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Neratinib

Cat. No. M1913

Neratinib Structure

Synonym: HKI-272

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50mg USD 200 In stock
100mg USD 350 In stock
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Quality Control
Biological Activity

Neratinib is a potent irreversible inhibitor of epidermal growth factor receptor and human epidermal receptor 2 with IC50 of 92 nM and 59 nM, respectively. Neratinib inhibits the proliferation of EGFR-dependent A431 cells with an IC50 of 81 nM. Neratinib (HKI-272) also weakly inhibits tyrosine kinases KDR and Src with IC50 of 0.8 μM and 1.4 μM, respectively. Likewise, Neratinib inhibited the photolabeling of ABCB1 with [(125)I]iodoarylazidoprazosin in a concentration-dependent manner (IC50 = 0.24 μM). Neratinib enhanced the sensitivity of ABCB1-overexpressing cells to ABCB1 substrates. It is noteworthy that Neratinib augmented the effect of chemotherapeutic agents in inhibiting the growth of ABCB1-overexpressing primary leukemia blasts and KBv200 cell xenografts in nude mice. Moreover, Neratinib increased doxorubicin accumulation in ABCB1-overexpressing cell lines and Rhodamine 123 accumulation in ABCB1-overexpressing cell lines and primary leukemia blasts. Neratinib (HKI-272) stimulated the ATPase activity of ABCB1 at low concentrations but inhibited it at high concentrations.

Protocol
Cell Experiment
Cell lines 3T3, 3T3/neu, A431, SK-Br-3, BT474, MDA-MB-435, and SW62 cells
Preparation method Cell Proliferation Assays.
Cells were plated in 96-well tissue culture plates (3T3, 3T3/neu, 5000 cells/well; A431, SK-Br-3, BT474, MDA-MB-435, and SW620, 10,000 cells/well). The following day, dilutions of compound (0.5 ng/ml–5 μg/ml) were added, and cells were cultured for 2 days (6 days for BT474). Cell proliferation was determined using sulforhodamine B, a protein binding dye. Briefly, cells were fixed with 10% trichloroacetic acid and washed extensively with water. Cells were then stained with 0.1% sulforhodamine B (Sigma-Aldrich) and washed in 5% acetic acid. Protein-associated dye was solubilized in 10 mm Tris, and absorbance was measured at 450 nm (Victor2). Inhibition of cell proliferation was calculated using the formula: percentage of inhibition = 100 − 100 (Td − To/Tc − To), where Td is the absorbance of drug treated cells, Tc is the absorbance of untreated cells, and To is the absorbance at the time of drug addition. To values were determined by plating cells separately and fixing them at the time of drug addition. The concentration of compound which inhibits cell proliferation by 50% (IC50) was determined from inhibition curves.
Concentrations 0.5 ng/ml-5 μg/ml
Incubation time 2 days
Animal Experiment
Animal models BT474, MCF-7, and SK-OV-3 tumour xenografts model in female athymic (nude) mice
Formulation 0.5% methocellulose-0.4% polysorbate-80 (Tween 80)
Dosages 40 or 60mg/kg daily for 20 days
Administration oral gavage
Conversion of different model animals based on BSA (Value based on data from FDA Draft Guidelines)
Species Mouse Rat Rabbit Guinea pig Hamster Dog
Weight (kg) 0.02 0.15 1.8 0.4 0.08 10
Body Surface Area (m2) 0.007 0.025 0.15 0.05 0.02 0.5
Km factor 3 6 12 8 5 20
Animal A (mg/kg) = Animal B (mg/kg) multiplied by  Animal B Km
Animal A Km

For example, to modify the dose of resveratrol used for a mouse (22.4 mg/kg) to a dose based on the BSA for a rat, multiply 22.4 mg/kg by the Km factor for a mouse and then divide by the Km factor for a rat. This calculation results in a rat equivalent dose for resveratrol of 11.2 mg/kg.

