Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Targeting the NAT10-HDAC4 positive feedback loop counteracts immunosuppression in breast cancer

doi: 10.1186/s13046-025-03638-7

Figure Lengend Snippet: Inhibition of NAT10 enhances the efficacy of anti–PD-L1 therapy in vivo . A Schematic diagram of the tumor model and treatment schedule with anti-PD-L1 administration. B–D Representative images ( B ), tumor growth curves ( C ), and tumor weights ( D ) of mouse 4T1 tumors from shNC, shNAT10, anti-PD-L1, or the combination of shNAT10 and anti-PD-L1 ( n = 6). E-F Representative mIF images of mouse tumor tissues showing staining for T cells ( E ), Tregs ( F ), and MDSCs ( F ) (scale bar, 20 μm). G - H ELISA analysis of GzmB (G) and IFN-γ (H) levels in the serum of mice from different groups. I Representative IHC staining of PD-L1, CD8, and GzmB in breast cancer tissues ( n = 220), with statistical analysis of staining intensity (scale bar, 200 μm). J Correlation of PD-L1 and CD8, and PD-L1 and GzmB protein levels in breast cancer tissues ( n = 220) assessed by IHC. All data are presented as mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: In the CD8+ T cell depletion assay, mice were administered anti-Mouse CD8α antibody (Clone 2.43) (MedChemExpress, #HY-P990790, China) intraperitoneally at a dose of 300 μg daily for three consecutive days.

Techniques: Inhibition, In Vivo, Staining, Enzyme-linked Immunosorbent Assay, Immunohistochemistry

Journal: Journal for immunotherapy of cancer

Article Title: Angiotensin receptor blocker attacks armored and cold tumors and boosts immune checkpoint blockade.

doi: 10.1136/jitc-2024-009327

Figure Lengend Snippet: Figure 1 AGTR1 highly expresses in armored and cold tumors and predicts low response to immune checkpoint blockade (ICB) therapy. (A) Heatmap showing the targets expression of concomitant medications in the three immuno-collagenic subtypes in the TCGA-BLCA cohort. (B) Boxplot showing high expression of AGTR1 in armored and cold tumors in the TCGA- BLCA cohort. Horizontal lines in the boxplots represent the median value, and the lower and upper hinges correspond to the first and third quartiles, respectively. Significance was calculated using the ANOVA with Tukey’s multiple-comparison test. ns, non-significance, ***p<0.001. (C) Prognostic value of AGTR1 in the TCGA-BLCA cohort. Subgroups for survival analysis were divided by the best cut-off point. Significance was calculated using the log-rank test. *p<0.05. (D) Representative images uncovering stromal and immune markers in the three immuno-collagenic subtypes in the in-house BLCA cohort. Staining data of HE, Masson, and PD-L1 IHC from our previous study 10 was used as controls. A total of 61 samples were analyzed due to 2 samples losses during multiple staining procedures. (E–G) Expression of PD-L1, CD8, and AGTR1 in the three immuno- collagenic subtypes in the in-house BLCA cohort. Data was presented as mean±SD. Significance was calculated using the Kruskal-Wallis test with the Dunn’s multiple-comparison test for (E). Significance was calculated using the ANOVA with Tukey’s multiple-comparison test for (F and G). ns, non-significance, *p<0.05, ***p<0.001. (H) Prognostic value of AGTR1 in the in- house BLCA cohort. Subgroups for survival analysis were divided by the value of 5%. Significance was calculated using the log-rank test. (I) Predictive value and prognostic value of AGTR1 in the IMvigor210 cohort. Subgroups for survival analysis were divided by the best cut-off point. Significance was calculated using the Student t-test (left) and the log-rank test (right). *p<0.05, ***p<0.001. (J, K) Predictive value of AGTR1 in the GSE135222 and the merged BRCA cohorts. Significance was calculated using the Mann-Whitney test (J) and the Student’s t-test (K). *p<0.05, ***p<0.001. ANOVA, analysis of variance; BLCA, bladder cancer; TCGA, The Cancer Genome Atlas.

Article Snippet: The PD- 1 in vivo mAb (catalog BE0273) was purchased from BioXCell (Lebanon, USA), and the CD8 in vivo mAb (catalog A2102) was purchased from Selleck (Shanghai, China).

