anti-cd47 neutralizing monoclonal antibody  (Bio X Cell)

Journal: Science Advances

Article Title: Zeb1 induces immune checkpoints to form an immunosuppressive envelope around invading cancer cells

doi: 10.1126/sciadv.abd7455

Figure Lengend Snippet: ( A ) H&E at P190 showing clusters of invading cells (arrow). ( B to B″ ) Zeb1 and PDL1 on an invading cluster. Arrows show the same position. ( C ) CD8 cells are excluded from invading clusters (arrow). ( D to D″ ) CD47 and Zeb1 on invading clusters. White arrows show the same position. ( E ) iNos + cells are excluded from invading clusters. ( F to F″ ) Arg1 + cells are closely apposed to Zeb1 hi cells in invading clusters. ( G ) CD8 cells and iNos+ cells surrounding tumors. ( H ) Microscopic views (~200 μm in diameter) were counted for three invasive clusters in three tumors (region 2). Regions at the edge of the tumor 300 μm on either side of an invading cluster were counted and averaged (region 1), as were regions in the central tumor (C). Error bars are SDs. Scale bars, 100 μm. ( I ) Real-time PCR. Primary cells cultured from K-Ras–driven mouse tumors (393P), and cells overexpressing Zeb1 and where Zeb1 was knocked down ( , , ). ( J ) Chromatin immunoprecipitation (ChIP) showing Zeb1 binding to the promoters of CD47 and PDL1 (see fig. S4). “Input” indicates a 1:10 dilution of chromatin before immunoprecipitation. Results are representative of three separate experiments.

Article Snippet: Cells were cocultured in the absence or presence of neutralizing anti-CD47 monoclonal antibody (mAb, 10 μg/ml) or control mAb (BioXCell) for 48 hours and then immunostained as described.

Techniques: Real-time Polymerase Chain Reaction, Cell Culture, Chromatin Immunoprecipitation, Binding Assay, Immunoprecipitation

Journal: Science Advances

Article Title: Zeb1 induces immune checkpoints to form an immunosuppressive envelope around invading cancer cells

doi: 10.1126/sciadv.abd7455

Figure Lengend Snippet: ( A to B″ ) Zeb1 + and PDL1 + cells at the IF and invading lung parenchyma (PI) at P220. ( C ) CD8 cells surrounding tumors at P220. ( D and D′ ) CD8 cells at the IF and PI, but not C. ( E ) PDL1 in a Zeb1 mutant background at P220. ( F and G ) CD8 + and PD1 + cells in Zeb1 mutant tumors at P220. ( H to I′ ) CD47 and Zeb1 on cells at the IF and PI. ( J ) Arg1 + cells surrounding tumors at P220. ( K and K′ ) Arg1 + cells adjacent to Zeb1 hi tumor cells (arrows). ( L to L″ ) Close proximity of CD47 + and Arg1 + cells at the IF at P220. ( M ) iNos + cells in normal lung surrounding tumors and airway infiltrate at P220. ( N ) Few iNos + cells are evident at the IF at P220. ( O ) iNos + cells in tumors in a Zeb1 mutant background. ( P to P‴ ) Most CD68 + macrophages in Zeb1 mutant tumors express iNos. ( Q ) Few Arg1 + cells are evident in Zeb1 mutant tumors. ( R ) Effect of Zeb1 on cells in the C, IF, and PI regions of tumors. Regions were identified and counted as in . Error bars are SDs. Scale bars, 100 μm. nd, not detected.

Article Snippet: Cells were cocultured in the absence or presence of neutralizing anti-CD47 monoclonal antibody (mAb, 10 μg/ml) or control mAb (BioXCell) for 48 hours and then immunostained as described.

Techniques: Mutagenesis

Journal: Science Advances

Article Title: Zeb1 induces immune checkpoints to form an immunosuppressive envelope around invading cancer cells

doi: 10.1126/sciadv.abd7455

Figure Lengend Snippet: ( A ) H&E section showing invasion of an AW at P220. ( B to B⁗ ) Coinduction of Zeb1 and CD47 at sites of AW invasion. ( C to E″ ) Close proximity of cells expressing CD47 and Arg1 at sites of AW invasion. ( F ) Close proximity of Zeb1 hi AC and Arg1 + macrophages at sites of AW invasion. ( G ) Quantification of cells expressing Zeb1, CD47, iNos, and Arg1 within AWs and surrounding AWs. Immunostaining in Zeb1 mutant lesions and for iNos is not shown. Three representative microscopic views (~200 μm in diameter) were counted for each region within a single tumor. In addition, results from three tumors from different lungs were averaged. As noted above, Zeb1 mutation prevents AD-to-AC transition, and persisting AD do not show evidence of AW invasion or induction of CD47 around AWs. In addition, note Zeb1 is not expressed in AD cells. Error bars are SDs. Scale bars, (A to D‴ and F) 100 μm and (E to E″) 25 μm.

Article Snippet: Cells were cocultured in the absence or presence of neutralizing anti-CD47 monoclonal antibody (mAb, 10 μg/ml) or control mAb (BioXCell) for 48 hours and then immunostained as described.

Techniques: Expressing, Immunostaining, Mutagenesis

Journal: Science Advances

Article Title: Zeb1 induces immune checkpoints to form an immunosuppressive envelope around invading cancer cells

doi: 10.1126/sciadv.abd7455

Figure Lengend Snippet: ( A ) Bone marrow macrophage (BMM) was cocultured with 393-P cells. Cells were treated with IL-4 to drive M2-like polarization. mRNA levels were quantified by real time PCR. ( B and C ) Cocultures were immunostained for Arg1 or CD206. White arrows show larger 393-P nuclei and yellow arrows show smaller BMM nuclei. ( D and E ) Arg1 and CD206 in BMM treated with IL4. ( F and G ) Cocultures in the presence of CD47 blocking antibody (10 μg/ml). ( H and I″ ) Nomarski images and immunostaining of adjacent macrophages and cancer cells. ( J ) Quantification of results in (A) to (I). Error bars are SDs. ( K ) Real-time PCR showing that IL-4 induces PDL1 mRNA in BMM. Three experiments are shown. ( L to M″ ) Arg1 + and PDL1 + cells at the IF and iNos + and PDL1 − cells in the central region of a mouse lung tumor at P220 (C). ( N ) Quantification of cells in (L) to (M″). Four tumors were analyzed. Scale bars, (H to I″) 10 μm and (other panels) 50 μm. ( O ) Left: A mouse lung tumor is shown with invading cells expressing the mesenchymal marker Col1a2 (green). Right: The image was colorized with red depicting inflammatory and green immunosuppressive areas.

Article Snippet: Cells were cocultured in the absence or presence of neutralizing anti-CD47 monoclonal antibody (mAb, 10 μg/ml) or control mAb (BioXCell) for 48 hours and then immunostained as described.

Techniques: Real-time Polymerase Chain Reaction, Blocking Assay, Immunostaining, Expressing, Marker