invivomab anti bioxcell be0001 2 cd3 human cd3  (Bio X Cell)

Journal: Science (New York, N.Y.)

Article Title: Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

doi: 10.1126/science.adx9954

Figure Lengend Snippet: ( A-B ) IL-2, IL-21, and IL-2/21-Trikine pSTAT signaling schematic (A) and dose-response on mouse CD8+ T cells (B). ( C-D ) IL-21, IL-2, and IL-21/2-Trikine pSTAT signaling schematic (C) and dose-response on mouse CD8+ T cells (D). ( E-F ) IL-10, IL-2, and IL-10/2-Trikine pSTAT signaling schematic (E) and dose-response on mouse CD8+ T cells (F). Values in (B, D, F) represent the percentage of CD8+ T cells that are pSTAT5+ (left) or pSTAT3+ (right). Errors bars indicate mean ± standard deviation (SD) of duplicate or triplicate wells. ( G ) The ratios of pSTAT3+ to pSTAT5+ CD8+ T cells following stimulation by IL-2/21-, IL-21/2-, or IL-10/2-Trikine for 15 minutes at Emax. ( H-L ) Pmel CD8+ T cells were activated with gp100 for 5 days and then starved overnight. Cells were then restimulated with 10 ng/mL anti-CD3 antibody and 10 nM indicated Trikines for 48 hours. MFI of IL7Ra (H), CD62L (I), PD-1 (J) and TIM-3 (K). CD8+ cell count (L). Results display 4 technical replicates and are representative of 2 independent experiments (B, D, F, G-L).

Article Snippet: For human T cell activation, cells were activated with plate-bound anti-human CD3ε (1 μg ml −1 , clone OKT-3, BioXCell) and soluble anti-human CD28 (5 μg ml −1 , clone 9.3, BioXCell).

Techniques: Standard Deviation, Cell Characterization

Journal: Science (New York, N.Y.)

Article Title: Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

doi: 10.1126/science.adx9954

Figure Lengend Snippet: ( A ) Schematic of IL-2, IL-21, and human-reactive IL-2/21-Trikine pSTAT signaling. ( B ) Dose-dependent pSTAT signaling of IL-2, IL-21, and human-reactive IL-2/21-Trikine on human CD8+ T cells. Errors bars indicate mean ± standard deviation (SD) of duplicate or triplicate wells. MFI, mean fluorescence intensity. Results are representative of 2 independent experiments. ( C ) Diagram of testing for human IL-2/21-Trikine signal on γ c knock-out YT-1 cells (left) and flow cytometry staining for γ c WT and knock-out YT-1 cells (right). ( D ) Human-reactive IL-2/21-Trikine pSTAT signaling in WT and γ c knock-out YT-1 cells. Errors bars indicate mean ± standard deviation (SD) of duplicate or triplicate wells. Results are representative of 2 independent experiments. ( E ) Schematic of in vitro repeated antigen challenge (RAC) experiment (top left) and target cell killing as measured by IncuCyte after 11 rounds of RAC (right). Results are representative of 2 independent experiments. Error bars represent mean ± SD. ( F ) Schematic of human melanoma TIL experiment. ( G-I ) CD8+ T cell counts (G) , percentage of CD8+ T cells with TEM or Teff phenotype (H), and percentage of CD3+CD8+ cells that are LAG-3+ (I) from human melanoma TILs treated with sIL-2, sIL-2 + IL-21, or IL-2/21-Trikine for 14 (G-H) or 21 (I) days. Errors bars indicate mean ± standard deviation (SD) of 4 replicate wells. Results are representative of 2 independent experiments. Schematics in A, C, E, and F created using BioRender.com .

Article Snippet: For human T cell activation, cells were activated with plate-bound anti-human CD3ε (1 μg ml −1 , clone OKT-3, BioXCell) and soluble anti-human CD28 (5 μg ml −1 , clone 9.3, BioXCell).

Techniques: Standard Deviation, Fluorescence, Knock-Out, Flow Cytometry, Staining, In Vitro

Journal: Science (New York, N.Y.)

