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| 25mg |
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Purity: ≥98%
PHA-767491 (formerly also called CAY10572) is a novel and potent ATP-competitive dual inhibitor of Cdc7/CDK9 with IC50 of 10 nM and 34 nM in cell-free assays, respectively. It exhibits approximately 20-fold selectivity against CDK1/2 and GSK3-β, 50-fold selectivity against MK2 and CDK5, and 100-fold selectivity against PLK1 and CHK2. These results suggest that it may have anticancer potential. Unlike 5-FU or gemcitabine, which only works in a few cell lines, PHA-767491 significantly induces apoptosis in a p53-independent manner in almost all cell lines. It also inhibits cell proliferation in a variety of human cell lines, with IC50 values ranging from 0.86 μM for SF-268 to 5.87 μM for K562. An important kinase called CDC7 stimulates replication origins to facilitate DNA replication. PHA-767491 inhibits the synthesis of DNA and modifies the replicative DNA helicase's phosphorylation at CDC7-dependent phosphorylation sites. PHA-767491, in contrast to existing DNA synthesis inhibitors, inhibits replication origin activation without impeding replication fork progression or causing a prolonged DNA damage response. In preclinical cancer models, PHA-767491 treatment induces apoptotic cell death in a variety of cancer cell types and inhibits tumor growth. PHA-767491 is the first known molecule to directly influence the mechanisms governing initiation rather than elongation in DNA replication, and its actions imply that inhibiting Cdc7 kinase may be a novel approach to the development of anticancer treatments.
| Targets |
CDK9 (IC50 = 34 nM); CDK2 (IC50 = 240 nM); CDK1 (IC50 = 250 nM); CDK5 (IC50 = 460 nM); GSK3-β (IC50 = 220 nM); Mk2 (IC50 = 470 nM); Plk1 (IC50 = 980 nM); Chk2 (IC50 = 1100 nM)
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| ln Vitro |
PHA-767491 has an IC50 of 0.64 µM in HCC1954 cells and 1.3 µM in Colo-205 cells, which means that it inhibits proliferation in both cell lines. PHA-767491 exhibits IC50 values of 18.6 nM, making it an effective DDK inhibitor in vitro. In HCC1954 cells, PHA-767491 (2 µM) totally eliminates Mcm2 phosphorylation within 24 hours[1]. When combined with 5-FU, PHA-767491 exhibits much greater cytotoxicity and significantly induces apoptosis, as evidenced by noticeably increased caspase 3 activation and fragmentation of poly(ADP-ribose) polymerase in HCC cells. By directly opposing the phosphorylation of Chk1 induced by 5-FU, PHA-767491 also suppresses the expression of the anti-apoptotic protein myeloid leukemia cell 1ine[2]. In a time- and dose-dependent manner, PHA-767491 (0-10 µM) reduces the viability of glioblastoma cells, with an IC50 of roughly 2.5 µM for U87-MG and U251-MG cells. In addition to inhibiting glioblastoma cell proliferation, migration, and invasion, PHA-767491 hydrochloride causes apoptosis in these cells[3].
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| ln Vivo |
PHA-767491 promotes in situ cell apoptosis and reduces Chk1 phosphorylation in tumor tissues sectioned from naked mice HCC xenografts[2].
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| Enzyme Assay |
For five minutes, 20 ng of purified human DDK is pre-incubated with DDK inhibitors at escalating concentrations. Next, in a buffer containing 50 mM Tris-HCl (pH 7.5), 10 mM MgCl2, and 1 mM DTT, 10 µCi (γ)-32P ATP and 1.5 µM cold ATP are added, and the mixture is incubated for 30 minutes at 30°C. The proteins are autoradiographed on HyBlot CL film and SDS-PAGEd after being denatured in 1X Laemmli buffer at 100°C. DDK's auto-phosphorylation is a measure of its kinase activity. Using ImageJ, 32P-labeled bands are quantified, and GraphPad is used to compute the IC50 values.
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| Cell Assay |
There are 2500 cells plated in each well of 96-well plates used for assays. Cells undergo treatment with small molecule inhibitors after 24 hours, and they are then incubated at 37°C for 72 hours. Next, the cells undergo lysis, and the CellTiter-Glo assay is employed to quantify the ATP content, which serves as a marker of metabolically active cells. Utilizing GraphPad software, IC50 values are determined. 100,000 cells are plated per well in six-well plates used for assays. Small molecule inhibitors are applied to the cells after a day, and they are then cultured for different lengths of time. Trypsinized cells are suspended in 5 milliliters of phosphate-buffered saline. After mixing 30 µL of this suspension with 30 µL of CellTiter-Glo reagent, it is incubated at room temperature for 10 minutes. The EnVision 2104 Multilabel Reader and the BioTek Synergy Neo Microplate Reader are used to measure luminosity.
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| Animal Protocol |
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| References |
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| Molecular Formula |
C12H11N3O
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| Molecular Weight |
249.7
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| Exact Mass |
213.09
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| Elemental Analysis |
C, 67.59; H, 5.20; N, 19.71; O, 7.50
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| CAS # |
845714-00-3
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| Related CAS # |
PHA-767491 hydrochloride;942425-68-5
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| Appearance |
Solid powder
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| SMILES |
C1CNC(=O)C2=C1NC(=C2)C3=CC=NC=C3
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| InChi Key |
DKXHSOUZPMHNIZ-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C12H11N3O/c16-12-9-7-11(8-1-4-13-5-2-8)15-10(9)3-6-14-12/h1-2,4-5,7,15H,3,6H2,(H,14,16)
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| Chemical Name |
2-pyridin-4-yl-1,5,6,7-tetrahydropyrrolo[3,2-c]pyridin-4-one
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| Synonyms |
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
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| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
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| Solubility (In Vivo) |
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| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 4.0048 mL | 20.0240 mL | 40.0481 mL | |
| 5 mM | 0.8010 mL | 4.0048 mL | 8.0096 mL | |
| 10 mM | 0.4005 mL | 2.0024 mL | 4.0048 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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