p70254  (MedChemExpress)

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: CircRNA CAR efficiently expressed CAR proteins and mediated remarkable tumor killing (A) Schematic representation of circRNA Anti-HER2-CAR circularization via group I intron autocatalysis. (B and C) Detecting the expression of CAR proteins in HEK293T cells after circRNA Anti-HER2-CAR transfection via western blot (B) and flow cytometry (C). (D) Comparative analysis of CAR expression levels from circRNA Anti-HER2-CAR , 1mΨ-mRNA Anti-HER2-CAR , and unmodified mRNA Anti-HER2-CAR in HEK293T cells. (E–G) Optimization of circRNA Anti-HER2-CAR encoding CAR in Jurkat (E), THP-1 (F), and J774A.1 (G). (H) Detection of CAR expression in primary T cells using flow cytometry. (I–K) Cytotoxic effects of primary T cells transfected with circRNA Anti-HER2-CAR on SK-OV-3 (I), B16F10-HER2 (J), and 4T1-HER2 (K) tumor cells. (L–N) Cytotoxic effects of Jurkat cells transfected with circRNA Anti-HER2-CAR on SK-OV-3 (L), B16F10-HER2 (M), and 4T1-HER2 (N) tumor cells. (O–Q) Cytotoxic effects of THP-1 cells transfected with circRNA Anti-HER2-CAR on SK-OV-3 (O), B16F10-HER2 (P), and 4T1-HER2 (Q) tumor cells. In (C)–(Q), data were presented as mean ± SEM ( n = 3). An unpaired two-sided Student’s t test was performed for comparison; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. See also ; .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: Expressing, Transfection, Western Blot, Flow Cytometry, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: Macrophages exhibited efficient tumor phagocytosis and pro-inflammatory polarization induced by circRNA CAR (A and B) Phagocytosis of THP-1 cells transfected with circRNA Anti-HER2-CAR against SK-OV-3 (A) and MC38-HER2 (B) cells. (C) Phagocytosis of J774A.1 cells transfected with circRNA Anti-HER2-CAR against CT26-HER2 cells. (D and E) Effects of circRNA Anti-HER2-CAR on iNOS (D) and CD206 (E) expression in J774A.1. (F and G) Effects of circRNA Anti-HER2-CAR on iNOS (F) and CD206 (G) expression in THP-1 cells. (H) Volcano plot illustrating differentially expressed genes in THP-1 cells. (I) Heatmap depicting gene expression patterns in THP-1 cells ( n = 2). (J) Bubble chart of relevant biological processes via Gene Ontology (GO) analysis ( n = 2). Bubble size represents the number of genes. In (A)–(G), data were presented as mean ± SEM ( n = 3). An unpaired two-sided Student’s t test was conducted for comparison; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. See also .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: Transfection, Expressing, Gene Expression, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: Screening for immunocyte-tropic LNPs that efficiently delivered circRNAs into immune cells in mice (A) Schematic representation of the LNP-circRNA complex. (B and C) The size distribution (B) and zeta potential (C) of the LNP-circRNA Luciferase complex. (D and E) Bioluminescence imaging in vivo (D) or ex vivo (E) of BALB/c mice intravenously injected with PBS or LNP-circRNA Luciferase . (F and G) Bioluminescence imaging in vivo (F) or ex vivo (G) of BALB/c mice intravenously injected with PBS or SORT-circRNA Luciferase . (H) Evaluation of targeting efficiency of SORT-circRNA Cre in the spleen of reporter mice. (I) Detection of anti-HER2-CAR expression at different time points in various immune cells of mouse spleen. In (H) and (I), data were presented as mean ± SEM, an unpaired two-sided Student’s t test was performed for comparison; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. See also .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: Zeta Potential Analyzer, Luciferase, Imaging, In Vivo, Ex Vivo, Injection, Expressing, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: CircRNA CAR efficiently inhibited tumor growth and improved survival time in mice (A) Schematic illustration of intravenous PBS, LNP-circRNA Ctrl , or SORT-circRNA Anti-HER2-CAR treatment in the CT26-HER2 tumor model. (B) Tumor growth curves for CT26-HER2 tumor-bearing mice treated as indicated in (A). (C) Schematic illustration of intratumoral PBS, LNP-circRNA Ctrl , or LNP-circRNA Anti-HER2-CAR treatment in the CT26-HER2 tumor model. (D and E) Tumor growth curves (D) and survival curves (E) of CT26-HER2 tumor-bearing mice treated as indicated in (C). (F) In vivo bioluminescence imaging of CT26-HER2 tumor-bearing mice treated as indicated in (C). (G) The quantified signal intensity of bioluminescence imaging in (F). (H) Schematic illustration of intratumoral PBS, LNP-circRNA Ctrl , or LNP-circRNA Anti-HER2-CAR treatment in the 4T1-HER2 tumor model. (I) Tumor growth curves of 4T1-HER2 tumor-bearing mice treated as indicated in (H). (J) Tumor growth curves of individual 4T1-HER2 tumor-bearing mice treated as indicated in (H). (K) Schematic illustration of intratumoral PBS, LNP-circRNA Ctrl , or LNP-circRNA CAR treatment in the MC38-HER2 tumor model. (L and M) Tumor growth curves (L) and survival curves (M) of MC38-HER2 tumor-bearing mice treated as indicated in (K). In (B), (D), (I), and (L), data were represented as the mean ± SEM, the tumor growth curves were calculated by two-way ANOVA analysis ( n = 6). In (E) and (M), data were represented as the mean ± SEM, the survival curves were calculated by Kaplan-Meier simple survival analysis ( n = 6). In (G), data were represented as the mean ± SEM, an unpaired two-sided Student’s t test was conducted for comparison. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001. See also .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: In Vivo, Imaging, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: In vivo panCAR reshaped the tumor microenvironment to a pro-inflammatory state (A) Changes in the proportion of immune cells of spleen via flow cytometry. CD8 + T cells, CD8 + Tem cells, CD8 + Tcm cells, or CD4 + T cells were gated from CD45 + cell population, and Treg cells were gated from CD4 + T cell population. (B) Flow cytometric analysis of changes in the proportion of infiltrating immune cells in tumors. CD8 + T cells, CD4 + T cells, or MHC II + macrophages were gated from CD45 + cell population, and Treg cells were gated from CD4 + T cell population. (C and D) H&E staining (C) or IHC staining (D) of tumor tissue sections obtained from CT26-HER2 tumor-bearing mice after PBS, SORT-LNP-circRNA Ctrl , or SORT-LNP-circRNA Anti-HER2-CAR treatment. The integrated density of IHC staining was quantified using ImageJ. (E) Heatmap of gene expression patterns of immune cells extracted from tumor tissues ( n = 2). (F) Bubble chart of relevant biological processes through GO analysis ( n = 2). The size of the bubbles represented the number of genes. (G) Gene set enrichment analysis (GSEA) showing enriched pathways in immune cells extracted from tumor tissues ( n = 2). In (A), (B), and (D), data were represented as the mean ± SEM; an unpaired two-sided Student’s t test was conducted for comparison; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. Each symbol represents an individual mouse. See also and .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: In Vivo, Flow Cytometry, Staining, Immunohistochemistry, Gene Expression, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: CircRNA vaccine synergistically boosted the anti-tumor activity of in vivo panCAR (A) Schematic illustration of circRNA VAC design. EPM, the endocytosis prevention motif; EABR, the ESCRT- and ALIX-binding region domain. (B) Flow cytometric analysis detecting the translation of circRNA HER2 in HEK293T cells ( n = 3). (C) Electron microscopy showing the vesicles secreted by HEK293T cells transfected with circRNA HER2-EPM-EABR . (D) Measurement of the endpoint titer of HER2-specific IgG with ELISA. (E) Schematic illustration of 4T1-HER2 tumor-bearing mice receiving PBS (intravenously), circRNA CAR (intravenously), circRNA VAC (intramuscularly), or circRNA CAR (intravenously) plus circRNA VAC (intramuscularly) combined therapy ( n = 5). (F) Tumor growth curves of overall mice treated as indicated in (E). (G) Tumor growth curves of individual mouse treated as indicated in (E). (H) Schematic illustration of B16F10-HER2 tumor-bearing mice receiving PBS (intravenously), circRNA CAR (intravenously), circRNA VAC (intramuscularly), or circRNA CAR (intravenously) plus circRNA VAC (intramuscularly) combined therapy ( n = 5). (I) Tumor growth curves of overall mice treated as indicated in (H). (J) Tumor growth curves of individual mouse treated as indicated in (H). (K) Survival curves of B16F10-HER2 tumor-bearing mice treated as indicated in (H). In (B), data were represented as the mean ± SEM; an unpaired two-sided Student’s t test was performed for comparison. In (D), data were represented as the geometric mean ± geometric SD; an unpaired two-sided Student’s t test was performed for comparison. In (F) and (I), data were represented as the mean ± SEM, and the tumor growth curves were calculated by two-way ANOVA analysis. In (K), the survival curves were calculated by Kaplan-Meier simple survival analysis. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. See also ; .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: Activity Assay, In Vivo, Binding Assay, Electron Microscopy, Transfection, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Cell Reports Medicine

Article Title: Synergically enhanced anti-tumor immunity of in vivo panCAR by circRNA vaccine boosting

doi: 10.1016/j.xcrm.2025.102250

Figure Lengend Snippet: In vivo panCAR enhanced the anti-tumor activity via antibody-mediated cellular cytotoxicity (A) Measurement of HER2-specific IgG, IgG1, IgG2A, IgG2B, or IgG2C-binding antibodies with ELISA ( n = 5 or 6). (B and C) HER2-specific antibodies in (A) mediated cellular cytotoxicity against SK-OV-3 (B) and MC38-HER2 (C) tumor cells in J774A.1. (D and E) HER2-specific antibodies in (A) mediated cellular cytotoxicity against SK-OV-3 (D) and MC38-HER2 (E) tumor cells in RAW 264.7. (F) Tumor growth curves of overall CT26 tumor-bearing mice treated with PBS, in vivo panCAR-VAC, or in vivo panCAR-VAC plus anti-NK1.1 antibodies to deplete NK cells ( n = 5). (G) Tumor growth curves of individual mouse treated as indicated in (F). (H) Survival curves of mice treated as indicated in (F). (I) Tumor growth curves of overall CT26 tumor-bearing mice treated with PBS, in vivo panCAR-VAC, or in vivo panCAR-VAC plus anti-CSF1R antibody to deplete macrophages ( n = 5). (J) Tumor growth curves of individual mouse treated as indicated in (I). (K) Survival curves of mice treated as indicated in (I). (L) The potential mechanism diagram of synergistic in vivo panCAR-VAC immunotherapy. In (A), data are shown as the mean ± SEM; In (B)–(E), data were represented as the mean ± SEM; an unpaired two-sided Student’s t test was performed for comparison. In (F) and (I), tumor growth curves were calculated by two-way ANOVA analysis ( n = 5). In (H) and (K), survival curves were calculated by Kaplan-Meier simple survival analysis ( n = 5). ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns, not significant. See also .

Article Snippet: The 96-well plates (Corning, #9018) were coated with recombinant human HER2 protein (MCE, #HY- P70254 ) diluted in coating solution (Bioss, #C04-01001) at 4°C overnight.

Techniques: In Vivo, Activity Assay, Binding Assay, Enzyme-linked Immunosorbent Assay, Comparison