>>Signaling Pathways>> Chromatin/Epigenetics>> PARP>>A-966492

A-966492

Catalog No.GC12390

A PARP1 and PARP2 inhibitor

Products are for research use only. Not for human use. We do not sell to patients.

A-966492 Chemical Structure

Cas No.: 934162-61-5

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5mg
US$76.00
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10mg
US$131.00
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50mg
US$414.00
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200mg
US$1,043.00
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Sample solution is provided at 25 µL, 10mM.

Product Documents

Quality Control & SDS

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Protocol

Kinase experiment:

The enzyme assay is conducted in buffer containing 50 mM Tris, pH 8.0, 1 mM dithiothreitol(DTT), and 4 mM MgCl2. PARP reactions contains 1.5 μM [3H]-NAD+ (1.6 μCi/mmol), 200 nM biotinylated histone H1, 200 nM slDNA, and 1 nM PARP-1 or 4 nM PARP-2 enzyme. Autoreactions utilizing SPA bead-based detection are carried out in 100 μL volumes in white 96-well plates. Reactions are initiated by adding 50 μL of 2X NAD+ substrate mixture to 50 μL of 2× enzyme mixture containing PARP and DNA. These reactions are terminated by the addition of 150 μL of 1.5 mM benzamide (appr 1×103-fold over its IC50). A 170 μL amount of the stopped reaction mixtures is transferred to streptavidin-coated Flash Plates, incubated for 1 hour, and counted using a TopCount microplate scintillation counter. Ki data are determined from inhibition curves at various substrate concentrations.

Cell experiment:

C41 cells are treated with A-966492 for 30 minutes in a 96-well plate. PARP are activated by damaging DNA with 1 mM H2O2 for 10 minutes. Cells are washed with ice-cold phosphate-buffered saline (PBS) once and fixed with prechilled methanol/acetone (7:3) at -20°C for 10 minutes. After they are air-dried, plates are rehydrated with PBS and blocked using 5% nonfat dry milk in PBS-Tween (0.05%) (blocking solution) for 30 minutes at room temperature. Cells are incubated with anti-PAR antibody 10H (1:50) in blocking solution at room temperature for 60 minutes followed by washing with PBS-Tween20 five times, and incubation with goat antimouse fluorescein 5(6)-isothiocyanate (FITC)-coupled antibody (1:50) and 1 μg/mL 40,6-diamidino-2-phenylindole (DAPI) in blocking solution at room temperature for 60 minutes. After washing with PBS-Tween20 5 times, analysis is performed using an fmax Fluorescence Microplate Reader set at the excitation and emission wavelength for FITC or the excitation and emission wavelength for DAPI. PARP activity (FITC signal) is normalized with cell numbers (DAPI).

Animal experiment:

A 0.2 cc amount of a 1:10 dilution of tumor brei in 45% Matrigel and 45% Spinner MEM is injected subcutaneously into the flank of female SCID mice on study day 0. Tumors are allowed to grow to the indicated size and then randomized to therapy groups (N=10 mice/group). PARP inhibitor therapy begin on day 14, with cisplatin treatment starting on day 16. At various intervals following tumor inoculation, the individual tumor dimensions are serially measured using calibrated microcalipers, and the tumor volumes are calculated.

References:

[1]. Penning TD, et al. Optimization of phenyl-substituted benzimidazole carboxamide poly(ADP-ribose) polymerase inhibitors: identification of (S)-2-(2-fluoro-4-(pyrrolidin-2-yl)phenyl)-1H-benzimidazole-4-carboxamide (A-966492), a highly potent and efficacious

Background

A-966492 is an inhibitor of PARP-1 with Ki value of 1nM [1].

PARP-1 belongs to the poly(ADP-ribose) polymerases (PARPs) family, it contributes to the resistance happened after cancer therapy. A-966492 is a potent inhibitor of both PARP-1 and PARP-2 (Ki value of 1.5nM) with good potency in C41 whole cells (EC50 value of 1nM). A-966492 shows excellent pharmaceutical properties with oral bioavailabilities of 34-72% and half-lives of 1.7-1.9 h. Additionally, A-966492 can crosses the blood-brain barrier. A-966492 is proved potent in a murine B16F10 syngeneic melanoma model and a BRCA1-deficient MX-1 breast carcinoma model. Meanwhile, it can enhance the efficacy of TMZ and carboplatin in these models [1].

References:
[1] Penning TD, Zhu GD, Gong J, Thomas S, Gandhi VB, Liu X, Shi Y, Klinghofer V, Johnson EF, Park CH, Fry EH, Donawho CK, Frost DJ, Buchanan FG, Bukofzer GT, Rodriguez LE, Bontcheva-Diaz V, Bouska JJ, Osterling DJ, Olson AM, Marsh KC, Luo Y, Giranda VL. Optimization of phenyl-substituted benzimidazole carboxamide poly(ADP-ribose) polymerase inhibitors: identification of (S)-2-(2-fluoro-4-(pyrrolidin-2-yl)phenyl)-1H-benzimidazole-4-carboxamide (A-966492), a highly potent and efficacious inhibitor. J Med Chem. 2010 Apr 22;53(8):3142-53.

Chemical Properties

Cas No. 934162-61-5 SDF
Chemical Name 2-[2-fluoro-4-[(2S)-pyrrolidin-2-yl]phenyl]-1H-benzimidazole-4-carboxamide
Canonical SMILES C1CC(NC1)C2=CC(=C(C=C2)C3=NC4=C(C=CC=C4N3)C(=O)N)F
Formula C18H17FN4O M.Wt 324.35
Solubility ≥ 32.4 mg/mL in DMSO, ≥ 3.16 mg/mL in EtOH with ultrasonic and warming, ≥ 16.53 mg/mL in Water with ultrasonic Storage Store at -20°C
General tips Please select the appropriate solvent to prepare the stock solution according to the solubility of the product in different solvents; once the solution is prepared, please store it in separate packages to avoid product failure caused by repeated freezing and thawing.Storage method and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored at -20°C, please use it within 1 month.
To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time.
Shipping Condition Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request.

Complete Stock Solution Preparation Table

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1 mg 5 mg 10 mg
1 mM 3.0831 mL 15.4154 mL 30.8309 mL
5 mM 0.6166 mL 3.0831 mL 6.1662 mL
10 mM 0.3083 mL 1.5415 mL 3.0831 mL
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