Oral attenuated Salmonella typhimurium vaccine against MG7-Ag mimotope of gastric cancer

Abstract

Aim: To develop an oral attenuated Salmonella typhimurium vaccine against gastric cancer and to evaluate its efficacy in mice.

Methods: A complementary sequence of Nco I site and a sequence coding for MG7-Ag mimotope were designed at the 5' terminus of forward primer. Using p1.2 II-HBCAg plasmid as template, PCR was performed to get a fusion gene of the mimotope and a HBcAg gene. The fusion gene was then subcloned into the plasmid pYA3341 complementary to Salmonella typhimurium X4550, and the recombinant plasmid was then transformed into attenuated Salmonella typhimurium X4550. Balb/c mice were orally immunized with the recombinant Salmonella typhimurium X4550. The mice were immunized every 2 wk to reinforce the immunity. At the 6th wk, serum titer of antibody was detected by ELISA, and at the 8th wk, cellular immunity was detected by (51)Cr release test. Ehrlich ascites carcinoma cells expressing MG7-Ag were used in tumor challenge assay as a model to evaluate the protective effect of the vaccine.

Results: Serum titer of antibody against MG7-Ag was significantly higher in mice immunized with the vaccine than in control groups (0.9538+/-0.043 vs 0.6531+/-0.018, P<0.01; 0.9538+/-0.043 vs 0.6915+/-0.012, P<0.01), while in vitro (51)Cr release assay of the splenocytes showed no statistical difference in the three groups. Two weeks after tumor challenge, 1 in 5 immunized mice was tumor free, while all the mice in the control group presented tumor.

Conclusion: Oral attenuated Salmonella typhimurium vaccine against the MG7-Ag mimotope of gastric cancer is immunogenic. It can induce significant humoral immunity against tumors in mice, and has some protective effects.

Figure 1

Incorporation of the MG7 epitope cDNA into HBcAg gene by PCR. An about 600 bp fragment was obtained (1,2). Lane M: DNA marker (100-3000 bp); lanes 1-2: the PCR product.

Figure 2

Identification of fusion gene vaccine. Lane M: DNA marker (100-3000 bp); lanes 1,2: the digestion product.

Figure 3

Western blot of the expression product of MG7/HBcAg fusion gene. Lane M: Protein marker (M_r); lane 1: X4550 transformed with empty pYA3341; lanes 2,3: X4550 transformed with pYA3341-MG7/HBcAg (after induction of IPTG for 2 and 4 h).