Development of an Ad7 cosmid system and generation of an Ad7|[Delta]|E1|[Delta]|E3HIVMN env|[sol]|rev recombinant virus

doi:doi:10.1038/sj.gt.3301903

Gene Therapy (2003) 10, 326–336. doi:10.1038/sj.gt.3301903

Development of an Ad7 cosmid system and generation of an Ad7E1E3HIV_MN env/rev recombinant virus

X Nan^1,3, B Peng¹, T-W Hahn^1,4, E Richardson¹, A Lizonova², I Kovesdi² and M Robert-Guroff¹

¹Basic Research Laboratory, National Cancer Institute, Bethesda, MD, USA
²GenVec, Gaithersburg, MD, USA

Correspondence: Dr M Robert-Guroff, National Cancer Institute, 41 Library Drive MSC 5055, Building 41 Room D804, Bethesda, MD 20892-5055, USA

³Present address: American Red Cross Holland Laboratory, Department of Immunology, 15601 Crabbs Branch Way, Rockville, MD 20855-2743, USA

⁴Present address: Kangwon National University, Chuncheon City, Kangwon-do 200701, Korea

Received 28 May 2002; Accepted 29 August 2002.

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Abstract

A strategy to circumvent immune responses to adenovirus (Ad) resulting from natural infection or repeated vector administrations involves sequential use of vectors from different Ad serotypes. To further develop an Ad-HIV recombinant AIDS vaccine approach, a replication-defective recombinant Ad from a non-subgroup C virus was required. Using a cosmid system, we generated an Ad7 Delta E1 Delta E3HIV_MN env/rev recombinant virus and compared expression of the inserted HIV genes with a similarly constructed replication-competent Ad7 Delta E3HIV_MNenv/rev recombinant. Ad7 Delta E1 Delta E3HIV_MNenv/rev expressed both HIV env and rev gene products. The envelope protein was correctly processed and functional, mediating syncytia formation of Ad7 Delta E1 Delta E3HIV_MN env/rev-infected cells and CD4⁺ T lymphocytes. Ad7 Delta E1 Delta E3HIV_MNenv/rev could be amplified on 293-ORF6 cells, containing the E4 ORF6 gene, shown earlier to support production of an Ad7 vector lacking the E1a gene. The utility of this cell line is now extended to the production of replication-defective Ad7 recombinants lacking E1a, E1b, and protein IX genes. Sequential immunizations with Ad-HIV recombinants based in different Ad serotypes have been shown to effectively elicit both humoral and cellular HIV-specific immune responses. The recombinant Ad7 Delta E1 Delta E3HIV_MNenv/rev will be useful in such AIDS vaccine strategies. Further, these studies have created new cosmid vectors that can be applied to generation of single- or double-deleted Ad7 recombinants with foreign genes inserted into the E1 and/or E3 regions.