Preparation of mB7.1-GPI and SEA-TM dual-anchored tumor cell vaccine and its antitumor effect

Abstract

To prepare the SEA-TM and mB7.1-GPI dual-anchored EL-4 cell vaccine and to investigate its antitumor effects. mB7.1-GPI-anchored EL-4 cell vaccine, SEA-TM-anchored EL-4 cell vaccine, SEA-TM and mB7.1-GPI dual-anchored EL-4 cell vaccine were prepared. In vitro the biological activities of these vaccines were measured using a lymphocyte proliferation assay and cytokine release assay on splenocytes derived from C57BL/6 mice. The splenocytes were co-cultured with EL-4 or EL-4/mB7.1-GPI or EL-4/SEA-TM or EL-4/SEA-TM + mB7.1-GPI (treated with Mitomycin C). Lymphocyte proliferation was determined with MTT assay, the concentrations of cytokines (IL-2 and IFN-gamma) were measured using a ELISA technique. Forty C57BL/6 mice were inoculated with EL-4 cells, after 3 days the mice were randomly divided into 5 groups with 8 in each and were treated with PBS, EL-4 cell vaccine, EL-4/mB7.1-GPI cell vaccine, EL-4/SEA-TM cell vaccine and EL-4/SEA-TM + mB7.1-GPI cell vaccine respectively, vaccines were injected three time with two-day interval. Animals were observed daily, tumor sizes were measured every third day. Twenty-five days after tumor challenge, 3 mice in each group were sacrificed and splenic lymphocytes were isolated to examine the activity of natural killer cells (NK) and cytolytic T lymphocytes (CTL). The survival of the remaining 5 mice in each group was observed till the 90th day. mB7.1-GPI or/and TM-SEA fusion protein was stably anchored onto the surface of EL-4 tumor cells. EL-4/mB7.1-GPI or EL-4/SEA-TM had a stronger ability to stimulate lymphocyte proliferation and IL-2 and IFN-gamma production than EL-4 (P < 0.05); while EL-4/SEA-TM + mB7.1-GPI showed a further increased ability than EL-4/mB7.1-GPI and EL-4/SEA-TM in stimulating lymphocyte proliferation and cytokine production in vitro (P < 0.05). Volume of tumor was smaller and survival time of mice was longer in EL-4/mB7.1-GPI vaccine group, EL-4/SEA-TM vaccine group and EL-4/SEA-TM + mB7.1-GPI vaccine group, comparing with PBS group and EL-4 cell vaccine group (P < 0.05). Tumor volume was much smaller and survival time of mice was much longer in EL-4/mB7.1-GPI + mB7.1-GPI vaccine group, comparing with EL-4/SEA-TM vaccine group and EL-4/mB7.1-GPI vaccine group (P < 0.05). Lymphocytes derived from the mice treated with EL-4/SEA-TM + mB7.1-GPI showed much higher NK activity and CTL activity than those derived from EL-4/mB7.1-GPI vaccine group and EL-4/SEA-TM vaccine group (P < 0.05), meanwhile the NK activity and CTL activity of EL-4/mB7.1-GPI vaccine group and EL-4/SEA-TM vaccine group was higher than EL-4 vaccine group (P < 0.05). mB7.1-GPI or/and SEA-TM fusion protein was stably anchored onto the surface of EL-4 tumor cells. The tumor cell vaccines prepared from these cells exhibited antitumor effect. The mB7.1-GPI and SEA-TM dual-anchored tumor cell vaccine had much stronger antitumor effect than the single-anchored tumor cell vaccine.