Journal: Cancers

Article Title: Drug-Induced Resistance and Phenotypic Switch in Triple-Negative Breast Cancer Can Be Controlled via Resolution and Targeting of Individualized Signaling Signatures

doi: 10.3390/cancers13195009

Figure Lengend Snippet: TNBC tissues are represented by different patient-specific signaling signatures, majority of which do not include EGFR. ( A ) Fold changes in expression levels of EGFR and pEGFR in TNBC and non-TNBC tumors are shown. Y1068 and Y1173 are EGFR phosphorylation sites; ( B ) Examples for patient-specific sets of active unbalanced processes are shown. Each sample harbors a set of 1–3 active unbalanced processes (PaSSS), represented schematically by a barcode. In each barcode active unbalanced processes are represented by black or gray squares, inactive white. Negative/positive amplitude denotes how the patients are correlated with respect to a particular process. Suggested PaSSS-based therapies appear below each barcode; ( C ) Heterogeneity index of 3 subgroups, represented by a ratio between the number of distinct PaSSSs and the number of samples in each subset, is shown for the TNBC subset of tissues, the entire set (3467 samples from 11 cancer types) and the subset of non-TNBC samples. (Abbreviations: TNBC—Triple Negative Breast Cancer, PaSSS—Patient-specific signaling signature, EGFR—Epidermal Growth Factor Receptor, VEGFR2—Vascular Endothelial Growth Factor Receptor 2, Her2—Human Epidermal growth factor Receptor 2, Src—Proto-oncogene tyrosine-protein kinase Src).

Article Snippet: The following conjugated antibodies were used: anti-p-EGFR (Y1068) (R&D Systems, Minneapolis, MN, USA, cat. no. IC3570G), anti-p-ERK2 (Thr202/Tyr204) (BioLegend, San Diego, CA, USA, cat. No. 675503), anti-p-S6 (Ser235/236) (BioLegend, cat. no. 608605), and anti-GAPDH (Santa Cruz Biotechnology, Dallas, Texas, USA, cat. no. sc-47724AF594).

Techniques: Expressing, Phospho-proteomics

Journal: JCI Insight

Article Title: Interim report on engineered NK cell trial in lung cancer refractory to immune checkpoint inhibitors

doi: 10.1172/jci.insight.186890

Figure Lengend Snippet: Utilizing the multicolor immunofluorescence histochemical staining, we examined lung tumor tissue from patient 006 on 3 occasions using DAPI (blue) for nuclear staining, pan-CK (gray) for lung parenchyma, CD45 (purple) for lymphocytes, CD57 (cyan) and CD56 (green) for NK cells, CD3 (yellow) for T cells, and EGFR (red) combined with CD56 or CD57 to identify NK cells transduced with the tEGFR-sIL15 construct. ( A ) FFPE tissue from a pretreatment tumor biopsy from patient 006 shows the presence of lung parenchyma (DAPI + pan-CK + ) but the absence of tEGFR + (red) transduced CD57 + or CD56 + NK cells. ( B ) FFPE tissue from the skinny needle tumor biopsy collected 24 hours following the fourth and final infusion of sIL15_TRACK NK cells without observable of tEGFR + (red) transduced CD57 + (cyan) or CD56 + (green) cells. ( C ) FFPE tissue from an excisional tumor biopsy collected 7 days following the fourth and final infusion of sIL15_TRACK NK cells shows sIL15_TRACK NK cells identified in DAPI + CD45 + cells by coexpression of tEGFR (red) with CD57 (cyan), as marked by white arrows. ( D ) Selected individual markers of the boxed region shown in C are shown. Scale bars: 10 μm ( A – C ); 10 μm ( D ).

Article Snippet: For assessment of tumor infiltration of NK cells, primary antibodies were as follows: EGFR (clone 31G7, Novus), CD3 (clone LN10, Leica), CD56 (clone MRQ-42, Cell Marque), CD45 (clone D9M81, Cell Signaling Technologies), CD57 (clone HNK-1, Biolegend), and pan-cytokeratin (clone AE1/AE3, Dako).

