Journal: International Journal of Biological Sciences

Article Title: Integrative analysis reveals a clinicogenomic landscape associated with liver metastasis and poor prognosis in hepatoid adenocarcinoma of the stomach

doi: 10.7150/ijbs.71449

Figure Lengend Snippet: Clinicopathological characteristics of HAS and stage-matched non-HAS patients

Article Snippet: Following that, the sections were incubated with anti-ERBB2 monoclonal primary antibody (Cell Signaling Technology, #2165), followed by a 30-min incubation with secondary antibody.

Techniques:

Journal: International Journal of Biological Sciences

Article Title: Integrative analysis reveals a clinicogenomic landscape associated with liver metastasis and poor prognosis in hepatoid adenocarcinoma of the stomach

doi: 10.7150/ijbs.71449

Figure Lengend Snippet: Potential targets and therapy for HAS. (A) Frequently altered genomic targets in HAS compared with non-HAS. (B) Comparison of ERBB2 expression between HAS and non-HAS. (C) Comparison of TMB between HAS and non-HAS. (D) Assessment of serum AFP and CEA levels in HAS patients before and after treatment. Three HAS patients received anti-ERBB2 therapy and five HAS patients received anti-PD-1 therapy. The line charts represent serum AFP/CEA level and histograms represent the CP of serum AFP/CEA level; details are shown as follows: CP (%) = (serum AFP or CEA levels before treatment - serum AFP or CEA levels after treatment) / serum AFP or CEA levels before treatment *100%. The values of 20 ng/mL and 5 ng/mL are defined as the serum AFP and CEA levels threshold, respectively. For each patient with elevated AFP/CEA level before treatment, 65% was defined as the cutoff for response to anti-ERBB2/anti-PD-1 therapy. Blue and red represent response and no response at the serum tumor biomarker levels, respectively. Grey means that this evaluation criterion is inapplicable for a patient with a normal AFP/CEA level before treatment. (E) Representative CT images of abdomen showing tumor lesions of two patients before and after treatment; one received anti-ERBB2 therapy and the other received anti-PD-1 therapy. The dotted line represents the metastatic liver lesion, and the arrow represents the primary gastric lesion. IHC, immunohistochemistry; AFP, alpha-fetoprotein; CEA, carcinoembryonic antigen; CP, change percent; Pt, patient.

Article Snippet: Following that, the sections were incubated with anti-ERBB2 monoclonal primary antibody (Cell Signaling Technology, #2165), followed by a 30-min incubation with secondary antibody.

Techniques: Expressing, Biomarker Assay, Immunohistochemistry

Journal: Cancers

Article Title: Quantifying the Effects of Combination Trastuzumab and Radiation Therapy in Human Epidermal Growth Factor Receptor 2-Positive Breast Cancer

doi: 10.3390/cancers14174234

Figure Lengend Snippet: HER2 quantification in BT474, SKBR3, and MDA-MB-231 cells. ( A ) Expression of HER2 protein (185 kDa) in each cell line. β-actin (42 kDa) was used as an internal control. Visible protein bands are seen in the BT474 and SKBR3 HER2+ cell lines, while MDA-MB-231 shows little to no expression. ( B ) HER2:β-actin ratio in each cell line. BT474 and SKBR3 cells have ratios of 1.41 and 1.46, respectively. MDA-MB-231 has a ratio of 0.08, confirming no HER2 amplification. The uncropped blots are shown in .

Article Snippet: The membrane was incubated in stripping buffer for 15 min, washed with TBST, and probed with Rabbit anti-human HER2/ErbB2 primary antibody (Cell Signaling Technology, Danvers, MA, USA) for 2 h at room temperature.

