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A 419259 Sale

(Synonyms: RK-20449) 目录号 : GC35209

An inhibitor of Src family kinases

A 419259 Chemical Structure

Cas No.:364042-47-7

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5mg
¥1,170.00
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10mg
¥1,633.00
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50mg
¥5,670.00
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Sample solution is provided at 25 µL, 10mM.

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实验参考方法

Kinase experiment:

In vitro kinase assays are performed on His(6)-tagged Lck (residues 62-509) and full-length c-Abl purified from Sf-9 cells, and commercial sources of Lyn, Src and PKC. Lck, Lyn, Src and Abl activities are measured with an ELISA-based assay. Flat bottom 96-well ELISA plates are incubated with a 200 μg/mL solution of Poly(Glu,Tyr) 4 : 1 substrate in phosphate buffered saline (PBS) for 1 h at 37°C and then washed with PBS containing 0.1% Tween-20 (PBS-T). Inhibitor dilutions are added to the washed plates already containing the appropriate enzyme in kinase assay buffer (250 mM Mopso, pH 6.75, 10 mM MgCl2, 2 mM MnCl2, 2.5 mM DTT, 0.02% BSA, 2 mM Na3VO4, 5% DMSO, 5 μM ATP). After incubation at room temperature for 20 min, plates are washed three times with PBS-T and plate-bound phosphotyrosine is detected with an anti-phosphotyrosine-HRP antibody conjugate and subsequent color development with K-Blue reagents. All assays are optimized to use the least amount of enzyme necessary for a reproducible signal-to-noise ratio[1].

Cell experiment:

K-562 cells are grown in RPMI 1640 supplemented with 10% fetal calf serum (FCS), and 50 g/mL Gentamycin. Meg-01 cells are cultured in Vitacell modified RPMI 1640 (ATCC), supplemented with 10% FCS and 50 μg/mL Gentamycin. The human GM-CSF-dependent myeloid leukemia cell line TF-1 is grown in RPMI 1640 supplemented with 10% FCS, 50 μg/mL Gentamycin, and 1 ng/mL of recombinant human GM-CSF. DAGM murine myeloid leukemia cells are cultured in RPMI 1640 supplemented with 10% FCS, 50 μg/mL Gentamycin, and 0.5 ng/mL recombinant IL-3. Concentrated stock solutions of PP2 (5 mM) and A-419259 (2 mM) are prepared in DMSO and stored at -20°C. Cellular proliferation is measured using the Live/Dead growth assay. This assay employs calcein-AM, a fluorogenic esterase substrate that is taken up by viable cells and hydrolyzed intracellularly, releasing a green fluorescent product. Briefly, 104 cells are plated per well in 96-well plates for each day of a 4-day time course. Various concentrations of PP2, A-419259 or vehicle control are added to the wells (five wells per concentration per day) and the plates are incubated at 37°C. At each time point, one plate is centrifuged at 1500 g for 10 min to pellet the cells. Cells are washed with phosphate buffered saline (PBS), and calcein-AM is added to each well to a final concentration of 1 μM. Plates are incubated in the dark at room temperature for 1 h. The plates are then read at 485/530 nm (excitation/emission) using a SpectraMax Gemini XS fluorescent plate reader and data are analysed with SoftMax Pro software[1].

References:

[1]. Wilson MB, et al. Selective pyrrolo-pyrimidine inhibitors reveal a necessary role for Src family kinases in Bcr-Abl signal transduction and oncogenesis. Oncogene. 2002 Nov 21;21(53):8075-88.
[2]. Pene-Dumitrescu T, et al. An inhibitor-resistant mutant of Hck protects CML cells against the antiproliferative and apoptotic effects of the broad-spectrum Src family kinase inhibitor A-419259.Oncogene (2008) 27, 7055-7069

产品描述

A-419259 is an inhibitor of Src family kinases, including Src, LCK, Lyn, and Hck (IC50s = 9, <3, <3, and 11.26 nM, respectively).1,2 It is selective for these kinases over c-Abl (IC50 = 3,000 nM) and PKC (IC50 = >33 μM).1 A-419259 inhibits growth of Philadelphia chromosome-positive (Ph+) K-562 and Meg-01 myeloid leukemia cells (IC50s = 0.1-0.3 and 0.1 μM, respectively), but not Ph- TF-1 and HEL cells. It induces apoptosis in K-562 cells in a concentration-dependent manner. A-419259 (300 nM) inhibits differentiation of murine embryonic stem cells while maintaining pluripotency.3 It reduces the total number of acute myeloid leukemia (AML) cells, as well as AML stem cells, in the bone marrow and spleen in mouse patient-derived xenograft (PDX) models of AML when administered at a dose of 30 mg/kg twice daily.4

1.Wilson, M.B., Schreiner, S.J., Choi, H.J., et al.Selective pyrrolo-pyrimidine inhibitors reveal a necessary role for Src family kinases in Bcr-Abl signal transduction and oncogenesisOncogene21(53)8075-8088(2002) 2.Pene-Dumitrescu, T., Peterson, L.F., Donato, N.J., et al.An inhibitor-resistant mutant of Hck protects CML cells against the antiproliferative and apoptotic effects of the broad-spectrum Src family kinase inhibitor A-419259Oncogene27(56)7055-7069(2008) 3.Meyn, M.A., III, Schreiner, S.J., Dumitresco, T.P., et al.SRC family kinase activity is required for murine embryonic stem cell growth and differentiationMol. Pharmacol.68(5)1320-1330(2005) 4.Saito, Y., Yuki, H., Kuratani, M., et al.A pyrrolo-pyrimidine derivative targets human primary AML stem cells in vivoSci. Transl. Med.5(181):181ra52(2013)

Chemical Properties

Cas No. 364042-47-7 SDF
别名 RK-20449
Canonical SMILES NC1=C2C(N([C@H]3CC[C@H](N4CCN(C)CC4)CC3)C=C2C5=CC=C(OC6=CC=CC=C6)C=C5)=NC=N1
分子式 C29H34N6O 分子量 482.62
溶解度 55 mg/mL in water 储存条件 Store at -20°C
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1 mM 2.072 mL 10.3601 mL 20.7202 mL
5 mM 0.4144 mL 2.072 mL 4.144 mL
10 mM 0.2072 mL 1.036 mL 2.072 mL
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