During the last 20 years, DNA-based immunization has been rapidly developed as a new approach to prime specific cellular and humoral immune responses to protein antigens . In mouse models, DNA vaccines have been successfully directed against many types of tumor with tumor protection reproducibly observed in an antigen-specific manner . However, conventional DNA vaccines with only the encoded antigen failed to mount an effective T-cell immunity in human trials, even when delivered by in vivo electroporation, which calls for a novel design for DNA vaccine [1, 2].
It is critical for DNA vaccination to be successful that encoded proteins are taken up, processed, and presented by dendritic cells (DC), the most potent antigen-presenting cells (APC) in vivo that initiate the adaptive immunity. Following intradermal or intramuscular injection of a plasmid DNA vaccine in mice, the encoded gene is expressed in transfected keratinocytes and myocytes at the site of injection  as well as a small number of DC [4–6]. Keratinocytes and myocytes are poorly effective in presenting antigen and priming naive immune cells due to lack of expression of MHC class II and costimulatory molecules, and do not have ready access to T cells in lymphoid tissues, as is the case for DC . It is thought that transduced DC initiate immune priming process, which can be further boosted by antigen released from other long-lived transfected cells [8, 9]. Therefore, targeting DNA vaccines to DC should improve the efficacy of DNA vaccines. In fact, a recent study demonstrated that DC-targeted DNA vaccines elicited much higher level of antibody and antigen-specific T cells, leading to effective protection against virus expressing encoded antigen .
Coupling of antigens to ligands or antibodies that specifically bind to DC receptors has been widely used as a means of DC targeting . Using this approach, a lowered requirement for antigen dose in stimulating immune responses in mice has been observed after targeting a variety of molecules, including MHC class II, DEC205, CD11c, Dectin-1/2, mannose receptor, and CD36 [12–17]. The studies have also shown that antibodies specific for the mannose receptor or DC-SIGN could effectively deliver antigen to human DCs, indicating that this strategy may also be applicable to human vaccination [18, 19].
Overexpression of the HER-2 receptor tyrosine kinase has been found in various human malignancies, including breast, ovarian and gastric carcinomas, non-small cell lung cancer, and salivary gland cancers, and has been associated with poor prognosis of patients [20, 21]. Endogenous HER2-specific CD4+ T cells and antibodies have been detected in patients with HER2-expressing cancers [22, 23], and in clinical trials, HER2-specific CD4+ and CD8+ T-cell responses could be induced by peptide vaccination [24, 25]. These studies provide strong supports for HER2 being an important tumor antigen for targeted immunotherapy. The clinically approved HER2-targeted immunotherapy involves infusion of humanized HER2-specific monoclonal antibody Herceptin; ref. . Although Herceptin has been shown to be effective in inhibiting tumor growth in a limited population of HER2-postive metastatic breast cancer patients, elicitation of an active and more comprehensive immune response that includes both antibody and T-cell responses may provide more effective protection .
Here, we prepared DC-targeting DNA vaccines by fusing tumor-associated antigen HER2/neu ectodomain (HER2/neu, residues 22 to 561 or 22 to 582) to single chain antibody fragment (scFv) from NLDC-145 (scFvNLDC-145), a monoclonal antibody binding the mouse DC-restricted surface molecule DEC-205, and evaluated the preventive and therapeutic effects of these DNA vaccines in HER2/neu-positive mouse breast tumor models. We further characterized the cellular mechanisms driving antitumor effect of DC-targeted DNA vaccines elucidating the basic processes necessary to achieve immune-mediated tumor rejection.