Techniques: Expressing, Amplification

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Analysis of HER2 expression level in breast cancer cells by flow cytometry and in vitro cell binding assay. ( A ) Flow cytometry in HER2-positive (JIMT-1) and negative (MDA-MB-231) breast cancer cells with pertuzumab. Gray-lined curve, isotype control; blue-lined curve, pertuzumab. ( B ) In vitro cell binding assay of 89 Zr-DFO-pertuzumab in breast cancer cells. ( C ) In vitro cell binding assay of 89 Zr-DFO-pertuzumab in JIMT-1 breast cancer cell by with or without (w/o) pre-treatment of trastuzumab (TRA) and herzuma (HER).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Expressing, Flow Cytometry, In Vitro, Cell Binding Assay

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Pharmacodynamic change of HER expression level by treatment of heat shock protein 90 inhibitor in JIMT-1 breast cancer cells. ( A ) Flow cytometric analysis of HER2 expression of JIMT-1. Pertuzumab used as the primary antibody and monoclonal anti-human FITC-conjugated IgG antibody as the secondary antibody. ( B ) In vitro cell binding assay of 89 Zr-DFO-pertuzumab. JIMT-1 cells were treated with various concentrations of heat shock protein 90 inhibitor (17-DMAG) for 24 h. ( C ) Correlation between flow cytometry analysis and cell binding assay. Correlation (r) = 0.9596.

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Expressing, In Vitro, Cell Binding Assay, Flow Cytometry

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Biodistribution of 89 Zr-DFO-pertuzumab in HER2-positive (JIMT-1) and -negative (MDA-MB-231) breast cancer xenograft models. 89 Zr-DFO-pertuzumab (1.6~1.8 MBq/50 μg/100 μL) was injected intravenously into ( A ) HER2-positive (JIMT-1) and ( B ) negative (MDA-MB-231) breast cancer xenograft models. Biodistribution was performed at 2 h, 1 day, 3 days, 5 days, and 7 days post-injection of 89 Zr-DFO-pertuzumab, and the amount of radioactivity of collected blood, organs, and tumor represented as percentage of the injected radioactivity dose/gram (%ID/g) was determined. Data were presented as mean ± sd ( n = 3).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Injection, Radioactivity

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Immuno-PET imaging of 89 Zr-DFO-pertuzumab in HER2-positive (JIMT-1) and -negative (MDA-MB-231) breast cancer xenograft models. PET images were acquired at 1 day, 5 days, and 7 days after injection of 89 Zr-DFO-pertuzumab in ( A ) HER2-positive (JIMT-1) and ( B ) -negative (MDA-MB-231) tumors. Tumors are indicated as yellow dotted circles. The tumor uptake of 89 Zr-DFO-pertuzumab was similar to that of the liver on day 1, but JIMT-1 tumor uptake increased, and liver uptake decreased in a time-dependent manner. At the same time, the uptake in the MDA-MB-231 tumor was relatively lower than that in the liver as time elapsed. All images were represented as the percentage of the injected radioactivity dose per gram of tissue (%ID/g).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Imaging, Injection, Radioactivity

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Immuno-PET imaging of 89 Zr-DFO-pertuzumab by trastuzumab treatment in JIMT-1 breast cancer xenograft model. ( A ) To evaluate the dynamics of HER2 expression level by treatment of isotype (ISO) and trastuzumab (TRA) using 89 Zr-DFO-pertuzumab, Immuno-PET imaging was performed in the JIMT-1 breast cancer xenograft model. Immuno-PET images were acquired 7 days after injection of 89 Zr-DFO-pertuzumab. ( B ) JIMT-1 tumor uptake of 89 Zr-DFO-pertuzumab as quantified using ASIPro display software. All images and quantitative data were expressed as a standardized uptake value (SUV).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Imaging, Expressing, Injection, Software

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Immuno-PET imaging of 89 Zr-DFO-pertuzumab by treatment of heat shock protein 90 inhibitor (17-DMAG) in JIMT-1 breast cancer xenograft model. ( A ) To evaluate the pharmacodynamics of HER2 expression level by treatment of 17-DMAG, Immuno-PET imaging was performed in the JIMT-1 breast cancer xenograft model. Mice were administered a total of 150 mg/kg of 17-DMAG over 24 h in three doses of 50 mg/kg each. PET images were acquired at 7 days post-injection of 89 Zr-DFO-pertuzumab. Immuno-PET tracer uptake was markedly reduced by treatment of 17-DMAG ( p < 0.0001). ( B ) JIMT-1 tumor uptake of 89 Zr-DFO-pertuzumab as quantified using ASIPro display software. All images and quantitative data were expressed as the standardized uptake value (SUV).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Imaging, Expressing, Injection, Software

