invivomab anti human cd8α  (Bio X Cell)

Journal: bioRxiv

Article Title: Quantitative extrapolation from single-tags (QuEST) immunofluorescence microscopy to derive TCR signalosome stoichiometries in human primary T cells

doi: 10.64898/2026.03.28.715001

Figure Lengend Snippet: a , Generation (top) and validation (bottom) of 1G4 TCR–GFP Jurkat cells. 1G4 TCR–GFP and CD8 were coexpressed, CD4 was eliminated by negative sorting. b , Generation (top) and validation (bottom) of CD28–GFP Jurkat cells. c , Generation (top) and validation (bottom) of Lck–GFP Jurkat cells. d , Generation (top) and validation (bottom) of ZAP-70–GFP Jurkat cells.

Article Snippet: Unlabelled primary antibodies : anti-human CD8α antibody (Cell Signaling Technology, Cat#85336), anti-human CD28 antibody (Cell Signaling Technology, Cat#38774S), anti-human CD45 antibody (Cell Signaling Technology, Cat#13917S), anti-human PD-1 antibody (Cell Signaling Technology, Cat#86163T), anti-human Lck antibody (Cell Signaling Technology, Cat#2787S), anti-human ZAP-70 antibody (Cell Signaling Technology, Cat#3165S), anti-human LAT antibody (Cell Signaling Technology, Cat#45533S), anti-human PLCγ1 antibody (Cell Signaling Technology, Cat#5690S), and anti-human phospho-ZAP-70 (Tyr319) antibody (Cell Signaling Technology, Cat#2701).

Techniques: Biomarker Discovery

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: SMARCA4 deficiency impairs CD8 + T cell function and confers resistance to PD-1 blockade in NSCLC (A) Progression-free survival (PFS) of eight patients with SMARCA4-deficient (BRG1-deficient) NSCLC treated with ICIs. (B) Representative axial CT images of a patient (P6) with SMARCA4-deficient NSCLC before and after ICIs treatment. (C) Change in the sum of target lesion diameters for six evaluable patients with SMARCA4-deficient NSCLC from baseline to first radiographic assessment. (D) Schematic of the experimental design for the orthotopic lung cancer model ( n = 8/group). (E) Representative in vivo bioluminescence images of mice from the indicated treatment groups at different time points. (F) Quantification of total bioluminescence flux from mice in each group over time. (G) Kaplan-Meier survival curves of mice from the four treatment groups. (H) Individual tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (I) UMAP visualization of major immune cell populations within the TME, analyzed by flow cytometry. (J) Quantitative comparison of the frequencies of indicated immune cell lineages between SMARCA4-WT and -KD tumors. (K and L) Representative flow cytometry plots showing the production of IFN-γ and TNF-α by tumor-infiltrating CD8 + T cells. (M) Quantification of the frequencies of IFN-γ + and TNF-α + cells among tumor-infiltrating CD8 + T cells. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one- or two-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Cell Function Assay, In Vivo, Flow Cytometry, Comparison

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: Attenuated IL-2/STAT5 signaling and an enhanced exhaustion phenotype in CD8 + T cells within SMARCA4-deficient tumors (A) Schematic of the workflow for transcriptomic profiling of CD8 + T cells. (B) Volcano plot displaying differentially expressed genes in CD8 + T cells from SMARCA4-KD versus WT tumors. (C–E) Pathway enrichment analyses of genes downregulated in CD8 + T cells from SMARCA4-KD tumors, including Gene Ontology (GO) terms, KEGG pathways, and Reactome pathways. (F) The gene set enrichment analysis (GSEA) plot. (G) Correlation matrix (pie chart) showing the association between IL-2 receptor subunits expression and key T cell exhaustion marker genes in tumor-infiltrating CD8 + T cells. (H) Radar plot comparing the normalized expression levels of genes encoding IL-2 receptor subunits and exhaustion markers in CD8 + T cells. (I) Quantification by flow cytometry of the expression frequencies of PD-1, TIGIT, and TIM-3 on tumor-infiltrating CD8 + T cells. (J and K) Representative multiplex immunofluorescence (mIF) images of SMARCA4-WT and -KD tumor sections stained for PanCK (cyan), CD8 (green), GZMB (white), PD-1 (red), TIGIT (orange), and TIM-3 (yellow). Scale bars, 20 μm. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Expressing, Marker, Flow Cytometry, Multiplex Assay, Immunofluorescence, Staining

