DNA vaccine strategies: candidates for immune modulation and immunization regimens Nicole A. Doria-Rose and Nancy L. Haigwood * Departments of Microbiology and Pathobiology, University of Washington and Viral Vaccines Program, Seattle Biomedical Research Institute, 4 Nickerson Street, Seattle, WA 98109, USA Accepted 25 April 2003 Abstract DNA vaccine strategies can differ greatly, with significant effects on the outcome of immunization. In this article, we discuss plasmid design strategies and vaccine regimens. Effectiveness against a pathogen can be affected by the choice of antigen and in- clusion of multiple antigens. Gene expression and the resulting immune response can be improved by gene modification and choice of promoters. In designing vaccine regimens, one must consider further dose, timing of doses, adjuvants, and routes of vaccination. Many vaccines are enhanced by combining DNA with other vaccines in ‘‘prime-boost’’ regimens, in which the second vaccine is often a recombinant viral vector or purified protein subunit. Prime-boost vaccines including DNA can elicit immune responses that differ in magnitude, quality, and balance of cellular and humoral responses from those elicited by single components and thus provide further enhancement for DNA immunizations. Ó 2003 Elsevier Science (USA). All rights reserved. Keywords: Antigen; Adjuvant; Plasmid; Promoter; Prime–boost 1. Introduction When plasmid vectors expressing human influenza virus proteins showed protection of mice from disease after live influenza challenge [1,2], it was a milestone event in the field of vaccinology. DNA vaccines offer many advantages over conventional immunization ap- proaches: they are simple to make and deliver, and they elicit both humoral and cellular immunity. Intracellular expression of native, oligomeric, surface-embedded an- tigens in the context of host cell surface proteins reca- pitulates the route that viral and parasitic pathogens use. Finally, multiple antigens can be delivered by combining several expression plasmids in the same im- munization. Thus, DNA vaccines hold the promise of delivering single or multiple native pathogen antigens combined with safety, effectiveness, and ease of pro- duction. Many features must be considered in designing an effective DNA immunization regimen. The choice of antigen(s), vector, delivery route, dose, timing, adju- vants, and boosting agents will all influence the outcome of vaccination because they affect the magnitude and quality of immunity elicited. In this article, we address issues of vaccine strategy pertinent to DNA vaccine design. In Sections 2–8, we focus on issues of plasmid design because gene expression is the sine qua non of DNA vaccines. In Sections 9–13, we consider the effects of regimen alterations, including ‘‘prime–boost’’ proto- cols that combine DNA with other vaccines. 2. Plasmid design The first choice in designing a DNA vaccine is the target antigen(s). One must select the pathogen genes and the form of the gene, whether secreted, intracellular, membrane bound, wild type, or mutated. Once the gene(s) is chosen, various modifications are commonly used to improve or alter the immunogenicity of the vaccines. Methods 31 (2003) 207–216 www.elsevier.com/locate/ymeth * Corresponding author. Fax: 1-206-284-0313. E-mail address: Nancy.Haigwood@sbri.org (N.L. Haigwood). 1046-2023/$ - see front matter Ó 2003 Elsevier Science (USA). All rights reserved. doi:10.1016/S1046-2023(03)00135-X