rmil12  (R&D Systems)

Journal: Journal of Extracellular Vesicles

Article Title: PlexinA1 (PLXNA1) as a novel scaffold protein for the engineering of extracellular vesicles

doi: 10.1002/jev2.70012

Figure Lengend Snippet: POIs fused with PLXNA1 truncations maintained their activities. (a) Schematic diagram of NanoLuc luciferase‐based assay of HepG2 cells adding PLXNA1 (863‐1316) EVs loaded with L7AE and mRNA of NanoLuc‐C/DBOX. (b) Relative levels of NanoLuc mRNA in HepG2 cell groups absorbed the EVs double‐loaded with NanoLuc‐C/DBOX and L7AE and the EVs loaded with NanoLuc‐C/DBOX. Mean ± SD. n = 3. ** p < 0.01. (c) Relative light units (RLUs) were measured by NanoLuc luciferase‐based assay in the indicating HepG2 cell groups. Mean ± SD. n = 3. ns p > 0.05, ** p < 0.01, **** p < 0.0001. (d) RLUs were measured by NanoLuc luciferase‐based assay in the indicating groups at different dosages. Mean ± SD. n = 3. (e) Detection of IFN‐γ secreted by mouse lymphocytes. The lymphocytes were incubated with rmIL12 and EV‐mIL12 at different dosages. EC50 and R squared of the line graphs were indicated. Mean ± SD. n = 3 in each dosage. (f) Mice bearing MC38 subcutaneous tumours were dosed intratumour into flank tumour three times every other day (Day 0, Day 2 and Day 4) with control (PBS), rmIL12 (100 ng), EVmIL12 (100 ng) and EVmIL12 (500 ng). The subcutaneous tumour volumes were measured every other day. Mean + SD. n = 6. ns p > 0.05, ** p < 0.01, **** p < 0.0001. (g) After dissection at Day 16th, the tumour volumes were measured. Mean ± SD. n = 6 in each group. ns p > 0.05, * p < 0.05, ** p < 0.01.

Article Snippet: Different doses of rmIL12 (10051‐ML‐050/CF, R&D Systems, USA) and EV‐derived mIL12 were respectively incubated with the mouse lymphocytes under conditions of 37°C and 5% CO 2 for 40 h. Following low‐speed centrifugation, the supernatants were collected and analyzed for IFN‐γ levels using an Elisa kit (SEKM‐0031, Solarbio, China).

Techniques: Luciferase, Incubation, Control, Dissection

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: a Generic treatment schedule for all tumor models. Red arrows indicate days when analyses were performed. Green arrows denote days of NHS-rmIL12 or rmIL12 treatment and “X” denotes day of first dosing. Blue denotes duration of Entinostat (ENT) dosing. b Individual tumor growth curves of EMT6-tumor-bearing mice treated as depicted in ( a ). Gray shaded area indicates ENT treatment and dashed line depicts NHS-rmIL12 or rmIL12 dosing. CR, complete response. c Graph shows survival and table shows median overall survival (mOS), P < 0.0001. PBS vs. ENT+NHS-rmIL12, P = 0.0003; PBS vs. ENT+rmIL12, P = 0.006; ENT vs. ENT+NHS-rmIL12, P = 0.0013; NHS-rmIL12 vs. ENT+NHS-rmIL12, P = 0.0001. d Individual tumor growth curves and ( e ) survival (inset indicates mOS) for MC38-tumor-bearing mice treated as depicted in ( a ). For survival, P < 0.0001. PBS or ENT or NHS-rmIL12 vs. ENT+NHS-rmIL12, P < 0.0001. f Individual tumor growth curves and ( g ), survival (inset indicates mOS) of CT26 tumor-bearing mice treated as depicted in ( a ). For survival, P = 0.0004. PBS vs. ENT+NHS-rmIL12, P = 0.0030; ENT vs. ENT+NHS-rmIL12, P = 0.0005; NHS-rmIL12 vs. ENT+NHS-rmIL12, P = 0.0120. h Naïve ( n = 5) and combination-treated mice previously cured from EMT6 tumors ( n = 9) were implanted with EMT6 in the right mammary fat pad and TS/A in the opposing left as depicted in the schematic. Mouse icon adapted from BioRender.com. All other elements in the schematic were inserted from Microsoft Powerpoint for Mac v 16.49 (Microsoft). Graph indicates tumor growth on each side as mean ± SEM. Table denotes number of mice with protective memory. Data from one independent experiment, except for CT26 and EMT6 tumor growth and survival with ENT+NHS−rmIL12 treatment, which is representative of two and three experiments conducted independently with similar results, respectively. For survival analysis, two-tailed Log-rank (Mantel–Cox) was used for comparisons. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Two Tailed Test

