Genetic instability of live, attenuated human immunodeficiency virus type 1 vaccine strains

Live, attenuated viruses have been the most successful vaccines in monkey m odels of human immunodeficiency virus type 1 (HIV-1) infection. However, th ere are several safety concerns about using such an anti-HIV vaccine in hum ans, including reversion of the vaccine strain to virulence and recombinati on with endogenous retroviral sequences to produce new infectious and poten tially pathogenic viruses. Because testing in humans would inevitably carry a substantial risk, we set out to test the genetic stability of multiply d eleted HIV constructs in perpetuated tissue culture infections. The Delta 3 candidate vaccine strain of HIV-1 contains deletions in the viral long ter minal repeat (LTR) promoter and the vpr and nef genes. This virus replicate s with delayed kinetics, but a profound enhancement of virus replication wa s observed after approximately 2 months of culturing. Analysis of the rever tant viral genome indicated that the three introduced deletions were mainta ined but a 39-nucleotide sequence was inserted in the LTR promoter region. This insert was formed by duplication of the region encoding three binding sites for the Sp1 transcription factor. The duplicated Sp1 region was demon strated to increase the LTR promoter activity, and a concomitant increase i n the virus replication rate was measured. In fact, duplication of the Sp1 sites increased the fitness of the Delta 3 virus (Vpr/Nef/U3) to levels hig her than that of the singly deleted Delta Vpr virus. These results indicate that deleted HIV-1 vaccine strains can evolve into fast-replicating varian ts by multiplication of remaining sequence motifs, and their safety is ther efore not guaranteed. This insight may guide future efforts to develop more stable anti-HIV vaccines.