Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Neutralization of acyl-CoA-binding protein attenuates glucocorticoid-mediated suppression of cancer immunosurveillance

doi: 10.1073/pnas.2518983123

Figure Lengend Snippet: Improvement of immunotherapy efficacy against EO771 tumor (breast cancer) by injection of an ACBP/DBI-neutralizing monoclonal antibody in the setting of corticosterone. ( A ) The heatmap depicts the gene expression levels of mouse splenocytes after 18 h of different treatments as indicated: vehicle; corticosterone (CORT); αDBI antibody (2.5 μg/mL) or its isotype control. Computed P -values are indicated in colored squares. Statistical analyses were performed using one-way ANOVA with Tukey correction. ( B ) EO771 cells (5 × 10 5 per mouse) were subcutaneously (s.c.) injected into the right lower quadrant of the abdomen near the mammary gland, in 6 to 8-wk-old female C57BL/6J (n = 9 to 10 mice per group). From day 7 onward (when tumors became palpable), mice received corticosterone (CORT, 0.1 mg/mL in drinking water with 0.66% ethanol), and were treated intraperitoneally (i.p.) with either vehicle, αDBI mAb (5 mg/kg) or its isotype every 2 d. In additional groups, mice received i.p. injections of αCD4/CD8a mAb or αPD1 mAb according to the schedule shown. Effects of CORT, αDBI mAb and αCD4/CD8a mAb are shown in panels C-E , while additional effects of αPD1 mAb are shown in panels F-H . Average tumor growth curves (mean ± SEM) are reported in C and F ; overall survival is reported in panels D and G . Statistical analyses were performed by mixed linear model of tumor sizes for longitudinal analysis and Cox regression for survival ( https://kroemerlab.shinyapps.io/TumGrowth/ ). The corresponding P -values are presented in the tables ( E and H ) and are formatted in bold font when below the 0.05 threshold.

Article Snippet: 10 mg/kg Oxaliplatin (Sigma, #Y0000271) was injected i.p. on day 8 and PBS was used as its corresponding vehicle control. αPD1 monoclonal antibody (BioXCell, #BE0273) or its isotype control was diluted in PBS and injected i.p. on day 16, 20, 23 at a dose of 200 μg per mouse.

Techniques: Injection, Gene Expression, Control

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Neutralization of acyl-CoA-binding protein attenuates glucocorticoid-mediated suppression of cancer immunosurveillance

doi: 10.1073/pnas.2518983123

Figure Lengend Snippet: Effect of combined ACBP/DBI-neutralizing monoclonal antibody and chemotherapy in TC-1 lung cancer, within the context of corticotherapy. ( A ) Viable TC-1 cells stably expressing luciferase (TC-1-Luc, 1 × 10 6 per mouse) were intravenously (i.v.) injected into 6 to 8-wk-old female C57BL/6J mice (n = 9 to 10 mice per group). Then mice received treatments as indicated: vehicle or free corticosterone (CORT) in the drinking water (0.1 mg/mL; ethanol/ water 0.66%); αDBI antibody or its isotype (5 mg/kg, i.p.); αPD1 (200 µg, i.p.) antibody or its isotype and oxaliplatin (OXA, 5 mg/kg, i.p.). Tumor size was monitored by luciferase activity. Representative time-lapse images (n = 3 per condition) are reported in B . Tumor growth curves are reported in C ; overall survival is reported in D . Statistical analyses were performed by mixed linear model of tumor sizes for longitudinal analysis and Cox regression analysis for survival ( https://kroemerlab.shinyapps.io/TumGrowth/ ). The computed P -values are presented in the table ( E ) and formatted in bold when below the 0.05 threshold.

Article Snippet: 10 mg/kg Oxaliplatin (Sigma, #Y0000271) was injected i.p. on day 8 and PBS was used as its corresponding vehicle control. αPD1 monoclonal antibody (BioXCell, #BE0273) or its isotype control was diluted in PBS and injected i.p. on day 16, 20, 23 at a dose of 200 μg per mouse.

