cxcl10  (R&D Systems)

Journal: Frontiers in Immunology

Article Title: Tumor-intrinsic interferon signaling drives pancreatic cancer resistance to tumor mucin1-targeted CAR T cell therapy

doi: 10.3389/fimmu.2025.1618415

Figure Lengend Snippet: Suppressing IFN signaling in PDA reduces ICAM-1 and CXCL10 levels. PDA cell lines were plated overnight for attachment. On the following day, PDA cells were treated with JAKi at 10μM and 25μM overnight. After JAKi treatment, it was removed, and tMUC1-CAR T cells or Mock T cells were added at E:T ratio of 5:1 in the absence of JAKi. The E:T ratio was calculated based on the initial number of PDA cells plated. After 24hr, co-culture supernatants were collected and assayed for cytokine levels of (A) ICAM-1 and (B) CXCL10 by ELISA. Data are presented as the mean ± SD of triplicate. Baseline levels of ICAM-1 and CXCL10 were low (<50pg/ml and <10pg/ml, respectively) in either CAR T cell-only or in PDA-only cultures, which were detected by ELISA. The statistical comparison was conducted between CAR T cell treatment in JAKi-pretreated PDA and CAR T cell treatment in PDA without JAKi pretreatment. *p<0.05, ****p<0.0001 (unpaired t test with Welch’s correction).

Article Snippet: On the following day, PDA cells were pre-incubated with PD-L1 blocking antibody (10μg/ml) or its isotype control antibody (10μg/ml; Cat# 400348; BioLegend), anti-ICAM-1 neutralizing antibody (5μg/ml; Cat# AF720; R&D Systems), anti-CXCL10 neutralizing antibody (5μg/ml; Cat# MAB266-100; R&D Systems), or fresh media alone for 2hr.

Techniques: Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Frontiers in Immunology

Article Title: Tumor-intrinsic interferon signaling drives pancreatic cancer resistance to tumor mucin1-targeted CAR T cell therapy

doi: 10.3389/fimmu.2025.1618415

Figure Lengend Snippet: Suppressing IFN signaling in CAR T cells slightly decreases ICAM-1 level. PDA cell lines were plated overnight for attachment. On the same day, tMUC1-CAR T cells were treated with or without JAKi at 10μM and 25μM overnight. On the following day, JAKi-pretreated CAR T cells were washed with fresh media to remove JAKi before being added to PDA cells at E:T ratio of 5:1 for co-culture. After 24hr, co-culture supernatants were collected and assayed for cytokine levels of (A) ICAM-1 and (B) CXCL10 by ELISA. Data are presented as the mean ± SD of triplicate. The statistical comparison was conducted between JAKi-pretreated CAR T cell group and media-pretreated CAR T cell group in PDA. *p<0.05, **p<0.01 (unpaired t test with Welch’s correction).

Article Snippet: On the following day, PDA cells were pre-incubated with PD-L1 blocking antibody (10μg/ml) or its isotype control antibody (10μg/ml; Cat# 400348; BioLegend), anti-ICAM-1 neutralizing antibody (5μg/ml; Cat# AF720; R&D Systems), anti-CXCL10 neutralizing antibody (5μg/ml; Cat# MAB266-100; R&D Systems), or fresh media alone for 2hr.

Techniques: Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Frontiers in Immunology

Article Title: Tumor-intrinsic interferon signaling drives pancreatic cancer resistance to tumor mucin1-targeted CAR T cell therapy

