her2 ecd fc protein  (Affibody)

Journal: iScience

Article Title: Identification of actionable targets for breast cancer intervention using a diversity outbred mouse model

doi: 10.1016/j.isci.2023.106320

Figure Lengend Snippet: TIEG1 regulates tumor immunity and tumor growth. Tumor growth was monitored using Kaplan-Meier tumor-free survival. p < 0.05 was considered significant. (A) C57BL/6 mice and C57BL/6 HER2 Tg mice with (n = 10) or without TIEG1 (n = 9) were electrovaccinated with pE2TM. (B and C) IFN-γ producing T cells were measured by ELISPOT assay. Data are represented as the average and SD in box and whisker plots. (D) Growth of implanted E0771/E2 tumor in HER2 Tg mice with or without TIEG1. (E) Onset age of spontaneous tumors in NeuT mice with or without TIEG1 (n = 9 and 7, respectively). (F) TIEG1 expression measured by qPCR. Data are represented as pooled averages +/− SEM. (G and H) TIEG1/Klf10 ( top panels ) and Cd3d (bottom panel) expression measured by scRNA sequencing.

Article Snippet: Recombinent HER2-ECD-Fc protein , SinoBiological , Cat#10004-H02H.

Techniques: Enzyme-linked Immunospot, Whisker Assay, Expressing, Sequencing

Journal: iScience

Article Title: Identification of actionable targets for breast cancer intervention using a diversity outbred mouse model

doi: 10.1016/j.isci.2023.106320

Figure Lengend Snippet:

Article Snippet: Recombinent HER2-ECD-Fc protein , SinoBiological , Cat#10004-H02H.

Techniques: Recombinant, Lysis, SYBR Green Assay, RNA Sequencing Assay, Sequencing, Single-cell Analysis, Knock-Out, Generated, Software, Enzyme-linked Immunospot

Journal: OncoTargets and therapy

Article Title: Identification of an Activating Mutation in the Extracellular Domain of HER2 Conferring Resistance to Pertuzumab

doi: 10.2147/OTT.S232912

Figure Lengend Snippet: The distinct binding epitopes of HER2-targeted monoclonal antibodies approved by the FDA. Trastuzumab binds to subdomain IV of the HER2 extracellular domain. Pertuzumab binds to an epitope in subdomain II, the dimerization domain of HER2.

Article Snippet: Cell culture supernatants containing the above fusion protein were collected at three days post transfection, and HER2-ECD-WT/S310F Fc fusion protein was purified by using a HiTrap Protein A column (GE Healthcare, PA, USA).

Techniques: Binding Assay

Journal: OncoTargets and therapy

Article Title: Identification of an Activating Mutation in the Extracellular Domain of HER2 Conferring Resistance to Pertuzumab

doi: 10.2147/OTT.S232912

Figure Lengend Snippet: The distribution and frequency of HER2 S310F mutation in tumours. ( A ) The distribution of recurrent mutations (more than five occurrences in an individual cancer type) in the extracellular domain of membrane proteins. The numbers mean the occurrences of each mutation in the same tumour. Different cancer types are represented by different colours. ( B ) The distribution of HER2 mutations based on COSMIC database. Abbreviations: BLCA, bladder urothelial carcinoma; CESC, cervical squamous cell carcinoma and endocervical adenocarcinoma; GBM, glioblastoma multiforme; LGG, brain lower grade glioma; SKCM, skin cutaneous melanoma; STAD, stomach adenocarcinoma; UCEC, uterine corpus endometrial carcinoma.

Article Snippet: Cell culture supernatants containing the above fusion protein were collected at three days post transfection, and HER2-ECD-WT/S310F Fc fusion protein was purified by using a HiTrap Protein A column (GE Healthcare, PA, USA).

Techniques: Mutagenesis

Journal: OncoTargets and therapy

Article Title: Identification of an Activating Mutation in the Extracellular Domain of HER2 Conferring Resistance to Pertuzumab

doi: 10.2147/OTT.S232912

Figure Lengend Snippet: Molecular modelling of the interaction between pertuzumab and HER2. ( A ) Structure visualization of the HER2-pertuzumab complex (PDB: 1S78), including the HER2 extracellular domain (bluish-green), pertuzumab’s light chain (purple) and heavy chain (green). The position of HER2 Ser310 was confirmed at the interface of HER2 and pertuzumab. Selected side chains from HER2 and pertuzumab are shown in stick representation, with carbons coloured by domain, nitrogens blue, oxygens red and hydrogens grey. The backbones are shown in ribbon representation. ( B ) Molecular modelling of pertuzumab bound to WT/S310F-mutant HER2. Hydrogen bonds are shown as red dashed lines. ( C ) The top 10 critical residues of the HER2-pertuzumab interaction predicted by the HawkDock server.

Article Snippet: Cell culture supernatants containing the above fusion protein were collected at three days post transfection, and HER2-ECD-WT/S310F Fc fusion protein was purified by using a HiTrap Protein A column (GE Healthcare, PA, USA).

