Neratinib

Manufactured by Selleck Chemicals

Sourced in United States

Neratinib is a laboratory reagent used in research applications. It functions as a tyrosine kinase inhibitor.

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Close spelling variants of this product

Neratinib (HKI-272) (3 citations)

34 protocols using neratinib

Antibody and Drug Reagents for EGFR, AXL, and MAPK Signaling

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Antibodies against EGFR (#2232, RRID:AB_331707), AXL (#8661, RRID:AB_11217435), phospho-ERK (#9101, RRID:AB_331646), ERK (#9102, RRID:AB_330744), PTEN (#9552, RRID:AB_10694066), β-actin (#4967, RRID:AB_330288), GAPDH (#2118, RRID:AB_561053), and vinculin, were obtained from Cell Signaling Technology. Anti-CBL (#05-440, RRID:AB_2290887) was obtained from MilliporeSigma. Anti-NF1 (A300-140A; RRID:AB_2149790) was obtained from Bethyl Antibodies. Vemurafenib, cobimetinib, neratinib, dabrafenib, trametinib, ZM336372, LY3009120, and belvarafenib were obtained from SelleckChem. Drugs for in vitro studies were dissolved in DMSO to create 10 or 100 mmol/L stock solutions and stored at −20°C.

Schreck K.C., Morin A., Zhao G., Allen A.N., Flannery P., Glantz M., Green A.L., Jones C., Jones K.L., Kilburn L.B., Nazemi K.J., Samuel D., Sanford B., Solomon D.A., Wang J., Pratilas C.A., Nicolaides T, & Mulcahy Levy J.M. (2021). Deconvoluting Mechanisms of Acquired Resistance to RAF Inhibitors in BRAFV600E-Mutant Human Glioma. Clinical Cancer Research, 27(22), 6197-6208.

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Evaluating Anti-Cancer Drug Efficacy

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Four lung cancer cell lines (A549, Calu-3, NCI-H2170 and NCI-H1781) and one normal human bronchial epithelial cell line (BEAS-2B) were used in the current study. Calu-3, H2170 and H1781 cells were received as gifts from Dr Adi F. Gazdar (University of Texas Southwestern Medical Center at Dallas, Dallas, TX, USA) (22 (link),23 (link)). A549 was purchased from American Type Culture Collection (Manassas, VA, USA). BEAS-2B was purchased from European Collection of Authenticated Cell Cultures (Public Health England, Porton Down, UK). All the cancer cell lines were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS), and the BEAS-2B cells were maintained in Dulbecco's modified Eagle's medium supplemented with 10% FBS. They were cultured in a humidified atmosphere containing 5% CO₂ at 37°C. Neratinib and erlotinib were purchased from Selleck Chemicals (Houston, TX, USA).

Ogoshi Y., Shien K., Yoshioka T., Torigoe H., Sato H., Sakaguchi M., Tomida S., Namba K., Kurihara E., Takahashi Y., Suzawa K., Yamamoto H., Soh J, & Toyooka S. (2019). Anti-tumor effect of neratinib against lung cancer cells harboring HER2 oncogene alterations. Oncology Letters, 17(3), 2729-2736.

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Comprehensive Molecular Inhibitor Characterization

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Erlotinib, NVP-AEW541 and afatinib were kindly provided by OSI Pharmaceuticals (Farmingdale, USA), Novartis (Basel, Switzerland) and Boehringer Ingleheim (Vienna, Austria), respectively. Paclitaxel and gemcitabine were acquired from Sigma-Aldrich (Dorset, UK) and Healthcare at Home (UK), respectively, while Iressa (gefitinib) and crizotinib were purchased from Tocris (Avonmouth, UK), respectively. Lapatinib, sapitinib, canertinib, neratinib, imatinib and dasatinib were all acquired from Selleckchem (USA).
Primary and secondary antibodies for flow cytometry were purchased from R&D Systems (Abingdon, UK), while the secondary FITC-conjugated rabbit anti-mouse mAb STAR9B was purchased from AbD Seroctec (Kidlington, UK). Primary antibodies for western blot were obtained from Cell Signaling Technology Inc. (Massachusetts, USA), apart from the anti-EGFR antibody F4, which was obtained from Sigma-Aldrich, while secondary antibodies were purchased from Li-Cor Inc (Nebraska, USA).

Stanley A., Ashrafi G.H., Seddon A.M, & Modjtahedi H. (2017). Synergistic effects of various Her inhibitors in combination with IGF-1R, C-MET and Src targeting agents in breast cancer cell lines. Scientific Reports, 7, 3964.

