Journal: Molecular Biology of the Cell

Article Title: ErbB4 tyrosine kinase inhibition impairs neuromuscular development in zebrafish embryos

doi: 10.1091/mbc.E18-07-0460

Figure Lengend Snippet: ErbB kinase inhibitors reduce motility of zebrafish embryos. Touch–response assays measuring motility of embryos treated from 8 to 48 hpf with the buffer control DMSO or with 10 µM of the indicated ErbB TKIs. (A) The colored lines demonstrate videorecorded tracks of swim bursts of individual embryos stimulated to move by a physical touch to the tail. (B, C) Quantification of the length of the entire swim burst (B) and the average speed during the burst (C) in the touch–response assays. Mean values are indicated above each column. DMSO, n = 18; erlotinib, n = 14; gefitinib, n = 11; osimertinib, n = 9; lapatinib, n = 7, afatinib, n = 7; neratinib, n = 6; canertinib, n = 7; dacomitinib, n = 9; AG 1478, n = 11. * p < 0.05.

Article Snippet: To block ErbB kinase activity in the embryos, 10 μM of tyrosine kinase inhibitors AG 1478 (Calbiochem), erlotinib, gefitinib, lapatinib, afatinib, neratinib, canertinib (all from Santa-Cruz Biotechnologies), osimertinib, and dacomitinib (both from Cayman Chemical), diluted in DMSO, or DMSO alone was applied in E3 medium at 8 hpf, except as indicated for experimentation shown in D. The final concentration of DMSO was 0.1% of the culture medium.

Techniques: Control

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: Increased progression of injury, impaired recovery and decreased survival after delayed (12 hr post-APAP) treatment with EGFRi. (A) Schematic showing experimental design. (B) Representative photomicrographs of H&E stained liver sections with necrotic area outlined, (C) serum ALT levels with percentage survival specified over bars and (D) percentage necrosis area based on H&E stained liver sections of mice treated with 300 mg/kg APAP followed by treatment with canertinib (80 mg/kg) or PBS, 12 hr post-APAP. All samples were collected and survival was recorded at 24 and 48 hr after APAP treatment (n=5-6). * indicate significant difference between groups at p<0.05. # indicates significant difference w.r.t. 24 hr time point within same treatment group.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Staining

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: Rapid and sustained activation of EGFR in mice and primary human hepatocytes after APAP-treatment. (A) Western blot analysis of phospho-EGFR (Tyr1068) and EGFR in total cell lysate and (B) ALT levels in serum, at various time points after administration of 300 mg/kg and 600 mg/kg APAP in mice (n=3-8). (C) Western blot analysis of phospho-EGFR and EGFR in total cell lysate and (E) ALT release in medium (represented as percentage of total ALT levels), at various time points after treatment of primary human hepatocytes (PHH) with 10 mM APAP. (D) Densitometric analysis showing activation of EGFR in PHH with data representing mean ± SEM of independent western blot analysis from 4 liver donors. * indicate significant difference between groups at p<0.05. # indicates significant difference w.r.t. 0 hr time point wherever indicated.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Activation Assay, Western Blot

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: Early treatment with EGFRi (1-h post-APAP) remarkably attenuated APAP-induced hepatotoxicity without altering APAP bioactivation and APAP-protein adducts formation. A, Western blot analysis of phospho-EGFR and EGFR in liver lysate, B, representative photomicrographs of H&E stained liver sections with necrotic area outlined, C, serum ALT levels, D, percentage necrosis area based on H&E stained liver sections, E, total glutathione levels in liver extract, and F, APAP-protein adducts levels in liver as measured by HPLC-ECD method. For all experiments mice (n = 3–12) were treated with 300 mg/kg APAP followed by canertinib (80 mg/kg) or PBS, 1-h post-APAP. All samples were collected at various time points up-to 24-h after APAP treatment. * indicate significant difference between groups at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Western Blot, Staining

