Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: Generation and characterization of CD19CAR T cells with different vectors. (A) Construction maps of piggyBac (PB) Mock, piggyBac -CD19CAR, and lentivirus (lv)-CD19CAR. (B) Rates of CAR T-cell positivity for vectors over time. CAR T cells were electroporated with piggyBac Mock or piggyBac -CD19CAR or infected with lv-CD19CAR. (C) Cytotoxicity of Mock T, CD19pbCAR T, and CD19lvCAR T cells against CD19 + Raji cells at days 10 and 15. (D) Cytokine levels in the medium after incubation of Mock T, CD19pbCAR T, and CD19lvCAR T cells with 1 × 10 5 Raji cells for 4 h. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Infection, Incubation

Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: Comparison effects of CD19pbCAR T cells and CD19lvCAR T cells against Raji cell-induced tumors in a xenograft model. (A) Luciferase imaging, showing systemic trafficking and tumor accumulation of Raji cells. 1 x 10 6 Raji-luc-C8 cells were injected into the tail veins of NPI mice, followed by the injection of the same volumes of PBS (vehicle), 1×10 7 CD19pbCAR T cells and 1×10 7 CD19lvCAR T cells. Fluorescence in each mouse was evaluated by the Caliper IVIS-lumina-XR on days 0, 4, 7, 11, and 14. (B) Measurements of tumor volumes at the indicated times. (C) Body weights of mice measured at the indicated times. (D) Numbers of surviving and dead mice determined until day 40. *p<0.05; **p<0.01; ***p<0.001.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Luciferase, Imaging, Injection, Fluorescence

Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: High concentration of CD19pbCAR T cells eliminated the growth of Raji cells in a murine model of hematological malignancy. (A) Luciferase imaging, showing systemic trafficking and tumor accumulation of Raji cells; 1 × 10 6 Raji-luc-C8 cells were injected into the tail veins of NPI mice 10 days, followed by the injection of medium (vehicle), Mock T cells, or 2.5 × 10 6 (low dose), 5.0 × 10 6 (middle dose), or 10 × 10 6 (high dose) CD19pbCAR T cells. Fluorescence in each mouse was evaluated by the Caliper IVIS-lumina-XR on days 0, 3, 7, and 14. (B) Numbers of surviving mice in the five groups. (C) Measurement of total fluorescence density of Raji tumor cells in each mouse at the indicated times. (D) Change in tumor volume in mice administered high dose CD19pbCAR T cells. **p<0.01; ***p<0.001.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Concentration Assay, Luciferase, Imaging, Injection, Fluorescence

Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: Distribution and specific accumulation of CD19pbCAR T cells. (A) Copies of pbCAR in tissues; 1 × 10 6 Raji tumor cells were injected into the tail veins of 21 NPI mice, followed 9 days later by intravenous injection of 1 ×10 7 CD19pbCAR T cells. Three mice each were euthanized on days 1, 3, 7, 10, 14, 22, and 29 for collection of tissues and blood. (B) Copies of pbCAR in the blood of the mice, as determined by qPCR, at the indicated times. (C) CD19pbCAR T cells specifically target transplanted tumor; 1 × 10 6 Raji tumor cells were injected subcutaneously into the right back sides of mice. When tumor volume reached 200 mm 3 , the mice were injected with 2x10 7 Mock T-luc or CD19pbCAR T-luc cells. Fluorescent images were taken at the indicated times. (D) Measurements of total body fluorescence and tumor volume. **p<0.01.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Injection, Fluorescence

Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: Phenotypes and specificities of CD19pbCAR T cells and CD19lvCAR T cells. (A) Proportions of CD4 and CD8 T cells measured on day 13 by flow cytometry. (B) Proportions of Tcm cells measured on days 10 and 13 by flow cytometry. (C) Expression of exhaustion markers (PD-1, LAG3, and TIM3) by flow cytometry on day 13. (D) Specificity of CD19pbCAR T cells, as shown by incubation with CD19 KO Raji cells. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Flow Cytometry, Expressing, Incubation

Journal: Frontiers in Immunology

Article Title: Evaluation of Nonviral piggyBac and lentiviral Vector in Functions of CD19chimeric Antigen Receptor T Cells and Their Antitumor Activity for CD19 + Tumor Cells

doi: 10.3389/fimmu.2021.802705

Figure Lengend Snippet: Proliferation and activities of CD19pbCAR T cells. (A) Proliferation of CD19pbCAR T cells 5-13 days after electroporation. (B) Cytotoxicity of Mock-T and CD19pbCAR T cells against Raji cells at different E:T ratios. (C) Cytokine levels of Mock-T and CD19pbCAR T cells 13 days after incubation with 1×105 Raji cells for 4 hours at different E:T ratios. *p<0.05; **p<0.01; ***p<0.001.

Article Snippet: Luciferase-expressing Raji cells (Raji-Luc-C8) were purchased from Beijing Idmo Co., Ltd. (Beijing, China).

Techniques: Electroporation, Incubation