Techniques: Expressing, Medications, Comparison, Staining, MANN-WHITNEY

Journal: Journal for immunotherapy of cancer

Article Title: Angiotensin receptor blocker attacks armored and cold tumors and boosts immune checkpoint blockade.

doi: 10.1136/jitc-2024-009327

Figure Lengend Snippet: Figure 5 ARB reverses CAFs-mediated T cell inhibition and exerts immune-dependent tumor suppressive role. (A) The positive rate of AGTR1 in CAFs in 10 BRCA samples in our published cohort. Based on the percentage of AGTR1+ cells in CAFs, BRCA patients were divided into AGTR1-positive and AGTR1-negative groups based on 10%. (B) Boxplot showing exhausted and cytotoxic signature scores in CD8+ T cells in AGTR1-positive and AGTR1-negative groups. Horizontal lines in the boxplots represent the median value, and the lower and upper hinges correspond to the first and third quartiles, respectively. Significance was calculated using the Student’s t-test. ***p<0.001. (C) Schematic protocol of co-culture of CAFs and T cells. (D, E) Differentially expressed genes and expression of activated markers in T cells identified by T cell activation PCR array. (F) Relative content of cytokines in supernatant from the co-culture system was assessed by ELISA assay. Data was presented as mean±SD. Significance was calculated with the Student’s t-test. **p<0.01. (G) Representative images uncovering CD8+ T cell infiltration in samples with low and high AGTR1 expression in the in-house BLCA cohort and quantitative analysis. Significance was calculated using the Pearson test. (H) Tumor growth curve of mice treated with PBS, losartan, and losartan+anti-CD8 antibody. (I) Representative images showing the tumors harvested from mice bearing Lewis cells treated with PBS, losartan, and losartan+anti-CD8 antibody, and weight of the harvested tumors. Data was presented as mean±SD. Significance was calculated with one-way ANOVA with Tukey’s multiple-comparison test. ns, non-significance, *p<0.05. BRCA, breast cancer; ANOVA, analysis of variance; ARB, angiotensin receptor blocker; BLCA, bladder cancer; CAFs, cancer-associated fibroblasts.

Article Snippet: The PD- 1 in vivo mAb (catalog BE0273) was purchased from BioXCell (Lebanon, USA), and the CD8 in vivo mAb (catalog A2102) was purchased from Selleck (Shanghai, China).

Techniques: Inhibition, Co-Culture Assay, Expressing, Activation Assay, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Journal for immunotherapy of cancer

Article Title: Angiotensin receptor blocker attacks armored and cold tumors and boosts immune checkpoint blockade.

doi: 10.1136/jitc-2024-009327

Figure Lengend Snippet: Figure 6 Angiotensin receptor blocker (ARB) shapes an inflamed tumor microenvironment (TME) and enhances immunotherapy in vivo. (A) Tumor growth curve of mice treated with PBS, losartan, anti-PD-1 antibody, and the combination. (B) Representative images showing the tumors harvested from mice bearing Lewis cells treated with PBS, losartan, anti- PD-1 antibody, and the combination, and weight of the harvested tumors. Data was presented as mean±SD. Significance was calculated with one-way ANOVA with Tukey’s multiple-comparison test. ns, non-significance, *p<0.05, ***p<0.001. (C) Representative images showing structure of heart, liver, and kidney from mice in different groups. (D) Representative images showing the levels of collagen, AGTR1, CD8, and Ki67 in tumor tissues from mice in different groups. (E) Representative results of flow cytometry analysis of total CD8+ T cells represented by CD3+CD8+ and quantitative analysis. Data was presented as mean±SD. Significance was calculated with one-way ANOVA with Tukey’s multiple-comparison test. ns, non-significance, **p<0.01, ***p<0.001. (F) Representative results of flow cytometry analysis of myeloid-derived suppressor cells (MDSC) represented by CD11b+Gr-1+ and quantitative analysis. Data was presented as mean±SD. Significance was calculated with one- way ANOVA with Tukey’s multiple-comparison test. ns, non-significance, ***p<0.001. PBS: phosphate buffer saline; ANOVA, analysis of variance.

Article Snippet: The PD- 1 in vivo mAb (catalog BE0273) was purchased from BioXCell (Lebanon, USA), and the CD8 in vivo mAb (catalog A2102) was purchased from Selleck (Shanghai, China).

Techniques: In Vivo, Comparison, Flow Cytometry, Derivative Assay, Saline

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: On CD28/CD40 Ligand Costimulation, Common γ -Chain Signals, and the Alloimmune Response 1

doi:

Figure Lengend Snippet: Acquisition of primed phenotype by dividing CD4+ and CD8+ T cells in vivo. A, CFSE-labeled B6AF1 cells were recovered from irradiated DBA/2 hosts 3 days after adoptive transfer. Cells were stained with CyChrome anti-mouse CD4 and anti-CD8 as well as PE-conjugated mAbs against CD25, CD44, and CD62L. Expression of the activation markers was analyzed by gating onto CFSE+CD4+ or CFSE+CD8+ cells. Cells stained with isotype control Abs were used as a control to set up the quadrants for analysis. B, Expression of CD25, CD44, and CD62L in each cell division was analyzed, and the mean fluorescence intensity (MFI) was calculated and plotted against the number of cell divisions. Representative data of five experiments shown.

Article Snippet: The following staining Abs were obtained from BD PharMingen (San Diego, CA): CyChrome anti-mouse CD4 (GK1.5), CyChrome anti-mouse CD8 α (clone 53-6.7), PE anti-mouse CD44 (clone IM7), PE anti-mouse CD62L (L-selectin, clone MEL-14), PE anti-mouse CD25 (clone PC61), PE anti-mouse IL-2 (clone JES6-5H4), PE anti-mouse TNF- α (clone MP6-XT22), PE anti-mouse IFN- γ (clone XMG 1.2), and PE isotype control Abs.

Techniques: In Vivo, Labeling, Irradiation, Adoptive Transfer Assay, Staining, Expressing, Activation Assay, Fluorescence

Journal: PLoS ONE

Article Title: Characterization of Developmental Pathway of Natural Killer Cells from Embryonic Stem Cells In Vitro

doi: 10.1371/journal.pone.0000232

Figure Lengend Snippet: (A) CD34 + CD45 − EB cells were sorted and cultured with OP9 and cytokines to generate ES-HPs. They were then cultured on OP9-DL1 stroma with cytokines for T cell differentiation. At different time points, cells were harvested and stained for CD44 and CD25 and analyzed by flow cytometer. Dead cells were stained with propidium iodide and OP9-DL1 cells expressing green fluorescent protein were gated out. The numbers show the percentages of cells in the quadrants. (B) ES-HP cells were cultured for T cell differentiation for 8 days with 5 ng/ml IL-7 as in (A). On day 8, IL-7 concentration was reduced to 1 ng/ml and cells were cultured for an additional 6 days. Cells were harvested and analyzed by flow cytometer for CD4 and CD8 expression. The numbers indicate the percentages of cells in the quadrants. (C) ES-HP cells were cultured for T cell differentiation with 5 ng/ml IL-7 for 2 weeks and an additional 4 days with 1 ng/ml IL-7, stained for TCRγδ and TCRβ and analyzed by flow cytometer. The numbers show the percentages of cells positively stained with the test antibodies (filled histogram) over control antibody (open histogram). (D) ES-HPs were generated from CD34 + CD45 − EB cells as in (A). The bulk and sorted CD45 + Lin − ES-HP cells were analyzed for T progenitor frequency by limiting dilution cultures as in , except that 30, 100, 300 and 900 cells per well for bulk and 10, 30 and 100 cells per well for CD45 + Lin − cells were plated. The results are average of two independent experiments. (E) CD45 + Lin − ES-HPs were sorted and cultured on OP9 cells with IL-7 and Flt3-L for B cell generation. After one week, cells were harvested and analysed for the expression of B220 and CD19 with flow cytometer. (F) One thousand sorted CD45 + Lin − ES-HPs were transferred into myeloid differentiation media as in . The number of myeloid and erythroid colonies were scored.

Article Snippet: PE-conjugated anti-mouse CD8α (53-6.7) mAb was purchased from Boehringer Mannheim Biochemica.

Techniques: Cell Culture, Cell Differentiation, Staining, Flow Cytometry, Expressing, Concentration Assay, Control, Generated