Techniques: Stable Transfection, Expressing, Luciferase, Injection, Activity Assay

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Neutralization of acyl-CoA-binding protein attenuates glucocorticoid-mediated suppression of cancer immunosurveillance

doi: 10.1073/pnas.2518983123

Figure Lengend Snippet: Modulation of antigen cross-presentation and Tsc22d3/TSC22D3 expression by ACBP/DBI neutralization in dendritic cells under glucocorticoid treatment. ( A ) The heatmap depicts the gene expression levels of mouse GM-CSF BMDCs isolated after 18 h of different treatments as indicated: vehicle; corticosterone (CORT, 1 μM); αDBI antibody (2.5 μg/mL) or its isotype (2.5 μg/mL). The computed P -values are presented in squares with different colors. Statistical analyses were performed by one-way ANOVA with Tukey correction. Tsc22d3 mRNA levels were measured by RT-qPCR in mouse BMDCs treated with corticosterone (CORT, 1 µM) in the presence or absence of flumazenil (1 µM) or bicuculline (10 µM) ( B ), or in Gabrg2 wild-type ( Gabrg2 WT/WT ) or Gabrg2 knock-in ( Gabrg2 F77I/F77I ) backgrounds ( C ). Tsc22d3/TSC22D3 expression was further assessed in mouse splenocytes ( D ) exposed to corticosterone (CORT, 1 µM), in human peripheral blood mononuclear cells (PBMCs; E ) and human monocyte-derived dendritic cells (moDCs; F ) exposed to hydrocortisone (HCS, 0.5 µM), each treated in the presence or absence of an ACBP/DBI-neutralizing antibody (αDBI; 2,5 µg/mL), as indicated. For panels ( B – F ), gene expression was quantified by RT-qPCR, normalized to housekeeping gene expression, and expressed as fold change relative to the corresponding untreated control condition within each experiment. Data are presented as columns with individual data points overlaid and shown as mean ± SEM. Statistical analyses were performed using ANOVA with correction for multiple comparisons; exact P values are indicated above the corresponding comparisons. ( G – I ) Upon CORT treatment, the efficacy of oxaliplatin (OXA; 5 mg/kg; i.p.) combined with either αPD1 antibody or its isotype control (200 µg/mouse; i.p.) and αDBI antibody or its isotype control (5 mg/kg) was assessed against MCA205 tumors in LysM-creX Tsc22d3 +/+ (n = 4 to 13 mice per group) ( H ) or LysM-creX Tsc22d3 −/− (n = 5 to 12 mice per group) ( I ) 8 to 10-wk-old male mice. Statistical analysis was performed using mixed linear model of tumor growth ( https://kroemerlab.shinyapps.io/TumGrowth/ ). Computed P -values are reported in panels H and I .

Article Snippet: 10 mg/kg Oxaliplatin (Sigma, #Y0000271) was injected i.p. on day 8 and PBS was used as its corresponding vehicle control. αPD1 monoclonal antibody (BioXCell, #BE0273) or its isotype control was diluted in PBS and injected i.p. on day 16, 20, 23 at a dose of 200 μg per mouse.

Techniques: Expressing, Neutralization, Gene Expression, Isolation, Quantitative RT-PCR, Knock-In, Derivative Assay, Control

Journal: Biomedicines

Article Title: Imaging Effector Memory T-Cells Predicts Response to PD1-Chemotherapy Combinations in Colon Cancer.

doi: 10.3390/biomedicines10102343

Figure Lengend Snippet: Figure 1. (A) Schema representing the dosing, measurement and imaging procedure. Mice (n = 10 per treatment arm) were treated with control IgG, αPD1, oxaliplatin (OXA), combination αPD1 + OXA, 5-fluorouracil (5-FU) or combination αPD1 + 5-FU post-tumour implantation. (B) Individual animal tumour volumes showing heterogeneity in tumour response. (C) Average tumour volume in each treatment arm, post-therapy response stratification. Data are displayed as means ± S.D. TNR, treated non-responder.

Article Snippet: Tumoured animals were treated with either a control (rat IgG2a isotype control, 5 mg/kg, IP (α-trinitrophenol mAb, n = 10)) or the immune checkpoint inhibitor αPD1 (rat IgG2a anti-mouse PD-1, 10 mg/kg, IP (αPD1 mAb RMP1-14, Bio-X-Cell, New Hampshire, USA, n = 15)) on days 6, 9 and 12 following tumour implantation.

