human edil3 elisa kit  (R&D Systems)

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: Humoral immune responses elicited to EDIL3 were associated with clinical outcomes in patients with metastatic melanoma receiving ipilimumab plus bevacizumab (Ipi-Bev). A, EDIL3-specific antibody FC measured by ELISA in posttreatment versus pretreatment plasma samples of 42 Ipi-Bev patients. CR/PR (green); SD, stable disease (blue); and PD, progressive disease (red). FC ≥1.5 considered clinically significant. B, Frequencies of EDIL3-specific antibody (FC ≥1.5) by clinical responses. C, Immunoblot analysis of EDIL3-specific Ig expression in pretreatment (Pre) and posttreatment (Post) patient's plasma samples of responders. Polyclonal anti-EDIL3 (Abcam) was used in the first lane. Densitometric quantification analysis for relative band intensities shown. D, Kaplan–Meier survival analyses for patients with FC ≥1.5 or <1.5 for EDIL3-specific antibody to Ipi-Bev treatment. E, Percentage of patients with EDIL3-specific antibody FC ≥1.5 in patient cohorts treated with Ipi-Bev ( n = 42), ipilimumab ( n = 34), PD-1 blockade ( n = 21), and nivolumab-ipilimumab ( n = 41) therapy.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot, Expressing

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: EDIL3 expression is associated with tumor T-cell immune exclusion gene signatures by TIDE analyses. A, Heat maps showing the correlation (Kendall rank) between EDIL3 , MFGE8 and CTNNB1 genes expression and TIDE-predicted signatures for CTL exclusion and dysfunction respectively within TCGA tumor types. B, EDIL3 , MFGE8 , and CTNNB1 log 2 (TPM+1) expression and TIDE-predicted value for immunotherapy response in TCGA-SKCM dataset with EDIL3 expression significantly higher among predicted nonresponders ( P = 7.4e-11, two-sided Mann–Whitney U test). In the box and whisker plots, the box delineates the first and third quartiles and is bisected by the median; whiskers extend to a maximum of 1.5 times the interquartile range.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Expressing, MANN-WHITNEY, Whisker Assay

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: EDIL3 expression associated with TGFβ signaling, EMT, and angiogenesis signatures. A and B, EDIL3 expression correlated with core biological pathways. EDIL3 expression ordered from low (left) to high (right). Rows display gene expression (z scores normalized) for genes grouped under pathway expression for TGFβ signaling, TGFβ signaling in fibroblast, EMT and angiogenesis signatures in TCGA-SKCM ( n = 469; A ) and CheckMate 064 ( n = 90; B ) databases. C, EDIL3 expression correlated with CAF FAP signature score using TIDE.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Expressing

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: EDIL3 is abundantly expressed in CAFs and is upregulated by TGFβ. A, Detection by ELISA of secreted EDIL3 in conditioned medium from NF and patient-derived CAFs (P4-CAF and CAF2). B and C, NF pretreated with or without LY2109761 (LY) followed by TGFβ treatment for 24 hours. EDIL3 expression was examined by B, ELISA of conditioned medium and C, immunoblot analyses of whole-cell lysates. D, qRT-PCR analysis of EDIL3 silencing by siRNA in P4-CAF. EDIL3 mediated regulation of TGFβ target genes Transgelin ( TAGLN; E ) and α-SMA ( ACTA2; F ). G, Immunoblot analysis of EDIL3 expression in P4-CAFs treated with VEGF-A (20 ng/mL) or/and bevacizumab (25 μg/mL) for 24 hours. β-actin was used as a loading control. All results are presented as means ± SD and represent three independent experiments. Statistical significance was determined by t test and indicated by P values or as **, P < 0.01; ****, P < 0.0001.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Derivative Assay, Expressing, Western Blot, Quantitative RT-PCR, Control

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: Circulating EDIL3 expression correlates with serum VEGF levels and angiogenesis signatures in TCGA-SKCM and CheckMate 064 databases. A, Spearman rank correlation analysis of pretreatment circulating serum levels of EDIL3 versus VEGF-A in Ipi-Bev–treated patients with melanoma ( n = 39). B, EDIL3 log 2 (TPM+1) transformed expression is significantly associated with high angiogenic signatures (Angio) in TCGA-SKCM and CheckMate 064 datasets (two-sided Mann–Whitney U test). C and D, EDIL3 (50 ng/mL) promoted migration of patient-derived endothelial cells similar to VEGF-A (20 ng/mL) assessed by wound healing assay. E–G, EDIL3 promoted tube formation ability of patient-derived endothelial cells similar to VEGF-A. All results are presented as means ± SD and represent three independent experiments. Statistical significance indicated by P values or as *, P < 0.05.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Expressing, Transformation Assay, MANN-WHITNEY, Migration, Derivative Assay, Wound Healing Assay

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: EDIL3 blocks immune–endothelial cell adhesion and inhibits T-cell migration. A, Expression of LFA-1(CD11a) on immune cells by flow cytometry. B, Schematic representation of immune–endothelial cell adhesion assay showing EDIL3 antagonizes LFA-1/ICAM-1–dependent adhesion. Graphics created with Biorender. C, rEDIL3 (10, 50, and 200 ng/mL) inhibits adhesion of activated T cells to TNFα-activated tumor endothelial cell's monolayer. D, rEDIL3 (50 ng/mL) blocks the immune–endothelial cell adhesion by interfering with the LFA-1 and ICAM-1 interaction. E, EDIL3 silencing in endothelial cells potentiated T-cell transendothelial migration. F, rEDIL3 partially blocked T-cell transendothelial migration induced by IP-10. All results are presented as means ± SD and represent three independent experiments. Statistical significance was determined by t test and indicated by P values or as *, P < 0.05; **, P < 0.01.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Migration, Expressing, Flow Cytometry, Endothelial Cell Adhesion Assay

Journal: Cancer Immunology Research

Article Title: EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-23-0171

Figure Lengend Snippet: EDIL3 blocks transendothelial CD8 + T-cell migration in microfluidic 3D cocultures. Representative images and quantification of migrated CD8 + T-cell numbers in adjacent extracellular matrix (region of interest) after 48 hours of coculture with, vasculature from patient-derived endothelial cells in the presence of the chemoattractant IP-10 ( A ) or tumor spheroids as the source of IP-10 with vasculature from patient-derived endothelial cells ( B ). rEDIL3 (50 ng/mL) pretreatment of CD8 + T cells blocked their transendothelial migration. All results are presented as means ± SD and represent three independent experiments. Statistical significance was determined by t test and indicated by P values or as **, P < 0.01; ****, P < 0.0001. C, Model of effects for anti-EDIL3 humoral responses in patients with advanced cancer: Tumor-derived EDIL3 mediates T-cell exclusion in the TME. EDIL3 binds to LFA-1 on T cells and prevents their adherence via ICAM-1 on TECs, thus impeding immune cell extravasation into the TME. ICB may augment a humoral response against EDIL3 accompanied by activated tumor endothelium and increased CD8 + T-cell infiltration. Patients with melanoma with anti-EDIL3 humoral immune responses demonstrated improved therapeutic response to treatment. In addition, immunosuppressive CAFs were identified as an alternate source for EDIL3 in the TME. Graphics created with Biorender.

Article Snippet: Secreted EGF-like repeats and discoidin I–like domains protein 3 (EDIL3) levels in patient plasma as well as in conditioned medium from cell culture were determined in duplicate using a human EDIL3 ELISA kit (R&D Systems; #DY6046-05), according to the manufacturer's instructions.

Techniques: Migration, Derivative Assay