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Establishment of Breast Cancer Cell Line Resistant Models

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The breast cancer cell lines BT474, SKBR3 and MCF7 were purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). All cell lines were maintained as recommended by the suppliers. All cell lines were authenticated using Human 9-Marker STR Profile and Interspecies Contamination Test by IDEXX BioAnalytics. Lapatinib, neratinib, tucatinib and palbociclib were purchased from Selleckchem (Houston, TX, USA). Trastuzumab was obtained from Roche Farma (Basel, Switzerland). A BT474-derived Lapatinib and Trastuzumab resistant (BT474-L^RT^R) cell line and a BT474-derived tucatinib and Trastuzumab resistant (BT474-Tu^RT^R) cell line were established by treating BT474 with the corresponding TKI plus Trastuzumab for 6 months, starting at low concentrations and increasing them at each passage. BT474-L^RT^R are maintained with 2 nM Lapatinib + 2 µg ml⁻¹ Trastuzumab, while BT474-Tu^RT^R are maintained with 2 nM tucatinib + 2 µg⁻¹ ml Trastuzumab.

Brasó-Maristany F., Griguolo G., Pascual T., Paré L., Nuciforo P., Llombart-Cussac A., Bermejo B., Oliveira M., Morales S., Martínez N., Vidal M., Adamo B., Martínez O., Pernas S., López R., Muñoz M., Chic N., Galván P., Garau I., Manso L., Alarcón J., Martínez E., Gregorio S., Gomis R.R., Villagrasa P., Cortés J., Ciruelos E, & Prat A. (2020). Phenotypic changes of HER2-positive breast cancer during and after dual HER2 blockade. Nature Communications, 11, 385.

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Characterizing Drug Response in Breast Cancer Cell Lines

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All cell lines were purchased from ATCC (Manassas, VA), cultured in the prescribed medium, and incubated at 37 °C in 5% CO₂. Cell line identity was confirmed by genotyping, and all cultures were tested to ensure absence of mycoplasma infection. SK-BR-3 selection conditions were 0.5 mg/mL for G418 and 0.25μg/mL for puromycin. Lapatinib, neratinib, the AKT inhibitor GSK690693, and the MEK inhibitors GSK1120212 and PD0325901 were purchased from Selleck Chemicals (Houston, TX).

Garay J.P., Smith R., Devlin K., Hollern D.P., Liby T., Liu M., Boddapati S., Watson S.S., Esch A., Zheng T., Thompson W., Babcock D., Kwon S., Chin K., Heiser L., Gray J.W, & Korkola J.E. (2021). Sensitivity to targeted therapy differs between HER2-amplified breast cancer cells harboring kinase and helical domain mutations in PIK3CA. Breast Cancer Research : BCR, 23, 81.

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Comprehensive Cancer Cell Panel Experiments

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The cell lines used in experiments included MFE-296, HEC50B, DU4475, BT-474, HCC1954, MDA-MB-468, SK-OV-3, KM12, HCC2998, SW480, HCT15. They were obtained through the MD Anderson Characterized Core Cell Line Facility and the MDACC CRC Moon Shot program. Cell lines were authenticated by fingerprinting using short tandem repeat testing. The absence of mycoplasma contamination was also verified. Cells were maintained in RPMI-1640 supplemented with 10% heat-inactivated FBS and 1% penicillin with streptomycin. The targeted compounds (PARP inhibitor – Olaparib, Chk2 inhibitor – BML-277, AKT inhibitor MK-2206, MET inhibitor – JNJ-38877605, HER2 inhibitor – Neratinib, AURKA inhibitor – Alisertib, CDK4/6 inhibitor – Ribociclib, MAP3K4 inhibitor – Doramapimod (directly inhibiting downstream signaling partner, p38/MAPK14), PIK3CA inhibitor – Buparlisib) were obtained from Selleckchem.

Li X., Dowling E.K., Yan G., Dereli Z., Bozorgui B., Imanirad P., Elnaggar J.H., Luna A., Menter D.G., Pilié P.G., Yap T.A., Kopetz S., Sander C, & Korkut A. (2022). Precision combination therapies based on recurrent oncogenic co-alterations. Cancer discovery, 12(6), 1542-1559.

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Evaluating EGF, NRG1b, and HGF Signaling

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Recombinant human epidermal growth factor (EGF) and neuregulin 1b (NRG1b) were purchased from R&D Systems (Minneapolis, MN). Recombinant human hepatocyte growth factor (HGF) was purchased from Peprotech (Rocky Hill, NJ). Collagen was obtained from Advanced Biomatrix (Carlsbad, CA) and fibronectin was obtained from Sigma (St. Louis, MO). Neratinib, erlotinib, tepotinib were purchased from SelleckChem (Houston, TX) and prepared at stock concentrations in fresh 100% DMSO (Amresco) before final dilution into assay medium. The monoclonal anti-HER2 antibody, 2C4, was expressed and purified from a mouse hybridoma cell line obtained from ATCC (Manassas, VA). Drugs for in vivo studies were prepared in 10% captisol.