Journal: Pharmaceutics

Article Title: Therapeutic Response Monitoring with 89 Zr-DFO-Pertuzumab in HER2-Positive and Trastuzumab-Resistant Breast Cancer Models

doi: 10.3390/pharmaceutics14071338

Figure Lengend Snippet: Western blot and immunofluorescence staining in JIMT-1 tumor treated with 17-DMAG. ( A ) Western blot analysis of HER2 expression in JIMT-1 tumor by treatment of 17-DMAG. JIMT-1 tumors were collected 7 days after 17-DMAG treatment. β -actin was used as an internal control. ( B ) Immunofluorescence staining of HER2 in control (CON, vehicle) and 17-DMA- treated JIMT-1 tumor tissues. Frozen section of JIMT-1 tumor was stained with CD31 (red), HER2 (green), and DAPI (blue). Images were acquired at 200× magnification. ( C ) Immunofluorescence staining of HER2 and CD31 was quantified using ImageJ software and quantitative data were expressed as relative fluorescence intensity (%). HER2 relative fluorescence intensity by 17-DMAG treatment was markedly reduced to 25% of control ( p < 0.0001).

Article Snippet: After gel transfer, the membrane was incubated with 5% ( w / v ) skim milk in TBS-T solution for blocking, gently shaken for 2 h at room temperature, and then treated with an anti-HER2 primary antibody (Cell signaling, Danvers, MA, USA) overnight at 4 °C.

Techniques: Western Blot, Immunofluorescence, Staining, Expressing, Software, Fluorescence

Journal: Advanced Science

Article Title: An Elaborate New Linker System Significantly Enhances the Efficacy of an HER2‐Antibody‐Drug Conjugate against Refractory HER2‐Positive Cancers

doi: 10.1002/advs.202102414

Figure Lengend Snippet: HER2 expression and cell cycle distribution in JIMT‐1 and N87 cells. a) HER2 expression in JIMT‐1 and N87 cells stained via ICC (red, Alexa 594‐stained HER2; blue, Hoechst‐stained nuclei, scale bar, 10 µm). Cell cycle distribution in b) JIMT‐1 and c) N87 cells treated with T‐DM1, LCB‐ADC1, or LCB‐ADC2 at 0.25 µg mL −1 for 72 h. Cells stained with propidium iodide (PI) were analyzed by flow cytometry.

Article Snippet: After thorough washing with PBS, the cells were incubated with anti‐HER2/ErbB2 rabbit polyclonal primary antibody (#2242S; Cell Signaling, Danvers, MA), anti‐ α ‐tubulin rabbit monoclonal primary antibody (#2125S; Cell Signaling), and Alexa 594 or 488 antirabbit IgG (Jackson Immuno Research, West Grove, PA).

Techniques: Expressing, Staining, Flow Cytometry

Journal: Advanced Science

Article Title: An Elaborate New Linker System Significantly Enhances the Efficacy of an HER2‐Antibody‐Drug Conjugate against Refractory HER2‐Positive Cancers

doi: 10.1002/advs.202102414

Figure Lengend Snippet: Therapeutic efficacy of LCB‐ADCs in HER2‐positive CDX models and microtubule interruption and mitotic arrest in CDX tumor tissue. a) Effect of LCB‐ADC1, LCB‐ADC2, T‐DM1 in JIMT‐1 cells. Drugs were i.v. injected at 2 mg kg −1 once (↑), n = 5. Data values are the mean ± standard deviation. *** p < 0.001 (T‐DM1 vs LCB‐ADC1 or 2). b) Effect of LCB‐ADC1, LCB‐ADC2, T‐DM1 in N87 cells. Drugs were i.v. injected at 2 mg kg −1 once (↑), n = 5. Data values are the mean ± standard deviation. *** p < 0.001 (T‐DM1 vs LCB‐ADC 2). HER2 expression in tumor tissue was detected by IHC (brown, DAB‐stained HER2; blue, hematoxylin‐stained nuclei, scale bar, 10 µm). c) In vivo microtubule interruption and mitotic arrest by LCB‐ADC1. Mice bearing N87 tumors were i.v. administered with T‐DM1, LCB‐ADC1 at 5 mg kg −1 once. After 10 d, the tumors were isolated, fixed, and stained with anti‐ α ‐tubulin (upper, blue, Hoechst‐stained nuclei; red, Alexa 594‐stained α ‐tubulin, scale bar; 10 µm) or antiphosphohistone H3 (PHH3) (lower, blue, hematoxylin‐stained nuclei; brown, 3,3′‐diaminobenzidine (DAB)‐stained PHH3, scale bar, 100 µm). d) Effect of LCB‐ADC1 in N87 cells. Drugs were i.v. injected at 0.2, 1 or 5 mg kg −1 , once a week for 4 weeks (↑), n = 8. Data values are the mean ± standard deviation. *** p < 0.001 (T‐DM1 vs LCB‐ADC1).