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: JNK activation, its mitochondrial translocation and signaling through other protein kinases not altered by EGFRi. Western blot analysis of phospho-JNK and JNK in (A) total cell lysate and (B) mitochondrial fraction with densitometric analysis of mitochondrial activated JNK (p-JNK) shown in (C). D, Western blot analysis of RIP3 and RIP1 in total cell lysate. E, Western blot analysis showing PKC activation (studied using antibody against phosphorylated PKC-substrates in mitochondria) with its densitometric analysis shown in (F). All analysis were done on liver samples collected at various time points after treatment with 300 mg/kg APAP + PBS or APAP + canertinib (80 mg/kg, 1-h post-APAP) (n = 3–4). *indicate significant difference between groups at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Activation Assay, Translocation Assay, Western Blot

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: APAP caused rapid mitochondrial translocation of EGFR and mitochondrial dysfunction, which was inhibited by EGFRi. A, Western blot analysis of phospho-EGFR and EGFR in mitochondrial fraction at 1.5 h after administration of APAP (300 mg/kg) in mice pretreated (2-h prior to APAP) with canertinib (80 mg/kg). B-E, Oxygen consumption rate analysis in freshly isolated mitochondria from mice (n = 3) treated with APAP (300 mg/kg) + PBS, APAP (300 mg/kg) + canertinib (80 mg/kg) or saline (control), using Seahorse extracellular flux analyzer. B and C, Canertinib was administered 2hr before APAP and analysis was done 1.5 h after APAP treatment. D and E, Canertinib was administered 1-h post-APAP and analysis was done 3 h after APAP treatment. Oxidative phosphorylation was manipulated with injection of ADP, oligomycin (oligo), FCCP, and antimycin A (anti-A). Respiration was sequentially measured in a coupled state with substrate present (basal respiration), followed by State 3 (phosphorylating respiration, in the presence of ADP and substrate), State 4o (leak state induced with the addition of oligomycin - inhibitor of ATP synthase), and then maximal uncoupler (FCCP)-stimulated respiration (State 3u). At the end, antimycin A (complex III inhibitor) was added to inhibit mitochondrial respiration completely * indicate significant difference w.r.t. to APAP + PBS group at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Translocation Assay, Western Blot, Isolation, Saline, Control, Phospho-proteomics, Injection

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: Decreased oxidative stress, mitochondrial protein nitration and release of endonucleases from mitochondria by EGFRi. A, Oxidized glutathione levels in total liver extract. B, Representative photomicrographs of liver sections stained for nitrotyrosine-protein adducts. C, Western blot analysis of nitrotyrosine adducts in mitochondrial fraction with its densitometric analysis shown in (D). E, Western blot analysis of AIF, endonuclease G and SMAC in cytosolic fraction with densitometric analysis of AIF as shown in (F). All analysis were done on liver samples collected at various time points after treatment with APAP (300 mg/kg) + PBS or APAP (300 mg/kg) + canertinib (80 mg/kg, 1-h post-APAP) (n = 3–5). *indicate significant difference between groups at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Nitration, Staining, Western Blot

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: (A–C) Role of glutathione depletion in rapid activation of EGFR by APAP. (D–F) Enhanced protection against APAP hepatotoxicity by combination of NAC and EGFRi (administered 4-h post-APAP). (A) Total glutathione levels in liver extract and (B) western blot analysis of phospho-EGFR and EGFR in total cell lysate from liver samples obtained 2 h after treatment with Phorone (200 mg/kg in corn oil) or corn oil (control) in mice (n = 5). (C) Densitomertic analysis showing EGFR activation based on western blot image shown in (B). (D) Representative photomicrographs of H&E stained liver sections with necrotic area outlined, (E) serum ALT levels and (F) percentage necrosis area based on H&E stained liver sections of mice treated with 300 mg/kg APAP followed by canertinib (80 mg/kg), NAC (500 mg/kg), combination of canertinib (80 mg/kg) and NAC (500 mg/kg) or PBS (control), 4-h post-APAP (n = 5). Samples were collected 24 h after APAP treatment. *indicate significant difference w.r.t. control group at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Activation Assay, Western Blot, Control, Staining