doi: 10.3389/fimmu.2025.1618415

Figure Lengend Snippet: Engagement of CAR T cell with PDA induces function loss of CAR T cells and up-regulation of immune checkpoints. (A) Loss of CAR T cell cytotoxicity. CAR T cells or Mock T cells were co-cultured with MiaPaCa-2 or HPAFII cells at E:T ratio of 5:1 or cultured with media alone for overnight. Then the live CAR T cells and Mock T cells were isolated from the first round co-culture, and they were added to the fresh MiaPaCa-2 or HPAFII plates at E:T ratio of 2:1 or 5:1 for 24hr as the second round of co-culture. At the end of culture, tumor cell lysis against MiaPaCa-2 (left panel) or against HPAFII (right panel) was determined using MTT assay. The percentage of lysis was calculated using the formula: [(OD of co-culture with Mock T cells– OD of co-culture with CAR T cells)/OD of co-culture with Mock T cells] ×100. The mock T cells and CAR T cells are from the same pair for calculation. Data are presented as the mean ± SD from quadruplicate. The statistical difference was conducted for cytolysis of CAR T cells retrieved from co-culture with PDA cells when compared with cytolysis of CAR T cells retrieved from culture with media alone. ****p<0.0001 (Multiple unpaired t tests with Welch’s correction). (B) Increase of CD25 expression on CAR T cells after engagement with PDA. CAR T cells were co-cultured with MiaPaCa-2 or HPAFII cells at E:T ratio of 5:1 or cultured with media alone for overnight. Then CAR T cells were stained and analyzed for CD25 expression on CAR-positive and CAR-negative live cells after gating on CD4 + T cells and CD8 + T cells. (C) Expression of ICs on CAR T cells. Cell culture was performed same as in (B) . IC expression on CAR-positive and CAR-negative live cells were displayed after gating on CD4 + T cells and CD8 + T cells. (D) The retaining of tMUC1 and up-regulation of PD-L1 on PDA. After rinsing off suspension cells in co-culture from (B) , adherent MiaPaCa-2 or HPAFII cells were stained and analyzed for tMUC1 and PD-L1. PDA cells cultured in media alone were included as baseline control. (E) Suppressing tumor IFN signaling blocks CAR T cells-induced PD-L1 increase. MiaPaCa-2 or HPAFII cells were pre-treated with JAKi at 25μM overnight. Then JAKi was removed, and CAR T cells were added in the absence of JAKi at E:T ratio of 5:1, calculated based on the initial number of PDA cells plated. After 24hr treatment with CAR T cells, live adherent PDA cells were stained and analyzed for PD-L1 expression. (F) Effectiveness of PD-L1 blocking antibody. CAR T cells-treated HPAFII cells were pre-incubated with PD-L1 blocking antibody at the indicated doses, followed by PD-L1-PE staining. (G) Blocking PD-L1 partially reversed HPAFII resistance to CAR T cell lysis. MiaPaCa-2 or HPAFII cells were treated with CAR T cells or Mock T cells in the presence of PD-L1 blocking antibody or its isotype control for 24hr. PDA cell lysis was determined using MTT assay. Data are presented as the mean ± SD from quadruplicate. The statistical difference was conducted for cytolysis of CAR T cells with PD-L1 blocking antibody when compared with cytolysis of CAR T cells with media alone. ****p<0.0001 (unpaired t tests with Welch’s correction). (H) Involvement of ICAM-1 and CXCL10 in CAR T cell cytotoxicity. MiaPaCa-2 or HPAFII cells were treated with CAR T cells or Mock T cells in the presence of anti-ICAM-1 or anti-CXCL10 antibodies for 24hr. PDA cell lysis was determined using MTT assay. Data are presented as the mean ± SD from quadruplicate. The statistical difference was conducted for cytolysis of CAR T cells with neutralizing antibody when compared with cytolysis of CAR T cells with media alone. **p<0.01 (unpaired t tests with Welch’s correction).

Article Snippet: On the following day, PDA cells were pre-incubated with PD-L1 blocking antibody (10μg/ml) or its isotype control antibody (10μg/ml; Cat# 400348; BioLegend), anti-ICAM-1 neutralizing antibody (5μg/ml; Cat# AF720; R&D Systems), anti-CXCL10 neutralizing antibody (5μg/ml; Cat# MAB266-100; R&D Systems), or fresh media alone for 2hr.

Techniques: Cell Culture, Isolation, Co-Culture Assay, Lysis, MTT Assay, Expressing, Staining, Suspension, Control, Blocking Assay, Incubation

Journal: Respiratory Research

Article Title: Tuberculous pleural effusion-induced Arg-1 + macrophage polarization contributes to lung cancer progression via autophagy signaling

doi: 10.1186/s12931-024-02829-8

Figure Lengend Snippet: CXL9 and CXL10 in TPE modulate M2 macrophage polarization. a Cytokine array analysis demonstrated differences between TPE and T. Spots with differentially regulated cytokines are identified with a red box and a blue box. Red boxes correspond to CD30, CXCL9, and CXCL10. Blue boxes correspond to ApoA1, YKL-40, IGFBP-2, and IGFBP-3. b Mouse BMDMs were treated with 20 µg recombinant CXCL9 and 20 µg recombinant CXCL10. c Mouse BMDMs were cultured with neutralizing antibodies for CXCL9 (20 µg) and CXCL10 (20 µg) after TPE treatment. Arg-1 mRNA, Ym-1 mRNA, and CXCR3 mRNA levels were measured. ApoA1: apolipoprotein A1; YKL-40: chitinase-3-like protein 1; IGFBP-2: insulin like growth factor binding protein 2; IGFBP-3: insulin like growth factor binding protein 3. The values represent the results of three experiments. ** p < 0.01, * p < 0.05