Article Snippet: After thorough washing with PBS, the cells were incubated with anti‐HER2/ErbB2 rabbit polyclonal primary antibody (#2242S; Cell Signaling, Danvers, MA), anti‐ α ‐tubulin rabbit monoclonal primary antibody (#2125S; Cell Signaling), and Alexa 594 or 488 antirabbit IgG (Jackson Immuno Research, West Grove, PA).

Techniques: Injection, Standard Deviation, Expressing, Staining, In Vivo, Isolation

Journal: Advanced Science

Article Title: An Elaborate New Linker System Significantly Enhances the Efficacy of an HER2‐Antibody‐Drug Conjugate against Refractory HER2‐Positive Cancers

doi: 10.1002/advs.202102414

Figure Lengend Snippet: Therapeutic efficacy of LCB‐ADCs in HER2‐positive GC PDX models. a) Effect of LCB‐ADC1, LCB‐ADC2, T‐DM1 in an HER2 +++ GC PDX model. The drugs were i.v. injected at 5 mg kg −1 once (↑), n = 5. b) Effect of LCB‐ADC1, LCB‐ADC3, T‐DM1 in an HER2 ++ GC PDX model. Drugs were i.v. injected at 5 mg kg −1 twice (↑), n = 5. Data values are a mean ± standard deviation. ** p < 0.01 (T‐DM1 vs LCB‐ADC3, single or repeated treatment). HER2 expression in tumor tissues was detected by IHC (brown, DAB‐stained HER2; blue, hematoxylin‐stained nuclei, scale bar, 10 µm). c,d) Mice bearing HER2 ++ GC patient‐derived tumors were i.v. administered with LCB‐ADC1, LCB‐ADC3, or T‐DM1 at 5 mg kg −1 , every other week for 4 weeks, n = 5. At the end of the experiment, c) tumors were isolated, photographed (scale bar, 1 cm) and d) weighed. Data values are mean ± standard deviation. * p < 0.05 (T‐DM1 vs LCB‐ADC1), ** p < 0.01 (T‐DM1 vs LCB‐ADC3).

Article Snippet: After thorough washing with PBS, the cells were incubated with anti‐HER2/ErbB2 rabbit polyclonal primary antibody (#2242S; Cell Signaling, Danvers, MA), anti‐ α ‐tubulin rabbit monoclonal primary antibody (#2125S; Cell Signaling), and Alexa 594 or 488 antirabbit IgG (Jackson Immuno Research, West Grove, PA).

Techniques: Injection, Standard Deviation, Expressing, Staining, Derivative Assay, Isolation

Journal: Advanced Science

Article Title: An Elaborate New Linker System Significantly Enhances the Efficacy of an HER2‐Antibody‐Drug Conjugate against Refractory HER2‐Positive Cancers

doi: 10.1002/advs.202102414

Figure Lengend Snippet: In vivo efficacy of LCB‐ADC3 at a low dose in an HER2 ++ GC PDX model. a) Tumor growth delay curves and b) survival rate when the tumor volume reached 1500 mm 3 . Mice bearing patient‐derived tumors were i.v. administered LCB‐ADC3 (0.5 or 1 mg kg −1 ) or T‐DM1 (4 mg kg −1 ), weekly for 4 weeks (↑), n = 5. Data values are the mean ± standard deviation. ** p < 0.01 (T‐DM1 vs LCB‐ADC3, 0.05 mg kg −1 ), *** p < 0.001 (T‐DM1 vs LCB‐ADC3, 1 mg kg −1 ).