Journal: Toxicological Sciences

Article Title: Dual Role of Epidermal Growth Factor Receptor in Liver Injury and Regeneration after Acetaminophen Overdose in Mice

doi: 10.1093/toxsci/kfw213

Figure Lengend Snippet: Impaired liver regeneration and cell cycle arrest by delayed (12-h post-APAP) treatment with EGFRi. A, Representative photomicrographs of PCNA stained liver sections with arrows indicating cells in S-phase with nuclear PCNA staining (brown). B, Total number of PCNA-positive cell per high power field (×40). C, Western blot analysis of cyclinD1, CDK4, phospho-Rb, and PCNA using total liver extract. Densitometric analysis of (D) cyclinD1, (E) CDK4, and (F) PCNA. Mice were treated with 300 mg/kg APAP. Canertinib (80 mg/kg) or PBS was administered 12-h post-APAP (n = 3–5). All samples were collected at 24 and 48 h after APAP treatment. * indicate significant difference between groups at P < .05.

Article Snippet: For studies with EGFR inhibitor (EGFRi), canertinib dihydrochloride salt (80 mg/kg, i.p.) (LC Laboratories, Woburn, Massachusetts) was dissolved in warm phosphate-buffered saline (PBS).

Techniques: Staining, Western Blot

Journal: Drug metabolism and drug interactions

Article Title: Inhibition of OATP-1B1 and OATP-1B3 by tyrosine kinase inhibitors

doi: 10.1515/dmdi-2014-0014

Figure Lengend Snippet: (A) Inhibitory potency of rifampicin toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of rifampicin (0.1–100 μM). Data are shown as mean ± SD. n = 4. (B) Inhibitory potency of pazopanib toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of pazopanib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (C) Inhibitory potency of nilotinib toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of nilotinib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (D) Inhibitory potency of vandetanib toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of vandetanib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (E) Inhibitory potency of canertinib toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of canertinib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (F) Inhibitory potency of erlotinib toward OATP-1B1. Intracellular accumulation of OATP-1B1 substrate [3H]ES in the presence of increasing concentrations of erlotinib (0.1–100 μM). Data are shown as mean ± SD. n = 4.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib, nilotinib and vandetanib were purchased from LC Laboratories (Woburn, MA, UDS).

Techniques:

Journal: Drug metabolism and drug interactions

Article Title: Inhibition of OATP-1B1 and OATP-1B3 by tyrosine kinase inhibitors

doi: 10.1515/dmdi-2014-0014

Figure Lengend Snippet: IC 50 of tested compounds toward OATP-1B1 and -1B3 transporter proteins.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib, nilotinib and vandetanib were purchased from LC Laboratories (Woburn, MA, UDS).

Techniques:

Journal: Drug metabolism and drug interactions

Article Title: Inhibition of OATP-1B1 and OATP-1B3 by tyrosine kinase inhibitors

doi: 10.1515/dmdi-2014-0014

Figure Lengend Snippet: (A) Inhibitory potency of rifampicin toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of rifampicin (0.1–100 μM). Data are shown as mean ± SD. n = 4. (B) Inhibitory potency of vandetanib toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of vandetanib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (C) Inhibitory potency of pazopanib toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of pazopanib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (D) Inhibitory potency of nilotinib toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of nilotinib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (E) Inhibitory potency of canertinib toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of canertinib (0.1–100 μM). Data are shown as mean ± SD. n = 4. (F) Inhibitory potency of erlotinib toward OATP-1B3. Intracellular accumulation of OATP-1B3 substrate [3H]CCK-8 in the presence of increasing concentrations of erlotinib (0.1–100 μM). Data are shown as mean ± SD. n = 4.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib, nilotinib and vandetanib were purchased from LC Laboratories (Woburn, MA, UDS).

