Brivanib (BMS-540215) is an ATP-competitive inhibitor against VEGFR2 with IC50 of 25 nM, moderate potency against VEGFR-1 and FGFR-1, but >240-fold against PDGFR-β. Phase 3.

VEGFR2:25 nM

Brivanib 对VEGFR1和FGFR-1具有抑制作用，其IC50分别为0.38 μM和0.148 μM。对于PDGFRβ、EGFR、LCK、PKCα或JAK-3，Brivanib显示出不敏感性，其IC50均超过1900 nM。此外，Brivanib能够抑制VEGF刺激下的HUVEC细胞增殖，IC50为40 nM，相较之下，在FGF刺激的HUVEC细胞中，IC50为276 nM。然而，Brivanib对肿瘤细胞系的活性较低。[1]

Brivanib 在无胸腺小鼠的 H3396 异种移植物上展现出抗肿瘤活性。在60和90 mg/kg（口服）的剂量下，Brivanib 完全抑制了肿瘤生长，TGI 分别为 85% 和 97%。[1] 此外，Brivanib 显著抑制了肝细胞癌（HCC）异种移植物中的肿瘤生长，这归因于 VEGFR2 的磷酸化减少。研究结果显示，在50 mg/kg和100 mg/kg的剂量下，06-0606 异种移植小鼠的肿瘤质量分别是对照组的 55% 和 13%。Brivanib 被认为在治疗 HCC 方面是有效的。[2]

In Vitro Kinase Assays: Recombinant proteins containing tyrosine kinases are expressed as GST fusion proteins using baculovirus expression vector system in Sf9 cells. All enzymes are stored at -80 °C. Brivanib is dissolved in DMSO and diluted by water/10% DMSO. The VEGFR2 kinase solution is composed by 8 ng GST-VEGFR2 enzyme, 75 μg/mL substrate, 1 μM ATP, and 0.04 μCi [γ-33P]-ATP in 50 μL buffer: 20 mM Tris (pH 7.0), 25 μg/mL BSA, 1.5 mM MnCl2, 0.5 mM dithiothreitol). Flk-1 kinase solution is composed by 10 ng GST-Flk-1 enzyme, 75 μg/mL substrate, 1 μM ATP, and 0.04 μCi [γ-33P]-ATP in 50 μL buffer: 20 mM Tris, pH 7.0, 25 μg/mL BSA, 4 mM MnCl2, 0.5 mM dithiothreitol). The reactions are incubated for 1 hour at 27 °C and terminated with cold trichloroacetic acid (TCA) to a final concentration of 15%. These TCA precipitates are collected onto unifilter plates and quantitated by liquid scintillation counter.

The cells are stimulated by VEGF or FGF at a concentration of 8 or 80 ng/mL. These cells are seeded in 96 well plates at a density of 2 × 103 and incubated for 24 hours. Brivanib at various dilutions are added to the cells for another 48 hours. Then 0.5 μCi of [3H] thymidine is added for 24 hours. After that the incorporated tritium is quantified using a β-counter. (Only for Reference)