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: SMARCA4 loss in tumor cells attenuates CD8 + T cell function via NF-κB-mediated suppression of ICAM1 (A) Flow cytometry analysis of surface expression of the exhaustion markers PD-1, TIGIT, and TIM-3 on human CD8 + T cells following co-culture. (B and C) Representative flow cytometry plots showing the production of IFN-γ and TNF-α and the surface expression of IL-2Rα (CD25) by CD8 + T cells following co-culture. (D) Quantification of the frequencies of IFN-γ + , TNF-α + , and IL-2Rα + cells among co-cultured CD8 + T cells. (E) Integrated single-nucleus RNA sequencing (snRNA-seq) analysis comparing IL2-STAT5 signaling activity. y axis: IL2-STAT5 signaling score. (F and G) Incoming and outgoing signaling patterns between major cell types in the TME, as inferred from snRNA-seq. (H) Specific cell-cell communication network illustrating the ICAM signaling pathway from tumor cells to CD8 + T cells in patients with SMARCA4-WT NSCLC. (I) Correlation analysis between SMARCA4 and ICAM1 mRNA expression in TCGA cohorts. (J) Immunohistochemistry staining and quantification of ICAM1 protein expression in tumor tissues from SMARCA4-WT ( n = 10) and -deficient ( n = 10) NSCLC patients. (K) Schematic illustrating the proposed link between SMARCA4 deficiency and impaired NF-κB activation. (L) Immunoblot analysis of ICAM1 and p65 protein levels in SMARCA4-WT H2122 cells treated with the NF-κB inhibitor PTDC or vehicle control. (M) ChIP-qPCR analysis showing NF-κB (p65) binding to a specific site within the ICAM1 promoter in SMARCA4-WT H2122 cells ( n = 3). (N) Dual-luciferase reporter assay in SMARCA4-WT H2122 cells ( n = 3). Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Cell Function Assay, Flow Cytometry, Expressing, Co-Culture Assay, Cell Culture, RNA Sequencing, Activity Assay, Immunohistochemistry, Staining, Activation Assay, Western Blot, Control, ChIP-qPCR, Binding Assay, Luciferase, Reporter Assay

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: The PD-1/IL-2 bsAb exerts potent anti-tumor efficacy in SMARCA4-deficient models (A) Schematic of the therapeutic experiment in humanized mice ( n = 6/group). (B) Representative images of excised tumors from each treatment group at the endpoint. (C) Tumor growth curves of individual mice in the indicated treatment groups. (D) Kaplan-Meier survival curves of mice from the four treatment groups. (E) Quantification by flow cytometry of tumor-infiltrating CD3 + CD8 + T cells. (F) Quantification of the percentage of tumor-infiltrating CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (G) Frequency of polyfunctional (TNF-α + IFN-γ + ) CD8 + T cells among tumor-infiltrating lymphocytes. (H) Schematic of the humanized patient-derived xenograft (PDX) model ( n = 5/group). (I) Representative images of excised PDX tumors from each treatment group. (J) Tumor growth curves of individual PDX-bearing mice. (K) Kaplan-Meier survival curves of PDX-bearing mice from the three treatment groups. (L) Quantification of tumor-infiltrating CD3 + CD8 + T cells in PDX tumors. (M) Quantification of the percentage of CD8 + T cells from PDX tumors expressing PD-1, TIGIT, and TIM-3. (N) Frequency of polyfunctional (TNF-α + IFN-γ + ) CD8 + T cells in PDX tumors. (O) Representative multiplex immunofluorescence (mIF) images of PDX tumor sections from different treatment groups, stained for PanCK (cyan), CD8 (green), GZMB (white), PD-1 (red), TIGIT (orange), and TIM-3 (yellow). Scale bars, 40 μm. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA or two-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Flow Cytometry, Expressing, Derivative Assay, Multiplex Assay, Immunofluorescence, Staining