Techniques: Immunofluorescence, Staining, Transduction, Construct

Journal: Frontiers in Cell and Developmental Biology

Article Title: BML-265 and Tyrphostin AG1478 Disperse the Golgi Apparatus and Abolish Protein Transport in Human Cells

doi: 10.3389/fcell.2019.00232

Figure Lengend Snippet: BML-265 and Tyrphostin AG1478 inhibit the transport of secretory proteins. HeLa cells transiently expressing Str-KDEL_TNF-SBP-EGFP (A) , SBP-EGFP-GPI (B) or SBP-EGFP-EGFR (C) were pre-treated with the indicated molecules at 10 μM for 1 h. Trafficking of the reporters was then induced by incubation with biotin for 1 h. The presence of the GFP-tagged reporters (green, upper panel) at the plasma membrane was then detected using an anti-GFP antibody on non-permeabilized cells (red, upper panel). The Golgi apparatus was visualized using immunostaining against GM130 (bottom panel). Scale bar: 10 μm.

Article Snippet: Antibodies to detect phosphorylated EGFR and total EGFR were purchased from Cell Signaling Technology (PhosphoPlus EGFR (Tyr1068) Antibody Duet, catalog number 11862S, dilution 1:1000).

Techniques: Expressing, Incubation, Immunostaining

Journal: Cancer letters

Article Title: Isthmin-1 promotes growth and progression of colorectal cancer through the interaction with EGFR and YBX-1.

doi: 10.1016/j.canlet.2024.216868

Figure Lengend Snippet: Fig. 7. Hypoxia elevates ISM1 expression in colon cancer. (A) Colon cancer gene expression data from the TCGA database were grouped by ISM1 expression to perform GSEA analysis. (B) Representative images of HE staining and IHC staining for ISM1 and HIF-1α expression in human colon cancerous tissues. Scale bar = 100 μm. (C) Western blot assay of ISM1 and HIF-1α protein levels in HT-29 cells under physical hypoxia for different period. Densitometry was performed for quanti fication (Right). (D) Effect of HIF-1α inhibitor (BAY87-2243, 10 μM) on ISM1 expression in HT-29 cells under hypoxia conditions. Densitometry was performed for quantification (Right). (E) Plasmids were used to overexpress HIF-1α (OE-HIF-1α) to observe the change of ISM1 expression. Densitometry was performed for quantification (Right). (F) The transcription binding sites of HIF-1α in the promoter region of ISM1 and the positions of the corresponding primers. (G) Dual luciferase reporter gene assay was used to evaluate the effect of HIF-1α on the ISM1 promoter. (H) Enrichment of HIF-1α on the ISM1 promoter as detected by ChIP under hypoxia. (I) The concentration of ISM1 in condition medium (CM) of HT-29 cells under physical hypoxia for different period by using the ELISA kit (n = 3). (J) HT-29 cells without hypoxia were stimulated with the conditional medium (CM) of HT-29 cells after 24 h hypoxia, and then the cells were collected at the following timepoints. EGFR phosphorylation level was detected by western blot assay. Densitometry was performed for quantification (Right). *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Antibodies against YBX1 (20339- 1-AP), EGFR (66455-1-Ig), HIF-1α (20960-1-AP), GAPDH (HRP-60004) and Tubulin (HRP-66031) were from Proteintech.

Techniques: Expressing, Gene Expression, Staining, Immunohistochemistry, Western Blot, Binding Assay, Luciferase, Reporter Gene Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Phospho-proteomics

Journal: Cancer letters

Article Title: Isthmin-1 promotes growth and progression of colorectal cancer through the interaction with EGFR and YBX-1.

doi: 10.1016/j.canlet.2024.216868

Figure Lengend Snippet: Fig. 8. Schematic depiction of ISM1 is transcriptionally regulated by HIF-1α and acts as a novel ligand of EGFR, promoting the progression and metastasis of colorectal cancer via the activation of EGFR signaling pathway.

Article Snippet: Antibodies against YBX1 (20339- 1-AP), EGFR (66455-1-Ig), HIF-1α (20960-1-AP), GAPDH (HRP-60004) and Tubulin (HRP-66031) were from Proteintech.

Techniques: Activation Assay