Icotinib (Conmana) is an orally available quinazoline-based inhibitor of epidermal growth factor receptor (EGFR), with potential antineoplastic activity.

EGFR:5 nM

Icotinib以剂量依赖的方式抑制EGFR活性，IC50值为5 nM，62.5 nM时完全抑制。Icotinib专一性抑制EGFR成员，包括野生型及突变体，抑制效能为61-99%。Icotinib还能剂量依赖性地阻断人表皮癌A431细胞中EGFR介导的细胞内酪氨酸磷酸化。同时，在对A431、BGC-823、A549、H460、HCT8、KB和Bel-7402细胞系进行的增殖实验中发现，细胞对Icotinib的相对敏感性依次为A431 > BGC-823 > A549 > H460 > KB > HCT8 > Bel-7402。Icotinib展现了广谱的抗肿瘤活性，尤其对表达较高水平EGFR的肿瘤效果显著。[1]

Icotinib 在不同类型的异种移植瘤中表现出抗肿瘤效果。在剂量为120 mg/kg的情况下，Icotinib 分别在A431、A549 和H460 异种移植瘤中抑制肿瘤生长51.5%、31.0% 和67.4%。[1]

Biochemical kinase assays: In the in vitro kinase assays, 2.4 ng/μL EGFR protein is mixed with 32 ng/μL Crk in 25 μL kinase reaction buffer containing 1 μM cold ATP and 1 μCi 32P-γ-ATP. The mix is incubated with Icotinib at 0, 0.5, 2.5, 12.5 or 62.5 nM on ice for 10 min followed by incubation at 30 °C for 20 min. After quenching with SDS sample buffer at 100 °C for 4 min, the protein mix is resolved by electrophoresis in a 10% SDS-PAGE gel. The dried gel is then exposed to the PhosphorImager to detect radioactivity. Quantification is performed by ImageQuant software. In this methodology the radioactive signal inversely correlates with kinase activity.

Cells (103 /well) are seeded into 96-well plates in RPMI-1640 medium containing 10% FBS and grown in a 5% CO2 incubator at 37 °C. After 24 h, cells are treated with Icotinib at 0, 0.78, 1.56, 3.125, 6.25, 12.5 or 25 μM for 96 h. Cell proliferation is calculated by subtracting the mean absorbance value on day 0 from the mean absorbance value on day 4.(Only for Reference)