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: STAT5 activation mediates the therapeutic effect of the PD-1/IL-2 bsAb in SMARCA4-deficient NSCLC (A) Representative flow cytometry plots showing the expression of the exhaustion markers PD-1, TIGIT, and TIM-3 on activated human CD8 + T cells. (B) Quantification of the mean fluorescence intensity (MFI) of the exhaustion markers PD-1, TIGIT, and TIM-3 on CD8 + T cells from the experiment in (A) ( n = 3). (C) Representative flow cytometry plots showing the production of IFN-γ and TNF-α by CD8 + T cells under the conditions described in (A). (D) Quantification of the frequencies of IFN-γ + and TNF-α + cells among CD8 + T cells ( n = 3). (E) Schematic of the adoptive T cell transfer experiment ( n = 8/group). (F) Representative in vivo bioluminescence images of mice from the indicated groups at different time points. (G) Tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (H) Kaplan-Meier survival curves of mice from the four treatment groups. (I) Quantification of the frequency of donor-derived CD45.2 + CD8 + T cells among total tumor-infiltrating lymphocytes. (J) Flow analysis of donor-derived CD45.2 + CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (K–M) Frequency of IFN-γ + (K–L) and TNF-α + (M) cells among donor-derived CD45.2 + CD8 + T cells. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA or two-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Activation Assay, Flow Cytometry, Expressing, Fluorescence, In Vivo, Derivative Assay

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: PD-1/IL-2 bsAb protects CD8 + T cells from macrophage phagocytosis via STAT5-mediated upregulation of CD47 (A) Schematic of the Cleavage Under Targets and Tagmentation (CUT&Tag) assays workflow. (B) Distribution of STAT5 binding signals relative to transcription start sites (TSS) in CD8 + T cells treated with or without the PD-1/IL-2 bsAb. (C) Genomic annotation of differentially enriched STAT5 binding peaks in the bsAb-treated group. (D) KEGG pathway enrichment analysis of genes associated with STAT5 binding peaks. (E and F) Strategy and Venn diagram for identifying potential STAT5 downstream genes. (G) ChIP-qPCR analysis of STAT5 binding to the promoter regions of selected candidate genes ( n = 3). (H) Genome browser tracks showing STAT5 binding signals at the CD47 locus in control and PD-1/IL-2 bsAb-treated CD8 + T cells. (I) Schematic of the macrophage phagocytosis assay. (J) Representative confocal microscopy images showing macrophages (red) engulfing CD8 + T cells (green). Scale bars, 20 μm. (K) Flow cytometry quantification of the percentage of macrophages that had phagocytosed CD8 + T cells under the indicated conditions ( n = 3). Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: Binding Assay, ChIP-qPCR, Control, Phagocytosis Assay, Confocal Microscopy, Flow Cytometry

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: CD47 protects CD8 + T cells from macrophage clearance to boost antitumor immunity in SMARCA4-deficient NSCLC (A) Schematic of the adoptive T cell therapy experiment ( n = 8/group). (B) Representative in vivo bioluminescence images of mice from the indicated treatment groups at different time points. (C) Tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (D) Individual tumor growth curves for mice in each treatment group. (E) Kaplan-Meier survival curves of mice from the four treatment groups. (F) Quantification by flow cytometry of donor-derived CD45.2 + CD8 + T cells among total tumor-infiltrating lymphocytes. (G) Representative flow cytometry plots for donor-derived CD45.2 + CD8 + T cells expressing the exhaustion markers PD-1, TIGIT, and TIM-3. (H) Quantification of the percentage of CD45.2 + CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (I) The production of TNF-α by donor-derived CD45.2 + CD8 + T cells. (J) The production of IFN-γ by donor-derived CD45.2 + CD8 + T cells. (K) Representative immunofluorescence images of tumor sections. White: CD8, green: CD47, red: F4/80. Scale bars, 70 μm. (L) Schematic model depicting the proposed mechanism of action. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by one-way ANOVA.

