invivomab anti mouse pd 1 cd279  (Bio X Cell)

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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: InVivoPlus anti-mouse PD-1 (CD279) , BioXcell , Cat#BE0146; RRID: AB_10949053.

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

Journal: Cell Reports Medicine

Article Title: 5-HT reuptake blockade induces pyroptosis in BRAF V600E -mutated melanomas via remodeling histone serotonylation

doi: 10.1016/j.xcrm.2025.102537

Figure Lengend Snippet: PH induces pyroptosis to potentiate PD-1 blockade (A) Representative immunofluorescence images of CD8 and GZMB staining in YUMM1.7 P - and YUMM1.7 DTR -derived tumors; right: quantification of indicated staining-positive cells. Scale bars, 10 μm. (B–D) YUMM1.7 DTR cells stably expressing the indicated shRNAs were subcutaneously injected into immunocompetent C57BL/6J mice. When tumor sizes reached 50 mm 3 (day 5, labeled as day 0 in C), mice were administrated with the indicated drugs (B). Tumor size in each group was recorded every 3 days (C). At the endpoint, the tumors were excised and weighed (D). PH, 25 mg/kg, intraperitoneal (i.p.) injection, daily. IgG2a or anti-PD-1 was diluted in saline and then given i.p. every 3 days. n = 7 tumors per group. (E) CD8 and GZMB staining in YUMM1.7 DTR allografts with the indicated treatment (left). Right: quantification of the indicated staining-positive cells. Scale bars, 10 μm. (F) Frequencies of intratumoral CD3ε + (left), CD4 + (middle), and CD8 + (right) cells among CD45 + cells. n = 7 tumors per group. (G) Frequencies of intratumoral GZMB + (left), PRF1 + (middle), and IFN-γ + (right) cells among CD8 + T cells. n = 7 tumors per group. Data are displayed as mean ± SEM. Statistical significance was determined by Student’s t test. ∗∗∗ p < 0.001, ∗∗ p < 0.01, ∗ p < 0.05, n.s., not significant. See also , , and .

Article Snippet: In vivo monoclonal anti-mouse PD-1 (CD279) (clone RMP1-14) , BioXcell , Cat# BE0146; RRID: AB_10949053.

Techniques: Immunofluorescence, Staining, Derivative Assay, Stable Transfection, Expressing, Injection, Labeling, Saline

Journal: Cell Reports Medicine

Article Title: Microbial-derived immunostimulatory small molecule augments anti-PD-1 therapy in lung cancer

