Journal: Clinical Cancer Research

Article Title: Targeting Neuropilin 1 as an Antitumor Strategy in Lung Cancer

doi: 10.1158/1078-0432.ccr-07-0001

Figure Lengend Snippet: Fig. 5. Cyclic 7-mer peptides bind NRP1and inhibit CL1-5 invasion and angiogenesis in vivo. A, the selected peptides reduce phosphorylation ofVEGFR2. HUVECs were pretreated with peptides for10 min followed by treatment withVEGF for 5 min. Phosphorylation ofVEGFR2 was determined byWestern blotting with an anti ^ phospho-VEGFR2 antibody.TotalVEGFR2 was determined byWestern blotting with an anti-VEGFR2 antibody. B, the effect of peptides on CL1-5 cells invasive activity. The invasive activity of cells was detected by invasion assay. CL1-5 cells (2.5 104) were seeded onTranswells coated with 30 Ag matrigel and incubated with peptides DG1or DG2 for 48 h.Then, the cells that had invaded the membrane were counted.Values were normalized to the relative invasion activity of the nontreated control cells. Experiments were done in triplicate, three independent times. In DG1- and DG2-treated cells, the invasion ability was statistically significantly different across the various concentrations of DG1or DG2 byANOVA (DG1, P < 0.001; DG2, P = 0.011). C, the effect of peptides on tumor angiogenesis in vivo. Immunohistochemical staining of the Matrigel plug sections with an anti-CD31antibody showed a significant decrease in CD31-positive vessels in plugs containing DG1peptide compared with mock-treated plugs. Original magnification, 200.The counts of microvessels surrounding the tumor nests were calculated. D, effect of peptides on tumorigenesis in vivo.Volumes of tumors from control CL1-5 cells (E) and DG1-treated cells (n) were measured at the indicated times as described in Materials and Methods. Means and 95% CI are shown (n = 5 mice per group). E, summary diagram showing thatVEGF165 can bind to NRP1and trigger the NRP1/VEGFR2/PI3K/Akt signaling pathways and result in tumor angiogenesis, cancer cell invasion, and tumorigenesis.The synthetic peptides DG1/DG2 can specifically block this signaling pathway and may have therapeutic potential.

Article Snippet: Human umbilical vascular endothelial cells (HUVEC) and culture media were purchased from Cell Applications, Inc.

Techniques: In Vivo, Phospho-proteomics, Activity Assay, Invasion Assay, Incubation, Membrane, Control, Immunohistochemical staining, Staining, Protein-Protein interactions, Blocking Assay

Journal:

Article Title: Development of a Frozen Cell Array as a High-Throughput Approach for Cell-Based Analysis

doi:

Figure Lengend Snippet: Binding of an anti-human Ep-CAM fluorescein-conjugated mAb to various cell lines analyzed by FACS. BKGEs (A2); human colorectal carcinoma cell lines COLO205 (A3), HCT116 (A4), and SW620 (A5); human prostate carcinoma cell lines PC3 (B1) and DU145 (B2); rat prostate tumorigenic cell line NRP154 (B3); HIAECs (B4); HUVECs (B5); lung HMVECs (C1); NHDFs (C2); human CASMCs (C3); neonatal NHEKs (C4); human malignant melanoma cell line A375 (C5); human breast carcinoma cell lines SkBr3 (D1), MCF7 (D2), and BT474 (D3); human rhabdomyosarcoma cell line A673 (D4); human liver carcinoma cell lines HepG2 (D5) and Hep3b (E1); T- and B-cell leukemia cell lines Jurkat (E2) and Bjab (E3), respectively; human lung carcinoma cell line SKMES1 (E4); and human neuroglioma cell line U87MG (E5) were incubated in the absence (thin line) or presence (thick line) of an anti-human Ep-CAM fluorescein-conjugated monoclonal antibody and analyzed by FACS.

Article Snippet: Normal cells included: human iliac artery endothelial cells (HIAECs), human umbilical vein endothelial cells (HUVECs), human microvascular endothelial cells from lung (HMVECs), normal human dermal fibroblasts (NHDFs), human coronary artery smooth muscle cells (CASMCs), and neonatal normal human epidermal keratinocytes (NHEKs) purchased from BioWhittaker/Clonetics (San Diego, CA) and the bovine kidney glomerular endothelial cell (BKGEs) purchased from VEC Technologies, Inc. (Rensselaer, NY, USA).

Techniques: Binding Assay, Incubation