DNA vaccination may work through direct transfection of antigen presenting cells (APC), or by secretion of the encoded protein by muscle or skin cells for uptake by APC. If cytokines are attached to the antigen, they may influence APC or responding T cells to drive the response toward a Th1 or Th2 direction, and/or potentiate it in an antigen-specific manner. To test this concept, expression vectors were constructed containing the ovalbumin (OVA) gene either alone, or linked to cytokine genes including GM-CSF, IFN-γ, IL-2, IL-4, IL-12, or a sequence encoding nine amino acids of IL-1β. These constructs expressed OVA-cytokine fusion proteins in vitro which retained cytokine bioactivity. mice were injected intramuscularly with the DNA constructs. Little if any OVA-specific antibody was produced in response to any of the DNA constructs, except for OVA-IL-4. However, lymphocytes from mice vaccinated with OVA-IL-12 and OVA-IL-1β constructs produced more IFN-γ and less IL-4 during in vitro restimulation assays than did other groups. All constructs elicited OVA-specific cytotoxic responses which were maintained or even increased over 16 weeks. The OVA-IL-12 and OVA-IL-1β peptide constructs elicited the strongest cytotoxic responses at 2 weeks postinjection. Cytotoxic responses were seen in all animals, even those lacking OVA-specific Ab, and were not related to Ab level. These studies indicate that the humoral, cytokine, and cytotoxic responses to DNA vaccination can be effectively altered by certain cytokine fusion constructs.
