pan dc isolation kit  (Miltenyi Biotec)

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: Patients with HER2+ breast cancer can have immunity to HSP90. (A) ELISA was used to examine HSP90-specific IgG immunity in 75 patients with HER2+ breast cancer (n=25, stages 1–3 only), triple-negative breast cancer (n=25) and hormone-positive breast cancer (n=25), and 25 healthy donors. (B) Subtype immunity of HSP90-specific IgG was evaluated in 32 patients with HER2+ breast cancer and 25 healthy donors. (C) In the IgG1 and IgG3 (B), patients with HER2+ breast cancer were classified by stages I–III and IV. Bold lines represent the mean levels of HSP90-specific IgG, IgG1, IgG2, IgG3 and IgG4 in each groups. **P<0.01, ***P<0.001 in Student’s t-test and in Tukey’s multiple comparison test of one-way analysis of variance. HER2+, human epidermal growth factor receptor 2-positive; HSP90, heat shock protein 90; N.S, not significant.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Enzyme-linked Immunosorbent Assay, Comparison

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: Human T-cell responses specific to HSP90 peptides can be recognized. (A) HSP90 peptides with the highest binding affinities across multiple MHC class II alleles. Colors represent final scores from five algorithms for each peptide sequences from dark red to light blue in the order of rank scores. Color strata are as follows: dark red: >9000, red: 8000–9000, orange: 7000–8000, light orange: 6000–7000, gold: 5000–6000, yellow: 4000–5000, light yellow: 3000–4000, light green: 2000–3000, and light blue: 1000–2000. (B, C) HSP90 peptides were profiled antigen-specific responses in human PBMCs from 10 healthy donors using IFN and IL-10 enzyme-linked immunospot assay. Per cent responding donors to HSP90 epitopes (B) and CSPW (C). White columns, IFN-γ; black columns, IL-10; horizontal bars indicate mean CSPW. CSPW, corrected spots per well; HSP90, heat shock protein 90; IFN, interferon; IFN-γ, interferon gamma; IL, interleukin; PBMC, peripheral blood mononuclear cell.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Binding Assay, Enzyme-linked Immunospot

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: HSP90-specific immunity inhibits tumor growth in MMTV neu -transgenic mice. (A) Experimental design and in vivo administration schedules for tumorigenesis (n=5 per groups) and T-cell depletion (n=5 per groups) in mouse models. (B) Images of the tumor and the mean tumor volume in control (PBS) and HSP90 peptide (p485 and p527) groups. (C) Two selected HSP90 peptides were evaluated HSP90-specific T-cell responses using IFN-γ ELISPOT assay. (D) Tumor growth is shown for individual mice in control (PBS+rat IgG), HSP90 peptides, HSP90 peptides+anti-CD4, HSP90 peptides+anti-CD8 and HSP90 peptides+anti-CD4 + anti-CD8 groups. Error bars represent SD. (E) Experimental design (n=3 per group) and schedule for cross-priming CD8+ T cells with HSP90 peptides. (F) HSP90 peptides pulsed splenic DCs with HSP90 peptides were evaluated using the cross-priming responses of antigen-specific CD8+ T cells using IFN-γ ELISPOT assay. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001 in Student’s t-test and in Tukey’s multiple comparison test of one-way and in Bonferroni post-tests of two-way analysis of variance. CSPW, corrected spots per well; DC, dendritic cell; ELISPOT, enzyme-linked immunospot; HSP, heat shock protein; HSP90, heat shock protein 90; IFN-γ, interferon gamma; MMC, mouse mammary carcinoma; N.S, not significant; TT, tetanus toxoid.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Transgenic Assay, In Vivo, Control, Enzyme-linked Immunospot, Comparison

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: Combination effect of HSP90 peptides and STING agonist improves antitumor effect and survival in MMTV neu -transgenic mice. (A) Experimental design showing in vivo administration schedules for therapy (n=3 per groups) and survival (n=6 per groups) due to the combination effect in mouse model. (B) Tumor volume in the control (PBS+dimethyl sulfoxide (DMSO), HSP90 peptides, STING agonist, and HSP90 peptides+STING agonist groups. (C) Kaplan-Meier survival curves of mice (six per groups) implanted with MMC cells and treated with the combination of HSP90 peptides and STING agonist. Statistical analysis of Kaplan-Meier survival curves was performed by log-rank (Mantel-Cox) test in comparison of survival curves using Prism program. (D) HSP90-specific T-cell responses and (E) potential epitope-spreading responses of HER2, c-MET and HIF-1α peptide were evaluated by enzyme-linked immunospot using splenocytes from experimental groups. (F) Intratumoral TCR repertoire analysis; mean productive rearrangements (left) and productive Simpson clonality of TCR by sequencing of TCRβ CDR region from genomic DNA of frozen mice tissues (n=3 per groups). Bars represent CSPW and error bars represent SD. *P 0.05, **P< 0.01, ***P<0.001 in Bonferroni post hoc tests of one-way and two-way analyses of variance. CSPW, corrected spots per well; HER2, human epidermal growth factor receptor 2; HSP, heat shock protein; HSP90, heat shock protein 90; MMC, mouse mammary carcinoma; N.S, not significant; STING, stimulator of interferon genes; TCR, T-cell receptor; TT, tetanus toxoid.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Transgenic Assay, In Vivo, Control, Comparison, Enzyme-linked Immunospot, Sequencing

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: Combination effect of HSP90 peptides/STING agonist/anti-CTLA-4 Ab induces immune environment and epitope spreading by effective antitumor immunity. (A) Experimental design (n=4 per groups) and schedule of the combination therapy of HSP90 peptide in the mouse model. (B) Tumor growth is shown for individual mice in control (PBS+hamster IgG+DMSO), HSP90 peptides, HSP90 peptides+anti-CTLA-4 Ab, STING agonist+anti-CTLA-4 Ab and HSP90 peptides+STING agonist+anti-CTLA-4 Ab groups. (C) HSP90-specific T-cell responses and (D) potential epitope-spreading responses of HER2, c-MET and HIF-1α peptide were evaluated by enzyme-linked immunospot using splenocytes from experimental groups. Bars represent CSPW and error bars represent SD. N.S, not significant. *P<0.05, **P<0.01, ***P<0.001 in Tukey’s multiple comparison test of one-way ANOVA and in Bonferroni post-tests of two-way ANOVA. Ab, antibody; ANOVA, analysis of variance; CTLA-4, cytotoxic T lymphocyte-associated antigen-4; HER2, human epidermal growth factor receptor 2; HSP, heat shock protein; HSP90, heat shock protein 90; MMC, mouse mammary carcinoma; STING, stimulator of interferon genes; TCR, T-cell receptor; TT, tetanus toxoid.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Control, Enzyme-linked Immunospot, Comparison

Journal: Journal for Immunotherapy of Cancer

Article Title: Novel peptide-based vaccine targeting heat shock protein 90 induces effective antitumor immunity in a HER2+ breast cancer murine model

doi: 10.1136/jitc-2022-004702

Figure Lengend Snippet: Multiplex IHC reveals immune system distribution of tumor microenvironment in tumor tissues. (A) Representative staining results from multiplex IHC assay of CD4, CD8, Foxp3, CTLA-4, CD11b, and Gr-1 in tumor tissues of mice. (B, C) Counts and percentage of total T cells per area (mm 2 ) of tumor-infiltrating T-cell phenotypes by multiplex IHC in tumor tissues of mice. (D, E) Boxplots showing median cell counts per area (mm 2 ) of CD8 + T cell:Treg (CD3 + CD4 + Foxp3 + ) ratio and MDSCs (CD11b + Gr-1 + ) obtained from five treatment conditions. Statistical significance derived from Kruskal-Wallis teats was compared using the stat compare means function, and the overall significance is indicated by the p value. ***P<0.0001 in χ 2 test of one-way analysis of variance. CTLA-4, cytotoxic T lymphocyte-associated antigen-4; HSP, heat shock protein; HSP90, heat shock protein 90; IHC, immunohistochemistry; MDSC, myeloid-derived suppressor cell; N.S, not significant; STING, stimulator of interferon genes; Treg, regulatory T.

Article Snippet: To assess the cross-priming of antigen-specific CD8 + T cells, HSP90 peptides pulsed splenic dendritic cells (DCs) (130-100-875; Miltenyi Biotec, California, USA) or splenic DCs were injected intravenously into the tail of MMTV neu -transgenic mice.

Techniques: Multiplex Assay, Staining, Derivative Assay, Immunohistochemistry