| 生物活性 | Afatinib (BIBW 2992) dimaleate is an orally active, potent and irreversible dual specificity inhibitor ofErbBfamily (EGFRandHER2), withIC50values of 0.5 nM, 0.4 nM, 10 nM and 14 nM forEGFRwt,EGFRL858R,EGFRL858R/T790Mand HER2, respectively. Afatinib dimaleate can be used for the research of esophageal squamous cell carcinoma (ESCC), non-small cell lungcancer(NSCLC) and gastriccancer[1][2][3][4]. |
| IC50& Target[1] | EGFR 0.5 nM (IC50) | HER2 14 nM (IC50) | EGFRL858R 0.4 nM (IC50) | EGFRL858R/T790M 10 nM (IC50) |
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体外研究
(In Vitro) | Afatinib dimaleate (100 nM) sufficiently prevents heregulin-stimulated HER3 phosphorylation[1].
Afatinib dimaleate (0-10000 nM) effectively inhibits anchorage-independent proliferation of NIH-3T3 cells ectopically expressing EGFR mutants, and inhibits cell proliferation of H1666, H3255, and NCI 1975 cells[1].
Afatinib dimaleate (48-72 h)shows growth inhibition in HKESC-1, HKESC-2, SLMT-1 and EC-1 cells[2].
Afatinib dimaleate (0-1 μM, 24-48 h) inhibits AKT and MAPK pathways, and inhibits EGFR and AKT phosphorylation in ESCC cell lines[2].
Afatinib dimaleate (0-1 μM, 16-48 h) induces G0/G1 cell cycle arrest in HKESC-2 and EC-1[2].
Afatinib dimaleate (0-1 μM, 24-48 h) effectively induces apoptotic cell death in HKESC-2 and EC-1[2].
Cell Proliferation Assay[1] | Cell Line: | NIH-3T3 cells, H1666, H3255, and NCI 1975 cells | | Concentration: | 0, 1, 10, 100, 1000, 10000 nM | | Incubation Time: | | | Result: | Effectively inhibited anchorage-independent proliferation of NIH-3T3 cells ectopically expressing EGFR mutants. Showed inhibition of anchorage independent cell proliferation of various lung cancer cell lines (H1666, H3255, and NCI 1975 cells), with IC50values of 60 nM, 0.7 nM and 99 nM, respectively. |
Cell Viability Assay[2] | Cell Line: | HKESC-1, HKESC-2, SLMT-1 and EC-1 cell lines | | Concentration: | | | Incubation Time: | 48 and 72 hours | | Result: | Observed over 95% of growth inhibition. The respective IC50concentrations at 48 hours (HKESC-1=0.078 μM, HKESC-2=0.115 μM, KYSE510=3.182 μM, SLMT-1=4.625 μM and EC-1=1.489 μM) and 72 hours (HKESC-1=0.002 μM, HKESC-2=0.002 μM, KYSE510=1.090 μM, SLMT-1=1.161 μM and EC-1=0.109 μM) were all in lower micro-molar range. |
Western Blot Analysis[2] | Cell Line: | HKESC-2 cells and EC-1 cells | | Concentration: | 0, 0.01, and 0.1 μM (HKESC-2 cells), 0, 0.1 and 1 μM (EC-1 cells) | | Incubation Time: | 24 and 48 hours | | Result: | Reduced the phosphorylation of EGFR and the endogenous expression level of HER2 receptors in ESCC cells. Suppressed AKT phosphorylation in a dose and time dependent manner. Significantly reduced the phosphorylation level of the downstream effectors of the AKT-mTOR axis especially in HKESC-2 cells. Inhibited the two major downstream pathways of the ErbB/HER axis, namely, AKT and MAPK pathways in ESCC cell lines. |
Cell Cycle Analysis[2] | Cell Line: | HKESC-2 cells and EC-1 cells | | Concentration: | 0, 0.01, and 0.1 μM (HKESC-2 cells), 0, 0.1 and 1 μM (EC-1 cells) | | Incubation Time: | 16, 24, and 48 hours | | Result: | Induced G0/G1 cell cycle arrest in both tested ESCC cell lines in a time and dose dependent manner. In HKESC-2 cells, the percentage of cells in G0/G1 phase was increased from 38.2% to 68.1% at 0.01 μM of afatinib and to 74.7% at 0.1 μM of afatinib, from 24 hours (82.4% G0/G1 arrest at 0.01 μM and 86.2% at 0.1 μM) to 48 hours (from 74.7% to 88.2% for 0.01 μM and 91.0% for 0.1 μM). In EC-1 cells, the percentage of cells arrested in the G0/G1 phase was increased from 59.1% to 66.6% and 72.2% at 24 and 48 hours respectively. |
