PIK-75 化学構造
分子量: 488.74

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Quality Control & MSDS

製品説明

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製品の説明

生物活性

製品説明 PIK-75はPI3Kアイソフォーム(5.8nMのIC50によるp110α)の選択的で競争的阻害剤で、そのうえ2nMのIC50で、強力にDNAPKを妨げます。
ターゲット p110α DNA-PK
IC50 5.8 nM [1] 2 nM [2]
In vitro試験 PIK-75 shows the impressive potency and isoform selectivity at p110α while the corresponding IC50 values are 1300 nM, 76 nM and 510 nM for other PI3K isoforms, p110β, -γ, and -δ, respectively. Furthermore, when binding to purified p110α, PIK-75 is a noncompetitive inhibitor with respect to ATP with Ki of 36 nM and competitive with respect to the substrate PI with Ki of 2.3 nM. [1] PIK-75 also shows potent inhibition of DNA-PK. [2] PIK-75 (1 μM) reduces cell survival by significantly decreasing mitochondrial activity in unstimulated nonasthmatic airway smooth muscle (ASM) cells, asthmatic ASM cells, and lung fibroblasts. While in TGFβ-stimulated ASM cells, PIK75 only decreases mitochondrial activity in asthmatic cells without effects in nonasthmatic cells. [3] A recent study shows that PIK-75 (10 nM) inhibits TNF-α-induced CD38 mRNA expression and significantly attenuates of TNF-α-induced ADP-ribosyl cyclase activity in human airway smooth muscle cells. [4]
In vivo試験 In the ErbB3WT tumor model, PIK-75 reduces in vitro chemotactic response to HRGβ1 and lowers pAkt levels by 40%. Besides, PIK-75 significantly reduces tumor cell motility and in vivo invasion in ErbB3WT primary tumors. [5] In the CD1 male mice, PIK-75 leads to serious impairments in the insulin tolerance test (ITT) and glucose tolerance test (GTT), and an increase in glucose production during a pyruvate tolerance test (PTT). [6]
臨床試験
特集 PI3K and DNA-PK inhibitor.

プロトコル (参考用のみ)

キナーゼアッセイ: [1]

Inhibition Assays The PI3K inhibitor PIK-75 is dissolved at 10 mM in dimethyl sulfoxide and stored at −20°C until use. PI3K enzyme activity is determined in 50 μL of 20 mM HEPES, pH 7.5, and 5 mM MgCl2 containing 180 μM phosphatidyl inositol, with the reaction started by the addition of 100 μM ATP (containing 2.5 μCi of [γ-32P]ATP). After a 30-minute incubation at room temperature, the enzyme reaction is stopped by the addition of 50 μL of 1 M HCl. Phospholipids are then extracted with 100 μL of chloroform/methanol [1:1 (v/v)] and 250 μL of 2 M KCl followed by liquid scintillation counting. Inhibitors are diluted in 20% (v/v) dimethyl sulfoxide to generate a concentration versus inhibition of enzyme activity curve, which is then analyzed with the use of Prism version 5.00 for Windows to calculate the IC50. For kinetic analysis, a luminescent assay measuring ATP consumption is used. PI3K enzyme activity is determined in 50 μL of 20 mM HEPES, pH 7.5, and 5 mM MgCl2 with PI and ATP at various concentrations. After a 60-minute incubation at room temperature, the reaction is stopped by the addition of 50 μL of Kinase-Glo followed by a further 15-minute incubation. Luminescence is then read using a Fluostar plate reader. Results are analyzed using Prism.

細胞アッセイ: [3]

細胞株 A2780, A2780/cp70, 2780AD, HCT116, HT29, WIL, CALU-3, MCF7, PC3 and HS852 cells
濃度 0-10 μM
反応時間 48 hours
実験の流れ Mitochondrial activity is assessed after stimulation with TGFβ with or without inhibitors for 48 hours using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay. Harvested washed cells are resuspended in DMEM-lO% FCS and aliquoted (500 μL) into 24-well cluster plates prior to serial dilution (1:2) in duplicates. To each well, 100 μL of an appropriate MTT concentration (dissolved in PBS and filtered through a 0.2 μm filter before use to remove any blue formazan product) is added immediately after diluting the cells, which are then incubated for 3.5 hours at 37 °C. The resulting blue formazan product is solubilized overnight (16 hours) at 37 °C by the addition of 500 μL of 10% sodium dodecyl sulfate (SDS) in 0.01 M HCl to each well. A sample (150 μL) from each duplicate well is transferred to a 96-well microplate, and the optical density determinedby automated spectrophotometry against a reagent blank (no cells). Absorbance is measured at a test wavelength of 570 nm and a reference wavelength of 690 nm. For each primary cell culture, results from three to six wells from each treatment are averaged, and data are expressed as absorbance 570 to 690 nm.

