Induction of specific anti-CEA responses by DNA vaccination with fusion sequence of CEA repeat epitopes and heterogenous HSP70 in mice

Objective To observe the immune responses stimulated by inoculation of DNA vaccine encoding a fusion protein of CEA tandem repeat epitopes and heterogenous HSP70 fragment as a basis for pioneering a novol DNA vaccine specific to carcinomas.Methods The recombinant sequence for two CEA epitopes was tandemly engaged and inserted into-upstream of in-frame-varied HSP70 of Mycobacterium tuberculosis to construct a gene vaccine.Balb/c mice were muscularly injected with recombinant DNA vaccine;negative control(mice were injected with normal saline),positive control(mice were injected with DNA vaccine harboring tandem CEA epitopes plus aluminium hydroxide adjuvant) and experimental group(mice were injected with PCITri CEA625-667-met HSP70) were set up.After 3 inoculations,the mice were sacrificed and their splenocytes and sera were collected.T cell subsets were analyzed by FCM.IFNγ in their cultural supernatants and seral IgG antibody against CEA were detected with ELISA.Results The amount of CD3+and CD4+T cells in the mouse spleen in negative control was 55.1%±6.1% and 30.2%±4.1% with no production of IFNγ by in vitro culture of spleenocytes,which could be considered as basal physiological parameters.The fused DNA vaccine induced vigorous T cell responses in mice with nearly 50-fold increases of spleen CD3+and CD4+T cells and enhanced IFNγ production up to 3 folds for non-specificity and 6 folds to specific challenge in cultured splenocytes under stimulation of CEA peptide(P0.01).The serum titer of IgG antibody against CEA epitope in negative control group was 0 and less than 1∶500 in positive control,whereas that in the mice immunized with the DNA vaccine encoding the fusion protein of the CEA epitopes and HSP70 was up to 1∶4000.Conclusion DNA vaccine for the fusion protein consisted of CEA repeat epitopes and heterogenous HSP70 fragment can induce a strong immune response in mice featured by stimulation of T helper cells.