Article Snippet: InVivoMAb anti-human CD8α , BioXcell , Cat#BE0004-2; RRID: AB_1107673.

Techniques: In Vivo, Flow Cytometry, Derivative Assay, Expressing, Immunofluorescence

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: SMARCA4 deficiency impairs CD8 + T cell function and confers resistance to PD-1 blockade in NSCLC (A) Progression-free survival (PFS) of eight patients with SMARCA4-deficient (BRG1-deficient) NSCLC treated with ICIs. (B) Representative axial CT images of a patient (P6) with SMARCA4-deficient NSCLC before and after ICIs treatment. (C) Change in the sum of target lesion diameters for six evaluable patients with SMARCA4-deficient NSCLC from baseline to first radiographic assessment. (D) Schematic of the experimental design for the orthotopic lung cancer model ( n = 8/group). (E) Representative in vivo bioluminescence images of mice from the indicated treatment groups at different time points. (F) Quantification of total bioluminescence flux from mice in each group over time. (G) Kaplan-Meier survival curves of mice from the four treatment groups. (H) Individual tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (I) UMAP visualization of major immune cell populations within the TME, analyzed by flow cytometry. (J) Quantitative comparison of the frequencies of indicated immune cell lineages between SMARCA4-WT and -KD tumors. (K and L) Representative flow cytometry plots showing the production of IFN-γ and TNF-α by tumor-infiltrating CD8 + T cells. (M) Quantification of the frequencies of IFN-γ + and TNF-α + cells among tumor-infiltrating CD8 + T cells. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one- or two-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Cell Function Assay, In Vivo, Flow Cytometry, Comparison

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: Attenuated IL-2/STAT5 signaling and an enhanced exhaustion phenotype in CD8 + T cells within SMARCA4-deficient tumors (A) Schematic of the workflow for transcriptomic profiling of CD8 + T cells. (B) Volcano plot displaying differentially expressed genes in CD8 + T cells from SMARCA4-KD versus WT tumors. (C–E) Pathway enrichment analyses of genes downregulated in CD8 + T cells from SMARCA4-KD tumors, including Gene Ontology (GO) terms, KEGG pathways, and Reactome pathways. (F) The gene set enrichment analysis (GSEA) plot. (G) Correlation matrix (pie chart) showing the association between IL-2 receptor subunits expression and key T cell exhaustion marker genes in tumor-infiltrating CD8 + T cells. (H) Radar plot comparing the normalized expression levels of genes encoding IL-2 receptor subunits and exhaustion markers in CD8 + T cells. (I) Quantification by flow cytometry of the expression frequencies of PD-1, TIGIT, and TIM-3 on tumor-infiltrating CD8 + T cells. (J and K) Representative multiplex immunofluorescence (mIF) images of SMARCA4-WT and -KD tumor sections stained for PanCK (cyan), CD8 (green), GZMB (white), PD-1 (red), TIGIT (orange), and TIM-3 (yellow). Scale bars, 20 μm. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Expressing, Marker, Flow Cytometry, Multiplex Assay, Immunofluorescence, Staining

