ON044580
Based on 1 Customer Validation
ON044580 is a non–ATP-competitive JAK2 and BCR-ABL kinases inhibitor. ON044580 inhibits wild-type (WT) and V617F mutant JAK2 with IC50 values of 1.23 μM, 1.09 μM, respectively. ON044580 also inhibits both WT and T315I mutant BCR-ABL kinases., ON044580 blocks the IL-3–mediated phosphorylation of JAK2 and STAT5, induces apoptosis in Imatinib (HY-15463)-resistant chronic myeloid leukemia (CML). ON044580 can be used for the study of CML, myelodysplasia (MDS) and other myeloproliferative neoplasms.
For research use only. We do not sell to patients.
- Purity : 99.11%
- CAS No.: 1035199-04-2
- Formula: C23H15BrFNO5S
- Molecular Weight:516.34
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Storage:
4°C, stored under nitrogen
* In solvent : -80°C, 6 months; -20°C, 1 month (stored under nitrogen)
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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Biological Activity
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Chemical Information
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Solvent & Solubility
- Protocols
- Purity & Documentation
- References
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Help & FAQs
Biological Activity
Description
IC50 & Target
[1]|
JAK2-WT 1.23 μM (IC50) |
JAK2-V617F 1.09 μM (IC50) |
WT BCR-ABL |
T315I-BCR-ABL |
Cellular Effect
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Cell Line
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Type | Value | Description | References |
|---|---|---|---|---|
| K562 | GI50 |
< 1 μM
Compound: 6c
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Growth inhibition of human K562 cells expressing p210Bcr/Abl after 96 hrs by trypan blue exclusion assay.
Growth inhibition of human K562 cells expressing p210Bcr/Abl after 96 hrs by trypan blue exclusion assay.
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20188579 |
| DU-145 | GI50 |
1 μM
Compound: 6c
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Growth inhibition of human DU145 cells after 96 hrs by trypan blue exclusion assay.
Growth inhibition of human DU145 cells after 96 hrs by trypan blue exclusion assay.
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20188579 |
| K562 | IC50 |
0.5 μM
Compound: 6c
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Inhibition of BCR/ABL p210 autophosphorylation in human K562 cells after 2 hrs by Western blot analysis.
Inhibition of BCR/ABL p210 autophosphorylation in human K562 cells after 2 hrs by Western blot analysis.
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20188579 |
| HEL | GI50 |
0.9 μM
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Inhibits the proliferation of HEL cells.
Inhibits the proliferation of HEL cells.
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20717479 |
| SET-2 | GI50 |
3.0 μM
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Inhibits the proliferation of SET-2 cells.
Inhibits the proliferation of SET-2 cells.
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20717479 |
| K562 | IC50 |
7.94 μM
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Inhibition of BCR-ABL kinase activity.
Inhibition of BCR-ABL kinase activity.
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20717479 |
In Vitro
ON044580 (0.25-10 μM; 30 min) inhibits recombinant wild-type JAK2 and JAK2-V617F kinase activities with IC50 values of 1.23 μM and 1.09 μM, respectively[1].
ON044580 (0-10 μM; 30 min) inhibits full-length JAK2 immunoprecipitated from Ba/F3:JAK2V617F cells with an IC50 of approximately 4 μM[1].
ON044580 (0-20 μM; 30 min) inhibits wild-type BCR-ABL and T315I mutant BCR-ABL kinase activities immunoprecipitated from K562 and 32D:p210T315I cells, with IC50 values of 7.94 μM and 2.08 μM, respectively[1].
ON044580 (0-20 μM; 2 h) inhibits JAK2 autophosphorylation and STAT5 phosphorylation in IL-3-stimulated Ba/F3:JAK2V617F cells in a concentration-dependent manner[1].
ON044580 (10 μM; 15-60 min) inhibits IL-3-mediated JAK2 phosphorylation within 15-30 minutes in Ba/F3:JAK2V617F cells[1].
ON044580 (0-10 μM; 2 h) inhibits STAT3 phosphorylation in U266 multiple myeloma cells (constitutive IL-6 receptor/JAK2/STAT3 pathway) in a dose-dependent manner[1].
ON044580 (0.5-10 μM; 72 h) inhibits the proliferation of JAK2V617F-expressing cells (Ba/F3:JAK2V617F, HEL, SET-2) with GI50 values of 0.25 μM, 0.9 μM, and 3.0 μM, respectively[1].
