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Research Area
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Description
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Cancer |
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Biological Activity
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Description
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AG-490 (Tyrphostin B42) is an inhibitor of EGFR, ErbB2 and JAK2 with IC50 of 0.1 μM, 13.5 μM, and ~10 μM, respectively. |
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Targets
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EGFR |
ErbB2 |
JAK2 |
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IC50 |
0.1 μM |
13.5 μM [1] |
~10 μM [2] |
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In Vitro
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AG-490 inhibits HER-2 driven cell proliferation with IC50 of 3.5 μM. [1] Corresponding to the specific dose-dependent inhibition of constitutively activated JAK2 in pre-B acute leukemia (ALL) cells, AG-490 (5 μM) almost completely blocks the growth of all ALL cells by inducing programmed cell death, with no deleterious effect on normal hematopoiesis. AG-490 does not inhibit the activities of Lck, Lyn, Btk, Syk, and Src. [2] AG-490 (60-100 μM) blocks the constitutive activation of Stat3sm, and inhibits spontaneous as well as interleukin 2-induced growth of mycosis fungoides (MF) tumor cells with IC50 values of 75 μM and 20 μM, respectively. [3] AG-490 potently inhibits IL-2-mediated human T cell growth with an IC50 of 25 μM by blocking the activities of JAK3 and STAT5a/b. [4] Although AG-490 alone has no effect on proliferation of FDrv210H cells at a concentration of 5 μM, AG-490 can synergize with STI571 to enhance its inhibitory effect on p210bcr-abl driven proliferation. [5] AG-490 significantly inhibits the constitutive activation of Stat3 in MOPC, MPC11, and S194 cells, leading to dramatic dose-dependent apoptosis. [6] AG-490 (100 μM) inhibits Akt phosphorylation, inhibits the activation of nuclear factor-κB, and causes the activation of GSK-3β, leading to the reduction of c-Myc. AG-490 (50 μM) can induce apoptosis of imatinib-resistant BaF3 cells expressing T315I and E255K mutants of Bcr-Abl. [7] AG-490 at 30 μM inhibits not only Epo-induced phosphorylation of wild-type JAK2 but also constitutive phosphorylation of the JAK2 V617F mutant. AG-490 also potently inhibits cytokine-independent cell growth induced by the JAK2 V617F mutant in BaF3 cells. [8] |
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In Vivo
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Administration of AG-490 drastically reduces the numbers of CD45+ and HLA-DR+ cells from 48 % and 46% in bone marrow of untreated mice, as well as 38% and 22% in the spleen of untreated mice to undetectale levels. [2] In vivo administration of AG-490 causes murine myeloma tumor cell apoptosis but does not inhibit IL-12-mediated macrophage activation and IFN-γ production by lymphocytes. [6] Consistent with the in vitro blocking of JAK2 V617F mutant activity, AG-490 treatment at 0.5 mg/day for 10 days effectively inhibits JAK2 V617F mutant-induced tumorigenesis and tumor cell invasion in nude mice. [8] Combined therapy with AG-490 and IL-12 induces greater antitumor effects than either agent alone in a murine myeloma tumor model. [6] |
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Clinical Trials
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Features
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Protocol
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Kinase Assay
[1]
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| In vitro kinase autophosphorylation |
AG-490 is dissolved in DMSO 10%-H2O-ethanol 45%. Crude membrane extracts (0.125 μg/mL) are preactivated with EGF (20 nM) in 50 mM HEPES buffer, pH 7.6, and 125 mM NaCl, for 15 minutes at 4 °C. Autophosphorylation activity of EGFR or ErbB2 kinase is assayed at 4 °C for 30 seconds in V-shaped 96-well plates. Membrane extracts (8 μL) are added to each well containing reaction mixture (12 μL, 50mM, HEPES, pH 7.4,125 mM NaCl, 12 mM M8Ac2, 2 mM MnCl2, 1 mM NaVO3, 1 μM ATP, and 1 μCi[γ-32P]ATP, final concentrations) and increasing concentrations of AG-490 (4 μL). After termination by addition of hot sample buffer, the samples are run on a 6% SDS-polyacrylamide gel electrophoresis minigel, the gels dried, and autoradiography performed during the linear exposure time period. The receptor bands are scanned densitrometrically, and the results analyzed by the Ez-Fit program. For the analysis of autophosphorylation of JAK2, JAK2 is immunoprecipitated by using anti-JAK2 antibody from lysates of G2 cells pretreated for 16 hours with increasing concentrations of AG-490 (0-50 μM). Immune complexes are then immunoblotted with anti-phosphotyrosine antibody. A dose-dependent inhibition of in vitro kinase activity is demonstrated by assessing JAK2 autophosphorylation. |
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Cell Assay
[2]
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| Cell Lines |
Pre-B ALL |
| Concentrations |
Dissolved in DMSO, final concentrations ~ 50 μM |
| Incubation Time |
16 hours |
| Methods |
Cells are exposed to different concentrations of AG-490 for 16 hours. For the determination of cell proliferation, [3H]tymidine (1 μCi) is added 6 hours or more before the cultures are terminated. Cells are then collected and samples counted in a liquid scintillation counter. |
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Animal Study
[2]
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| Animal Models |
SCID mice intravenously injected with ALL cells |
| Formulation |
Dissolved in DMSO |
| Doses |
0.85 mg + 0.5 mg daily |
| Administration |
Continuous pump infusion supplemented with daily intraperitoneal injections |
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References |
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[1] Gazit A, et al. J Med Chem, 1991, 34(6), 1896-1907.
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[2] Meydan N, et al. Nature, 1996, 379(6566), 645-648.
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[3] Nielsen M, et al. Proc Natl Acad Sci U S A, 1997, 94(13), 6764-6769.
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[4] Kirken RA, et al. J Leukoc Biol, 1999, 65(6), 891-899.
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[5] Sun X, et al. Blood, 2001, 97(7), 2008-2015.
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[6] Burdelya L, et al. Mol Cancer Ther, 2002, 1(11), 893-899.
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[7] Samanta AK, et al. Cancer Res, 2006, 66(13), 6468-6472.
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[8] Abe M, et al. Int Immunopharmacol, 2009, 9(7-8), 870-877.
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