For research use only.

Catalog No.S7198 Synonyms: GSK-3 Inhibitor IX, 6-bromoindirubin-3-oxime

11 publications

BIO Chemical Structure

CAS No. 667463-62-9

BIO is a specific inhibitor of GSK-3 with IC50 of 5 nM for GSK-3α/β in a cell-free assay, shows >16-fold selectivity over CDK5, also a pan-JAK inhibitor with IC50 of 30 nM for Tyk2. BIO induces apoptosis in human melanoma cells.

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Selleck's BIO has been cited by 11 publications

2 Customer Reviews

  • Lysates of HCT116p53KO cells were harvested 24 hs after treatment with different GSK3 inhibitors and GSK3A/B activation/inactivation checked by western blot: a mix of pSer21-GSK3A and pSer9-GSK3B antibodies and antibody cross-reacting with both pTyr279-GSK3A and pTyr216-GSK3B were used to assess the specificity of the inhibitor for GSK3A. BIO: 6-bromoindirubin-3'-oxime, TWS: TWS119, SB2: SB216763, SB4: SB415286.

    PLoS One 2014 9(7), e100947. BIO purchased from Selleck.

    (e and f) Confocal immunofluorescence staining. U87 GSLCs and U251 GSLCs were seeded in suspension and treated with control, Tet, BIO or ICG-001 for 48 h. Blue fluorescence represents DAPI, red fluorescence represents β-catenin, and green fluorescence represents TUNEL. Scale bars, 40 μm

    Int J Oncol, 2017, 50(1):101-110. BIO purchased from Selleck.

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Biological Activity

Description BIO is a specific inhibitor of GSK-3 with IC50 of 5 nM for GSK-3α/β in a cell-free assay, shows >16-fold selectivity over CDK5, also a pan-JAK inhibitor with IC50 of 30 nM for Tyk2. BIO induces apoptosis in human melanoma cells.
Features The first pharmacological agent shown to maintain self-renewal in human and mouse embryonic stem cells.
GSK-3 [1]
(Cell-free assay)
TYK2 [4]
(Cell-free assay)
CDK5/p35 [1]
(Cell-free assay)
CDK2/CyclinA [1]
(Cell-free assay)
CDK1/CyclinB [1]
(Cell-free assay)
5 nM 30 nM 0.08 μM 0.30 μM 0.32 μM
In vitro