ON044580 (0.5 μM; 2-24 h) induces PARP cleavage in Ba/F3:JAK2V617F cells at 24 h[1].
ON044580 (0.1-0.5 μM; ex vivo, 48 h) selectively reduces aneuploid (monosomy 7) cells while maintaining/increasing diploid cells in bone marrow mononuclear cells[1].
ON044580 (1-10 μM; 2 h short-term exposure followed by 96 h recovery) inhibits growth of Ba/F3:JAK2V617F, HEL, and K562 cells in a concentration-dependent manner, while normal mouse bone marrow cells remain unaffected[1].
ON044580 (0.5-10 μM; ex vivo, 48 h) induces apoptosis in primary CML cells from blast crisis patients refractory to Imatinib (HY-15463)[1][2].
ON044580 (0.25-10 μM; 30 min) inhibits recombinant Abl kinase activity by 50% at 5 μM and 75% at 10 μM[2].
ON044580 (0.1-10 μM; 16 h) reduces BCR-ABL protein levels and pTyr-JAK2 in Imatinib-sensitive and Imatinib-resistant (T315I, E255K) BCR-ABL+ cells in a dose-dependent manner[2].
ON044580 (10 μM; 6-16 h) reduces levels of BCR-ABL downstream signaling molecules including STAT3, pTyr705-STAT3, pSer727-STAT3, and Akt in BCR-ABL+ 32D cells[2].
ON044580 (0-10 μM; 16 h) reduces HSP90α transcript levels in BCR-ABL+ 32D cells[2].
ON044580 (0.25-10 μM; 16 h) reduces HSP90 protein levels in Imatinib-sensitive and Imatinib-resistant (T315I, E255K) BCR-ABL+ cells[2].
ON044580 (10 μM; 3 h) disrupts the BCR-ABL/JAK2/HSP90 high molecular weight network complex in BCR-ABL+ 32D cells[2].
ON044580 (0-10 μM; 48 h) induces apoptosis in Imatinib-sensitive and Imatinib-resistant (T315I, K562-R) BCR-ABL+ cells at concentrations of 1-5 μM[2].
ON044580 (0.1-0.5 μM; 2 weeks) inhibits colony formation of Imatinib-sensitive and Imatinib-resistant (T315I, E255K) BCR-ABL+ cells in soft agar[2].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only. Further protocols information, click here.
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Cell Line:Ba/F3:JAK2V617F cells (+IL-3)
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Concentration:0.1, 0.5, 1, 5, 10 μM
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Incubation Time:2 h
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Result:Inhibited JAK2 autophosphorylation and STAT5 phosphorylation in a concentration-dependent manner.
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Cell Line:Ba/F3:JAK2V617F cells (+IL-3)
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Concentration:10 μM
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Incubation Time:15-60 min
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Result:Inhibited IL-3-mediated JAK2 phosphorylation within 15-30 minutes.
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Cell Line:Ba/F3:JAK2V617F, HEL, SET-2 cells
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Concentration:0.5, 1, 2.5, 5, 10 μM
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Incubation Time:72 h
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Result:Inhibited the proliferation of JAK2V617F-expressing cells (Ba/F3:JAK2V617F, HEL, SET-2) with GI50 values of 0.25 μM, 0.9 μM, and 3.0 μM, respectively.
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Cell Line:Ba/F3:JAK2V617F
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Concentration:0.5 μM
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Incubation Time:2-24 h
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Result:Induced PARP cleavage.
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Cell Line:BCR-ABL+ 32D cells
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Concentration:10 μM
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Incubation Time:6-16 h
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Result:Reduces levels of BCR-ABL downstream signaling molecules including STAT3, pTyr705-STAT3, pSer727-STAT3, and Akt.
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Cell Line:BCR-ABL+ 32D cells
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Concentration:1, 5, 10 μM
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Incubation Time:16 h
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Result:Reduced HSP90 protein levels.
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Cell Line:BCR-ABL+ 32D cells
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Concentration:1, 5, 10 μM
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Incubation Time:16 h
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Result:Reduced HSP90α transcript levels.
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Cell Line:BCR-ABL+ cells (WT, T315I, K562-R)
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Concentration:2.5, 5, 10 μM
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Incubation Time:48 h
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Result:Induced apoptosis in Imatinib-sensitive and Imatinib-resistant (T315I, K562-R) BCR-ABL+ cells.