Chemical Information
Molecular Weight 557.04
Formula C30H29ClN6O3
CAS Number 698387-09-6
Purity 100.00%
Solubility DMSO
Storage at -20°C
Customer Product Validations & Biological Datas
Source ACS Chem Biol (2016). Figure 5. Neratinib (Abmole Bioscience)
Method FECH inhibition assay
Cell Lines K562 Cells
Concentrations 1 μM
Incubation Time 6 days
Results In line with the Kd values obtained from kinobead experiments, Vemurafenib, CUDC 101, Cyc-116, Linsitinib, MK-2461, Neratinib, GSK-690693, and Crenolanib decreased FECH binding to the PPIX beads by 50% or more compared to the DMSO control.
Rating
Source ACS Chem Biol (2016). Figure 4. Neratinib (Abmole Bioscience)
Method FECH inhibition assay
Cell Lines K562 Cells
Concentrations 1 μM
Incubation Time 6 days
Results We then subjected MK-2461, Neratinib, and Linsitinib to the same assay (1 μM for 6 days), and each drug clearly led to reduced heme levels indicating inhibition of FECH activity in K562 cells with Neratinib showing the most potent effect (60% reduction).
Rating
Source ACS Chem Biol (2016). Figure 3. Neratinib (Abmole Bioscience)
Method FECH binding
Cell Lines K562 Cells
Concentrations
Incubation Time
Results The basis of this assay is that proteins bound to a drug molecule are stabilized against heat induced unfolding and aggregation, and the extent of this protection is dose dependent.
Rating
Source ACS Chem Biol (2016). Figure 2. Neratinib (Abmole Bioscience)
Method FECH binding
Cell Lines
Concentrations
Incubation Time
Results Figure 2b−f show the full dose response data for five selected inhibitors, and the complete set of these plots can be found in Supporting Information Figure S1 and Table S1.
Rating
Source Tianjin Med J (2016). Neratinib, Figure 4. (AbMole, USA)
Method MTT实验
Cell Lines A549 细胞
Concentrations 10 μmol/L
Incubation Time 24 h
Results 与阴性对照组比较,在作用48、72、96 h 后 Neratinib 及3-MA 能有效抑制A549 细胞的增殖(F 分别为15.213 4.965、5.366,均P<0.05),见图4。
Rating
Source Tianjin Med J (2016). Neratinib, Figure 3. (AbMole, USA)
Method Transwell invasion
Cell Lines A549 细胞
Concentrations 10 μmol/L
Incubation Time 24 h
Results Neratinib 处理组、3-MA 处理组穿膜细胞数分别为40±8、59±10,均显著低于阴性对照组的113±13,差异均有统学意义(F=18.246,P<0.05),见图3。提示HER2 低表达及细胞自噬活性水平降低能抑制肺癌细胞系A549 细胞侵袭。
Rating
Source Tianjin Med J (2016). Neratinib, Figure 2. (AbMole, USA)
Method Wound healing 实验
Cell Lines A549 细胞
Concentrations 10 μmol/L
Incubation Time 24 h
Results Neratinib 及3-MA 对A549 细胞迁徙能力的影响Neratinib 处理组、3-MA 处理组24 h 迁徙率分别为11.1%±1.4% 10.5%±1.2%,均低于阴性对照组的42.9%±2.4%,差异均有统计学意义(F=27.430,P<0.05),见图2。提HER2 低表达及细胞自噬活性水平能降低抑制肺癌细胞系A549 细胞迁徙。
Rating
Source Tianjin Med J (2016). Neratinib, Figure 1. (AbMole, USA)
Method Western blot
Cell Lines A549 细胞
Concentrations 10 μmol/L
Incubation Time 24 h
Results Neratinib 及3-MA 对A549 细胞自噬相关蛋白 Beclin-1 及LC3B(Ⅱ/Ⅰ)表达的影响与阴性对照 组相比,3-MA 处理组的Beclin-1 蛋白及LC3BⅡ/ Ⅰ表达水平明显降低;Neratinib 处理组的HER2、Beclin-1 蛋白及LC3BⅡ/Ⅰ表达水平明显降低(P<0.05),见图1。提示在人源肺癌A549 细胞中,改变 HER2 的表达能调控细胞自噬的活性。
Rating
Product Citations
References

Neratinib reverses ATP-binding cassette B1-mediated chemotherapeutic drug resistance in vitro, in vivo, and ex vivo.
Zhao XQ, et al. Mol Pharmacol. 2012 Jul;82(1):47-58. PMID: 22491935.

Antitumor activity of HKI-272, an orally active, irreversible inhibitor of the HER-2 tyrosine kinase.
Rabindran SK, et al. Cancer Res. 2004 Jun 1;64(11):3958-65. PMID: 15173008.

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  Catalog
Abmole Inhibitor Catalog 2017




Keywords: Neratinib, HKI-272 supplier, EGFR/HER2, inhibitors

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