BIO (6-bromoindirubin-3'-oxime) is a specific inhibitor of glycogen synthase kinase-3 (GSK-3), with IC50 of 5 nM for GSK-3α/β, shows >16-fold selectivity over CDK5. BIO interacts within the ATP binding pocket of these kinases, reduces β-catenin phosphorylation on a GSK-3-specific site in cellular models, closely mimicks Wnt signaling in Xenopus embryos. [1] In human and mouse embryonic stem cells, BIO maintains the undifferentiated phenotype and sustains expression of the pluripotent state-specific transcription factors Oct-3/4, Rex-1 and Nanog. BIO-mediated Wnt activation is functionally reversible, as withdrawal of the compound leads to normal multidifferentiation programs in both human and mouse embryonic stem cells. [2] BIO promotes proliferation in mammalian cardiomyocytes. [3]6BIO is also a pan-JAK inhibitor, with IC50 values of 0.03, 1.5, 8.0, 0.5 μM for TYK2, JAK1, JAK2 and JAK3. BIO selectively inhibits phosphorylation of STAT3 and induces apoptosis of human melanoma cells. [4]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
human SH-SY5Y cells NGO1V5dHfW6ldHnvckBie3OjeR?= NWDVb2FMUW6qaXLpeIlwdiCxZjDHV2s{NW2nZHnheIVlKGKndHGtZ4F{\WmwIIDoc5NxcG:{eXzheIlwdiCrbjDoeY1idiCVSD3TXVV[KGOnbHzzJIlvKHC{ZYPlcoNmKG:oIF3HNVMzKGK7IGfld5Rmem5iYnzveEBidmGueYPpd{whUUN3ME2wMlI6KM7:TR?= M2\PS|E5QDF4MUGw
human K562 cells MVPQdo9tcW[ncnH0bY9vKGG|c3H5 MVK3NkBp M{DKbGFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgbJVu[W5iS{W2NkBk\WyuczDh[pRmeiB5MjDodpMh[nliTWTUJIF{e2G7LDDJR|UxRTFwMzFOwG0> NYOxSVZqOTl5OEOxOFk>
human IMR90 cells MnLlVJJwdGmoZYLheIlwdiCjc4PhfS=> MmXSO|IhcA>? MV7BcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IHj1cYFvKEmPUkmwJINmdGy|IHHmeIVzKDd{IHjyd{BjgSCPVGSgZZN{[XluIFnDOVA:OS57IN88US=> NE\ZN5kyQTd6M{G0PS=>
human HCT116 cells MVjQdo9tcW[ncnH0bY9vKGG|c3H5 NXLSd2M{PzJiaB?= NU\LcXI4SW62aYDyc4xq\mW{YYTpeoUh[WO2aY\peJkh[WejaX7zeEBpfW2jbjDIR3QyOTZiY3XscJMh[W[2ZYKgO|IhcHK|IHL5JG1VXCCjc4PhfUwhUUN3ME21MlIh|ryP NFiyUGEyQTd6M{G0PS=>
human HL60 cells Mn;TVJJwdGmoZYLheIlwdiCjc4PhfS=> Mn7IOUBl[Xm| NEXEVJpCdnSrcILvcIln\XKjdHn2[UBi[3Srdnn0fUBi\2GrboP0JIh2dWGwIFjMOlAh[2WubIOgZYZ1\XJiNTDkZZl{KGK7IF3UWEBie3OjeR?= MYKxPVc5OzF2OR?=
human HuH7 cells MXXQdo9tcW[ncnH0bY9vKGG|c3H5 NHHQZoM4OiCq MX3BcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IHj1cYFvKEi3SEegZ4VtdHNiYX\0[ZIhPzJiaILzJIJ6KE2WVDDhd5NigQ>? M{P0SlE6Pzh|MUS5
human SH-SY5Y cells MXvDfZRwfG:6aXRCpIF{e2G7 MXi0PEBp M4W5O2N6fG:2b4jpZ4l1gSCjZ3HpcpN1KGi3bXHuJHNJNVO\NWmgZ4VtdHNiYX\0[ZIhPDhiaILzJIJ6KE2WUzDy[YR2[3Srb36gZZN{[XluIFnDOVA:QSEQvF2= NUjXclFbOTh6MU[xNVA>
human NB39 cells M3XYUGN6fG:2b4jpZ:Kh[XO|YYm= NVfoXHZ7PDhiaB?= MmO0R5l1d3SxeHnjbZR6KGGpYXnud5QhcHWvYX6gUmI{QSClZXzsd{Bie3Onc4Pl[EBieyClZXzsJJZq[WKrbHn0fUBi\nSncjC0PEBpenNiYomgUXRVKGG|c3H5 MYeyNVgxOjl2Nx?=
human SK-N-SH cells MXPDfZRwfG:6aXRCpIF{e2G7 MoHkOFghcA>? M1LYOWN6fG:2b4jpZ4l1gSCjZ3HpcpN1KGi3bXHuJHNMNU5vU1igZ4VtdHNiYYPz[ZN{\WRiYYOgZ4VtdCC4aXHibYxqfHliYX\0[ZIhPDhiaILzJIJ6KE2WVDDhd5NigQ>? MoftNlE5ODJ7NEe=
mouse HT22 cells M4LyWGZ2dmO2aX;uJIF{e2G7 NVr0dVlyOTBizszN NHLXVoQzPCCq NGTLOVFKdmirYnn0bY9vKG:oIFfTT|MudWWmaXH0[YQh[mW2YTDjZZNmcW5icHjvd5Bpd3K7bHH0bY9vKGmwIH3veZNmKEiWMkKgZ4VtdHNiYYSgNVAhfU1iYX\0[ZIhOjRiaILzJIJ6KFenc4Tldo4h[myxdDDhcoFtgXOrczDpckBxemW|ZX7j[UBw\iCPR{GzNi=> NIrLUHczOjl7OES0Ny=>
HEK293 cells NXjZbndXTnWwY4Tpc44h[XO|YYm= NYjZeo9IOC53LUGg{txO MnXoTY5pcWKrdHnvckBw\iCqdX3hckBIW0t|IHHjeIl3cXS7IHnuJGhGUzJ7MzDj[YxteyClb370ZYlvcW6pIITo[UBYdnRxYnX0ZU1k[XSnbnnuJIFkfGm4YYTl[EBz\XCxcoTldkBxW3WyZYLUU3BHVEGVSDCoV3RHOjl|IHPlcIx{MSCjdDCwMlUhfG9iMTD1UUBjgSCZboSgdoVxd3K2ZYKg[4Vv\SCjc4PhfS=> MUKyOFY6PzJ2NB?=