doi: 10.1016/j.xcrm.2025.102519

Figure Lengend Snippet: Screening of R-derived Bacteroides isolates for IFNγ stimulation reveals a six-isolate stimulatory consortium (A) Graph shows 30 representative isolates from the Bacteroides genus. Each bar represents the fold change in percent of CD8 + T cells producing IFNγ over unstimulated controls from primary splenocytes exposed to cell-free bacterial supernatant (1:100) from individual isolates. The last 6 isolates pictured are defined as stimulatory based on criteria defined in the methods section. Each bar is one biological replicate. (B) Fold change in percent of CD8 + T cells producing IFNγ over unstimulated controls ( n = 3) by PMA-/ionomycin-positive control ( n = 7), 6-consort cell-free supernatant ( n = 5), or further 3 kDa molecular weight-cutoff-filtered small molecules ( n = 5) as compared to culture medium control (MEGA, n = 5). Each point is a biological replicate. Values are from three independent experiments. (C) Bar plot shows the relative abundance of the 6 Bacteroides species present in the 6-consort fecal samples 2 weeks post-colonization ( n = 5) as determined by 16S rRNA gene sequencing, with each bar representing a single mouse. Each species is labeled according to the species-level taxonomy of corresponding isolates present in the 6-consort. (D) Growth curve of LLC subcutaneous allograft tumors after 6-consort or human fecal microbiota transplant from R ( n = 4) or NR ( n = 6) pooled feces (NR feces n = 5, R feces n = 6, and 6-consort n = 5) into germ-free mice treated with anti-PD-1 monoclonal antibody injection. Each point is tumor volume mean ± SEM. Mixed-effects model with the Geisser-Greenhouse correction. (E) Mean ± SD of intratumoral IFNγ+ CD8 + frequency of dissociated tumors from human microbiota-colonized R ( n = 6), NR ( n = 5), and 6-consort mice ( n = 5) at day 20 (endpoint). Each point is a biological replicate. (F) Representative images of resected LLC-tumor-bearing lungs from NR feces-colonized mice (one per cage with n = 3 cages and 2–3 animals per cage) after treatment with anti-PD-1. Below each lung image is the corresponding H&E histology. Each image features a 2-mm scale bar. (G) Representative images of resected LLC-tumor-bearing lungs from R feces-colonized mice (one per cage with n = 3 cages and 2–3 animals per cage) after treatment with anti-PD-1. Below each lung image is the corresponding H&E histology. (H) Representative images of resected LLC-tumor-bearing lungs from 6-consort-colonized mice (one per cage with n = 3 cages and 2–3 animals per cage) after treatment with anti-PD-1. Below each lung image is the corresponding H&E histology. (I) Mean ± SD of weight of resected lung in grams from NR feces ( n = 6), R feces ( n = 8), and 6-consort ( n = 6) mice. Each point is a biological replicate. (J) Mean ± SD of number of discrete tumor nodules as counted from each resected lung from NR feces ( n = 6), R feces ( n = 8), and 6-consort ( n = 6) mice. Each point is a biological replicate. (K) Mean ± SD of intratumoral IFNγ+ CD8 + frequency from dissociated lung tumors from NR feces ( n = 6), R feces ( n = 8), and 6-consort mice ( n = 6). Each point is a biological replicate. Statistical differences between groups excluding tumor growth curves were determined using multiple t tests with Sidak-Bonferroni correction for multiple comparisons. p values for each comparison indicated in figures. ns: p > 0.05.

Article Snippet: Anti-Mouse CD279 (PD-1) (Clone RMP1-14) , Leinco , Cat# P362-50mg; RRID:AB_2737557.

Techniques: Derivative Assay, Positive Control, Molecular Weight, Control, Sequencing, Labeling, Injection, Comparison

Journal: Cell Reports Medicine

Article Title: Microbial-derived immunostimulatory small molecule augments anti-PD-1 therapy in lung cancer

doi: 10.1016/j.xcrm.2025.102519

Figure Lengend Snippet: 6-consort enhances anti-PD-1 treatment in vivo through IFNγ (A) Growth curve of LLC subcutaneous allograft tumors after human fecal microbiota transplant from NR pooled feces (1 × 10 7 CFU; n = 6 patients) or 6-consort (1 × 10 7 CFU) into germ-free mice (n = 4–6) treated with anti-PD-1 or combination anti-PD-1/anti-IFNγ monoclonal antibody injection. Anti-IFNγ was administered every other day starting the day of LLC tumor implantation until endpoint. Each point is tumor volume mean. p value calculated using mixed-effects model with the Geisser-Greenhouse correction. (B) Mean ± SD of serum IFNγ concentration in pg/mL at endpoint. p value calculated by multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. (C) Mean ± SD of intratumoral IFNγ+ CD8 + frequency of resected subcutaneous allograft tumors from human microbiota-colonized NR mice with or without anti-IFNγ depletion ( n = 4 and n = 6, respectively) and 6-consort mice with or without IFNγ depletion ( n = 5 and n = 5, respectively). p value calculated by multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Note: 6-consort + anti-PD-1 n = 5 due to insufficient events measured during flow cytometry. Each point is a biological replicate. (D) Growth curve of LLC subcutaneous allograft tumors after colonization with 6-consort in germ-free mice treated with anti-PD-1 or untreated. Each point is tumor volume mean ± SEM. p value calculated using mixed-effects model with the Geisser-Greenhouse correction. (E) Mean ± SD of tumor weight at endpoint for 6-consort mice treated with anti-PD-1 monoclonal antibody or untreated. p value calculated by multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. p values for each comparison indicated in figures, ns: p > 0.05.

