444 SIV Promoter Exchange Results in a Highly Attenuated Strain that Strongly Protects Against Pathogenic Challenge Virus P. Blancou*1,2, N. Chenciner2, R. Ho Tsong Fang3, V. Monceaux3, M-C. Cumont3, D. Guetard2, B. Hurtrel3, S. Wain-Hobson2 1Harvard Univ, Boston, MA; 2URM Inst Pasteur, Paris, France; and 3PIL Inst Pasteur, Paris, France
Background: Among the many simian immunodeficiency virus (SIV) immunogens only live attenuated viral vaccines have afforded strong protection to a natural pathogenic isolate. To date there has been a general inverse correlation between attenuation and protection. As the viral promoter is crucial to the tempo of replication it was reasoned that promoter exchange might confer a novel means of attenuating SIV. Here, the core enhancer and promoter sequences of the SIV macaque 239 strain have been exchanged for those of the human cytomegalovirus immediate early promoter.
Methods: Virus stock were made on CEMx174. Challenges were performed with 10 animal infectious dose 50. Viral load was assessed by bDNA assay with a threshold of 10^3 copies/ml. ELISpot assays were performed on fresh PBMC with whole inactivated HIV-2 and pooled Gag or Nef peptides. Cycling cells were quantitated by immunocytochemistry on lymph node biopsy. The percentage of CD4+ T-lymphocytes was assessed by flow cytometry.
Results: Compared to parental virus infection in macaques, primary viremia was reduced more than 1000-fold to undetectable levels with little sign of an increase of cycling cells in lymph nodes, CD4+ depletion or altered T-cell activation markers in peripheral blood. Despite such drastic attenuation high titres of antibody and vigorous ELISpot reactivity to SIV peptides were detected. When challenged intravenously at 4 or 6 months with the uncloned pathogenic SIVmac251 strain, viremia was curtailed by ~1000-fold at peak height and thereafter.
Conclusions: These findings show that promoter exchange constitutes a novel means to highly attenuate SIV while conferring protection against challenge virus.