MacNeil I.A., Khan S.A., Sen A., Soltani S.M., Burns D.J., Sullivan B.F, & Laing L.G. (2022). Functional signaling test identifies HER2 negative breast cancer patients who may benefit from c-Met and pan-HER combination therapy. Cell Communication and Signaling : CCS, 20, 4.

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Quantitative Fluorescent Imaging of Drug Combinations

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Lapatinib, neratinib, crizotinib, trametinib, capecitabine (Selleckchem), pertuzumab, and trastuzumab (OHSU Pharmacy) were used at the concentrations indicated in figure legends. DMSO (ThermoFisher) and human IgG isotype control (Abcam) concentrations were equivalent to the highest dose of the respective drug used in each experiment. Treatment durations were as indicated in respective figure legends. Cells were treated in 96 and 384 multiwell plates with soluble drug and ligand combinations added to their medium, then fixed for fluorescent imaging and quantification (described below). Each treated cell line was seeded at an experimentally determined concertation so that untreated control wells would reach 80% confluency by the end of the treatment period. Drug combination studies and CTG assays in Figure 4F were performed as previously reported (Heiser et al., 2012 (link)) (Kuo et al., 2009 (link)) in randomized replicates.

Watson S.S., Dane M., Chin K., Tatarova Z., Liu M., Liby T., Thompson W., Smith R., Nederlof M., Bucher E., Kilburn D., Whitman M., Sudar D., Mills G.B., Heiser L.M., Jonas O., Gray J.W, & Korkola J.E. (2018). Microenvironment-Mediated Mechanisms of Resistance to HER2 Inhibitors Differ between HER2+ Breast Cancer Subtypes. Cell systems, 6(3), 329-342.e6.

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Gastric Cancer Cell Line Panel for Targeted Therapies

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Twelve gastric cancer cell lines (ECC10, GCIY, KATO‐III, MKN7, MKN74, NCI‐N87, NUGC3, NUGC4, OCUM‐1, SH‐10‐TC, SNU‐16, and SNU‐216) were used in this study. ECC10, GCIY, and MKN7 were provided by Riken BRC through the National Bio‐Resource Project of Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan. NUGC3, MKN74, and OCUM‐1 were provided by Japanese Collection of Research Bioresources/Health Science Research Resources Bank (JCRB/HSRRB), Osaka, Japan. KATO‐III, NCI‐N87, NUGC4, SNU‐16, SH‐10‐TC were purchased from ATCC (Manassas, VA, USA), and SNU‐216 was obtained from the Korean Cell Line Bank. All the cells, other than GCIY and OCUM‐1, were cultured in RPMI‐1640 media supplemented with 10% FBS. GCIY and OCUM‐1 were cultured in minimum essential media (Sigma‐Aldrich) with 15% FBS and DMEM with 0.5 mmol/L sodium pyruvate and 10% FBS, respectively. Afatinib, neratinib, and PPP were purchased from Selleckchem and MedChem Express. Gefitinib was purchased from Sykkinase. Trastuzumab and pertuzumab were obtained from Chugai Pharmaceutical Co.

Yoshioka T., Shien K., Namba K., Torigoe H., Sato H., Tomida S., Yamamoto H., Asano H., Soh J., Tsukuda K., Nagasaka T., Fujiwara T, & Toyooka S. (2018). Antitumor activity of pan‐HER inhibitors in HER2‐positive gastric cancer. Cancer Science, 109(4), 1166-1176.

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Comprehensive Toolkit for Cancer Research

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RSL3 (S8155), neratinib (S2150), lapatinib (S2111), gefitinib (S1025), ML210 (S0788), afatinib (S1011), dacomitinib (S2727), sapitinib (S2192), Z-VAD-FMK (S7023), necrostatin-1 (S8037), 3-Methyladenine (3-MA, S2767), tucatinib (S8362), liproxstatin-1 (S7699), erastin (S7242), deferoxamine mesylate (DFO, S5742), deferiprone (S4067), ferrostatin-1 (S7243), cobimetinib (S8041), Trastuzumab (A2007) and AZD6738 (S7693) were obtained from Selleck Chemicals. L-glutathione (G6013) and N-Acetyl-l-cysteine (A9165) were purchased from Sigma-Aldrich. T-DM1 (HY-P9921) was obtained from MedChemExpress.

Park S.Y., Jeong K.J., Poire A., Zhang D., Tsang Y.H., Blucher A.S, & Mills G.B. (2023). Irreversible HER2 inhibitors overcome resistance to the RSL3 ferroptosis inducer in non-HER2 amplified luminal breast cancer. Cell Death & Disease, 14(8), 532.

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