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: SMARCA4 loss in tumor cells attenuates CD8 + T cell function via NF-κB-mediated suppression of ICAM1 (A) Flow cytometry analysis of surface expression of the exhaustion markers PD-1, TIGIT, and TIM-3 on human CD8 + T cells following co-culture. (B and C) Representative flow cytometry plots showing the production of IFN-γ and TNF-α and the surface expression of IL-2Rα (CD25) by CD8 + T cells following co-culture. (D) Quantification of the frequencies of IFN-γ + , TNF-α + , and IL-2Rα + cells among co-cultured CD8 + T cells. (E) Integrated single-nucleus RNA sequencing (snRNA-seq) analysis comparing IL2-STAT5 signaling activity. y axis: IL2-STAT5 signaling score. (F and G) Incoming and outgoing signaling patterns between major cell types in the TME, as inferred from snRNA-seq. (H) Specific cell-cell communication network illustrating the ICAM signaling pathway from tumor cells to CD8 + T cells in patients with SMARCA4-WT NSCLC. (I) Correlation analysis between SMARCA4 and ICAM1 mRNA expression in TCGA cohorts. (J) Immunohistochemistry staining and quantification of ICAM1 protein expression in tumor tissues from SMARCA4-WT ( n = 10) and -deficient ( n = 10) NSCLC patients. (K) Schematic illustrating the proposed link between SMARCA4 deficiency and impaired NF-κB activation. (L) Immunoblot analysis of ICAM1 and p65 protein levels in SMARCA4-WT H2122 cells treated with the NF-κB inhibitor PTDC or vehicle control. (M) ChIP-qPCR analysis showing NF-κB (p65) binding to a specific site within the ICAM1 promoter in SMARCA4-WT H2122 cells ( n = 3). (N) Dual-luciferase reporter assay in SMARCA4-WT H2122 cells ( n = 3). Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Cell Function Assay, Flow Cytometry, Expressing, Co-Culture Assay, Cell Culture, RNA Sequencing, Activity Assay, Immunohistochemistry, Staining, Activation Assay, Western Blot, Control, ChIP-qPCR, Binding Assay, Luciferase, Reporter Assay

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: The PD-1/IL-2 bsAb exerts potent anti-tumor efficacy in SMARCA4-deficient models (A) Schematic of the therapeutic experiment in humanized mice ( n = 6/group). (B) Representative images of excised tumors from each treatment group at the endpoint. (C) Tumor growth curves of individual mice in the indicated treatment groups. (D) Kaplan-Meier survival curves of mice from the four treatment groups. (E) Quantification by flow cytometry of tumor-infiltrating CD3 + CD8 + T cells. (F) Quantification of the percentage of tumor-infiltrating CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (G) Frequency of polyfunctional (TNF-α + IFN-γ + ) CD8 + T cells among tumor-infiltrating lymphocytes. (H) Schematic of the humanized patient-derived xenograft (PDX) model ( n = 5/group). (I) Representative images of excised PDX tumors from each treatment group. (J) Tumor growth curves of individual PDX-bearing mice. (K) Kaplan-Meier survival curves of PDX-bearing mice from the three treatment groups. (L) Quantification of tumor-infiltrating CD3 + CD8 + T cells in PDX tumors. (M) Quantification of the percentage of CD8 + T cells from PDX tumors expressing PD-1, TIGIT, and TIM-3. (N) Frequency of polyfunctional (TNF-α + IFN-γ + ) CD8 + T cells in PDX tumors. (O) Representative multiplex immunofluorescence (mIF) images of PDX tumor sections from different treatment groups, stained for PanCK (cyan), CD8 (green), GZMB (white), PD-1 (red), TIGIT (orange), and TIM-3 (yellow). Scale bars, 40 μm. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA or two-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Flow Cytometry, Expressing, Derivative Assay, Multiplex Assay, Immunofluorescence, Staining

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: STAT5 activation mediates the therapeutic effect of the PD-1/IL-2 bsAb in SMARCA4-deficient NSCLC (A) Representative flow cytometry plots showing the expression of the exhaustion markers PD-1, TIGIT, and TIM-3 on activated human CD8 + T cells. (B) Quantification of the mean fluorescence intensity (MFI) of the exhaustion markers PD-1, TIGIT, and TIM-3 on CD8 + T cells from the experiment in (A) ( n = 3). (C) Representative flow cytometry plots showing the production of IFN-γ and TNF-α by CD8 + T cells under the conditions described in (A). (D) Quantification of the frequencies of IFN-γ + and TNF-α + cells among CD8 + T cells ( n = 3). (E) Schematic of the adoptive T cell transfer experiment ( n = 8/group). (F) Representative in vivo bioluminescence images of mice from the indicated groups at different time points. (G) Tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (H) Kaplan-Meier survival curves of mice from the four treatment groups. (I) Quantification of the frequency of donor-derived CD45.2 + CD8 + T cells among total tumor-infiltrating lymphocytes. (J) Flow analysis of donor-derived CD45.2 + CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (K–M) Frequency of IFN-γ + (K–L) and TNF-α + (M) cells among donor-derived CD45.2 + CD8 + T cells. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA or two-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Activation Assay, Flow Cytometry, Expressing, Fluorescence, In Vivo, Derivative Assay