Article Title: Rewiring STAT signaling from the cell surface with Trikine immunotherapeutics

doi: 10.1126/science.adx9954

Figure Lengend Snippet: ( A-B ), C57BL/6 mice bearing established s.c. B16F10 tumors received i.p. injections of PBS, mono-IL-10, or IL-10/2-trikine (equivalent to 3 μg functional IL-10) starting on day 5, administered every other day until day 19 (n = 9 animals); the experimental timeline (A) and average tumor growth curves (B). Results are combined from 2 independent experiments. ( C-L ), C57BL/6 mice bearing established s.c. B16F10 tumors received i.p. injections of cytokine (3 μg functional cytokine) or left NT on days 9 and 11 (n = 5 animals). Mice were sacrificed on day 13, tumors were collected for flow cytometry analysis. (C), Average frequencies of CD45 + cells among single-live cells. (D- F), Counts of CD8 + T cells (D), CD4 + T cells (E), NK cells (F), myeloid cells (G), macrophages (H), monocytes (I), CD103 + dendritic cells (J), CD11b + dendritic cells (K), and B cells (L) per mg of tumor tissue. Results are representative of 2 independent experiments. ( M-O ), C57BL/6 mice bearing established subcutaneous C2 pancreatic tumors were treated intraperitoneally with cytokine (5.6 μg functional cytokine per dose) or left untreated (NT), administered every other day from day 6 to day 20 (n = 5 mice per group); experimental timeline (M), average tumor growth curves (N) and survival curves (O) of female mice in experiment described in (M). Results are representative of 2 independent experiments. Indicated statistical significance is IL-10/2-Trikine vs. PBS, IL-10/2-Trikine vs. ICB, and IL-10/2-Trikine vs. mono-IL-10. Indicated statistical significance in (O) is mono-IL-10 vs. PBS, is IL-10/2-Trikine vs. PBS, IL-10/2-Trikine vs. ICB, and IL-10/2-Trikine vs. mono-IL-10. Data represent mean ± s.e.m. and are analyzed by one-way (C to L) or two-way (B, N) ANOVA with Tukey’s post-test. Schematics in A and M created using BioRender.com . ( P-T ) C57BL/6 mice were inoculated subcutaneously. with 2.5 × 10 5 6694c2vTRP1 cells and treated with PBS, mono-IL-10 (26 μg/mouse, n = 5), or IL-10/2-Trikine (30.6 μg/mouse, n = 5) intraperitoneally on days 8 and 10 post-inoculation. Tumors were harvested on day 14 and CD45+ cells were isolated by magnetic bead separation, individually labeled with hashtag antibodies, pooled by treatment group, and subjected to Single Cell 5' RNA and cell surface protein sequencing. (P) UMAP of tumor-infiltrating CD3 + T and NK cells colored by unsupervised clusters. (Q) Bar graph shows cell cluster composition across treatment groups. Statistical significance was assessed by unpaired t test. Black asterisks indicate comparisons between PBS and IL-10/2-Trikine; white asterisks indicate comparisons between IL-10 and IL-10/2-Trikine. *P < 0.05, **P < 0.01. (R) Gene Set Enrichment Analysis (GSEA) of differentially expressed genes (DEGs) between all clusters from IL-10/2-Trikine–treated and mono–IL-10–treated groups using MSigDB Hallmark gene sets. (S) UMAP of tumor-infiltrating T/NK cells colored by a STAT5 target gene expression score ( Gzmb , Prf1 , Nkg7 , Gzma , Klrd1 , Klrb1c , Klra7 , Ccr5 , Ly6a , Thy1 ). (T) Mean STAT5 scores per mouse across all clusters, normalized per mouse. Each dot represents one mouse; error bars indicate standard deviation. Statistical significance was assessed by Wilcoxon test. **P < 0.01. scRNAseq data are the result of one independent experiment.

Article Snippet: For human T cell activation, cells were activated with plate-bound anti-human CD3ε (1 μg ml −1 , clone OKT-3, BioXCell) and soluble anti-human CD28 (5 μg ml −1 , clone 9.3, BioXCell).

Techniques: Functional Assay, Flow Cytometry, Isolation, Labeling, Single Cell, Sequencing, Targeted Gene Expression, Standard Deviation