Apoptosis Analysis[2] | Cell Line: | HKESC-2 cells and EC-1 cells | | Concentration: | 0, 0.01, and 0.1 μM (HKESC-2 cells), 0, 0.1 and 1 μM (EC-1 cells) | | Incubation Time: | 24 and 48 hours | | Result: | Effectively induced cell death by triggering apoptotic mechanisms in ESCC cell lines. Showed a stronger expression level of cleaved Poly (ADP-ribose) polymerase (PARP) in these cell lines. |
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体内研究
(In Vivo) | Afatinib dimaleate (0-20 mg/kg, Orally, daily for 25 days) shows dramatic tumor regression and downregulation of EGFR, HER2, HER3 and AKT phosphorylation[1].
Afatinib dimaleate (15 mg/kg, Orally, in a schedule of 5 days on plus 2 days off, for two weeks) strongly inhibits the growth of HKESC-2 tumor[2].
| Animal Model: | Athymic NMRI-nu/nu female mice (21–31 g, five to six-week-old, transgenic murine lung cancer model and xenograft models)[1] | | Dosage: | 15 mg/kg, 20 mg/kg | | Administration: | Orally, daily for 25 days | | Result: | Resulted in dramatic tumor regression with a cumulative treated/control tumor volume ratio (T/C ratio) of 2% in a standard xenograft model of the epidermoid carcinoma cell line A431, and downregulation of EGFR and AKT phosphorylation. Induced regression of large tumors in this HER2-driven model, effectively controlled xenograft tumor formation by the NCIH1975 cell line, expressing EGFR L858R/T790M, with a T/C value of 12% for doses of 20 mg/kg. Induced more than 50% percent tumor reduction after a 4-week treatment period. Downregulated EGFR, HER2 and HER3 phosphorylation. |
| Animal Model: | Six weeks old female athymic nude mice (nu/nu) (16-20 g)[2] | | Dosage: | 15 mg/kg | | Administration: | Oral gavage in a schedule of 5 days on plus 2 days off, for two weeks | | Result: | Strongly inhibited the growth of HKESC-2 tumor. Average tumor sizes of vehicle and treatment at end point are 348 ± 24 mm3and 108 ± 36 mm3respectively. |
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| 运输条件 | Room temperature in continental US; may vary elsewhere. |
| 储存方式 | 4°C, sealed storage, away from moisture *In solvent : -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture) |
| 溶解性数据 | In Vitro: H2O : 50 mg/mL(69.63 mM;Need ultrasonic) DMSO : ≥ 35 mg/mL(48.74 mM) *"≥" means soluble, but saturation unknown. 配制储备液 | 1 mM | 1.3926 mL | 6.9630 mL | 13.9260 mL | | 5 mM | 0.2785 mL | 1.3926 mL | 2.7852 mL | | 10 mM | 0.1393 mL | 0.6963 mL | 1.3926 mL |
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储备液的保存方式和期限：-80℃, 6 months; -20℃, 1 month (sealed storage, away from moisture)。-80℃ 储存时，请在 6 个月内使用，-20℃ 储存时，请在 1 个月内使用。 In Vivo: 请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照In Vitro方式配制澄清的储备液，再依次添加助溶剂： ——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用；
以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶 1. 请依序添加每种溶剂： PBS Solubility: 100 mg/mL (139.26 mM); Clear solution; Need ultrasonic
*以上所有助溶剂都可在本网站选购。 |
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