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: PD-1/IL-2 bsAb protects CD8 + T cells from macrophage phagocytosis via STAT5-mediated upregulation of CD47 (A) Schematic of the Cleavage Under Targets and Tagmentation (CUT&Tag) assays workflow. (B) Distribution of STAT5 binding signals relative to transcription start sites (TSS) in CD8 + T cells treated with or without the PD-1/IL-2 bsAb. (C) Genomic annotation of differentially enriched STAT5 binding peaks in the bsAb-treated group. (D) KEGG pathway enrichment analysis of genes associated with STAT5 binding peaks. (E and F) Strategy and Venn diagram for identifying potential STAT5 downstream genes. (G) ChIP-qPCR analysis of STAT5 binding to the promoter regions of selected candidate genes ( n = 3). (H) Genome browser tracks showing STAT5 binding signals at the CD47 locus in control and PD-1/IL-2 bsAb-treated CD8 + T cells. (I) Schematic of the macrophage phagocytosis assay. (J) Representative confocal microscopy images showing macrophages (red) engulfing CD8 + T cells (green). Scale bars, 20 μm. (K) Flow cytometry quantification of the percentage of macrophages that had phagocytosed CD8 + T cells under the indicated conditions ( n = 3). Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by unpaired Student’s t test or one-way ANOVA where appropriate.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: Binding Assay, ChIP-qPCR, Control, Phagocytosis Assay, Confocal Microscopy, Flow Cytometry

Journal: Cell Reports Medicine

Article Title: Dual PD-1/IL-2Rα targeting restores CD8 + T cell fitness via STAT5/CD47 axis in SMARCA4-deficient NSCLC

doi: 10.1016/j.xcrm.2026.102633

Figure Lengend Snippet: CD47 protects CD8 + T cells from macrophage clearance to boost antitumor immunity in SMARCA4-deficient NSCLC (A) Schematic of the adoptive T cell therapy experiment ( n = 8/group). (B) Representative in vivo bioluminescence images of mice from the indicated treatment groups at different time points. (C) Tumor growth curves, as measured by bioluminescence, for mice in each treatment group. (D) Individual tumor growth curves for mice in each treatment group. (E) Kaplan-Meier survival curves of mice from the four treatment groups. (F) Quantification by flow cytometry of donor-derived CD45.2 + CD8 + T cells among total tumor-infiltrating lymphocytes. (G) Representative flow cytometry plots for donor-derived CD45.2 + CD8 + T cells expressing the exhaustion markers PD-1, TIGIT, and TIM-3. (H) Quantification of the percentage of CD45.2 + CD8 + T cells expressing PD-1, TIGIT, and TIM-3. (I) The production of TNF-α by donor-derived CD45.2 + CD8 + T cells. (J) The production of IFN-γ by donor-derived CD45.2 + CD8 + T cells. (K) Representative immunofluorescence images of tumor sections. White: CD8, green: CD47, red: F4/80. Scale bars, 70 μm. (L) Schematic model depicting the proposed mechanism of action. Data are represented as mean ± SD (error bars) from biological replicates. Statistical analyses, n.s., no significance. Statistical significance was determined by one-way ANOVA.

Article Snippet: CD8 + T cell dependency was assessed by co-administering a depleting anti-human CD8α antibody (BioXcell, 15 μg/mouse) with PD-1/IL-2 bsAb in the SMARCA4-KD model. Depletion (initiated one day before PD-1/IL-2 bsAb and maintained twice weekly) was confirmed by flow cytometry (>90% reduction in circulating human CD3 + CD8 + T cells).