Article Snippet: Anti-Mouse CD279 (PD-1) (Clone RMP1-14) , Leinco , Cat# P362-50mg; RRID:AB_2737557.

Techniques: In Vivo, Injection, Tumor Implantation, Concentration Assay, Flow Cytometry, Comparison

Journal: Cell Reports Medicine

Article Title: Microbial-derived immunostimulatory small molecule augments anti-PD-1 therapy in lung cancer

doi: 10.1016/j.xcrm.2025.102519

Figure Lengend Snippet: Synthetic cis- Bac429 induces stimulation of IFNγ production (A) Structure of synthetic compound Bac429 in the cis configuration. (B) Structure of synthetic compound Bac429 in the trans configuration. (C) Structure of synthetic saturated analog control compound sat -Bac429. (D) Mean ± SD for IFNγ mRNA expression in murine splenocytes stimulated with cis- Bac429, trans- Bac429, or sat -Bac429 at a concentration range between 10 and 100 μM or PMA/ionomycin ( n = 3 replicates/condition), with relative fold expression compared to unstimulated controls. DMSO was used as a solvent vehicle for unstimulated controls. p values calculated with multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. (E) Representative flow cytometry plots for analysis of IFNγ+ CD8 + T cells from murine splenocytes. (F) Mean ± SD for IFNγ stimulation quantified by flow cytometry from primary splenic CD8 + T cells by cis- Bac429, trans- Bac429, and sat -Bac429 at a concentration range between 10 and 100 μM or PMA/ionomycin ( n = 3 replicates/condition). DMSO was used as a solvent vehicle for unstimulated controls. p values calculated with multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. (G) Mean ± SD for percent of live cells quantified by flow cytometry from the experiment described in (F). p values calculated with multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. (H) Representative flow cytometry plots for analysis of IFNγ+ CD8 + T cells from healthy human PBMCs. (I) Mean ± SD for IFNγ stimulation quantified by flow cytometry from primary human PBMCs CD8 + T cells by cis- Bac429, trans- Bac429, and sat -Bac429 at a concentration range between 10 and 100 μM or PMA/ionomycin ( n = 5 human donors). p values calculated with multiple t tests with Sidak-Bonferroni correction for multiple comparisons. DMSO was used for unstimulated solvent vehicle control. Each point is a biological replicate. (J) Mean ± SD for percent of live cells quantified by flow cytometry from the bioassay for IFNγ stimulation by cis- Bac429, trans- Bac429, and sat -Bac429 at a concentration range between 10 and 100 μM or PMA/ionomycin ( n = 5 human donors). p values calculated with multiple t tests with Sidak-Bonferroni correction for multiple comparisons. (K) Growth curve of LLC subcutaneous allograft tumors after 6-consort ( n = 8) or sterile culture media control ( n = 6) in germ-free mice treated with anti-PD-1 monoclonal antibody injection. Each point is tumor volume mean ± SEM. Mixed-effects model with the Geisser-Greenhouse correction; 6- consort vs. GF p < 0.001. (L) Mean ± SD of intratumoral IFNγ+ CD8 + frequency of dissociated tumors from 6-consort mice ( n = 8) or sterile culture media control ( n = 6) in germ-free mice at day 22 (endpoint). t test: 6-consort vs. GF p = 0.035. Each point is a biological replicate. (M) Mean ± SD of pmol/g Bac429 by QTOF MS/MS from stool of germ-free mice colonized with 6-consort ( n = 8) or sterile culture media control ( n = 7) at baseline pre-treatment with anti-PD-1 or at endpoint post-treatment. Multiple t tests with Sidak-Bonferroni correction for multiple comparisons. Each point is a biological replicate. (N) Mean ± SD of pmol/g Bac429 by QTOF MS for serum, lung, spleen, and tumor of germ-free mice colonized with 6-consort ( n = 8) or sterile culture media control ( n = 6) at endpoint. t test: p > 0.05 for all comparisons. ND indicates not detected. p values for each comparison indicated in figures; ns: p > 0.05.

