Journal: Neoplasia (New York, N.Y.)

Article Title: Tumor neoantigens as key drivers of significant anti - tumor immunity in triple - negative breast cancer mouse models

doi: 10.1016/j.neo.2025.101205

Figure Lengend Snippet: PETC and TdL play key roles in regulating the antitumor response of T cells. (A) Schematic illustration of the CD3+ T-cell-depleted or IgG isotype control mouse model used to identify the effects of treatment with 4T1 TdL. (B) Five mice in each group, treated with either anti-CD3 or an IgG isotype control, received a total of three treatments with 4T1 TdL or control formulations, and the tumor volumes were subsequently measured. Significance was evaluated via two-way ANOVA, with ****p < 0.0001. (C) FACS analysis was performed to evaluate the presence of CD80+CD86+ (left) or CD80+MHCII+ (right) BMDCs after 4T1 TdL pulsing. The Mann–Whitney test was conducted, with *p < 0.05; ns indicates not significant, with error bars representing the standard error of the mean (SEM). We evaluated the functionality and cytotoxicity of T cells isolated from mice pre-exposed to 4T1 cells or treated with 4T1 TdL (n=3). To confirm the antigen-specific immune responses of all the T cells isolated from the immunized mice, we cocultured the T cells with DCs pulsed with 4T1 TdL in vitro. (D) Cytotoxicity was evaluated via the LDH assay, considering the numerical changes in the ratio of effector (T) cells isolated from the spleen to target (4T1) cells (n=3). Two-way ANOVA with Tukey's HSD was performed, and the results were statistically significant at ****p < 0.0001. TNF-α and IFN-γ secretion from CD4+ and CD8+ T cells isolated from tumors (n=3) was measured via ELISpot assays. ELISpot assays were used to measure the levels of TNF-α and IFN-γ secretion (E), while intracellular FACS analysis was used to assess the representative expression (Granzyme B, perforin, and CRTAM) in cytotoxic T lymphocytes (CTLs) from CD4+ and CD8+ T cells isolated from tumors from mice pre-exposed to 4T1 cells (F) and treated with 4T1 TdL (G) (n=3). The Mann–Whitney test was performed; statistical significance is denoted by *p < 0.05, and "ns" indicates not significant. Error bars represent the standard error of the mean (SEM).

Article Snippet: Following the protocol for the 4T1 TdL-treated syngeneic mouse model in a therapeutic setting, 100 μg of anti-mouse PD-L1 (B7-H1) (#BP0101; Bio X cell) or a rat IgG2b isotype control (#BP0090; Bio X cell) was administered five times on the day following the administration of 4T1 TdL.

Techniques: Control, MANN-WHITNEY, Isolation, In Vitro, Lactate Dehydrogenase Assay, Enzyme-linked Immunospot, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: Tumor neoantigens as key drivers of significant anti - tumor immunity in triple - negative breast cancer mouse models

doi: 10.1016/j.neo.2025.101205

Figure Lengend Snippet: Treating 4T1 syngeneic mice with both PDL1 antibody and therapeutic TdL greatly improves effectiveness. (A) Scheme of combination immunotherapy involving 4T1 TdL and an anti-PDL1 antibody, along with its control formulation, after the tumor size reached 100 mm3. (B) Tumor growth in mice with untreated CTL, 4T1 therapeutic TdL with IgG control, or anti-PDL1 antibody (n=5). Two-way ANOVA with Tukey's HSD was conducted, with statistical significance denoted by *p < 0.05, **p < 0.01 and ***p < 0.001. (C) UMAP plot illustrating the characterized cell clusters originating from tumors treated with anti-PDL1 inhibitor and 4T1 therapeutic TdL + anti-PDL1 inhibitor, encompassing natural killer (NK) cells, neutrophils, monocytes, macrophages, innate lymphoid cells (ILCs), dendritic cells (DCs), CD8+ T cells, B cells, and CD4+ T cells. Each sample was subjected to analysis for anti-PDL1 inhibitor (n=1) and 4T1 therapeutic TdL+ anti-PDL1 inhibitor (n=1), resulting in a total of 1,515 and 1584 immune cells normalized to 100%, respectively. (D) UMAP of the expression of genes such as SPARC, RGS1, XIST, CCL7, LRG1, S100A8, and S100A9 within tumor cell clusters. A violin plot of gene expression levels is displayed, with the median within the range box. Sparc is associated with the nonimmune component, whereas Rgs1, Xist, and CCl7 are associated with macrophages and monocytes. Lrg1, S100a8, and S1009 are associated with neutrophils.

Article Snippet: Following the protocol for the 4T1 TdL-treated syngeneic mouse model in a therapeutic setting, 100 μg of anti-mouse PD-L1 (B7-H1) (#BP0101; Bio X cell) or a rat IgG2b isotype control (#BP0090; Bio X cell) was administered five times on the day following the administration of 4T1 TdL.

Techniques: Control, Formulation, Expressing, Gene Expression