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: a Representative hematoxylin and eosin (H&E) images of EMT6 tumors treated with PBS or Entinostat (ENT) with areas of necrosis indicated in red. Necrosis quantification in EMT6 ( n = 7) and MC38 ( n = 4) tumors after 14 days on ENT. EMT6: P = 0.0253; MC38: P = 0.0309. b EMT6 tumor-bearing mice ( n = 4) were treated as indicated in the schematic, and % increase in fluorescence (Fl) intensity over the average autofluorescence of PBS- and ENT-treated mice was quantified. P = 0.0422. c Graphs show quantification of human IgG in the sera ( n = 5, P < 0.0001) or TME supernatant ( n = 9, P < 0.0001) from individual EMT6 tumor-bearing mice on day 21 treated as indicated in Fig. . Correlation of human IgG TME supernatant levels with tumor volume change between days 14 and 20, n = 9. Serum levels ( n = 6) of ( d ) IL-12p70 ( P < 0.0001), ( e ) TNFα ( P < 0.0001) and ( f ) IFNγ (d17 and d25, P < 0.0001; d21, P = 0.0025) at days 17, 21 and 25 post-tumor implant from EMT6 tumor-bearing mice treated as in Fig. . g Correlation between change in tumor volume from day 15 to day 20 with IFNγ serum levels on day 21, NHS-rmIL12, n = 7; ENT+NHS-rmIL12, n = 6. Correlation plots show values from individual mice. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. Data in panels ( a , b ) and day 17 cytokine levels are from one experiment. All remaining data are representative of pooled data from two experiments conducted independently with similar results. Unpaired Student’s t test ( a , b ), Pearson’s correlation coefficient ( c right panel, g ), ordinary one-way ANOVA with Tukey’s multiple comparisons test ( c – f ). Statistics are all two-sided. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. Scale bar: 1 mm. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Fluorescence

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: EMT6 tumor-bearing mice were treated with PBS, Entinostat (ENT), NHS-rmIL12, or ENT+NHS-rmIL12 (combo) as depicted in Fig. . Two days after the last NHS-rmIL12 dose (day 21), tumor-infiltrating CD45 + cells were profiled by scRNAseq ( n = 5 tumors/group) and flow cytometry. Shown are merged UMAP plots of ( a ) identified immune subsets, ( b ) selected marker gene expression, and ( c ) frequencies of lymphoid subsets from scRNAseq analysis. Each dot represents one single cell and is colored according to cell clusters ( a ) or in red ( b ). d Representative flow cytometric analysis of FoxP3 expression on CD4 + T cells in EMT6 tumors. Flow cytometry quantification of ( e ) Tregs (d17: n = 6; d21: PBS and NHS-rmIL12, n = 11; ENT and combo, n = 12, P = 0.0055; d25: n = 11, except combo, n = 12, P < 0.0001), ( f ) CD8 + T cells (PBS and combo, n = 12; ENT, n = 10; NHS-rmIL12, n = 11; P = 0.0013) and ( g ) ratio of CD8 + T cells to Tregs in EMT6 tumors on days 17 ( n = 6), 21 (PBS and NHS-rmIL12, n = 11; ENT and combo, n = 12; P < 0.0001), and 25 ( n = 12 except ENT, n = 10; P < 0.0001) post-tumor implant. h – j MC38-tumor-bearing mice were treated as indicated in Supplementary Fig. . Quantification of ( h ) number of Tregs ( P < 0.0001), ( i ) CD8 + T cells and ( j ) ratio of CD8 + T cells to Tregs ( P = 0.0009) in MC38 tumors ( n = 5, except combo, n = 4) at day 21 post-tumor implant by flow cytometry. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. One-way ANOVA with Tukey’s multiple comparisons test. Statistics are all two-sided. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. The data from scRNA-seq, EMT6 flow cytometry on day 17, and MC38 flow cytometry are from single independent experiments. All day 21 and remaining day 25 data are representative of pooled data from two experiments conducted independently with similar results. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Flow Cytometry, Marker, Gene Expression, Expressing

