles CC1(C(=O)NC2=C(O1)C=CC(=N2)NC3=NC(=NC=C3F)NC4=CC(=C(C(=C4)OC)OC)OC)C.C1=CC=C(C=C1)S(=O)(=O)O

Mechanism of Action

Features
Lead drug candidate for rheumatoid arthritis.
Targets/IC50/Ki
Flt3
(Cell-free assay)
Syk
(Cell-free assay)
41 nM
In vitro

R406 is a potent inhibitor of immunoglobulin E (IgE)- and IgG-mediated activation of Fc receptor signaling. This compound inhibits the anti-IgE-induced production and release of LTC4 and cytokines and chemokines, including TNFα, IL-8, and GM-CSF. It inhibits phosphorylation of Syk substrate linker for activation of T cells in mast cells and B-cell linker protein/SLP65 in B cells. This chemical binds to the ATP binding pocket of Syk and inhibits its kinase activity as an ATP-competitive inhibitor with Ki of 30 nM. It blocks Syk-dependent FcR-mediated activation of monocytes/macrophages and neutrophils and Bcr-mediated activation of B lymphocytes.

This compound significantly induces chronic lymphocytic leukemia (CLL) cell apoptosis in nurselike cells cocultures and blocks CCL3 and CCL4 secretion by CLL cells in response to B-cell antigen receptor (Bcr) triggering.

It is a potent inhibitor of platelet signaling and functions initiated by FcγRIIA cross-linking by specific antibodies or by sera from HIT patients.

Kinase Assay
In Vitro Fluorescence Polarization Kinase Assay
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 mL containing 5mM HS1 peptide substrate, 4 mM 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 mL 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, this compound is tested at eleven concentrations and curve-fitting is performed by non-linear regression analysis.
In vivo

R406 reduces cutaneous reverse passive Arthus reaction by approximately 86% at 5 mg/kg in prophylactic treated mice. This compound also shows efficacy in inhibiting paw inflammation in antibody-induced arthritis mouse models.

It does not adversely affect macrophage or neutrophil function in innate immune responses and has minimal functional immunotoxicity notwithstanding its lymphocytopenic effect.

References
  • [4] https://pubmed.ncbi.nlm.nih.gov/17490694/

Applications

Methods Biomarkers Images PMID
Western blot p-RPS6 / T-RPS6 / p-4E-BP1 / T-4E-BP1 p-MEK / T-MEK / p-ERK / T-ERK p-c-RAF / T-c-RAF p-AKT / T-AKT / p-mTOR / T-mTOR
S2194-WB1
23535559
Growth inhibition assay Cell viability (U87, U251 cells) Cell viability (GSC lines)
S2194-viability1
31043589

Clinical Trial Information

(data from https://clinicaltrials.gov, updated on 2024-05-22)

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

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Handling Instructions

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Frequently Asked Questions

Question 1:
What’s the difference between S1533 and S2194?

Answer:
S1533 and S2194 are two different forms of this compound. S1533 is the free base form, containing only its molecule without an acid added to it. S2194 has an additional C6H6O3S acid on it which makes the molecule a salt form. The free base and salt forms have same biology activities. Free base has a lower molecular weight and salt form has a better solubility in DMSO.

Signaling Pathway Map