Techniques: In Vivo, Flow Cytometry, Derivative Assay, Expressing, Immunofluorescence

Journal: Immunity

Article Title: Positioning and reversible suppression of CCR7 + dendritic cells in perivascular tumor niches shape cancer immunity

doi: 10.1016/j.immuni.2025.11.020

Figure Lengend Snippet: (A) Heatmaps depict the enrichment of immune and non-immune cell types in the immediate neighborhood of CCR7 + DCs in NSCLC spatial transcriptomic data ( n = 4). (B) (Left) Representative FOV displaying CCR7 + DCs (HLA-DR + LAMP3 + ; yellow) located near BVs (CD31 + PDPN − ; magenta) and Tregs (CD4 + FOXP3 + ; white) in one HNSCC sample using high-plex whole-tissue imaging. Scale bar represents 20 μm. (Right) Box plots display the frequencies of BV-associated, LV-associated, and non-vessel-associated CCR7 + DCs close (<5 μm) to Tregs among all tumor CCR7 + DCs with nearby Tregs. Wilcoxon test, whiskers represent min to max; * p < 0.05. (C) Correlations between CCR7 + DCs and Tregs within CD45 + cells, as determined by scRNA-seq in multiple human cancer types. Spearman rank correlation; significant correlations are shown with a fitted red line. (D) (Left) Scheme outlining the analyses of CCR7 + DCs and Tregs in NSCLC samples. Patients with numerous (>5) CCR7 + DC clusters ( n = 12) were selected for downstream analyses. (Right) Frequency of CCR7 + DCs (CD11c + LAMP3 + ) with at least one nearby (<50 μm) Treg (CD4 + FOXP3 + ) in each individual patient. Numbers of FOVs analyzed per sample are as follows: NR01, n = 126; NR06, n = 455; NR09, n = 180; NR12, n = 79; NR26, n = 293; R11, n = 122; R15, n = 205; R35, n = 175; R37, n = 459; R45, n = 276. (E) (Left) Scheme outlining the analysis of tumor biopsies from HNSCC patients before immunotherapy (pre-IO). Patients were divided into non-responders (NR, n = 5) and responders (R, n = 5) based on the assessment of clinical response at 6 months. (Right) CCR7 + DC shortest distance to Tregs, T CONV , and CD8 + T cells in NR versus R tumors. Data are shown for all CCR7 + DCs compiled (NR tumors, n = 1,457 cells; R tumors, n = 1,324 cells). Unpaired t test, whiskers represent min to max; **** p < 0.0001. Numbers of FOVs analyzed per sample as in (D). (F) (Left) Scheme outlining the analyses of CCR7 + DC-CD8 + T cell niches. (Right) Frequencies of CCR7 + DC-CD8 + T cell niches with or without Tregs in their proximity (<100 μm). Two-way ANOVA with multiple comparisons, whiskers represent min to max; * p < 0.05. Numbers of FOVs analyzed per sample as in (D). (G) Representative FOV displaying CCR7 + DCs (FSCN1 + cells; FSCN1 in yellow) located near BVs (CD31 + LYVE-1 − cells; CD31 in magenta) and Tregs (FOXP3 + cells; FOXP3 in white) in untreated MC38 tumors. Scale bar represents 50 μm. (H) Correlations between the numbers of CCR7 + DCs and Tregs per mg of tumor tissue, as determined by fluorescence-activated cell sorting (FACS) analyses of MC38 and D4M3. A tumors. Spearman rank correlation; significant correlations are shown with a fitted red line. (I) Box plots show the frequencies of tumor CCR7 + DCs close (<5 μm) to Tregs that are associated to BVs or LVs in MC38 tumors ( n = 7). Whole-tumor sections were analyzed. Paired t test, whiskers represent min to max; **** p < 0.0001. See also and .

Article Snippet: Unconjugated rabbit anti-human CD8α (Clone D8A8Y) , Cell Signaling Technology , Cat#85336.

