Dendritic cell vaccine with Ag85A enhances anti-colorectal carcinoma immunity

Abstract

Dendritic cells (DCs) are able to trigger T-cell activation and thus have been considered important for vaccine production against cancers. Vaccines containing DCs have been reported to be effective for developing immunity against cancer cells. The interactions between DCs and auxiliary agents are critical in the development of second-generation vaccines. In the present study, it was evaluated whether Ag85A-mixed DCs could enhance anti-tumor immunity in laboratory mice with colorectal carcinoma. Functional and phenotypic analyses of the effects of Ag85A-mixed DCs were conducted via flow cytometry and measurement of T-cell proliferation. In addition, interferon (IFN)-γ production was assessed. The therapeutic efficacy of DC vaccination for colorectal carcinoma treatment in mice was investigated. It was identified that Ag85A-mixed DCs exhibited strong upregulation of CD80, CD86 and major histocompatibility complex class II. Cytotoxic T-lymphocytes with CT26-primed Ag85A-DCs were indicated to induce stronger responses against CT26 tumor cells and trigger IFN-γ production. Furthermore, the Ag85A-mixed DC vaccine exerted a considerable inhibitory effect on tumor progression in mice as compared with the control group. Therefore, DCs in combination with the Ag85A gene may reinforce anti-colorectal carcinoma immunity. The current study provides a novel potential strategy for cancer treatment by enhancing immunity via Ag85A-mixed DC vaccination.

Keywords: antigen 85A; colorectal carcinoma; dendritic cells; major histocompatibility complex class II.

Figure 1.

Determination of Ag85A mRNA expression levels by RT-qPCR. Ag85A expression in DC 2.4 cells transfected with pc-DNA3.1, DC2.4 plus pcDNA3.1 (mock-DC) or non-transfected DC 2.4 (Con) for 24 h was determined by RT-qPCR. Data are presented as the mean ± standard error of the mean from three independent tests. **P<0.01 vs. mock-DC group. DC, dendritic cell; Ag85A, antigen 85A; Con, control; RT-qPCR, reverse transcription-quantitative polymerase chain reaction.

Figure 2.

Ag85A transfection enhances the expression level of primary surface markers in DCs. Expression levels of (A) CD80, (B) CD86 and (C) MHC class II in Ag85ADCs, mock-DCs and controls were assessed via flow cytometry. Data are presented as the mean ± standard error of the mean from three independent tests. **P<0.01 vs. mock-DC group. DC, dendritic cell; Ag85A, antigen 85A; Con, control; MHC, major histocompatibility complex.

Figure 3.

Ag85A transfection promotes T-cell proliferation. T-cell proliferation was measured by Cell Counting Kit 8 analysis on days 1, 2 and 3. Data are presented as the mean ± standard error of the mean from three independent tests. *P<0.05 vs. mock-DC group. DC, dendritic cell; Ag85A, antigen 85A; Con, control.

Figure 4.

Ag85A transfection increases tumor-specific cytotoxicity of T-cells in vitro. (A) Cytotoxic T-lymphocyte activity was measured by targeting CT26 tumor cells in CT26 lysate-pulsed Ag85A-DCs (Ag85A DC), CT26 lysate-pulsed mock-DCs (mock-DCs) and CT26 lysate (Con) at an effector-to-target ratio of 50:1. (B) IFN-γ production was assessed via ELISA. (C and D) Frequencies of CD8⁺ IFN-γ⁺ T cells were analyzed via flow cytometry. Data are presented as the mean ± standard error of the mean from three independent tests; *P<0.05 vs. control group; ^#P<0.05 vs. mock-DC group. DC, dendritic cell; Ag85A, antigen 85A; Con, control; IFN, interferon.

Figure 5.

Administration of Ag85A-DC vaccine into tumors suppresses tumor development in mice. (A) Images, (B) weights and (C) volumes of resected orthotopic xenograft lab mice at 21 days after administration of tumor cells into the Ag85A-DC vaccine group, mock-DC vaccine group, and CT26 lysate group. Data are presented as the mean ± standard error of the mean, n=6. *P<0.05 vs. control; ^#P<0.05 vs. mock-DC vaccine group. DC, dendritic cell; Ag85A, antigen 85A; Con, control.