SELECTION OF VIRUS VARIANTS AND EMERGENCE OF VIRUS ESCAPE MUTANTS AFTER IMMUNIZATION WITH AN EPITOPE VACCINE

In this report, we assessed the evolution of the cytotoxic T-lymphocyt e (CTL) response induced by an epitope vaccine, In two macaques immuni zed with a mixture of lipopeptides derived from simian immunodeficienc y virus (SIV) Nef and Gag proteins, CTL responses were directed agains t the same, single epitope of the Nef protein (amino acids 128 to 137) presenting an alanine at position 136 (Nef 128-137/136A). However, af ter 5 months of SIV infection, peripheral blood mononuclear cells from both macaques lost their ability to be stimulated by autologous SIV-i nfected cells while still retaining their capacity to generate cytotox ic responses after specific Nef 128-137/136A peptide stimulation. The sequences of the pathogenic viral isolate used for the challenge showe d a mixture of several variants. Within the Nef epitopic sequence from amino acids 128 to 137, 82% of viral variants expressed the epitopic peptide Nef 128-137/136A; the remaining variants presented a threonine at position 136 (Nef 128-137/136T), In contrast, sequence analysis of cloned proviral DNA obtained from both macaques 5 months after SN cha llenge showed a different pattern of quasi-species variants; 100% of c lones presented a threonine at position 136 (Nef 128-137/136T), sugges ting the disappearance of viral variants with an alanine at this posit ion under antiviral pressure exerted by Nef 128-137/136A-specific CTLs . In addition, 12 months after SIV challenge, six: of eight clones fro m one macaque presented a glutamic acid at position 131 (Nef128-137/13 1E+136T), which was not found in the infecting isolate. Furthermore, C TLs generated very early after SIV challenge were able to Lyse cells s ensitized with the Nef 128-137/136A epitope. In contrast, lysis was si gnificantly less effective or even did not occur when either the selec ted peptide Nef 128-137/136T or the escape variant peptide Nef 128-137 /131E+136T was used in a target cell sensitization assay. Dose analysi s of peptides used to sensitize target cells as well as a major histoc ompatibility complex (MHC)-peptide stability assay suggested that the selected peptide Nef 128-137/136T has an altered capacity to bind to t he MHC. These data suggest that CTL pressure leads to the selection of viral variants and to the emergence of escape mutants and supports th e fact that immunization should elicit broad CTL responses.