Techniques: CCK-8 Assay

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Cellular accumulation of pazopanib (100 nM) in MDCKII cells. (A) Intracellular accumulation in WT (black bar) and MDR1 transfected cells (open bars) in the absence and presence of zosuquidar (1 μM), specific P-gp inhibitor and selected inhibitors canertinib (5 μM) and erlotinib (5 μM). (B) Cellular accumulation of pazopanib in Bcrp1 transfected cells (open bars) relative to parental WT cells (black bars) in the absence and presence of a specific Bcrp1 inhibitor Ko143 (200 nM) and canertinib (5 μM) and erlotinib (5 μM). Results are expressed as mean ± S.D., n = 4 (*p < 0.05).

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Transfection

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Transepithelial transport of pazopanib (5 μM) across MDCKII cell monolayers. Bi-directional transport was carried out in MDCKII parental, MDR1 (A) and murine Bcrp1 (B) transfected cell monolayers. Results are expressed as mean ± S.D., n = 3–4 wells.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Transfection

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Permeability values of pazopanib across MDCK-MDR1 cell monolayers.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Permeability

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Permeability values of pazopanib across MDCK-Bcrp1 cell monolayers.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Permeability

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Pazopanib brain concentration after a single intravenous dose of 5 mg/kg through tail vein in FVB-wild type mice at 60 min time point in the absence and presence of P-gp inhibitor LY335979 (zosuquidar, 25 mg/kg), Bcrp1 inhibitor Ko143 (10 mg/kg) and dual P-gp/Bcrp1 inhibitor elacridar (GF120918). Results are expressed as mean ± S.E.M., n = 3 (*p < 0.05).

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Concentration Assay

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Plasma and brain pharmacokinetic parameters obtained by noncompartmental analysis of the concentration-time profile data after an i.v. bolus dose of pazopanib in absence and presence of elacridar (GF120918) in FVB wild type mice.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Clinical Proteomics, Concentration Assay

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Influence of erlotinib and canertinib on plasma concentration profile of pazopanib after an i.v. bolus injection in FVB-wild type mice. Erlotinib and canertinib were administered intravenously 30 min prior to pazopanib through tail vein. Values are presented as mean ± S.E., n = 3.

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Clinical Proteomics, Concentration Assay, Injection

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Modulation of P-gp and Bcrp1 mediated efflux by erlotinib and canertinib: upper panels, brain concentration of pazopanib either alone or in the presence of elacridar, canertinib and erlotinib (10 mg/kg each) at t = 30 min (A) and t = 60 min (B) after an intravenous dose of 5 mg/kg pazopanib in FVB wild type mice. Lower panels, brain-plasma concentration ratio of pazopanib either alone or in the presence of elacridar, canertinib and erlotinib at t = 30 min (C) and t = 60 min (D) after an intravenous dose of 5 mg/kg in FVB wild type mice. Results are expressed as mean ± S.E., n = 3 (*p < 0.05 and **p < 0.01).

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Concentration Assay, Clinical Proteomics

Journal: International journal of pharmaceutics

Article Title: Enhanced brain accumulation of pazopanib by modulating P-gp and Bcrp1 mediated efflux with canertinib or erlotinib

doi: 10.1016/j.ijpharm.2012.05.038

Figure Lengend Snippet: Brain to plasma (Cb/Cp) concentration ratio of pazopanib at 15, 30, 60 and 120 min post dose in FVB wild type mice in the absence (vehicle treated, black bars) and presence of a P-gp and Bcrp1 dual inhibitor elacridar (gray bars). Results are expressed as mean ± S.E.M., n = 3 (*p < 0.05).

Article Snippet: Chemicals Pazopanib, erlotinib, canertinib and vandetanib were purchased from LC Laboratories (Woburn, MA).

Techniques: Clinical Proteomics, Concentration Assay