Journal: Vaccine

Article Title: Tumor-associated antigens identified early in mouse mammary tumor development can be effective vaccine targets.

doi: 10.1016/j.vaccine.2019.05.024

Figure Lengend Snippet: (A) TgMMTV-neu mice received three vaccinations with empty vector and each of the seven early tumor antigens. MMC tumor cells were implanted on day 0. (B) TgMMTV-neu mice received three vaccinations with empty vector ● and the five late stage tumor antigens that have human homologs. MMC tumor cells were implanted on day 0 ***p<0.001. (C) Vaccination Schema. (D)MMC syngeneic tumor lysate but not lysate from FVB splenocytes induces an IFN-g T cell immune response in mice vaccinated with the early and late stage tumor antigens. IFN-g secreting cells quantified as precursor frequency (y-axis) for FVB mice (n=5) vaccinated with the plasmids for the early tumor antigens (light gray) or FVB mice (n=5) vaccinated with the plasmids for the late stage antigens (dark gray) as compared to FVB mice (n=5) vaccinated with empty control vector plasmid (white). The positive control is concanavalin A (CONA). * p<0.05 (E) Vaccination with the early and late stage tumor antigens induces a CD8+ T cell immune response. Granzyme B ELISA was performed on the pooled supernatants from the ELISPOT assay. ** p=0.01 * p<0.05

Article Snippet: The syngeneic mouse mammary tumor cell line MMC and human cell lines MCF10F (non-malignant breast), HCC1500 (ER positive HER2 negative), HCC70 (ER negative HER2 negative), and SKBR3 (ER negative HER2 positive) were seeded in two 96 well plates (ATCC, Manassas VA).

Techniques: Plasmid Preparation, Control, Positive Control, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot

Journal: Vaccine

Article Title: Tumor-associated antigens identified early in mouse mammary tumor development can be effective vaccine targets.

doi: 10.1016/j.vaccine.2019.05.024

Figure Lengend Snippet: Using four pooled siRNA for each target in the syngeneic MMC mouse tumor cell line, grey bars are the early stage antigens measuring (A) survival and (B) caspase 3/7 activity as compared to cells transfected with liposomes but no siRNA (control). For the late stage tumor antigens, the grey bars show the late stage tumor antigens measuring (C) survival and (D) caspase 3/7 activity as compared to cells transfected with liposomes but no siRNA (control). White bars include liposome transfected cells (control), non-targeting control siRNA transfected cells (negative control), and untransfected cells (untx). Apoptosis inducing siRNA transfected cells is the black bar (positive control) ** p<0.01 **** p<0.0001 as compared to control.

Article Snippet: The syngeneic mouse mammary tumor cell line MMC and human cell lines MCF10F (non-malignant breast), HCC1500 (ER positive HER2 negative), HCC70 (ER negative HER2 negative), and SKBR3 (ER negative HER2 positive) were seeded in two 96 well plates (ATCC, Manassas VA).

Techniques: Activity Assay, Transfection, Liposomes, Control, Negative Control, Positive Control

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: Three days after orthotopically injecting E0771 or AT-3 breast cancer cells plus mouse mammary fibroblasts into syngeneic C57BL/6J mice, we randomly assigned animals to treatment with anti–PD-1 antibody or PBS vehicle every 3 days for 4 doses total. Graphs show mean values ± SEM (symbols) and calculated logistic regression (smooth line) for E0771 ( A ) or AT-3 ( D ) tumors ( n = 6 control; n = 8 anti–PD-1) treated with anti–PD-1 antibody or PBS. ( B and E ) Growth of E0771 and AT3 tumor growth, respectively, for individual mice over time. Three tumors from the anti–PD-1 group failed to grow tumors and are overlapped on the x axis of panel B . We analyzed differences in tumor growth data by logistic regression. Survival curves demonstrate that anti–PD-1 treatment significantly prolonged survival for mice with E0771 tumors ( C ) but not with AT-3 ( F ), as analyzed by the Mantel-Cox test.

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Control

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: We injected C57BL/6J mice intraperitoneally with cholesterol labeled with BODIPY and euthanized animals 24 hours later to collect and dissociate tumors for flow cytometry ( n = 5 each for EO771 and AT-3). Plots for ( A ) CD8 + and ( B ) CD4 + T cells show accumulation of BODIPY-cholesterol in cells from individual tumors from EO771 and AT-3 tumors relative to vehicle only or isotype antibody control. ( C ) CD8 + and ( D ) CD4 + T cells in EO771 tumors showed significantly higher fold change accumulation of fluorescent cholesterol relative to FMO control. ( E ) CD8 + , but not ( F ) CD4 + , T cells in EO771 tumors also expressed higher levels of PD-1. ** P < 0.01,*** P < 0.001, **** P < 0.0001 for differences between means using nonparametric Mann-Whitney tests ( C and D ; n = 5 mice per group), while differences between T cell population percentages were assessed using 2-tailed Student’s t test ( E and F ).

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Injection, Labeling, Flow Cytometry, Control, MANN-WHITNEY

Journal: JCI Insight

Article Title: Evaluating immunotherapeutic outcomes in triple-negative breast cancer with a cholesterol radiotracer in mice

doi: 10.1172/jci.insight.175320

Figure Lengend Snippet: When tumors reached approximately 70 mm 2 , we treated mice with anti–PD-1 antibody or control for 4 days. Mice were then injected with 100 μCi of eFNP-59 followed by T cell isolation protocols. ( A ) Graph shows uptake of eFNP-59 per microgram of spleen tissue measured by scintigraphy. Symbols show individual mice with annotations for mean values and standard deviations. ( B ) We isolated CD4 + and CD8 + tumor-infiltrating lymphocytes (TILs) by positive selection with immunomagnetic beads and determined accumulation of eFNP-59 normalized to total cell protein with a BCA assay. Representative data from 2 experimental replicates, with statistical comparisons by 1-way ANOVA with Dunn’s multiple-comparison test. * P < 0.05; ** P < 0.01.

Article Snippet: We implanted 5 × 10 5 EO771 or AT-3 mouse mammary tumor cell lines and 1 × 10 5 mouse mammary fibroblasts orthotopically into fourth inguinal mammary fat pads of 6- to 8-week-old female C57BL/6J female mice (The Jackson Laboratory) ( ).

Techniques: Control, Injection, Cell Isolation, Isolation, Selection, BIA-KA, Comparison