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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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Linifanib (ABT-869) is a novel, potent ATP-competitive RTK inhibitor for KDR, CSF-1R, Flt-1 and Flt-3 with IC50 of 4 nM, 3 nM, 3 nM and 3 nM, respectively. |
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Targets
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KDR |
CSF-1R |
Flt-1 |
Flt-3 |
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IC50 |
4 nM |
3 nM |
3 nM |
4 nM [1] |
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In Vitro
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Linifanib shows inhibitory to Kit, PDGFRβ and Flt4 with IC50 of 14 nM, 66 nM and 190 nM in kinases assay. Linifanib also inhibits ligand-induced KDR, PDGFRβ, Kit, and CSF-1R phosphorylation with IC50 of 2 nM, 2 nM, 31 nM and 10 nM at cellular level and this cellular potency could be affected by serum protein. Linifanib suppresses VEGF-stimulated HUAEC proliferation with IC50 of 0.2 nM. While Linifanib has weak activity against tumor cells which are not induced by VEGF or PDGF, except for MV4-11 leukemia cells (with constitutively active form of Flt3) with IC50 of 4 nM. Linifanib could cause a decrease in S and G2-M phases with a corresponding increase in the sub-G0-G1 apoptotic population in MV4-11 cells. [1] Linifanib binds to the ATP-binding site of CSF-1R with Ki of 3 nM. [2] Linifanib (10 nM) exhibits a reduced phosphorylation of Akt at Ser473 and decreased phosphorylation of GSK3βat Ser9 in Ba/F3 FLT3 ITD cell lines. [3] |
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In Vivo
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Linifanib (0.3 mg/kg) results in complete inhibition of KDR phosphorylation in lung tissue. Linifanib also inhibits the edema response with ED50 of 0.5 mg/kg. Linifanib (7.5 and 15 mg/kg, bid) significantly inhibits both bFGF- and VEGF-induced angiogenesis in the cornea. Linifanib inhibits tumor growth in flank xenograft models including HT1080, H526, MX-1 and DLD-1 with ED75 from 4.5-12 mg/kg. Linifanib also shows efficacy in A431 and MV4-11 xenografts at low dose levels. Linifanib (12.5 mg/kg bid) reveals a decrease of microvasculure density in MDA-231 xenograft. Linifanib shows a Cmax and AUC24 hours with 0.4 μg/mL and 2.7 μg?hour/mL in HT1080 fibrosarcoma model. [1] |
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Clinical Trials
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Linifanib is in a Phase III evaluation for hepatocellular carcinoma. |
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Features
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Combination Therapy
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Description
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Combination of AB869 with Rapamycin treatment reduces the tumors to the lowest volume, and is significantly better than single agent treatment. Combination treatment of ABT869 with Rapamycin shows synergistic effect on expression levels of p27 in subcutaneous Huh7 and SK-HEP-1 xenograft models. [4] In combination, ABT869 augments the effects seen with Paclitaxel in both (MDA-231 and MDA-435LM) of the orthotopic models. [5] |
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Protocol
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Kinase Assay
[1]
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| Kinase assays |
Potencies (IC50 values) are determined by assays of active kinase domains cloned and expressed in baculovirus using the FastBacbaculovirus expression system or obtained commercially. For tyrosine kinase assays, a biotinylated peptide substrate containing a single tyrosine is used with 1 mM ATP, anEu-cryptate–labeled anti-phosphotyrosine antibody (PT66), and Strepavidin-APC in a homogeneous time-resolved fluorescence assay. Serine/threonine kinases are assayed using 5 μM ATP, [33P]ATP, and a biotinylated peptide substrate with peptide capture and incorporation of 33P determined using a SA-Flashplate. Linifanib is assayed at multiple concentrations prepared by serial dilution of a DMSO stock solution of Linifanib. The concentration resulting in 50% inhibition of activity is calculated using nonlinear regression analysis of the concentration response data. |
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Cell Assay
[1]
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| Cell Lines |
HUAEC, HT-29, HT1080, A431, MDA-435, MDA-231, H526, DLD-1, 9L and MV4-11 cells |
| Concentrations |
0-100 μM |
| Incubation Time |
72 hours |
| Methods |
Cells are seeded into 96-well plates at 2.5 × 103 per well and incubated with serum-free medium for 24 hours. Linifanib and VEGF (final, 10 ng/mL) are added and incubated for 72 hours in serum-free medium. For carcinoma cell lines, 3 × 103 cells/well are plated overnight in full growth medium. Linifanib is added to the cells in full growth medium and incubated for 72 hours. For leukemia cells, generally 5 × 104 per well are plated in full growth medium, Linifanib is added, and incubated for 72 hours. The effects on proliferation are determined by addition of Alamar Blue (final solution, 10%), incubation for 4 hours at 37 °C in a CO2 incubator and analysis in a fluorescence plate reader (544 nm, excitation: 590 nm, emission). |
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Animal Study
[1]
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| Animal Models |
H526, DLD-1, MDA-231, MDA-435LM, HCT-116, H526, DLD-1, MDA-231, MDA-435LM, MV4-11 and MX-1 xenografts are established in mice. |
| Formulation |
2% ethanol, 5% Tween 80, 20% PEG400, 73% saline |
| Doses |
~ 10 mg/kg |
| Administration |
Oral administration |
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References |
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[1] Albert DH, et al. Mol Cancer Ther, 2006, 5(4), 995-1006.
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[2] Guo J, et al. Mol Cancer Ther, 2006, 5(4), 1007-1013.
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[3] Hernandez-Davies JE, et al. Mol Cancer Ther, 2011, 10(6), 949-959.
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[4] Jasinghe VJ, et al. J Hepatol. 2008, 49(6), 985-997.
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[5] Albert DH, et al. Mol Cancer Ther. 2006, 5(4), 995-1006.
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