Techniques: Mutagenesis

Journal: OncoTargets and therapy

Article Title: Identification of an Activating Mutation in the Extracellular Domain of HER2 Conferring Resistance to Pertuzumab

doi: 10.2147/OTT.S232912

Figure Lengend Snippet: The binding affinities of anti-HER2 antibodies for WT/S310F-mutant HER2. ( A ) The binding affinities of pertuzumab and trastuzumab for NIH3T3 cells expressing WT (HER2 WT) or S310F-mutant (HER2 S310F) HER2. NIH3T3 cells were used as a negative control. ( B ) The affinities of serial concentrations of pertuzumab and trastuzumab for WT (NIH3T3-HER2 WT) or S310F-mutant (NIH3T3-HER2 S310F) HER2 cells. ( C ) Comparisons of pertuzumab and trastuzumab interacting with WT (HER2-ECD-WT) or S310F-mutant (HER2-ECD-S310F) HER2 ECD protein by ELISA as described in the materials and methods. ( D ) The kinetic parameters for trastuzumab and pertuzumab derived from the fitted data models according to SPR analyses.

Article Snippet: Cell culture supernatants containing the above fusion protein were collected at three days post transfection, and HER2-ECD-WT/S310F Fc fusion protein was purified by using a HiTrap Protein A column (GE Healthcare, PA, USA).

Techniques: Binding Assay, Mutagenesis, Expressing, Negative Control, Enzyme-linked Immunosorbent Assay, Derivative Assay

Journal: OncoTargets and therapy

Article Title: Identification of an Activating Mutation in the Extracellular Domain of HER2 Conferring Resistance to Pertuzumab

doi: 10.2147/OTT.S232912

Figure Lengend Snippet: Effect of S310F mutation on the antitumour activity of pertuzumab. ( A ) NIH3T3 cells were used as the parental cell line and negative control, pCMV3-HER2 WT/S310F-mutant and pMH3-HER3 constructs were cotransfected into NIH3T3 cells. ( B ) Different ability of pertuzumab to inhibit HER2 phosphorylation signalling in WT/S310F-mutant HER2 cell lines after stimulation with or without HRG. NIH3T3 cells co-expressing WT/S310F HER2 and HER3 were incubated with culture medium or pertuzumab/trastuzumab for 2 h and then stimulated with or without HRG. Western blotting was performed using antibodies for the indicated proteins. ( C ) HER2 expression levels of MCF7 cells and MCF7 cells transfected with WT (MCF7-HER2 WT) or S310F-mutant (MCF7-HER2 S310F) HER2 gene, respectively. ( D ) Cell viability of MCF7, MCF7-HER2 WT and MCF7-HER2 S310F cell lines treated with culture medium (basal), HRG, HRG+pertuzumab or HRG+trastuzumab. Cell viability (%) is represented as the mean ± SD. One-way analysis of variance (ANOVA) was applied for statistical comparisons on multiple groups, ***p < 0.001. Abbreviation: NS, no significance.

Article Snippet: Cell culture supernatants containing the above fusion protein were collected at three days post transfection, and HER2-ECD-WT/S310F Fc fusion protein was purified by using a HiTrap Protein A column (GE Healthcare, PA, USA).

Techniques: Mutagenesis, Activity Assay, Negative Control, Construct, Expressing, Incubation, Western Blot, Transfection

Journal: Cancer cell

Article Title: Unbiased Combinatorial Screening Identifies a Bispecific IgG1 that Potently Inhibits HER3 Signaling via HER2-Guided Ligand Blockade.

doi: 10.1016/j.ccell.2018.04.003

Figure Lengend Snippet: Figure 3. Selectivity of PB4188 for HER2/HER3 Signaling (A–D) PathHunter assays to measure ligand-induced dimerization of EGFR/HER2 (A), HER2/HER4 (B), HER2/HER3 (C), and EGFR/HER3 (D). Cells were stim- ulated with an EC80 concentration of EGF (A) or HRG (B–D) and titrations of the indicated antibodies. (E) Human stem cell-derived cardiomyocytes were incubated with the indicated antibodies in combination with doxorubicin. Cell viability was determined by measuring cellular ATP. (F) SKBR-3 cells cultured in serum-free medium were supplemented with 12.5 nM HRG and the indicated antibodies. Twenty-four hours later, protein lysates were analyzed with PathScan array. (G) Phosphorylated (p prefixed) and total levels of signaling proteins after HRG stimulation (12.5 nM) in N87 cells analyzed by western blot. T, trastuzumab; P*, pertuzumab*; T + P*, equimolar combination of trastuzumab plus pertuzumab*. Data in (A–D) are represented as means ± SEM. Boxes in (E) show the middle quartile (25%–75%); horizontal bars represent the median; whiskers show the maximum and minimum values in the dataset.

Article Snippet: Recombinant Fc-tagged human EGFR, HER2, HER3 and HER4 extracellular domain (ECD) proteins were purchased from R&D Systems (Cat# 344-ER, 1129-ER, 348-RB, 1131-ER).

Techniques: Concentration Assay, Derivative Assay, Incubation, Cell Culture, Western Blot