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: EMT6 tumor-bearing mice were treated with PBS, Entinostat (ENT), NHS-rmIL12 or ENT+rmNHS-IL12 as depicted in Fig. . a Selected list of gene pathways upregulated on CD8 + TILs from combination-treated mice relative to PBS controls by scRNASeq. b Representative image of ELISPOT wells of day 21 splenocytes from each treatment group cultured with EMT6 tumor cells. Graphs show IFNγ spot forming cells (SFC) from splenocytes of individual mice ( n = 6) on day 21 (EMT6, P = 0.0008; TS/A, P = 0.7239) or ( c ) day 25 ( P = 0.0008) post-tumor implant incubated with EMT6 or TS/A tumor cells. Data expressed per 5 × 10 5 splenocytes/well. d Correlation between day 21 and 25 ELISPOT data and tumor volume change between days 14 to 18 and days 21 to 24 for each combination-treated mouse, respectively. e Graph shows IFNγ/TNFα double producing, bifunctional CD8 + splenic T cells on day 25 post-tumor implant ( n = 12, P < 0.0001). f Correlation of cytokine data in panel ( e ) to tumor volume change between days 14 to 18 for each combination-treated mouse. Expression of ( g ) granzyme B ( P < 0.0001) and ( h ) Ki67 (d17, P = 0.0054; d21 and d25, P < 0.0001) in CD8 + TILs as a measure of T-cell activation on days 17 ( n = 6), 21 ( n = 12), and 25 ( n = 12) post-tumor implant. i Frequency of IFNɣ and TNFα double producing, bifunctional CD8 + TILs without ex vivo stimulation on days 21 ( P < 0.0001) and 25 ( P < 0.0001) post-tumor implant, n = 12. Correlation plots show values from individual mice. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. For correlations, Pearson’s correlation coefficient was performed. One-way ANOVA with Tukey’s multiple comparisons test. Statistics are all two-sided. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. scRNA-seq, day 25 ELISPOT and day 17 data are from single independent experiments. Day 21 and remaining day 25 data are representative of pooled data from two experiments conducted independently with similar results. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Enzyme-linked Immunospot, Cell Culture, Incubation, Expressing, Activation Assay, Ex Vivo

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: EMT6-tumor-bearing mice were treated with PBS, Entinostat (ENT), NHS-rmIL12 or ENT+NHS-rmIL12 (combo) as depicted in Fig. . Two days after the last NHS-rmIL12 dose (day 21), tumor-infiltrating CD45 + cells were analyzed by scRNAseq and flow cytometry. UMAP plots show ( a ) expression of designated chemokine genes associated with T-cell chemotaxis, and ( b ) major immune subset clusters identified by scRNAseq. Each dot represents one single cell and is colored in red ( a ) or according to major immune cell subsets identified ( b ). c CCR5 expression on splenic CD44 hi CD8 + T cells by flow cytometry on days 21 ( n = 12, P < 0.0001) and 25 ( n = 6, P = 0.0001) post-tumor implant. Proteomic quantification of ( d ) CCR5- and CXCR3-associated chemokines ( n = 6; d25: CCL3, P = 0.0066; CCL4, P = 0.0111; CCL5, P = 0.0047; CXCL9, P = 0.0019; CXCL10, P = 0.0051), and ( e ) designated cytokines (IFNγ: PBS, n = 10; ENT, n = 6; NHS-rmIL12, n = 5; combo, n = 8; P = 0.0019; GM-CSF: n = 6, except PBS and combo, n = 12; P = 0.0005; IFNβ: n = 6, except PBS and combo, n = 12; P = 0.0008) in tumor microenvironment (TME) supernatant on designated days post-tumor implant. Lower panels show UMAP gene expression clusters for the respective encoding genes. f Upregulated GO/HALLMARK gene pathways elicited by combination therapy in combined neutrophil gene clusters relative to PBS controls. g Proteomic quantification of designated cytokines (left panels, IL1α: PBS, n = 12; ENT and NHS-rmIL12, n = 6; combo, n = 9; P = 0.0016; IL1β: PBS, n = 12; ENT and NHS-rmIL12, n = 6; combo, n = 11; P = 0.0015), correlation between cytokine levels and tumor volume change between days 14 and 18 for each combination-treated mouse (middle panels), and UMAP gene expression of designated encoding genes (right panels). Correlation plots show values from individual mice. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. Pearson’s correlation coefficient ( g , middle panels), and ordinary one-way ANOVA with Tukey’s multiple comparisons test ( c – g ). Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. Statistics are all two-sided. The data from RNA-seq, and TME chemokine proteins on day 25 are from single independent experiments. All day 21 and remaining day 25 data are representative of pooled from two experiments conducted independently with similar results. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Flow Cytometry, Expressing, Chemotaxis Assay, Gene Expression, RNA Sequencing

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: EMT6 tumor-bearing mice were treated as depicted in Fig. . a Frequencies of TAM populations identified by scRNAseq. b Top 10 upregulated gene pathways on TAM gene clusters identified by scRNAseq as not M1- or M2-like from combination-treated mice relative to PBS controls. c Relative frequency of M1- and M2-like TAMs by scRNAseq. d Top 10 upregulated gene pathways on M1-like TAM gene clusters identified by scRNAseq from combination-treated mice relative to PBS controls. e Flow cytometry quantification of TAM infiltration on days 21 ( n = 12, except ENT, n = 11; P = 0.0056) and 25 ( n = 12, except ENT and NHS-rmIL12, n = 11; P = 0.0015) post-tumor implant. Expression of M1-like markers ( f ) IA/IE (d21: PBS and NHS-rmIL12, n = 12; ENT, n = 11; combo, n = 8; P < 0.0001; d25: n = 12, except ENT, n = 11; P < 0.0001), ( g ) CD38 + CD206 - (d21: n = 12, except PBS and combo, n = 11; P < 0.0001; d25: n = 12, except PBS, n = 11; P < 0.0001) and ( h ) NOS2 (d21: n = 10, except NHS-rmIL12, n = 11; P < 0.0001; d25: n = 6, P < 0.0001) on TAMs. Representative histograms of NOS2 flow cytometry data from day 21 post-tumor implant. Expression of M2-like markers ( i ) CD38 - CD206 + (d21: n = 12, except PBS and combo, n = 11; P < 0.0001; d25: n = 12, except NHS-rmIL12, n = 11; P < 0.0001) and ( j ) Arg1 (d21: n = 11, except combo, n = 10; P = 0.0085; d25: n = 12, P < 0.0001) on TAMs. Correlation of M1-like or M2-like TAM frequency to tumor volume change between days 14 to 18 for each combination-treated mouse using the markers NOS2/Arg1 ( k ) or CD38/CD206 ( l ). M1 to M2 ratios using the markers CD38/CD206 (d21: PBS and combo, n = 10; ENT, n = 9; NHS-rmIL12, n = 11; P = 0.0004; d25: PBS and combo, n = 11; ENT, n = 10; NHS-rmIL12, n = 12; P < 0.0001) ( m ) or NOS2/Arg1 (d21: n = 11, except combo, n = 10; P < 0.0001; d25: PBS and combo, n = 12; ENT and NHS-rmIL12, n = 11; P = 0.0002) ( n ). TAM infiltration ( n = 5) ( o ) and M1/M2 ratios on day 21 post-tumor implant using the markers CD38/CD206 ( n = 5, except combo, n = 4; P < 0.0001) ( p ) or NOS2/Arg1 ( n = 5, except combo, n = 3; P < 0.0001) ( q ) in MC38 tumors treated as indicated in Supplementary Fig. . Correlation plots show values from individual mice. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. Pearson’s correlation coefficient ( l ), one-way ANOVA with Tukey’s multiple comparisons test ( e – j , m – q ). Statistics are all two-sided. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. scRNA-seq and MC38 data are from single independent experiments. EMT6 day 21 and 25 data representative of pooled from two experiments conducted independently with similar results. Combo, ENT+NHS-rmIL12. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Flow Cytometry, Expressing

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: a Tumor growth of EMT6 tumor-bearing athymic nu/nu mice (PBS and ENT, n = 10; NHS-rmIL12, n = 8; combo, n = 9; P < 0.0001) treated with Entinostat (ENT) and/or NHS-rmIL12 as indicated in the schematic. b Tumor growth ( P < 0.0001) and ( c ) survival ( P < 0.0001) in syngeneic Balb/c mice after treatment with ENT+NHS-rmIL12 as in Fig. with or without CD8 depletion, n = 9. Inset indicates median overall survival (mOS). For all tumor growth curves, gray shaded area indicates Entinostat treatment and dashed lines are NHS-rmIL12 doses. CD8 depletion period is indicated with orange shading. d Top 10 pathways downregulated in TAM gene clusters from CD8-depleted mice treated with combination therapy vs. undepleted. e Expression by scRNAseq of select M1 ( Cd38, Nos2 ) and M2-associated ( Mrc1, Arg1 ) genes in CD45 + cells isolated from CD8-depleted or undepleted EMT6 tumors. Each red dot represents one single cell. f Effect of CD8 depletion in the relative frequency of M1- and M2-like TAMs by scRNAseq. Mice from depletion study were analyzed 21 days after tumor implant and ( g ) CD8 tumor-infiltrating lymphocytes (TIL) (PBS, n = 6; combo, n = 4, combo/CD8dep, n = 5; P = 0.0028), ( h ) M1-like CD38 + CD206 - TAMs (PBS and combo/CD8dep , n = 6 ; combo, n = 5; P < 0.0001), ( i ) M2-like CD38 - CD206 + TAMs (PBS and combo/CD8dep, n = 6; combo, n = 5; P < 0.0001), and ( j ) the ratio of M1/M2 TAMs using (i) CD38/CD206 (PBS and combo/CD8dep, n = 6; combo, n = 5; P = 0.0030) or (ii) NOS2/Arg1 (PBS and combo/CD8dep, n = 6; combo, n = 4; P < 0.0001) was quantified by flow cytometry. k – l Graphs show quantification of ( k ) IL12p70 (PBS and combo, n = 10; ENT and NHS-rmIL12, n = 6; combo/CD8dep, n = 12; P < 0.0001), TNFα (PBS, n = 11; ENT and NHS-rmIL12, n = 6; combo, n = 8; combo/CD8dep, n = 12; P = 0.0011), IFNγ (PBS, n = 10; ENT, n = 6; NHS-rmIL12, n = 5; combo, n = 11; combo/CD8dep, n = 12; P = 0.0002), and ( l ) IFNβ (PBS, n = 11; ENT and NHS-rmIL12, n = 6; combo, n = 10; combo/CD8dep, n = 12; P = 0.0009), GM-CSF (PBS, n = 11; ENT and NHS-rmIL12, n = 6; combo, n = 11; combo/CD8dep, n = 12; P = 0.0005), IL1α (PBS, n = 12; ENT and NHS-rmIL12, n = 6; combo, n = 10; combo/CD8dep, n = 12; P = 0.0009), and IL1β (PBS, n = 12; ENT and NHS-rmIL12, n = 6; combo, n = 11; combo/CD8dep, n = 12; P = 0.0004) in tumor microenvironment (TME) supernatant on day 21 post-tumor implant in EMT6 tumor-bearing Balb/c mice. m , n Effect of EMT6 tumor CD8 depletion on designated chemokines’ ( m ) gene expression by RNAseq and ( n ) CCL3 ( P = 0.0021), CCL4 ( P = 0.0146), CCL5 ( P = 0.0007), CXCL9 ( P = 0.0014), and CXCL10 ( P = 0.0002) protein levels in TME supernatant (PBS, n = 11; ENT, n = 6; NHS-rmIL12, n = 5; combo and combo/CD8dep, n = 10). Each dot represents one single cell ( e ). Correlation plots show values from individual mice. Truncated violin plots show values from individual mice with contours denoting kernel density distributions; dashed line, median; and dotted line, interquartile range. For survival, log-rank (Mantel–Cox) was used for comparisons. One-way ANOVA with Tukey’s multiple comparisons test was used for all comparisons except two-way ANOVA was used for average tumor growth comparison. Statistics are all two-sided. Grey = p < 0.05, red = p < 0.01, blue = p < 0.001, black = p < 0.0001. Data from single independent experiments ( b – f , m ) or pooled from two experiments conducted independently with similar results ( a , g – l , n ). Combo, ENT+NHS-rmIL12. Source data are provided as a Source Data file.

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Expressing, Isolation, Flow Cytometry, Gene Expression, Comparison

Journal: Nature Communications

Article Title: Tumour-targeted interleukin-12 and entinostat combination therapy improves cancer survival by reprogramming the tumour immune cell landscape

doi: 10.1038/s41467-021-25393-x

Figure Lengend Snippet: a Heatmap of Spearman’s correlations between genes encoding key immune markers responsible for the antitumor efficacy of NHS-rmIL12 plus Entinostat treatment in murine models, and immune infiltration in various human cancers. Data analysis from designated TCGA datasets was generated using the open-source platform TIMER2.0 and heat maps generated using GraphPad Prism 7. red = Spearman Rho >0 corresponds to positive correlation; blue = Spearman Rho <0 corresponds to negative correlation; not significant Spearman Rho are crossed out. b Corresponding dot plot of two-tailed Spearman’s correlation of the TCGA breast cancer data set (BRCA, n = 1100). Line represents best-fitting regression line with 95% confidence interval (grey shading). c Correlation between the key immune signature (IL12B, IFNG, CD8B, GZMA, CD38) observed in murine models with clinical overall survival of human patients with various cancer types. Kaplan–Meier curves were generated using the open-source platform GEPIA2, with a median high/low 50% cutoff. BLCA, bladder cancer; BRCA, breast cancer; CESC, cervical squamous cell cancer; COAD, colon adenocarcinoma; ESCA, esophageal carcinoma; HNSC, head and neck squamous carcinoma; KIRP, kidney renal papillary cell carcinoma; LGG, low-grade glioma; LIHC, liver hepatocellular carcinoma; LUAD, lung adenocarcinoma; LUSC, lung squamous cell adenocarcinoma; OV, ovarian cancer; PAAD, pancreatic adenocarcinoma; PRAD, prostate adenocarcinoma; SARC, sarcoma; SKCM, skin cutaneous melanoma; STAD, stomach adenocarcinoma. Mouse and human icons ( a , c ) were imported from powerpoint (Microsoft Powerpoint for Mac v 16.49, Microsoft).

Article Snippet: Recombinant murine IL-12 (rmIL12) was obtained from Peprotech.

Techniques: Generated, Two Tailed Test