Techniques: Imaging, Fluorescence, FACS

Journal: Immunity

Article Title: Positioning and reversible suppression of CCR7 + dendritic cells in perivascular tumor niches shape cancer immunity

doi: 10.1016/j.immuni.2025.11.020

Figure Lengend Snippet: (A) (Left) Scheme outlining the experimental setup for bulk RNA-seq analyses of tumor-derived CCR7 + DCs. (Right) GO pathway enrichment analyses performed on differentially expressed genes (DEGs) in CCR7 + DCs in MC38 tumors ( n = 4) from Treg-depleted ( FoxP3 -DTR) compared with Treg-sufficient (WT) mice. Bar plot indicates the −log 10 raw binomial p -values of the top 10 most enriched pathways in CCR7 + DCs. (B) (Left) Experimental setup for ex vivo stimulation of OT-I CD8 + T cells with tumor CCR7 + DCs. (Right) Percentage of OT-I CD8 + T cells that proliferated after 5-day culture with OVA 257–264 peptides-loaded CCR7 + DCs isolated from WT or Treg-depleted tumors. As a control, CCR7 + DCs without OVA 257–264 peptides were used. Two-way ANOVA with multiple comparisons, whiskers represent min to max; ** p < 0.01. (C) (Left) Relative gene expression levels analyzed by bulk RNA-seq. Each dot represents one mouse ( n = 4), whiskers represent mean to max. Unpaired t test with multiple comparisons; * p < 0.05. (Right) Representative histogram of CD40 protein expression and relative mean fluorescence intensity (MFI) measured by FACS and expressed both as normalized values and absolute MFI. Each dot represents one mouse ( n = 18), whiskers represent min to max. Unpaired t test; ** p < 0.01. (D) Analyses of cDCs in tumor-draining lymph nodes. Absolute cell counts (left, n = 10) and MFI of CD40 expression (right, n = 18) measured by FACS in migratory cDCs (CCR7 + CD8α − ) from WT or Treg-depleted mice. Whiskers represent mean to max. (E) (Left) Experimental setup for ex vivo analyses of tumor CCR7 + DCs isolated from anti-PD-1-treated mice that received or not αCD25 NIB mAbs. (Right) CD40 protein expression measured by FACS and expressed both as normalized values and absolute MFI. Each dot represents one mouse ( n = 4 WT and n = 6 FoxP3-DTR), whiskers represent min to max. Unpaired t test; ** p < 0.01. (F) (Left) Overall survival analyses of MC38 tumor-bearing mice treated, or not treated, with αPD-1 and αCD25 NIB mAbs, and in which CD4 + or CD8 + cells were depleted or not ( n = 8 or 9 mice/group). Log-rank Mantel-Cox test; * p < 0.05, *** p < 0.001, and *** p < 0.0001. (Right) Percentage of tumor-free mice on day 60 in the indicated treatment groups. (G) (Left) Experimental setup for ex vivo stimulation of OT-I CD8 + T cells with tumor CCR7 + DCs as in (B). The DCs were obtained from mice receiving anti-PD-1 immunotherapy and that were treated or not with αCD25 NIB mAbs. (Right) Percentage of OT-I CD8 + T cells that proliferated after 5-day culture with OVA 257–264 peptide-loaded CCR7 + DCs. Each dot represents one mouse ( n = 8 and n = 7), whiskers represent min to max. Two-way ANOVA with multiple comparisons; * p < 0.05. (H) (Left) Scheme outlining bone marrow chimeras with inducible Cd40 -deficiency in cDCs and the treatment schedule. (Right) Growth curves of MC38 tumors inoculated in zDC iDTR : Cd40 WT and zDC iDTR : Cd40 KO bone marrow chimeras treated with αPD-1, αCD25 NIB , or αPD-1 + αCD25NIB combination ( n = 8–10 mice/group). Mean with SEM. Two-way ANOVA with multiple comparisons; * p < 0.05 and **** p < 0.0001. See also and .

Article Snippet: Unconjugated rabbit anti-human CD8α (Clone D8A8Y) , Cell Signaling Technology , Cat#85336.

Techniques: RNA Sequencing, Derivative Assay, Ex Vivo, Isolation, Control, Gene Expression, Expressing, Fluorescence