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Cell Line:BCR-ABL+ cells (WT, T315I, E255K)
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Concentration:0.01, 0.05, 0.1, 0.5, 1 μM
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Incubation Time:2 weeks
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Result:Inhibited colony formation.
Chemical Information
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CAS No. 1035199-04-2
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Appearance Solid
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Molecular Weight 516.34
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Formula C23H15BrFNO5S
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Color Off-white to light yellow
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SMILES
O=C(C1=CC=C(C(O)=O)C=C1)/C(SCC2=CC=C(Br)C=C2)=C\C3=CC=C(F)C([N+]([O-])=O)=C3
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
4°C, stored under nitrogen
* In solvent : -80°C, 6 months; -20°C, 1 month (stored under nitrogen)
Solvent & Solubility
In Vitro:
DMSO : 50 mg/mL (96.84 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (stored under nitrogen). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (stored under nitrogen). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
In Vivo:
Select the appropriate dissolution method based on your experimental animal and administration route.
- For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
- To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
- The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.
Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 2.5 mg/mL (4.84 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:
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Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
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%DMSO +
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
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%+
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+%Tween-80 + +
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%Saline +
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Working solution concentration: 0.22 mg/mL
Method for preparing stock solution: mg drug dissolved in μL DMSO. Stock solution concentration: mg/mL. * In solvent : -80°C, 6 months; -20°C, 1 month (stored under nitrogen)
1. Take μL DMSO stock solution;
2. Add μL .
μL , mix evenly;
3. Then add μL Tween 80, mix evenly;
4. Then add μL
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
Protocols
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Kinase activity and phosphorylation assays
Kinase activity assays measure the ability of kinases to transfer phosphate groups from ATP to specific substrates, while phosphorylation assays detect the presence and levels of phosphorylated proteins. Common methods include radiolabeled ATP incorporation (e. g. ,), ADP release detection via bioluminescence (e. g. ,[3]), enzyme-linked immunosorbent assays (ELISA) for phospho-specific epitopes (e. g. ,[6]), and microtiter-based formats for high-throughput screening (e. g. ,[8]). The ADP-Glo assay quantifies kinase activity by measuring ADP produced during phosphorylation using a luciferase-based system. Radiometric assays involve autoradiography or scintillation counting after incorporation of 32P-labeled ATP into substrate proteins. ELISA-based approaches rely on phospho-specific antibodies to detect activated kinases in cell lysates or purified samples.
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Western Blot
Western blotting (WB) is a commonly used experimental method in molecular biology, biochemistry, and immunogenetics for identifying and quantifying target proteins. It combines gel electrophoresis with immunoassay, enabling researchers to analyze protein expression, post-translational modifications, and molecular weight.
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Apoptosis
Apoptosis, also called programmed cell death, is generally characterized by distinct morphological characteristics.
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TUNEL staining for apoptotic DNA fragmentation
TUNEL staining detects DNA strand breaks by using terminal deoxynucleotidyl transferase to add labeled nucleotides to exposed 3′-OH DNA termini, generating either microscopic staining in fixed cells or tissue sections, or fluorescence/cytometric signal in cell suspensions. TUNEL positivity reflects DNA fragmentation but should not be interpreted alone as definitive apoptosis, because TUNEL can also label necrotic, autolytic, mechanically damaged, or DNA-repair-associated DNA breaks.
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Cell Cytotoxicity Assay
Cytotoxicity assays are usually based on the assessment of cell membrane damage, which can also be indirectly detected by measuring cell viability. Detection methods include MTT assay, CKK-8 assay, LDH assay and ATP assay, etc.
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Annexin V plus membrane-impermeant dye apoptosis staining
Annexin V-based apoptosis assays rely on the detection of phosphatidylserine (PS) externalization from the inner leaflet of the plasma membrane to the outer leaflet, an early biochemical hallmark of apoptosis. Fluorescently labeled Annexin V binds PS in a calcium-dependent manner, enabling identification of early apoptotic cells by flow cytometry or fluorescence microscopy. When combined with a membrane-impermeant DNA-binding dye (e. g. , propidium iodide), this approach allows discrimination between viable (Annexin V−/dye−), early apoptotic (Annexin V+/dye−), and late apoptotic or necrotic (Annexin V+/dye+) cell populations by assessing membrane integrity and PS exposure.
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Apoptosis Solutions
Apoptosis is a regulated, generally non-lytic cell-death pathway that removes unwanted, damaged, infected, or abnormal cells through coordinated morphological changes, caspase activation, DNA fragmentation, and membrane remodeling. The intrinsic apoptosis pathway is controlled mainly by mitochondrial outer membrane permeabilization, BCL-2 family proteins, cytochrome c release, apoptosome formation, caspase-9 activation, and downstream executioner caspase-3/7 activation. The extrinsic apoptosis pathway is initiated by death receptors such as Fas, TNFR, and TRAIL receptors, which recruit adaptor proteins and activate caspase-8 before engaging executioner caspases or mitochondrial amplification through BID cleavage. Apoptosis is linked to many phenotypes, including cancer cell killing, tissue homeostasis, immune regulation, neurodegeneration, infection response, and treatment-induced cytotoxicity; unresolved questions include how apoptosis interacts with necroptosis, pyroptosis, ferroptos
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Protocol for Kinase activity and phosphorylation assays
Kinase activity assays measure transfer of phosphate from ATP to a protein or peptide substrate, generating phosphorylated substrate, ADP, or incorporated radiolabeled phosphate as the readout; phosphorylation assays measure site-specific phosphorylation in cells or tissues as a proxy for kinase-pathway activation, inhibition, or substrate regulation. Phosphorylation can be detected by phospho-specific Western blot, immunoprecipitation kinase assay, phospho-immunofluorescence, phospho-flow cytometry, luminescent ADP detection, radiolabeled ATP incorporation, or reporter-based pathway assays, and these readouts can be applied to cancer cells, primary neurons, mouse tumors, organoids, inflammatory macrophages, ferroptosis studies, and mitophagy studies when the kinase target is biologically relevant.
Purity & Documentation
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Data Sheet (285 KB)
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SDS (251 KB)
- English - EN (251 KB)
- Français - FR (251 KB)
- Deutsch - DE (251 KB)
- Norwegian - NO (251 KB)
- Español - ES (251 KB)
- Swedish - SV (251 KB)
- Italian - IT (251 KB)
- Korean - KR (251 KB)
- Portuguese - PT (251 KB)
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Handling Instructions (2659 KB)
References
[1]. Jatiani SS, et al. A Non-ATP-Competitive Dual Inhibitor of JAK2 and BCR-ABL Kinases: Elucidation of a Novel Therapeutic Spectrum Based on Substrate Competitive Inhibition. Genes Cancer. 2010 Apr;1(4):331-45. [Content Brief]
[2]. Samanta AK, et al. Destabilization of Bcr-Abl/Jak2 Network by a Jak2/Abl Kinase Inhibitor ON044580 Overcomes Drug Resistance in Blast Crisis Chronic Myelogenous Leukemia (CML). Genes Cancer. 2010 Apr;1(4):346-59. [Content Brief]
Complete Stock Solution Preparation Table
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (stored under nitrogen). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.
| Optional Solvent | Concentration Solvent Mass | 1 mg | 5 mg | 10 mg | 25 mg |
|---|---|---|---|---|---|
| DMSO | 1 mM | 1.9367 mL | 9.6835 mL | 19.3671 mL | 48.4177 mL |
| 5 mM | 0.3873 mL | 1.9367 mL | 3.8734 mL | 9.6835 mL | |
| 10 mM | 0.1937 mL | 0.9684 mL | 1.9367 mL | 4.8418 mL | |
| 15 mM | 0.1291 mL | 0.6456 mL | 1.2911 mL | 3.2278 mL | |
| 20 mM | 0.0968 mL | 0.4842 mL | 0.9684 mL | 2.4209 mL | |
| 25 mM | 0.0775 mL | 0.3873 mL | 0.7747 mL | 1.9367 mL | |
| 30 mM | 0.0646 mL | 0.3228 mL | 0.6456 mL | 1.6139 mL | |
| 40 mM | 0.0484 mL | 0.2421 mL | 0.4842 mL | 1.2104 mL | |
| 50 mM | 0.0387 mL | 0.1937 mL | 0.3873 mL | 0.9684 mL | |
| 60 mM | 0.0323 mL | 0.1614 mL | 0.3228 mL | 0.8070 mL | |
| 80 mM | 0.0242 mL | 0.1210 mL | 0.2421 mL | 0.6052 mL |
ON044580 Related Classifications
HY-164393 Related Classifications
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