... Click to View More Cell Line Experimental Data

Methods Test Index PMID
Western blot
p-AKT / AKT / p21 / p27 ; 

PubMed: 27510556     

In BIO treatment of cardiac fibroblasts for 1 day, the protein levels of phosphorylated Akt, p21, and p27 were assessed.

p-β-catenin / β-catenin ; 

PubMed: 27211495     

ERKi and GSK3i differently regulate PI3K/Akt and β-catenin pathways. U0126 increased while BIO decreased Akt phosphorylation after 24 hours of treatment. BIO prevented β-catenin phosphorylation leading to intact β-catenin accumulation, while U0126 only had a marginal effect.

FoxO3a / FoxO1 / p-FoxO3a / p-FoxO1 ; 

PubMed: 27211495     

U0126 markedly increased the phosphorylation of Akt downstream targets FOXO1 and FOXO3a, while BIO decreased it.

27510556 27211495
pAKT / p21 / p27 ; 

PubMed: 27510556     

Protein expressions of pAkt, p21 and p27 were determined by immunofluorescence staining in cardiac fibroblasts treated with BIO for 1 day. *p < 0.05, t-test. Scale bar = 100 μm.


PubMed: 24286133     

Detection of TNF-α protein expression by immunofluorescence microscopy. Nucleus pulposus cells were cultured with or without 1.0 μM BIO for 24 h, fixed, and stained with an antibody against c-fos. Left: cells stained with an antibody to TNF-α; middle: cells stained with 4′,6-diamidino-2-phenylindole (DAPI), to identify healthy nuclei; right: cells stained with an antibody to TNF-α and with DAPI. Scale bar, 50 μm (original magnification, 20×).

E-cadherin / Nanog ; 

PubMed: 25538040     

hESCs were treated with either vehicle (Veh) or BIO for 6 hours. Twenty-four hours after removal of BIO, hESCs exhibit thick and compact colonies that express higher levels of E‐cadherin (E‐cad) and Nanog.


PubMed: 25538040     

hESCs were treated with either vehicle (Veh) or BIO for 6 hours. Twenty‐four hours after removal of BIO, hESCs exhibit thick and compact colonies that express higher levels of E‐cadherin (E‐cad) and Oct3/4. In contrast, long-term BIO treatment leads to opposing results. Scale bar = 20 µm.

27510556 24286133 25538040
Growth inhibition assay
Cell proliferation ; 

PubMed: 27510556     

The increase in cardiomyocyte proliferation after treatment with BIO for 4 days was also confirmed by MTT assay.