Article Snippet: After thorough washing with PBS, the cells were incubated with anti‐HER2/ErbB2 rabbit polyclonal primary antibody (#2242S; Cell Signaling, Danvers, MA), anti‐ α ‐tubulin rabbit monoclonal primary antibody (#2125S; Cell Signaling), and Alexa 594 or 488 antirabbit IgG (Jackson Immuno Research, West Grove, PA).

Techniques: In Vivo, Derivative Assay, Standard Deviation

Journal: Advanced Science

Article Title: An Elaborate New Linker System Significantly Enhances the Efficacy of an HER2‐Antibody‐Drug Conjugate against Refractory HER2‐Positive Cancers

doi: 10.1002/advs.202102414

Figure Lengend Snippet: Mitotic arrest of HER2‐positive GC PDX tumor tissue. a,b) Mice bearing HER2‐positive PDX tumors were i.v. administered with a) LCB‐ADC2, b) LCB‐ADC3 or T‐DM1 at 5 mg kg −1 once, n = 5. After 1, 4, and 7 d, the tumors were isolated, fixed, and stained with antiphosphohistone H3 (PHH3) (blue, hematoxylin‐stained nuclei; brown, DAB‐stained PHH3, scale bar, 100 µm). Data values are a mean ± standard deviation. *** p < 0.001 (T‐DM1 vs LCB‐ADC2 or 3).

Article Snippet: After thorough washing with PBS, the cells were incubated with anti‐HER2/ErbB2 rabbit polyclonal primary antibody (#2242S; Cell Signaling, Danvers, MA), anti‐ α ‐tubulin rabbit monoclonal primary antibody (#2125S; Cell Signaling), and Alexa 594 or 488 antirabbit IgG (Jackson Immuno Research, West Grove, PA).

Techniques: Isolation, Staining, Standard Deviation

Journal: Scientific Reports

Article Title: PTEN is a predictive biomarker of trastuzumab resistance and prognostic factor in HER2-overexpressing gastroesophageal adenocarcinoma

doi: 10.1038/s41598-021-88331-3

Figure Lengend Snippet: Characteristics of patients and chemotherapy regimen with trastuzumab.

Article Snippet: Membranes were probed with specific primary antibodies against HER2 (polyclonal, #2242), PTEN (clone 138G6, #9559), pan-Akt (clone C67E7, #4691), phosphorylated Akt (Ser473, clone D9E, #4060), p44/p42 mitogen-activated protein kinase [MAPK; ERK1/2] (clone 137F5, #4695), phosphorylated p44/p42 MAPK (ERK1/2) (Thr202/Tyr204, clone D13.14.4E, #4370), S6 ribosomal protein (clone 5G10, #2217), phosphorylated S6 ribosomal protein (Ser235/236, clone D57.2.2E, #4858) (Cell Signaling Technology), and horseradish peroxidase (HRP)-conjugated secondary antibody (Dako, Santa Clara, CA).

Techniques:

Journal: Scientific Reports

Article Title: PTEN is a predictive biomarker of trastuzumab resistance and prognostic factor in HER2-overexpressing gastroesophageal adenocarcinoma

doi: 10.1038/s41598-021-88331-3

Figure Lengend Snippet: Univariate and multivariate analyses of overall survival progression free survival.

Article Snippet: Membranes were probed with specific primary antibodies against HER2 (polyclonal, #2242), PTEN (clone 138G6, #9559), pan-Akt (clone C67E7, #4691), phosphorylated Akt (Ser473, clone D9E, #4060), p44/p42 mitogen-activated protein kinase [MAPK; ERK1/2] (clone 137F5, #4695), phosphorylated p44/p42 MAPK (ERK1/2) (Thr202/Tyr204, clone D13.14.4E, #4370), S6 ribosomal protein (clone 5G10, #2217), phosphorylated S6 ribosomal protein (Ser235/236, clone D57.2.2E, #4858) (Cell Signaling Technology), and horseradish peroxidase (HRP)-conjugated secondary antibody (Dako, Santa Clara, CA).

Techniques: