hepa1 6 cells  (ATCC)

Journal: Cell Reports Medicine

Article Title: CAR-T triggers TAM reeducation and adaptive anti-tumor response via TREM2 deficiency or CD40 agonist

doi: 10.1016/j.xcrm.2025.102539

Figure Lengend Snippet: TREM2 + TAMs hinder the therapeutic effectiveness of GPC3-CAR-T cells (A) UMAP visualization of scRNA-seq data from tumor biopsies following IL15-GPC3-CAR-T therapy (data from Steffin et al., Nature 2025; GEO: GSE253352 ). (B) The heatmap displayed the signature differentially expressed genes across distinct myeloid cell subpopulations. (C) The bar chart showed the proportions of myeloid cell subsets in the PD and PR groups. (D) The boxplot demonstrated the statistical differences among various myeloid subpopulations. (E) In vitro cytotoxicity measured by lactate dehydrogenase (LDH) release at indicated effector cell vs. target cell ratios (E:T ratios) using Hepa1-6 cells overexpressing human GPC3 or vector control were co-cultured with Ctrl-CAR-T (targeted CD19) or GPC3-CAR-T cells. (F) Subcutaneous tumor growth and survival in C57BL/6J mice ( n = 6/group) inoculated with GPC3-Hepa1-6 cells and treated with Ctrl- or GPC3-CAR-T cells (1 × 10 6 ) via tail vein. (G) Orthotopic liver tumor model: tumor weight at day 28 and survival in mice receiving intrahepatic GPC3-Hepa1-6 implants followed by CAR-T treatment. (H) Flow cytometric analysis of immune and stromal cell populations in tumors from subcutaneous and orthotopic models following GPC3-CAR-T treatment. (I) Expression of TAM-associated genes (Trem2, Spp1, Cd274, Arg1, Tgfb1, and Cd206) in F4/80 + TAMs sorted from tumors of GPC3-CAR-T-treated mice, measured by RT-qPCR. (J) Schematic of the orthotopic HCC model for CAR-T therapy in Trem2 -WT/KO mice. (K) Tumor representative photographs of different groups were shown. Dot plots showed liver weight, liver-to-body weight ratio, per liver, n = 8. (L) UMAP plots of individual groups displayed distinct intratumoral cell populations. n = 2 for all treatment groups on scRNA sequence. (M) Proportion of the distinct intratumoral cell populations. Data were represented by mean ± SEM. ns, no significance, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: Hepa1-6 cell line , ATCC , Cat#CRL-1830.

Techniques: In Vitro, Plasmid Preparation, Control, Cell Culture, Expressing, Quantitative RT-PCR, Sequencing

Journal: Cell Reports Medicine

Article Title: CAR-T triggers TAM reeducation and adaptive anti-tumor response via TREM2 deficiency or CD40 agonist

doi: 10.1016/j.xcrm.2025.102539

Figure Lengend Snippet: The combination of CD40 agonism with GPC3-CAR-T therapy held promising application prospects in the treatment of HCC (A) Schematic procedure of HCC model constructed by Hepa1-6 cell orthotopic injection. GPC3-CART and CD40 agonist (FGK45) were injected. (B) Tumor representative photographs of different groups were shown. (C) Dot plots showed liver weight of mice in different groups. n = 8. (D) Survival curves of each group of mice. n = 8. (E and F) Multiple immunofluorescence images of TAMs, CD8 + , and CD4 + T cells in tumor tissue between groups. Scale bar, 50 μm. Quantification of cells numbers from three random fields per mouse ( n = 8 mice/group). Data were represented by mean ± SEM. ∗∗∗ p < 0.001. (G and H) Flow cytometry and quantification plots of IFN-γ + CD8 + T cells and IFN-γ + CD4 + T cells, as well as IFN-γ − CD8 + T cells and IFN-γ - CD4 + T cells from TILs in vivo . N = 8. Data were represented by mean ± SEM. ∗ p < 0.01, ∗∗∗ p < 0.001. (I) Schematic procedure of a therapeutic breast cancer liver metastasis model constructed by 4T1 cell expressing hTrop2 protein. Murine hTrop2-CAR-T and CD40 agonist (FGK45) were injected. (J) Tumor representative photographs of different groups were shown. (K) Dot plots showed liver weight of mice in different groups. n = 8. Data were represented by mean ± SEM. ns, no significance, ∗∗∗ p < 0.001. (L) Survival curves of each group of mice. n = 8. (M) Schematic representation of the transendothelial migration assay. TAMs were isolated from human HCC tissue. Sotigalimab (20 ng/mL) and h-IFN-γ (40 ng/mL) were added. Flow cytometry quantification of migrated CD8 + T cells (CellTrace Far Red labeled, n = 5). Data were represented by mean ± SEM. ns, no significance, ∗∗ p < 0.01, ∗∗∗ p < 0.001. (N) Schematic representation of the transendothelial migration assay. DTS was isolated from human GPC3 + HCC tissue. GPC3-CAR-T was added at indicated ratio, with or without sotigalimab. Flow cytometry quantification of migrated CD8 + T cells (CellTrace Far Red labeled, n = 5). Data were represented by mean ± SEM. ns, no significance, ∗∗∗ p < 0.001.

Article Snippet: Hepa1-6 cell line , ATCC , Cat#CRL-1830.

Techniques: Construct, Injection, Immunofluorescence, Flow Cytometry, In Vivo, Expressing, Migration, Isolation, Labeling