Techniques: Imaging, Control

Journal: Biomedicines

Article Title: Imaging Effector Memory T-Cells Predicts Response to PD1-Chemotherapy Combinations in Colon Cancer.

doi: 10.3390/biomedicines10102343

Figure Lengend Snippet: Figure 2. (A) Selected MIP images demonstrating [18F]AlF-NOTA-KCNA3P uptake into tumours responding to αPD1, oxaliplatin (OXA), combined αPD1 + OXA, 5-fluorouracil (5-FU), combined αPD1 + 5-FU or non-responding tumours (TNRs). Tumour borders are delineated by yellow dotted lines. (B) Graphical representation of [18F]AlF-NOTA-KCNA3P tumour uptake values in each treatment arm (n = 6–10 mice/group; ** p < 0.01, *** p < 0.001 compared to TNRs; $ p < 0.05 compared to 5-FU; data shown as the means %ID/g ± S.E.M.). (C) Individual [18F]AlF-NOTA-KCNA3P tumour values comparing TRs and TNRs (n = 46, **** p < 0.0001).

Article Snippet: Tumoured animals were treated with either a control (rat IgG2a isotype control, 5 mg/kg, IP (α-trinitrophenol mAb, n = 10)) or the immune checkpoint inhibitor αPD1 (rat IgG2a anti-mouse PD-1, 10 mg/kg, IP (αPD1 mAb RMP1-14, Bio-X-Cell, New Hampshire, USA, n = 15)) on days 6, 9 and 12 following tumour implantation.

Techniques:

Journal: Oncoimmunology

Article Title: DNA prime and peptide boost immunization elicits robust neoantigen-specific CD8 + T cell responses and therapeutic protection in mouse tumor models

doi: 10.1080/2162402X.2025.2606497

Figure Lengend Snippet: DNA prime-peptide boost immunization strategy against endogenous neoepitopes confers therapeutic protection in MC38 tumor model. C57BL/6 mice were intradermally challenged with MC38 cells and 3 d later were intradermally immunized with a DNA vaccine encoding Adpgk (299–307) and p15E (134–141) neoepitopes. Seven days later mice received peptide boost immunization. Additional groups included mice treated with αPD-1 monotherapy or a combination of DNA/peptide vaccination and αPD-1. αPD-1 monoclonal antibody was administered intraperitoneally at 200 µg per dose on days 12, 15, and 18 post-tumor challenge. Unvaccinated and DNA immunized mice were used as controls. (a–b) Individual tumor growth (a) and cumulative event curve (tumor rejection) (b). (c–d) CD8 + T-cell responses against neoepitopes were evaluated in blood 7 d after the last vaccination by flow cytometry. Peripheral blood cells were stimulated ex vivo with cognate peptides, followed by intracellular cytokine staining. (c) Representative plots showing the expression of TNF- α and IFN- γ in total CD8 + T cells. Unstimulated control (upper panels), Adpgk (299–307) peptide (middle panels) and p15E (134–141) peptide (bottom) samples are shown. (d) Quantification of IFN-γ + Adpgk (299–307) - and p15E (134–141) -specific CD8 + T cells in peptide stimulated samples. (e) Representative flow cytometry plots showing co-expression of IFN- γ and TNF- α (left), and granzyme B (GZB, right) in IFN-γ⁺ Adpgk (299–307)- and p15E (134–141) -specific CD8⁺ T cells from peptide-stimulated samples of DNA/peptide-vaccinated mice. Histograms (right panels) depict GZB expression in IFN-γ⁻ (gray) and IFN-γ⁺ (blue) CD8⁺ T cells from a representative DNA prime-peptide boost vaccinated mouse. Data in (a–b) is from four independent experiments for Control ( n = 21), DNA ( n = 5) and DNA/Peptide ( n = 20) and two independent experiments for αPD-1 ( n = 10) and DNA/Peptide + αPD-1 ( n = 8). Data in (c-d) is from seven independent experiments for control ( n = 19), DNA ( n = 10) and DNA/Peptide ( n = 23) and two independent experiments for αPD-1 ( n = 10) and DNA/Peptide + αPD-1 ( n = 10). * p < 0.05, ** p < 0.01 by Mantel‒Cox test in (b). *** p < 0.001, **** p < 0.0001 by Kruskal-Wallis unpaired test.

Article Snippet: Anti–PD-1 (αPD-1) monoclonal antibody (clone RMP1-14, BioXCell, catalog #BE0146) was administered at a dose of 200 μg per mouse on days 12, 15, and 18 after tumor challenge. αPD-1 therapy was evaluated both as monotherapy and in combination with the DNA prime–peptide boost immunization regimen described above.

Techniques: Flow Cytometry, Ex Vivo, Staining, Expressing, Control

Journal: Gut

Article Title: Ginseng polysaccharides alter the gut microbiota and kynurenine/tryptophan ratio, potentiating the antitumour effect of antiprogrammed cell death 1/programmed cell death ligand 1 (anti-PD-1/PD-L1) immunotherapy.

doi: 10.1136/gutjnl-2020-321031

Figure Lengend Snippet: Figure 1 GPs potentiate the antitumour effect of the αPD-1 mAb in conventional and humanised PD-1 knock-in (HuPD-1) LLC-bearing mice. (A) Schematic diagram of the GPs plus αPD-1 mAb treatment in LLC-bearing mice. (B) Tumour growth curves were shown in each group. Conventional C57 BL/6J mice were received GPs on day 0 and intraperitoneally injected with αPD-1 mAb on day nine after tumour inoculation. (C) Tumour weight of the conventional LLC-bearing mice. (D) Survival curve of the conventional LLC-bearing mice. (E) Tumour growth curves were shown in each group in huPD-1 knock-in LLC-bearing mouse model. (F) Tumour weight of the HuPD-1 knock-in tumour-bearing mice. Data were representative of one or three independent experiments with n=5–10 per group. Error bars represent the mean±SEM. Tumour growth curves were assessed by two-way ANOVA with Sidak correction. Tumour weight was assessed by one-way ANOVA. Log-rank (Mantel-Cox) tests were performed for survival data. *P<0.05, **p<0.01, ***p<0.001. ANOVA, analysis of variance; GPs, ginseng polysaccharides; LLC, Lewis lung cancer; mAb, monoclonal antibody; PD-1, programmed death 1.

Article Snippet: A total of 24 mice from different litters were equally divided into four groups: Vehicle (treated with PBS), αPD-1 monoclonal antibody (mAb) (250 μg/mouse, clone: RMP1-14, Bio X Cell), GPs (200 mg/kg) and GPs plus αPD-1 mAb group.

Techniques: Knock-In, Injection

Journal: Gut

Article Title: Ginseng polysaccharides alter the gut microbiota and kynurenine/tryptophan ratio, potentiating the antitumour effect of antiprogrammed cell death 1/programmed cell death ligand 1 (anti-PD-1/PD-L1) immunotherapy.

doi: 10.1136/gutjnl-2020-321031

Figure Lengend Snippet: Figure 2 The cytotoxic CD8+ T cell population increases, whereas the Treg cell population decreases after treatment with the combination therapy. In conventional LLC-bearing mice, 24 days after tumour inoculation, the mice were treated with the αPD-1 mAb, GPs, or a combination of GPs and the αPD-1 mAb. Immune cells in the blood, spleen and tumour were analysed. (A–E) CD8+/CD4+ T cell ratio, expression of IFN-γ, TNF-α, granzyme B (GZMB) among CD8 +T cells and FOXP3 CD4+ T cells in the blood, spleen and tumour tissues. (F) Representative IHC profiles of CD4, CD8, IFN-γ, TNF-α and GZMB in tumour tissues at ×400 magnification. Scale bar=50 µm. (G–K) Quantitative analysis of positive area per field by image J software (NIH) with ImmunoRatio Plugin. Data presented as means of 15 fields and expressed as mean±SD (n=6). All experiments were repeated two or three times. Each symbol represents an individual animal. Data represent the mean±SD and were analysed by one-way ANOVA. *P<0.05, **p<0.01, ***p<0.001. ANOVA, analysis of variance; GPs, ginseng polysaccharides; IFN-γ, interferon-γ; IHC, immunohistochemistry; LLC, Lewis lung cancer; mAb, monoclonal antibody; TNF-α, tumour necrosis factor-α; Treg, regulatory T cells.