Article Snippet: Recombinant CXCL9 and CXCL10 and neutralizing antibodies against CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques: Recombinant, Cell Culture, Binding Assay

Journal: Respiratory Research

Article Title: Tuberculous pleural effusion-induced Arg-1 + macrophage polarization contributes to lung cancer progression via autophagy signaling

doi: 10.1186/s12931-024-02829-8

Figure Lengend Snippet: Schema of mechanism by which TPE-induced Arg-1 + macrophage polarization contributes to lung cancer malignancy. CXCL9 and CXCL10 are involved in TPE-induced macrophage M2 Arg-1 + polarization. TPE-Arg-1 + MФ CM induces A549 proliferation by upregulating autophagy and E-cadherin signaling. ABH reversed the effect of TPE-Arg-1 + MФ CM leading to cancer growth suppression by downregulating autophagy and E-cadherin signaling. ABH: 2(S)-amino-6-boronohexanoic acid (an arginase inhibitor)

Article Snippet: Recombinant CXCL9 and CXCL10 and neutralizing antibodies against CXCL9 and CXCL10 were purchased from R&D Systems.

Techniques:

Journal: Cellular and Molecular Immunology

Article Title: The BCL-2 inhibitor APG-2575 resets tumor-associated macrophages toward the M1 phenotype, promoting a favorable response to anti-PD-1 therapy via NLRP3 activation

doi: 10.1038/s41423-023-01112-y

Figure Lengend Snippet: APG-2575 remodels the transcriptomic landscape of macrophages and CD8+ T cells and promotes CD8+ T-cell infiltration via the chemokines CCL5 and CXCL10. A UMAP plot from merged data of tumor-infiltrating Mo/MF populations. B UMAP plots with annotated clusters of Mo/MF cells from the control and APG-2575 groups. C A heatmap showing the differentially expressed genes (rows) among Mo/MF subpopulations (columns). Representative genes from each cluster are highlighted (right). D Ratios of the proportions of Mo/MF clusters across different regimens. E GSEA using genes differentially expressed between IL-4-activated RAW264.7 cells with control or APG-2575 treatment. F ‒ I Scatter plot showing the results of Pearson correlation analysis of CCL5 and CXCL10 expression and the infiltration of CD8+ T cells and M1 macrophages in TCGA cohorts. J Growth of LLC tumors in C57BL/6 mice treated with the indicated regimens. ( K – M ) Flow cytometric analysis of CD8+, CD8 + GZMB+ and CD8+ TNF-α+ T cells in C57BL/6 mouse xenograft tumors treated with the indicated regimens. N ‒ Q Immunohistochemical staining of CD8 and GZMB in C57BL/6 mouse xenograft tumors

Article Snippet: Murine or human T-cell migration was assessed with medium, APG-2575-treated supernatant alone, supernatant plus 10 μg/mL anti-CCL5 neutralizing antibodies, and/or 10 μg/mL anti-CXCL10 neutralizing antibody (R&D Systems).

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Cellular and Molecular Immunology

Article Title: The BCL-2 inhibitor APG-2575 resets tumor-associated macrophages toward the M1 phenotype, promoting a favorable response to anti-PD-1 therapy via NLRP3 activation

doi: 10.1038/s41423-023-01112-y

Figure Lengend Snippet: Graphical summary of the results. APG-2575 can synergize with ICIs through a mechanism involving the repolarization of TAMs from the M2 to the M1 phenotype, further enhancing CD8+ T-cell recruitment into the TME via the augmentation of CCL5 and CXCL10 secretion and thereby improving tumor immunosuppression

Article Snippet: Murine or human T-cell migration was assessed with medium, APG-2575-treated supernatant alone, supernatant plus 10 μg/mL anti-CCL5 neutralizing antibodies, and/or 10 μg/mL anti-CXCL10 neutralizing antibody (R&D Systems).

Techniques: