Dna Vaccines

A. Concepts of DNA Vaccines

Recent work from a number of laboratories has demonstrated that the injection of a DNA plasmid containing foreign genes for proteins of a pathogen or cancer antigens directly into a host results in the subsequent expression of the foreign gene in that host and the presentation of the specific encoded proteins to the immune system (Fig. 1). DNA vaccine constructs are produced as small circular vehicles or plasmids. These plasmids are constructed with a promoter site that starts the transcription process, an antigenic DNA sequence, and a messenger RNA stop site containing the poly A tract necessary for conversion of the messenger RNA sequence into the antigen protein by the ribosomal protein manufacturing machinery (Fig. 2). The concept of genetic immunization provides that both DNA and RNA that encode specific proteins can be used to generate specific immune responses. Because DNA and RNA are both nucleic acids, the term nucleic acid vaccine has also been used to describe this process.

B. History of DNA Inoculation

The ability of genetic material to deliver genes for therapeutic purposes and its use in gene therapy has been appreciated for some time. Early experiments describing DNA inoculation into living cells were DNA transfer experiments performed by a number of investigators in the 1950s and 1960s (3-5). These reports describe the ability of DNA preparations isolated from tumors or viral infections to induce tumors or virus infection following injection into animals. Importantly, many such inoculated animals developed antibody responses to the proteins encoded within the injected DNA sequences. Over the next 20 years, a number of scattered reports focusing on gene function or gene therapy techniques provided evidence that injection of viral DNA or plasmids containing foreign gene resulted in antibody production and was related to the DNA inoculations. In 1985, Dobensky reported that plasmids containing insulin DNA sequences could produce insulin following inoculation into a living animal for some period of time. Longer-term expression of foreign genes was described in 1990 by Wolff and colleagues following plasmid inoculation in vivo (6). These 2 separate observations demonstrated that DNA in the absence of vectors could deliver proteins that might have biological relevance. Although both studies focused on the use of this technology for gene replacement strategies, other studies were already underway in several laboratories using this same technology for vaccine applications.

In 1992, Johnston and colleagues at the Southwest Foundation reported that injection of DNA encoding human growth hormone into mice resulted in transient hormone production followed by the development of antibodies in the inoculated animals specific for the human growth hormone gene (7). This important work used a ''genetic gun'' or gene gun to shoot gold particles covered with DNA through the skin layers of mice. Although these investigators were actually studying the use of this technology for a gene therapy replacement strategy, they described this development of antibody responses due to this unusual immunization procedure as genetic immunization. Simultaneously with the publication by Johnston and colleagues, a vaccine meeting held at the Cold Spring Harbor Laboratory, Long Island, New York, in September 1992, described the use of DNA immunization to generate humoral and cellular immune responses against a human pathogen, as well as protection from both tumors and viral challenges in animal systems.

Investigators from Merck and Vical reported on the development of immune responses to intramuscular injected plasmid encoding pathogen proteins. They observed that both antibody responses as well as CTL responses were induced to influenza viral gene products by this immunization technique. Furthermore, vaccinated mice were able to resist lethal viral challenge. Harriet Robinson and her colleagues at the University of Massachusetts reported on the use of the gene gun to deliver influenza virus genes in DNA plasmids, indicating that both antibody and T lymphocyte responses were produced in vaccinated mice and in chickens. In challenge studies, these responses were protective. The use of the gene gun allowed investigators to deliver very low nanogram amounts of DNA at the site of injection and still observe immune responses. David Weiner and his colleagues at the University of Pennsylvania reported the direct injection of DNA encoding the genes for the human immunodeficiency virus (HIV). Again both antibodies and T lymphocyte responses specific for the viral gene products were observed in experimental animals. As HIV does not infect mice, an in vivo mouse model was used where tumor cells, which are normally lethal to the mice, were constructed to express HIV proteins. Animals who were vaccinated with the HIV DNA vaccine were demonstrated to be immune to these HIV antigen-expressing tumor cells. Although the audience was skeptical of the ability of nonliving genetic material to produce useful immune responses, the large amount of data presented by each of these groups representing several years of successful work in diverse systems could no longer be ignored by the scientific and vaccine community. DNA vaccines were officially born.

Following these initial reports, DNA vaccination and the generation of antibody and T lymphocyte responses, as well as protective responses in a variety of animal models, have been reported in the scientific literature for many human pathogens such as hepatitis B virus, rabies virus, herpes simplex virus, hepatitis C virus, human T cell leukemia virus, human papilloma virus, and tuberculosis (8-10).

C. Potential Advantages of DNA Vaccines

As summarized in Table 1, nucleic acid immunization may afford several potential advantages over traditional vaccina-

Figure 1 Induction of antigen-specific humoral and cellular immune responses following DNA immunization.
Schematic The Constructed Promoter
Figure 2 A diagram of a DNA vaccine construct consisting of a mammalian expression vector. The plasmids are constructed with a promoter, an antigenic DNA sequence, and a messenger RNA stop site containing the poly A tract.

Table 1 Summary of Immune Modulation by Molecular Adjuvants

Immune modulation Molecular -

Table 1 Summary of Immune Modulation by Molecular Adjuvants

Immune modulation Molecular -

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