Naked DNA and vaccine design.

O ver the past decade, the revolution in molecular and structural biology has opened up the immune system to detailed analysis. The genes encoding critical molecules of the immune system have been identified and the structure of these genes has been determined. Immunologists now not only have detailed information on the structure of the immunoglobutin receptor on B cells, but also information on the structure and function of the T cell antigen receptor complex and the molecules encoded by genes of the major histocompatibility complex (MHC). Indeed, the three dimensional structure of both the MHC class I and, more recently, the MHC class II molecules has been solved by X-ray crystallographic analysis 1,2. These structural studies have directly established that the antigen receptor on T cells recognizes short peptide fragments of foreign antigenic proteins, such as bacterial and viral polypeptides, bound to MHC class I and class II molecules on the surface of antigenpresenting cells. This new insight into the nature of antigen recognition by T cells has led to the development of many novel strategies to manipulate the immune system and to treat autoimmune diseases. One area where the revolution in immunology has not, as yet, had its full impact is in vaccine development. At the moment, Jenner's approach of immunizing with a live, attenuated virus remains the most effective means of producing sustained immunity to most viruses. Part of the reason why live viruses are effective vaccines is that they efficiently activate the CD8 + precursor cells that give rise to the major antiviral effector cells: the MHC class I restricted cytolytic T cells. The reason for this lies in the cell biology of the synthesis, assembly and transport of the MHC class I molecule. MHC class I molecules capture viral peptides during their synthesis and assembly in the endoplasmic reticulum (ER). The MHC molecules transport the bound viral peptides from the ER to the cell surface, where the complex is displayed to the antigen receptor on CD8 ÷ cytolytic T cell precursors and activated effector cytolytic T cells. To charge nascent MHC class I molecules, viral polypeptides must gain access to the cytoplasm of the cell. Here, the viral polypeptides are fragmented and then transported as peptide fragments into the ER. The most efficient way for viral polypeptides to gain access to the cytoplasm of an antigenpresenting cell for proteolytic fragmentation is for them to be expressed de n o v o in infected cells. Recently, a novel vaccination strategy has been developed using naked DNA encoding a viral polypeptide as an immunogen. This strategy is based on the fortuitous observation by Wolff et al. 3 that direct intramuscular inoculation of mice with plasmid DNA containing the gene for a marker protein driven by a strong eukaryotic promoter resulted in persistent expression of the gene in striated muscle at the site of injection. This finding extended earlier work on the uptake and expression of viral and eukaryotic DNA by direct DNA inoculation into experimental animals". The result of Wolff et al. has prompted researchers at Merck Research Laboratories and Vical to use the same strategy of DNA uptake and expression as a method of vaccination against influenza virus. As reported recently, Ulmer et al. v have injected mice with DNA encoding the type A influenza A/PR/8/34 nucleocapsid protein (NP) linked to the human cyto-