Article Snippet: A total of 24 mice from different litters were equally divided into four groups: Vehicle (treated with PBS), αPD-1 monoclonal antibody (mAb) (250 μg/mouse, clone: RMP1-14, Bio X Cell), GPs (200 mg/kg) and GPs plus αPD-1 mAb group.

Techniques: Expressing, Software, Immunohistochemistry

Journal: Gut

Article Title: Ginseng polysaccharides alter the gut microbiota and kynurenine/tryptophan ratio, potentiating the antitumour effect of antiprogrammed cell death 1/programmed cell death ligand 1 (anti-PD-1/PD-L1) immunotherapy.

doi: 10.1136/gutjnl-2020-321031

Figure Lengend Snippet: Figure 3 Combination treatment maintains gut homeostasis by modulating gut microbiota and enhancing gut immunity. (A) Relative abundance of top 15 genera in different treatment groups. (B) LEfSe analysis for differential abundant taxa detected between αPD-1 mAb and combination group. Threshold parameters were set as p=0.05 for the Mann-Whitney U test and multiclass analysis=all against all. LDA score >2.0. (C) Tumour growth curve from four groups treated with vehicle, αPD-1 mAb, GP and combination GP and αPD-1 mAb, respectively, on day 9 after tumour inoculation and antibiotics (ABX) were administrated 2 weeks before tumour inoculation and continued until the end of the experiment. (D) Tumour weight of ABX-treated tumour-bearing mice. Data are representative with n=6 per group. (E, F) Histomorphology of the colon in LLC-bearing mice and inflammation score was evaluated. Score 0: normal colon mucosa with intact epithelium; score 1: scattered inflammatory cell infiltrates in the mucosa; score 2: diffuse mucosal infiltrates without submucosal spreading and intact epithelial layer; score 3: moderate infiltration of inflammatory cells into mucosa and submucosa with epithelial hyperplasia and goblet cell loss; score 4: marked inflammatory cell infiltrates in mucosa and submucosa accompanied by crypt abscesses and loss of goblet cells and crypts; score 5: marked inflammatory cell infiltrates within the mucosa spreading to the submucosa going along with crypt loss and haemorrhage. Original magnification ×100; scale bars 100 µm; black arrowhead-inflammatory cell infiltrates within mucosa (solid) and submucosa (dotted); yellow arrowhead-goblet cell loss. (G, H) Heatmaps showing differential genes and gene ontology (GO) functional analysis in IELs of small intestine between αPD-1 versus combination group and GP versus combination group. (I, J) Heatmaps showing differential genes and GO functional analysis in IELs of small intestine between GP versus combination group and GP versus combination group. Top 20 significantly enriched go terms in cellular compares, molecular function and biological process are presented. GO terms with padj <0.05 are significant enrichment. (K, L) Levels of RORγt+ Treg and Th17 cells in colon laminar propria. data represent mean±SD and analysed by Mann-Whitney U test or Kruskal-Wallis test. *P<0.05, **p<0.01. GPs, ginseng polysaccharides; IELs, intraepithelial lymphocytes; LLC, Lewis lung cancer; mAb, monoclonal antibody; Treg, regulatory T cells.

Article Snippet: A total of 24 mice from different litters were equally divided into four groups: Vehicle (treated with PBS), αPD-1 monoclonal antibody (mAb) (250 μg/mouse, clone: RMP1-14, Bio X Cell), GPs (200 mg/kg) and GPs plus αPD-1 mAb group.

Techniques: MANN-WHITNEY, Functional Assay

Journal: Gut

Article Title: Ginseng polysaccharides alter the gut microbiota and kynurenine/tryptophan ratio, potentiating the antitumour effect of antiprogrammed cell death 1/programmed cell death ligand 1 (anti-PD-1/PD-L1) immunotherapy.

doi: 10.1136/gutjnl-2020-321031

Figure Lengend Snippet: Figure 6 GPs reinstate the response to αPD-1 mAb in LLC tumour-bearing mice transplanted microbiota from non-responders. (A) Schematic diagram of FMT experiment. (B) Tumour growth curves in each group. error bars represent the mean±SEM Tumour growth curves were assessed by two-way ANOVA with Sidak correction. (C–G) CD8+/CD4+ T cell ratio, expression of IFN-γ, TNF-α, granzyme B (GZMB) among CD8 +T cells and Foxp3+ CD4+ T cells in the blood, spleen and tumour tissues. (H) Representative IHC profiles of CD4, CD8, IFN-γ, TNF-α and GZMB in tumour tissues at ×400 magnification. scale bar=50 µm. (I) Quantitative analysis of positive area per field by image J software (NIH). Data presented as mean of 15 fields and expressed as mean±SD (n=3) and were analysed by one-way ANOVA. Each symbol represents an individual animal. *P<0.05, **p<0.01, ***p<0.001. (J) Kynurenine/tryptophan ratio in mice plasma which transferred microbiota from NRs. (K) Representative IHC profiles of IDO in tumour tissues from microbiota colonised mice. (L) Quantitative analysis of positive area per field of IDO expression by image J software (NIH). (M) Relative abundance of Bacterorides in αPD-1 mAb and combination group. (N) Relative abundance of Bacteroides vulgatus in αPD-1 mAb and combination group. (O) Relative abundance of Parabacteroides distasonis in αPD-1 mAb and combination group. ANOVA, analysis of variance; GPs, ginseng polysaccharides; IFN-γ, interferon-γ; mAb, monoclonal antibody; IHC, immunohistochemistry; NR, non-responder; TNF-α, tumour necrosis factor-α.

Article Snippet: A total of 24 mice from different litters were equally divided into four groups: Vehicle (treated with PBS), αPD-1 monoclonal antibody (mAb) (250 μg/mouse, clone: RMP1-14, Bio X Cell), GPs (200 mg/kg) and GPs plus αPD-1 mAb group.

Techniques: Expressing, Software, Clinical Proteomics, Immunohistochemistry

Journal: Gut

Article Title: Ginseng polysaccharides alter the gut microbiota and kynurenine/tryptophan ratio, potentiating the antitumour effect of antiprogrammed cell death 1/programmed cell death ligand 1 (anti-PD-1/PD-L1) immunotherapy.

doi: 10.1136/gutjnl-2020-321031

Figure Lengend Snippet: Figure 7 GPs combined with αPD-1 mAb improve the response rate by reinstating the gut microbiota. (A) GPs potentiated the antitumour effect with αPD-1 mAb via enhancing CD8 T cells function, increasing the production of IFN-γ and TNF-α and reducing the suppression effect of Treg in circulating system, which might be addressed by reshaping gut microbiota and thus influencing tryptophan metabolism and SCFAs. Combination treatment enriched the abundance of Muribaculum when administrated into LLC-bearing mice. Combination treatment upregulated the expression of epithelium protecting genes, like CLCA3, TFF3, AGR2, Zg16, Pla2g10 and Guca2a. The metabolites SFCAs and kynurenine trafficked into circulating system and enhanced the immune function thus suppressing tumour growth and prolonging survival. (B) Combination therapy reinstated gut microbiota which helps to revert gut microbiota from non-responders to responders, thus potentiating the response to αPD-1 mAb. 16S SMRT sequencing found Parabacteroides distasonis and Bacteroides vulgatus were higher in abundance in responders to pembrolizumab. When transplanted the gut microbiota from non-responders to germ-free mice by faecal microbiota transplantation (FMT) and then inoculated LLC tumour cells after colonisation, combination therapy was administrated to the mice. A rich abundance of B. vulgatus and P. distasonis were found in combination group by 16S PacBio SMRT sequencing compared with αPD-1 mAb and vehicle group. Meanwhile, combination therapy significantly suppressed tumour growth by enhancing the function of CD8+ T cells, increasing the production of IFN-γ, TNF-α, and granzyme B and decreasing Treg cells. GPs, ginseng polysaccharides; IFN-γ, interferon-γ; LLC, Lewis lung cancer; mAb, monoclonal antibody; IHC, immunohistochemistry; NRs, non- responder; SFCAs, short-chain fatty acids; TNF-α, tumour necrosis factor-α.

Article Snippet: A total of 24 mice from different litters were equally divided into four groups: Vehicle (treated with PBS), αPD-1 monoclonal antibody (mAb) (250 μg/mouse, clone: RMP1-14, Bio X Cell), GPs (200 mg/kg) and GPs plus αPD-1 mAb group.

Techniques: Expressing, Sequencing, Transplantation Assay, Immunohistochemistry