Th17-related mRNA expression levels and Th17 cells in human glioma.

<p>(A) IL-17A mRNA levels are displayed as the percent change between the glioma group (Glioma) and normal human brain (Normal), normalized to GAPDH. (B) CD3ε mRNA levels are displayed as the percent change between the glioma groups (Glioma), astrocytoma (grade 2; Astro), oligodendroglioma (grade 3; Oligo), glioblastoma multiforme (GBM) and normal human brain (Normal), normalized to GAPDH. (C) IL-17A mRNA levels are displayed as the percent change between the glioma groups (Glioma), astrocytoma (grade 2; Astro), oligodendroglioma (grade 3; Oligo), glioblastoma multiforme (GBM) and normal human brain (Normal), normalized to CD3ε. (D) Analysis of GBM for CD4⁺IgG1⁺ (isotype control) or CD4⁺IL-17A⁺ cells. Flow cytometry data are representative of at least 2 independent GBM patient specimens. Bar heights represent means (±SEM).</p

Human primer sequences.

<p>The gene symbols used above indicate: IL-17A =  interleukin-17A; CD3e =  CD3 antigen, epsilon polypeptide; GAPDH  =  glyceraldehyde 3-phosphate dehydrogenase.</p

Mouse primer sequences.

<p>The gene symbols used above indicate: IL-17A  =  interleukin-17A; TGF-b  =  transforming growth factor-beta; IL-23 =  interleukin-23; IL-6 =  interleukin-6; CD3e =  CD3 antigen, epsilon polypeptide; GAPDH  =  glyceraldehyde 3-phosphate dehydrogenase.</p

Th17-related mRNA expression levels in mouse brain with or without a GL261 cell injection.

<p>(A) PCR products for IL-17A and GAPDH from control (Normal) mouse brain or 3 week post-operative wild-type (WT) mouse brain that received an intracranial (IC) injection of PBS, WT mouse brain that received an IC injection of 4×10⁵ GL261 cells (Glioma) or recombinase activating gene 1-deficient (Rag1^−/−) mouse brain that received an IC injection of 4×10⁵ GL261 cells (Rag1^−/−). Individual mice are differentiated by numbers 1–3. (B) IL-23 mRNA levels are displayed as the percent change between the experimental group (Exp.) and the mouse brain that received an IC injection of PBS (PBS), normalized to GAPDH. (C) TGF-β1 mRNA levels are displayed as the percent change between the experimental group (Exp.) and the mouse brain that received an IC injection of PBS (PBS), normalized to GAPDH. (D) IL-6 mRNA levels are displayed as the percent change between the experimental group (Exp.) and the mouse brain that received an IC injection of PBS (PBS), normalized to GAPDH. (E) CD3ε mRNA levels are displayed as the percent change between the experimental group (Exp.) and the mouse brain that received an IC injection of PBS (PBS), normalized to GAPDH. Bar heights represent means (±SEM). ND = not detectable.</p

Th17 cell phenotype in dLN and mouse brain with or without a GL261 cell injection.

<p>(A) Analysis of draining lymph nodes for CD4⁺IgG2A⁺ (isotype control) or CD4⁺IL-17A⁺ cells in mice that received an intracranial (IC) injection of PBS or 4×10⁵ GL261 cells (Glioma). (B) Analysis of brains for CD4⁺IgG2A⁺ (isotype control) or CD4⁺IL-17A⁺ cells in mice that received an IC injection of PBS or 4×10⁵ GL261 cells (Glioma). (C) Average CD4⁺IL-17A⁺ cell frequency in dLNs or brains (±SEM). (D) Analysis of brains for CD4⁺IL-17A⁺IgG1⁺ (isotype control) or CD4⁺IL-17A⁺IFN-γ⁺ cells (1^st row), CD4⁺IL-17A⁺IgG_2A⁺ (isotype control) or CD4⁺IL-17A⁺FoxP3⁺ cells (2^nd row) and CD4⁺IL-17A⁺IgG_2B⁺ (isotype control) or CD4⁺IL-17A⁺IL-4⁺ cells (4^th row) in mice that received an intracranial injection of PBS or 4×10⁵ GL261 cells (Glioma). (E) Average CD4⁺IL-17A⁺IFN-γ⁺, CD4⁺IL-17A⁺FoxP3⁺, and CD4⁺IL-17A⁺IL-4⁺ cell frequency in brains (±SEM). Data are representative of at least 2 independent experiments. * and ** denotes significant differences at <i>p</i>≤0.05 and <i>p</i>≤0.01, respectively.</p

A mouse brain that received an intracranial injection of GL261 cells.

<p>At three weeks post-operatively. (A) Hematoxylin and eosin staining of mouse glioma at 3 weeks post-operatively. Scale bar  = 1 mm. (B) GFAP (green), CD11b (red) and DAPI (blue) immunofluorescence along the border between mouse brain parenchyma and glioma. Scale bar  = 50 µM. (C) CD11c (green), CD11b (red), and DAPI (blue) immunofluorescence in the mouse glioma. Scale bar  = 50 µM. (D) CD4 (green) and FoxP3 (red) immunofluorescence in the mouse glioma. Tailed arrows indicate CD4⁺FoxP3⁻ cells, while untailed arrows indicate CD4⁺FoxP3⁺ T regulatory cells. Scale bar  = 25 µM.</p

Suppl Tables I and II from Development of a Function-Blocking Antibody Against Fibulin-3 as a Targeted Reagent for Glioblastoma

Supplemental Table I: Antibodies used for Western blotting and immunohistochemistry (IHC); Supplemental Table II: Sequences used for qRT-PCR primers.</p

Figures S1 to S8 from Development of a Function-Blocking Antibody Against Fibulin-3 as a Targeted Reagent for Glioblastoma

Figure S1: Generation of mAb428.2 antibody; Figure S2: Affinity of mAb428.2 for fibulin-3; Figure S3: mAb428.2 binds to cell-surface and interstitial fibulin-3; Figure S4: mAb428.2 does not cause acute toxicity; Figure S5: Locally-delivered mAb428.2 targets fibulin-3 signaling and tumor growth; Figure S6: IV-delivered mAb428.2 inhibits tumor growth; Figure S7: Effects of mAb428.2 against intracranial tumors; Figure S8: Biodistribution of mAb428.2 in naïve and tumor-bearing mice.</p

Legends of Figs S1 to S8 unmarked from Development of a Function-Blocking Antibody Against Fibulin-3 as a Targeted Reagent for Glioblastoma

Legends of Figs S1 to S8, unmarked version</p

Supplementary Figure 1 from Suppression of Human Glioma Xenografts with Second-Generation IL13R-Specific Chimeric Antigen Receptor–Modified T Cells

PDF file, 136K, Bar graph representing IL2 secretion by untransduced or IL13-CAR transduced T cells upon co-culturing.</p