Article Snippet: Anti-Mouse CD279 (PD-1) (Clone RMP1-14) , Leinco , Cat# P362-50mg; RRID:AB_2737557.

Techniques: Control, Expressing, Concentration Assay, Solvent, Flow Cytometry, Bioassay, Sterility, Injection, Tandem Mass Spectroscopy, Comparison

Journal: Cell Reports Medicine

Article Title: Microbial-derived immunostimulatory small molecule augments anti-PD-1 therapy in lung cancer

doi: 10.1016/j.xcrm.2025.102519

Figure Lengend Snippet: Intratumoral administration of cis- Bac429 in combination with anti-PD-1 therapy reduces tumor volume and enhances systemic anti-tumor immunity (A) Growth curve of left-flank LLC subcutaneous allograft tumors in mice receiving intratumoral injection of either cis- Bac429 (2.15 mg/kg) or DMSO vehicle at the same time as anti-PD-1 or isotype treatment in right-flank tumors. Each point represents tumor volume mean ± SEM (n = 5–7 mice/group). p values calculated using mixed-effects model with Geisser-Greenhouse correction. (B) Growth curve of right-flank LLC subcutaneous allograft tumors in mice receiving intratumoral injection of either cis- Bac429 (2.15 mg/kg) or DMSO vehicle at the same time as anti-PD-1 or isotype treatment in right-flank tumors. Each point represents tumor volume mean ± SEM (n = 5–7 mice/group). p values calculated using mixed-effects model with Geisser-Greenhouse correction. (C) Mean ± SD of intratumoral IFNγ+ CD8 + frequency at endpoint for left-flank LLC subcutaneous tumors in mice treated with an anti-PD-1 or isotype monoclonal antibody that received intratumoral injections in the right flank of either cis- Bac429 or DMSO (n = 5–7 mice/group). Each point is a biological replicate. (D) Mean ± SD of intratumoral IFNγ+ CD8 + frequency at endpoint for right-flank LLC subcutaneous tumors in mice treated with an anti-PD-1 or isotype monoclonal antibody that received intratumoral injections in the right flank of either cis- Bac429 or DMSO (n = 5–7 mice/group). Each point is a biological replicate. (E) Growth curve of left-flank LLC subcutaneous allograft tumors in mice receiving intratumoral injection of either cis- or sat- Bac429 (2.15 mg/kg) or DMSO alone at the same time as anti-PD-1 treatment or saline control in right flank tumors. Each point represents tumor volume mean ± SEM (n = 3–7 mice/group). p values calculated using mixed-effects model with Geisser-Greenhouse correction. (F) Growth curve of right-flank LLC subcutaneous allograft tumors in mice receiving intratumoral injection of either cis- or sat- Bac429 - Bac429 (2.15 mg/kg) or DMSO alone at the same time as anti-PD-1 treatment or saline control. Each point represents tumor volume mean ± SEM (n = 3–7 mice/group). p values calculated using mixed-effects model with Geisser-Greenhouse correction. (G) Mean ± SD of intratumoral IFNγ+ CD8 + frequency at endpoint for left-flank LLC subcutaneous tumors in mice treated with an anti-PD-1 monoclonal antibody or saline control that received intratumoral injections in the right flank of either cis- Bac429, sat -Bac429, or DMSO (n = 3–7 mice/group). Each point is a biological replicate. (H) Mean ± SD of intratumoral IFNγ+ CD8 + frequency at endpoint for right-flank LLC subcutaneous tumors in mice treated with an anti-PD-1 monoclonal antibody or saline control that received intratumoral injections in the right flank of either cis- Bac429, sat -Bac429, or DMSO (n = 3–7 mice/group). Each point is a biological replicate. Statistical differences between groups excluding tumor growth curves were determined using multiple t tests with Sidak-Bonferroni correction for multiple comparisons. p values for each comparison indicated in figures; ns: p > 0.05.

Article Snippet: Anti-Mouse CD279 (PD-1) (Clone RMP1-14) , Leinco , Cat# P362-50mg; RRID:AB_2737557.

Techniques: Injection, Saline, Control, Comparison