In vivo BIO suppresses melanoma tumor growth in a mouse xenograft model. [4]


Kinase Assay:[1]
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Kinase assay:

Kinase activities are assayed in Buffer A or C at 30°C, at a final ATP concentration of 15 μM. Blank values are subtracted and activities calculated as pmoles of phosphate incorporated during a 10 min incubation. Controls are performed with appropriate dilutions of dimethylsulfoxide. In a few cases phosphorylation of the substrate is assessed by autoradiography after SDS-PAGE. GSK-3α/β is purified from porcine brain by affinity chromatography on immobilized axin. It is assayed, following a 1/100 dilution in 1 mg BSA/ml 10 mM DTT, with 5 μl 40 μM GS-1 peptide, a specific GSK-3 substrate, (YRRAAVPPSPSLSRHSSPHQSpEDEEE), in buffer A, in the presence of 15 μM [γ-32P] ATP (3,000 Ci/mmol; 1 mCi/ml) in a final volume of 30 μl. After 30 min incubation at 30°C, 25 μl aliquots of supernatant are spotted onto 2.5 × 3 cm pieces of Whatman P81 phosphocellulose paper, and 20 seconds later, the filters are washed five times (for at least 5 min each time) in a solution of 10 ml phosphoric acid/liter of water. The wet filters are counted in the presence of 1 ml ACS scintillation fluid.
Cell Research:[1]
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  • Cell lines: COS1, Hepa or SH-SY5Y cells
  • Concentrations: ~10 μM
  • Incubation Time: 12 or 24 h
  • Method: COS1, Hepa (wild-type, CEM/LM AhR deficient and ELB1 ARNT deficient), or SH-SY5Y cells are grown in 6 cm culture dishes in Dulbecco's Modified Medium (DMEM) containing 10% fetal bovine serum. For treatment, IO (5 μM), BIO (5 or 10 μM), MeBIO (5 or 50 μM), LiCl (20 or 40 mM), or mock solution (DMSO, 0.5% final concentration) is added to medium when cell density reaches ∼70% confluence. After 12 (SH-SY5Y) or 24 hours, the cells, while still in plate, are lysed with lysis buffer (1% SDS, 1 mM sodium orthovanadate, 10 mM Tris [pH 7.4]). The lysate is passed several times through a 26G needle, centrifuged at 10,000 × g for 5 min, and adjusted to equal protein concentration. About 8 μg of each sample is loaded for immunoblotting. Enhanced chemiluminescence is used for detection. The following primary antibodies are used: mouse anti-β-catenin CT (Upstate Biotechnolgies, Clone 7D8, recognizes total β-catenin), mouse anti-phospho-β-catenin (Upstate Biotechnologies, Clone 8E7, recognizes dephosphorylated β-catenin), mouse anti-GSK-3 β, mouse anti-GSK-3 phosphoTyr216, rabbit anti-AhR (Aryl hydrocarbon receptor), and rabbit anti-actin.
    (Only for Reference)
Animal Research:[4]
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  • Animal Models: mouse
  • Dosages: 50 mg/kg
  • Administration: Oral gavage
    (Only for Reference)

Solubility (25°C)

In vitro DMSO 71 mg/mL (199.34 mM)
Water Insoluble
Ethanol '6 mg/mL
In vivo Add solvents to the product individually and in order(Data is from Selleck tests instead of citations):
30% PEG400+0.5% Tween80+5% propylene glycol
For best results, use promptly after mixing.
30 mg/mL

* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.

Chemical Information

Molecular Weight 356.17


CAS No. 667463-62-9
Storage powder
in solvent
Synonyms GSK-3 Inhibitor IX, 6-bromoindirubin-3-oxime
Smiles C1=CC=C2C(=C1)C(=C(N2)C3=C(NC4=C3C=CC(=C4)Br)O)N=O

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Clinical Trial Information

NCT Number Recruitment interventions Conditions Sponsor/Collaborators Start Date Phases
NCT04263519 Not yet recruiting Drug: Tacrolimus Alzheimer Disease|Mild Cognitive Impairment Massachusetts General Hospital July 1 2021 Phase 2
NCT04482595 Not yet recruiting Drug: BIO 300 Oral Suspension|Drug: Placebo Pulmonary Fibrosis Humanetics Corporation|NYU Langone Health|National Institute of Allergy and Infectious Diseases (NIAID) September 2020 Phase 2

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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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Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID