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anti tim3 antibody  (Proteintech)


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    Proteintech anti tim3 antibody
    Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of <t>TIM3,</t> PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.
    Anti Tim3 Antibody, supplied by Proteintech, used in various techniques. Bioz Stars score: 95/100, based on 46 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/anti+tim3+antibody/TIM3+Antibody/pmc12674200-170-140-146
    Average 95 stars, based on 46 article reviews
    anti tim3 antibody - by Bioz Stars, 2026-10
    95/100 stars

    Images

    1) Product Images from "Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects"

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects

    Journal: International Journal of Oncology

    doi: 10.3892/ijo.2025.5823

    Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of TIM3, PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.
    Figure Legend Snippet: Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of TIM3, PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.

    Techniques Used: Western Blot, Expressing, Flow Cytometry, Fluorescence, Cell Culture, Control, Activation Assay

    Compound 053 enhances the in vivo antitumor effect of GEM in the 4T1 xenograft model. (A) Mouse treatment plan. (B) H&E staining of heart, liver, spleen, lung and kidney. Scale bar, 100 μ m. (C) Tumor morphology of mouse removed at 21 days of treatment. (D) The tumor size was measured with a caliper every 3 days and the volume was calculated (n=5). (E) Protein levels of CD3, CD4, CD8, 4-1BB, OX40, PD-1 and TIM3 were detected using immunohistochemical staining in tumor tissues of mice after 21 days of treatment. Nuclei are localized in blue and target proteins are localized in brown. Scale bar, 100 μ m. (F) Body weight was measured every 3 days (n=5). 053, FZU-0045-053; GEM, gemcitabine; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3.
    Figure Legend Snippet: Compound 053 enhances the in vivo antitumor effect of GEM in the 4T1 xenograft model. (A) Mouse treatment plan. (B) H&E staining of heart, liver, spleen, lung and kidney. Scale bar, 100 μ m. (C) Tumor morphology of mouse removed at 21 days of treatment. (D) The tumor size was measured with a caliper every 3 days and the volume was calculated (n=5). (E) Protein levels of CD3, CD4, CD8, 4-1BB, OX40, PD-1 and TIM3 were detected using immunohistochemical staining in tumor tissues of mice after 21 days of treatment. Nuclei are localized in blue and target proteins are localized in brown. Scale bar, 100 μ m. (F) Body weight was measured every 3 days (n=5). 053, FZU-0045-053; GEM, gemcitabine; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3.

    Techniques Used: In Vivo, Staining, Immunohistochemical staining

    Compound 053 modulates PD-L1 on tumor cells and co-stimulatory/co-inhibitory molecules on tumor-infiltrating T cells in the 4T1 xenograft model. (A) Flow cytometry was performed to analyze PD-L1 surface expression on tumor cells. The bar graph shows quantitative analysis of relative fluorescence intensity. (B) PD-1 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (C) TIM3 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) 4-1BB expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) ICOS expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) OX-40 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. (G) Flow cytometry was used to analyze the differentiation of CD3 + CD4 + T cells and CD3 + CD8 + T cells. 053, FZU-0045-053; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; GEM, gemcitabine; PD-L1, programmed death-ligand 1.
    Figure Legend Snippet: Compound 053 modulates PD-L1 on tumor cells and co-stimulatory/co-inhibitory molecules on tumor-infiltrating T cells in the 4T1 xenograft model. (A) Flow cytometry was performed to analyze PD-L1 surface expression on tumor cells. The bar graph shows quantitative analysis of relative fluorescence intensity. (B) PD-1 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (C) TIM3 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) 4-1BB expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) ICOS expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) OX-40 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. (G) Flow cytometry was used to analyze the differentiation of CD3 + CD4 + T cells and CD3 + CD8 + T cells. 053, FZU-0045-053; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; GEM, gemcitabine; PD-L1, programmed death-ligand 1.

    Techniques Used: Flow Cytometry, Expressing, Fluorescence, Control

    Mechanism of compound 053 regulating autophagy and immune response to inhibit breast cancer. 053 can inhibit autophagy and promote apoptosis of breast cancer cells. 053 also downregulated PD-L1 expression in breast cancer cells, while promoting T cell activation and proliferation by upregulating co-stimulatory molecules (4-1BB, OX40 and ICOS) and downregulating co-inhibitory molecules (TIM-3 and PD-1) on the surface of the T cells. (By Figdraw). 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; PD-L1, programmed death-ligand 1.
    Figure Legend Snippet: Mechanism of compound 053 regulating autophagy and immune response to inhibit breast cancer. 053 can inhibit autophagy and promote apoptosis of breast cancer cells. 053 also downregulated PD-L1 expression in breast cancer cells, while promoting T cell activation and proliferation by upregulating co-stimulatory molecules (4-1BB, OX40 and ICOS) and downregulating co-inhibitory molecules (TIM-3 and PD-1) on the surface of the T cells. (By Figdraw). 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; PD-L1, programmed death-ligand 1.

    Techniques Used: Expressing, Activation Assay

    Related Articles

    Incubation:

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects
    Article Snippet: .. The sections were then blocked with 5% BSA for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: Anti-Ki67 antibody (cat. no. GB111499-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-LC3 antibody (cat. no. 81004-1-RR; 1:1,000; Proteintech Group, Inc), anti-p62 antibody (cat. no. GB11239-1-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L1 antibody (cat. no. GB150059-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L2 antibody (cat. no. 27406-1-AP; 1:4,000; Proteintech Group, Inc), anti-CD3 antibody (cat. no. GB11014-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD4 antibody (cat. no. GB11064-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-4-1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti-CD134/OX40 antibody (cat. no. 32621-1-AP; 1:500; Proteintech Group, Inc), anti-PD1 antibody (cat. no. GB153744-100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti-TIM3 antibody (cat. no. 11872-1-AP; 1:1,000; Proteintech Group, Inc). .. Subsequently, the samples were incubated with HRP-conjugated goat anti-rabbit IgG (cat. no. GB23303; 1:5,000; Wuhan Servicebio Technology Co., Ltd) or HRP-conjugated goat anti-mouse IgG (cat. no. GB23301; 1:5,000; Wuhan Servicebio Technology Co., Ltd) at 37°C for 1 h. Following this, staining was performed with DAB chromogen for 15 min (cat. no. G1211; Wuhan Servicebio Technology Co., Ltd), and hematoxylin was used as a counterstain for 3 min at room temperature.

    Article Title: Discovery of the late autophagy inhibitor FZU‑0045‑053 and its anti‑breast cancer and immunomodulatory effects.
    Article Snippet: .. The sections were then blocked with 5% BSA for 1 h at room temperature and subse‐ quently incubated overnight at 4 ̊C with the following primary antibodies: Anti‐Ki67 antibody (cat. no. GB111499‐100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐LC3 antibody (cat. no. 81004‐1‐RR; 1:1,000; Proteintech Group, Inc), anti‐p62 antibody (cat. no. GB11239‐1‐100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐PD‐L1 antibody (cat. no. GB150059‐100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐PD‐L2 antibody (cat. no. 27406‐1‐AP; 1:4,000; Proteintech Group, Inc), anti‐CD3 antibody (cat. no. GB11014‐100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐CD4 antibody (cat. no. GB11064‐100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti‐4‐1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti‐CD134/OX40 antibody (cat. no. 32621‐1‐AP; 1:500; Proteintech Group, Inc), anti‐PD1 antibody (cat. no. GB153744‐100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti‐TIM3 antibody (cat. no. 11872‐1‐AP; 1:1,000; Proteintech Group, Inc). .. Subsequently, the samples were incubated with HRP‐conjugated goat anti‐rabbit IgG (cat. no. GB23303; 1:5,000; Wuhan Servicebio Technology Co., Ltd) or HRP‐conjugated goat anti‐mouse IgG (cat. no. GB23301; 1:5,000; Wuhan Servicebio Technology Co., Ltd) at 37 ̊C for 1 h. Following this, staining was performed with DAB chromogen for 15 min (cat. no. G1211; Wuhan Servicebio Technology Co., Ltd), and hematoxylin was used as a counter‐ stain for 3 min at room temperature.



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    Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of <t>TIM3,</t> PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.
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    Cell Signaling Technology Inc tim3
    Synergistic therapeutic effect of BX471 (CCR1 inhibitor) combined with PD-1 blockade. A Schematic representation of syngeneic models established with B16F10 melanoma cells for combination therapy. Following tumor resection and single-cell dissociation, immune cells of interest were analyzed by flow cytometry. B Tumor growth curves of B16F10 cells treated with BX471 and PD-1 blockade (ICB) either in combination or as monotherapies. C - H ) Flow cytometry analysis of immune cell populations in tumors treated with either BX471, PD-1 blockade (ICB), a combination of both, or placebo. Cell populations analyzed include: CCR1 + macrophages ( C ); M2 macrophages ( D ); M1 macrophages ( E ); <t>TIM3</t> + CD8 + T cells ( F ); PD-1 + CD8 + T cells ( G ) and CD69 + CD8 + T cells ( H ). Data with error bars are shown as mean ± SEM. ns, not significant; *, P <0.05; **, P <0.01; ***, P <0.001; ****, P <0.0001 as determined by unpaired Student t test (B-H)
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    Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of TIM3, PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.

    Journal: International Journal of Oncology

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects

    doi: 10.3892/ijo.2025.5823

    Figure Lengend Snippet: Compound 053 modulates co-stimulatory and co-inhibitory molecules on T cell surfaces. (A) The protein levels of 4-1BB, OX40 and CD40L after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (B) The protein levels of TIM3, PD-1 and VISTA after 5 μ M ADM, GEM (80 μ M), 053 (5 μ M) or CQ (30 μ M) treatment were analyzed by western blotting. (C) After T cells were treated with 053 (0-10 μ M) for 24 h, ICOS expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of ICOS expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) After T cells were treated with 053 (0-10 μ M) for 24 h, OX-40 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of OX-40 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (G) After T cells were treated with 053 (0-10 μ M) for 24 h, 4-1BB expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (H) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of 4-1BB expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (I) After T cells were treated with 053 (0-10 μ M) for 24 h, TIM3 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (J) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of TIM3 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. (K) After T cells were treated with 053 (0-10 μ M) for 24 h, PD-1 expression on T cell surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (L) T cells were co-cultured with MDA-MB-231 cells at a 1:1 ratio and treated with 053 (0-10 μ M) for 24 h, followed by flow cytometric analysis of PD-1 expression on T cell surfaces. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. 053, FZU-0045-053; ADM, adriamycin; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; CD40L, CD40 ligand; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; VISTA, V-domain Ig suppressor of T cell activation; GEM, gemcitabine; CQ, chloroquine.

    Article Snippet: The sections were then blocked with 5% BSA for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: Anti-Ki67 antibody (cat. no. GB111499-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-LC3 antibody (cat. no. 81004-1-RR; 1:1,000; Proteintech Group, Inc), anti-p62 antibody (cat. no. GB11239-1-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L1 antibody (cat. no. GB150059-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L2 antibody (cat. no. 27406-1-AP; 1:4,000; Proteintech Group, Inc), anti-CD3 antibody (cat. no. GB11014-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD4 antibody (cat. no. GB11064-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-4-1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti-CD134/OX40 antibody (cat. no. 32621-1-AP; 1:500; Proteintech Group, Inc), anti-PD1 antibody (cat. no. GB153744-100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti-TIM3 antibody (cat. no. 11872-1-AP; 1:1,000; Proteintech Group, Inc).

    Techniques: Western Blot, Expressing, Flow Cytometry, Fluorescence, Cell Culture, Control, Activation Assay

    Compound 053 enhances the in vivo antitumor effect of GEM in the 4T1 xenograft model. (A) Mouse treatment plan. (B) H&E staining of heart, liver, spleen, lung and kidney. Scale bar, 100 μ m. (C) Tumor morphology of mouse removed at 21 days of treatment. (D) The tumor size was measured with a caliper every 3 days and the volume was calculated (n=5). (E) Protein levels of CD3, CD4, CD8, 4-1BB, OX40, PD-1 and TIM3 were detected using immunohistochemical staining in tumor tissues of mice after 21 days of treatment. Nuclei are localized in blue and target proteins are localized in brown. Scale bar, 100 μ m. (F) Body weight was measured every 3 days (n=5). 053, FZU-0045-053; GEM, gemcitabine; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3.

    Journal: International Journal of Oncology

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects

    doi: 10.3892/ijo.2025.5823

    Figure Lengend Snippet: Compound 053 enhances the in vivo antitumor effect of GEM in the 4T1 xenograft model. (A) Mouse treatment plan. (B) H&E staining of heart, liver, spleen, lung and kidney. Scale bar, 100 μ m. (C) Tumor morphology of mouse removed at 21 days of treatment. (D) The tumor size was measured with a caliper every 3 days and the volume was calculated (n=5). (E) Protein levels of CD3, CD4, CD8, 4-1BB, OX40, PD-1 and TIM3 were detected using immunohistochemical staining in tumor tissues of mice after 21 days of treatment. Nuclei are localized in blue and target proteins are localized in brown. Scale bar, 100 μ m. (F) Body weight was measured every 3 days (n=5). 053, FZU-0045-053; GEM, gemcitabine; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3.

    Article Snippet: The sections were then blocked with 5% BSA for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: Anti-Ki67 antibody (cat. no. GB111499-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-LC3 antibody (cat. no. 81004-1-RR; 1:1,000; Proteintech Group, Inc), anti-p62 antibody (cat. no. GB11239-1-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L1 antibody (cat. no. GB150059-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L2 antibody (cat. no. 27406-1-AP; 1:4,000; Proteintech Group, Inc), anti-CD3 antibody (cat. no. GB11014-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD4 antibody (cat. no. GB11064-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-4-1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti-CD134/OX40 antibody (cat. no. 32621-1-AP; 1:500; Proteintech Group, Inc), anti-PD1 antibody (cat. no. GB153744-100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti-TIM3 antibody (cat. no. 11872-1-AP; 1:1,000; Proteintech Group, Inc).

    Techniques: In Vivo, Staining, Immunohistochemical staining

    Compound 053 modulates PD-L1 on tumor cells and co-stimulatory/co-inhibitory molecules on tumor-infiltrating T cells in the 4T1 xenograft model. (A) Flow cytometry was performed to analyze PD-L1 surface expression on tumor cells. The bar graph shows quantitative analysis of relative fluorescence intensity. (B) PD-1 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (C) TIM3 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) 4-1BB expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) ICOS expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) OX-40 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. (G) Flow cytometry was used to analyze the differentiation of CD3 + CD4 + T cells and CD3 + CD8 + T cells. 053, FZU-0045-053; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; GEM, gemcitabine; PD-L1, programmed death-ligand 1.

    Journal: International Journal of Oncology

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects

    doi: 10.3892/ijo.2025.5823

    Figure Lengend Snippet: Compound 053 modulates PD-L1 on tumor cells and co-stimulatory/co-inhibitory molecules on tumor-infiltrating T cells in the 4T1 xenograft model. (A) Flow cytometry was performed to analyze PD-L1 surface expression on tumor cells. The bar graph shows quantitative analysis of relative fluorescence intensity. (B) PD-1 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (C) TIM3 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (D) 4-1BB expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (E) ICOS expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. (F) OX-40 expression on tumor-infiltrating T cells surfaces was analyzed by flow cytometry. The bar graph shows quantitative analysis of relative fluorescence intensity. Data are presented as mean ± SEM (n=3). Statistical significance was determined by one-way ANOVA followed by Dunnett's multiple comparisons test to compare all experimental groups against a single control group. * P<0.05, ** P<0.01, *** P<0.001, **** P<0.0001 vs. 0 μ M 053. (G) Flow cytometry was used to analyze the differentiation of CD3 + CD4 + T cells and CD3 + CD8 + T cells. 053, FZU-0045-053; 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; GEM, gemcitabine; PD-L1, programmed death-ligand 1.

    Article Snippet: The sections were then blocked with 5% BSA for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: Anti-Ki67 antibody (cat. no. GB111499-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-LC3 antibody (cat. no. 81004-1-RR; 1:1,000; Proteintech Group, Inc), anti-p62 antibody (cat. no. GB11239-1-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L1 antibody (cat. no. GB150059-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L2 antibody (cat. no. 27406-1-AP; 1:4,000; Proteintech Group, Inc), anti-CD3 antibody (cat. no. GB11014-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD4 antibody (cat. no. GB11064-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-4-1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti-CD134/OX40 antibody (cat. no. 32621-1-AP; 1:500; Proteintech Group, Inc), anti-PD1 antibody (cat. no. GB153744-100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti-TIM3 antibody (cat. no. 11872-1-AP; 1:1,000; Proteintech Group, Inc).

    Techniques: Flow Cytometry, Expressing, Fluorescence, Control

    Mechanism of compound 053 regulating autophagy and immune response to inhibit breast cancer. 053 can inhibit autophagy and promote apoptosis of breast cancer cells. 053 also downregulated PD-L1 expression in breast cancer cells, while promoting T cell activation and proliferation by upregulating co-stimulatory molecules (4-1BB, OX40 and ICOS) and downregulating co-inhibitory molecules (TIM-3 and PD-1) on the surface of the T cells. (By Figdraw). 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; PD-L1, programmed death-ligand 1.

    Journal: International Journal of Oncology

    Article Title: Discovery of the late autophagy inhibitor FZU-0045-053 and its anti-breast cancer and immunomodulatory effects

    doi: 10.3892/ijo.2025.5823

    Figure Lengend Snippet: Mechanism of compound 053 regulating autophagy and immune response to inhibit breast cancer. 053 can inhibit autophagy and promote apoptosis of breast cancer cells. 053 also downregulated PD-L1 expression in breast cancer cells, while promoting T cell activation and proliferation by upregulating co-stimulatory molecules (4-1BB, OX40 and ICOS) and downregulating co-inhibitory molecules (TIM-3 and PD-1) on the surface of the T cells. (By Figdraw). 4-1BB, tumor necrosis factor receptor superfamily member 9; OX40, tumor necrosis factor receptor superfamily member 4; ICOS, inducible T-cell co-stimulator; PD-1, programmed cell death protein 1; TIM3, T-cell immunoglobulin and mucin-domain containing-3; PD-L1, programmed death-ligand 1.

    Article Snippet: The sections were then blocked with 5% BSA for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: Anti-Ki67 antibody (cat. no. GB111499-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-LC3 antibody (cat. no. 81004-1-RR; 1:1,000; Proteintech Group, Inc), anti-p62 antibody (cat. no. GB11239-1-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L1 antibody (cat. no. GB150059-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-PD-L2 antibody (cat. no. 27406-1-AP; 1:4,000; Proteintech Group, Inc), anti-CD3 antibody (cat. no. GB11014-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD4 antibody (cat. no. GB11064-100; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-CD8 antibody (cat. no. GB11068; 1:1,000; Wuhan Servicebio Technology Co., Ltd), anti-4-1BB/CD137 antibody (cat. no. A2025; 1:200; ABclonal Biotech Co., Ltd), anti-CD134/OX40 antibody (cat. no. 32621-1-AP; 1:500; Proteintech Group, Inc), anti-PD1 antibody (cat. no. GB153744-100; 1:400; Wuhan Servicebio Technology Co., Ltd) and anti-TIM3 antibody (cat. no. 11872-1-AP; 1:1,000; Proteintech Group, Inc).

    Techniques: Expressing, Activation Assay

    Synergistic therapeutic effect of BX471 (CCR1 inhibitor) combined with PD-1 blockade. A Schematic representation of syngeneic models established with B16F10 melanoma cells for combination therapy. Following tumor resection and single-cell dissociation, immune cells of interest were analyzed by flow cytometry. B Tumor growth curves of B16F10 cells treated with BX471 and PD-1 blockade (ICB) either in combination or as monotherapies. C - H ) Flow cytometry analysis of immune cell populations in tumors treated with either BX471, PD-1 blockade (ICB), a combination of both, or placebo. Cell populations analyzed include: CCR1 + macrophages ( C ); M2 macrophages ( D ); M1 macrophages ( E ); TIM3 + CD8 + T cells ( F ); PD-1 + CD8 + T cells ( G ) and CD69 + CD8 + T cells ( H ). Data with error bars are shown as mean ± SEM. ns, not significant; *, P <0.05; **, P <0.01; ***, P <0.001; ****, P <0.0001 as determined by unpaired Student t test (B-H)

    Journal: Cell Communication and Signaling : CCS

    Article Title: Blocking CCR1 + macrophages overcomes resistance to immune checkpoint inhibitors in melanoma

    doi: 10.1186/s12964-025-02444-0

    Figure Lengend Snippet: Synergistic therapeutic effect of BX471 (CCR1 inhibitor) combined with PD-1 blockade. A Schematic representation of syngeneic models established with B16F10 melanoma cells for combination therapy. Following tumor resection and single-cell dissociation, immune cells of interest were analyzed by flow cytometry. B Tumor growth curves of B16F10 cells treated with BX471 and PD-1 blockade (ICB) either in combination or as monotherapies. C - H ) Flow cytometry analysis of immune cell populations in tumors treated with either BX471, PD-1 blockade (ICB), a combination of both, or placebo. Cell populations analyzed include: CCR1 + macrophages ( C ); M2 macrophages ( D ); M1 macrophages ( E ); TIM3 + CD8 + T cells ( F ); PD-1 + CD8 + T cells ( G ) and CD69 + CD8 + T cells ( H ). Data with error bars are shown as mean ± SEM. ns, not significant; *, P <0.05; **, P <0.01; ***, P <0.001; ****, P <0.0001 as determined by unpaired Student t test (B-H)

    Article Snippet: For human melanoma tissue, the primary antibodies included CCR1 (Absin, abs133089, 1:200), CD68 (Proteintech, 66231-2-Ig, 1:3000), CD8 (Proteintech, 66231-2-Ig, 1:250), and TIM3 (CST, 45208, 1:400).

    Techniques: Flow Cytometry

    Spatial niches of CCR1 + Macrophages and CD8 + Tex Cell in the melanoma tumor microenviroment (TME) and their association with ICB therapy response. A In the melanoma spatial map, the spot-level scores representing the abundance of CD8 + Tex cells and CCR1 + macrophage show a concordant pattern suggesting that the two cell types are co-localised. B Representative mIHC images demonstrating the spatial niches of CCR1 + macrophages and TIM3 + CD8 + T cells in human melanoma tissues. CCR1 is indicated in cyan, CD68 in green, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 20 μm. C Representative mIHC images used to calculate the nearest distance between CCR1 + macrophages and TIM3 + CD8 + T cells in human melanoma tissues. CCR1 is indicated in cyan, CD68 in green, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 100 μm. D Representative mIHC images demonstrating the spatial niches of CCR1 + macrophages and TIM3 + CD8 + T cells in mouse melanoma tissues from different treatment groups. CCR1 is indicated in green, F4/80 in white, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 100 μm. Data with error bars are shown as mean ± SEM. ns, not significant; *, P <0.05; **, P <0.01; ***, P <0.001; ****, P <0.0001 as determined by unpaired Student t test

    Journal: Cell Communication and Signaling : CCS

    Article Title: Blocking CCR1 + macrophages overcomes resistance to immune checkpoint inhibitors in melanoma

    doi: 10.1186/s12964-025-02444-0

    Figure Lengend Snippet: Spatial niches of CCR1 + Macrophages and CD8 + Tex Cell in the melanoma tumor microenviroment (TME) and their association with ICB therapy response. A In the melanoma spatial map, the spot-level scores representing the abundance of CD8 + Tex cells and CCR1 + macrophage show a concordant pattern suggesting that the two cell types are co-localised. B Representative mIHC images demonstrating the spatial niches of CCR1 + macrophages and TIM3 + CD8 + T cells in human melanoma tissues. CCR1 is indicated in cyan, CD68 in green, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 20 μm. C Representative mIHC images used to calculate the nearest distance between CCR1 + macrophages and TIM3 + CD8 + T cells in human melanoma tissues. CCR1 is indicated in cyan, CD68 in green, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 100 μm. D Representative mIHC images demonstrating the spatial niches of CCR1 + macrophages and TIM3 + CD8 + T cells in mouse melanoma tissues from different treatment groups. CCR1 is indicated in green, F4/80 in white, CD8 in yellow, TIM3 in red and DAPI in blue. Scale bars, 100 μm. Data with error bars are shown as mean ± SEM. ns, not significant; *, P <0.05; **, P <0.01; ***, P <0.001; ****, P <0.0001 as determined by unpaired Student t test

    Article Snippet: For human melanoma tissue, the primary antibodies included CCR1 (Absin, abs133089, 1:200), CD68 (Proteintech, 66231-2-Ig, 1:3000), CD8 (Proteintech, 66231-2-Ig, 1:250), and TIM3 (CST, 45208, 1:400).

    Techniques: