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Journal of Virology, November 1999, p. 9145-9152, Vol. 73, No. 11
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Evaluation of Novel Human Immunodeficiency Virus Type 1 Gag DNA Vaccines for Protein Expression in Mammalian Cells and Induction of Immune Responses

Jian-Tai Qiu,1 Ruijiang Song,2 Markus Dettenhofer,1 Chunjuan Tian,1 Thomas August,2 Barbara K. Felber,3 George N. Pavlakis,3,* and Xiao-Fang Yu1,*

Department of Molecular Microbiology and Immunology, The Johns Hopkins School of Hygiene and Public Health,1 and Department of Pharmacology and Molecular Sciences, The Johns Hopkins School of Medicine,2 Baltimore, Maryland 21205, and National Cancer Institute-Frederick Cancer Research and Development Center, ABL-Basic Research Program, Frederick, Maryland 21702-12013

Received 20 May 1999/Accepted 2 August 1999

Human immunodeficiency virus (HIV)-specific cytotoxic T lymphocytes (CTL) are an important parameter of host defenses that limit viral replication after infection. Induction of effective CTL against conserved viral proteins such as Gag may be essential to the development of a safe and effective HIV type 1 (HIV-1) vaccine. DNA vaccination represents a novel strategy for inducing potent CD8+ CTL responses in vivo. However, expression of HIV-1 structural proteins by DNA vectors has been hampered by a stringent requirement for coexpression with other viral components, such as Rev and RRE. Furthermore, even with Rev and RRE present, the level of expression of HIV-1 Gag, Pol, or Env is very low in murine cells. These problems have limited our ability to address the key issue of how to generate effective CTL responses to Gag in a mouse model. To overcome this problem, we compared several novel DNA expression vectors for HIV-1 Gag protein expression in primate and mouse cells and for generating immune responses in mice after DNA vaccination. A DNA vector containing wild type HIV-1 gag coding sequences did not induce detectable Gag expression in any of the cells tested. Attempts to increase nuclear export of Gag expression RNA by adding the constitutive transport element yielded only a moderate increase in Gag expression in monkey-derived COS cells and an even lower increase in Gag expression in HeLa cells or several mouse cell lines. In contrast, silent-site mutations in the HIV-1 gag coding sequences significantly increased Gag expression levels in all cells tested. Furthermore, this construct induced both Gag-specific antibody and CTL responses in mice after DNA vaccination. Using this construct, we achieved stable expression of HIV-1 Gag in the mouse cell line p815, which can now be used as a target cell for measuring HIV-1 Gag-specific CTL responses in immunized mice. The DNA vectors described in this study should make it possible to systematically evaluate the approaches for maximizing the induction of CTL responses against HIV-1 Gag in mouse and other animal systems.

* Corresponding author. Mailing address for George N. Pavlakis: ABL-Basic Research Program, Bldg. 535, Rm. 210, NCI-FCRDC, Frederick, MD 21702-1201. Phone: (301) 846-1474. Fax: (301) 846-6368. E-mail: pavlakis{at}ncifcrf.gov. Mailing address for Xiao-Fang Yu: Department of Molecular Microbiology and Immunology, The Johns Hopkins School of Hygiene & Public Health, 615 N. Wolfe St., Baltimore, MD 21205. Phone: (410) 955-3768. Fax: (410) 614-8263. E-mail: xfyu{at}jhsph.edu.

Journal of Virology, November 1999, p. 9145-9152, Vol. 73, No. 11
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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