動物実験: [5]

動物モデル MTLn3 cells are injected into the right fourth mammary fat pad from the head of female severe-combined immunodeficient/NCr mice.
製剤 PIK-75 is dissolved in DMSO and then diluted in PBS.
投薬量 ≤1 μM
投与方法 Administered via i.p.

Conversion of different model animals based on BSA (Value based on data from FDA Draft Guidelines)

SpeciesMouseRatRabbitGuinea pigHamsterDogMonkeyBaboon
Weight (kg)0.020.151.80.40.0810312
Body Surface Area (m2)0.0070.0250.150.050.020.50.240.6
Km factor361285201220
Animal A (mg/kg) = Animal B (mg/kg) multiplied by  Animal B Km
Animal A Km

For example, to modify the dose of resveratrol used for a mouse (22.4 mg/kg) to a dose based on the BSA for a rat, multiply 22.4 mg/kg by the Km factor for a mouse and then divide by the Km factor for a rat. This calculation results in a rat equivalent dose for resveratrol of 11.2 mg/kg.

Rat dose (mg/kg) = mouse dose (22.4 mg/kg) ×  mouse Km(3)  = 11.2 mg/kg
rat Km(6)
1

参考

化学情報

Download PIK-75 SDF
分子量 488.74
化学式

C16H14BrN5O4S.HCl

CAS No. 372196-77-5
保管 2年-20℃
6月-80℃in solvent
別名 N/A
溶解度 (25°C) * In vitro DMSO 3 mg/mL (6.13 mM)
<1 mg/mL (<1 mM)
エタノール <1 mg/mL (<1 mM)
* <1 mg/ml means slightly soluble or insoluble.
* 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.
化学名 (E)-N'-((6-bromoH-imidazo[1,2-a]pyridin-3-yl)methylene)-N,2-dimethyl-5-nitrobenzenesulfonohydrazide hydrochloride

カスタマーフィードバック (5)


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Rating
Source Cell Death Differ, 2014, 21(3), 491-502. PIK-75 purchased from Selleck
Method Crystal violet staining
Cell Lines A549 cells
Concentrations 200 nM
Incubation Time 1 h
Results When treated with PIK-75 A549 cells became sensitive to apoptosis induction by TRAIL, even at concentrations of TRAIL as low as 10 ng/ml. Intriguingly, when examining clonogenic survival, it observed that this novel combination almost completely obliterated clonogenic survival of A549 cells.

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Rating
Source Cell Death Differ, 2014, 21(3), 491-502. PIK-75 purchased from Selleck
Method Western blot
Cell Lines A549 cells
Concentrations 100 nM
Incubation Time 0-24 h
Results It next investigated whether known components of the TRAIL–DISC and the downstream apoptosis pathway it activates are regulated by PIK-75 or SNS-032 treatment. Whereas the majority of the DISC components and downstream pro- and anti-apoptotic proteins remained unchanged, cFlip and Mcl-1 protein levels were rapidly suppressed by pharmacological CDK9 inhibition by SNS-032 or PIK-75.

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Rating
Source Mol Carcinog, 2012, 52, 667-75 . PIK-75 purchased from Selleck
Method MTT assay
Cell Lines MCF7 cells
Concentrations 0.01-1 μM
Incubation Time 48 h
Results Knockdown of BRCA1 can sensitize the MCF7 cells to Perifosine in a dose-dependent manner (Figure A). BRCA1-KD also sensitizes the MCF7 cells to dual PI3K/mTOR inhibitors, such as PI-103 or BEZ235 (Figure B, D). Another inhibitor, PIK-75 which specifically inhibits PI3Ka and PI3Kg, but not mTOR, also showed similar effects on proliferation of BRCA1-KD MCF7 cells (Figure C).

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Rating
Source J Pharmacol Exp Ther, 2014, 348(3), 432-41. PIK-75 purchased from Selleck
Method Western blot
Cell Lines RH30 cells
Concentrations 0.1 uM
Incubation Time 0-24 h
Results PIK-75 robustly inhibited PI3K signaling, indicated by decreased phosphorylation of AKT at Thr308 and Ser473. However, inhibition of AKT phosphorylation by PIK-75 was ready to recover, and the level of phosphorylated Akt restored within 1.5 hour. However, inhibition of phosphorylated p70S6K1 by PIK-75 recovered quickly, which might be attributed to the following reasons: first, inhibition of PI3Kα might be compensated by other isoforms of PI3K, which results in phosphorylation of Akt and S6K1; second, mTOR may also be activated by other upstream regulators upon PI3K inhibition, which lead to S6K1 phosphorylation.

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Rating
Source Dr. Yong-Weon Yi from Georgetown University Medical Center. PIK-75 purchased from Selleck
Method MTT assays
Cell Lines MDA-MB-231/MDA-MB-436 cells
Concentrations 0-10 μM
Incubation Time 72 h
Results PIK-75 potently inhibited the survival of MDA-MB-231 and MDA-MB-436 cells in a dose-dependent manner.

文献中の引用 (11)

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