Human papillomavirus 16-specific T cell responses in classic HPV-related vulvar intra-epithelial neoplasia. Determination of strongly immunogenic regions from E6 and E7 proteins.

Cell-mediated immunity directed against human papillomavirus 16 (HPV-16) antigens was studied in 16 patients affected with classic vulvar intra-epithelial neoplasia (VIN), also known as bowenoid papulosis (BP). Ten patients had blood lymphocyte proliferative T cell responses directed against E6/2 (14-34) and/or E6/4 (45-68) peptides, which were identified in the present study as immunodominant among HPV-16 E6 and E7 large peptides. Ex vivo enzyme-linked immunospot-interferon (IFN)-gamma assay was positive in three patients who had proliferative responses. Twelve months later, proliferative T cell responses remained detectable in only six women and the immunodominant antigens remained the E6/2 (14-34) and E6/4 (45-68) peptides. The latter large fragments of peptides contained many epitopes able to bind to at least seven human leucocyte antigen (HLA) class I molecules and were strong binders to seven HLA-DR class II molecules. In order to build a therapeutic anti-HPV-16 vaccine, E6/2 (14-34) and E6/4 (45-68) fragments thus appear to be good candidates to increase HPV-specific effector T lymphocyte responses and clear classic VIN (BP) disease lesions.

Temporal loss of Nef-epitope CTL recognition following macaque lipopeptide immunization and SIV challenge.

To address the subtle interactions between antiviral cytotoxic T-cell (CTL) immune responses and the evolution of viral quasispecies variants in vivo, we performed a longitudinal study in a simian immunodeficiency virus (SIV)-infected rhesus macaque that had a long experimental SIV infection before developing simian AIDS. Before being infected with SIV, this animal was immunized with a mixture of seven lipopeptides derived from SIV Nef and Gag proteins and showed a bispecific antiviral CTL response directed toward Nef 169-178 and 211-225 peptides. After SIV infection, CTL activity against the Nef 169-178 epitope was no longer detectable, as assessed from peripheral blood mononuclear cells stimulated by autologous SIV. CTL activity against the 211-225 epitope was lost after 3 months, and an additional CTL response to the amino acids 112-119 Nef epitope emerged. Analysis of the Nef proviral sequence revealed the presence of immune escape variants first in the 211-225 epitope and much later in the 112-119 epitope. In contrast, epitope 169-178 showed only two mutations among all viral sequencing performed. We conclude that in this macaque, bispecific CTL exerted a strong selective pressure and escape virus mutants finally emerged. We identified CTL recognizing a conserved Nef epitope 112-119 (SYKLAIDM), essential for viral replication, which could be associated with a prolonged AIDS-free period. These results stress the importance of the induction of broader multispecific CTLs directed against highly conserved and functional T-cell epitopes by vaccination, with the aim of keeping HIV infection in check.

Long-term humoral and cellular immunity induced by a single immunization with replication-defective adenovirus recombinant vector.

This study examines the suitability of replication-defective adenovirus vectors for engineering recombinant vaccines. The immunological abilities and limitations of E1-deleted adenoviruses containing the lacZ gene (Ad-beta-gal) were investigated by examining the humoral and cellular immune responses to the beta-galactosidase protein. BALB/c mice (H-2d) were given in a single injection of recombinant adenovirus. The cytotoxic T lymphocyte (CTL) response of spleen cells was evaluated. Recognized target cells were H-2d-derived tumor cells transfected by the lac Z gene, or incubated with the 876-884 beta-galactosidase peptide known to be restricted by the Ld molecule of the major histocompatibility complex. A long-lasting beta-galactosidase-specific cytotoxic T cell response was obtained. By contrast, CTL from mice immunized with the Ld-restricted peptide were less specific for the endogenous epitope presented by the transfectants expressing beta-galactosidase. Ad-beta-gal-immunized mice were also protected against an intra-cerebral challenge with a recombinant vaccinia virus expressing the lac-Z gene. These results suggest that Ad-beta-gal-induced CTL have protective abilities in vivo. The induction of beta-galactosidase-specific T helper lymphocytes and humoral IgG responses were also examined. A proliferative response occurred only late after immunization and the primed T lymphocytes produced interleukin-2, but no interleukin-4. A humoral IgG response to the beta-galactosidase protein was detected 15-30 days after a single immunization and remained stable for 6 months without boosting. Lastly, we followed the evolution of the immune response over the course of successive immunizations. The magnitude and kinetics of the cellular and humoral responses were similar to those obtained after a single immunization. Consistent with these observations, an adenovirus-specific neutralizing antibody response was detected as early as the second immunization. Thus, a single immunization with a replication-defective adenovirus recombinant vector induces long-lasting humoral and cellular immune responses specific to the transgene product.

Cytotoxic T-cell response and AIDS-free survival in simian immunodeficiency virus-infected macaques.

OBJECTIVES: To determine whether cytotoxic T lymphocytes have a beneficial effect during infection with the simian immunodeficiency virus (SIV) in macaques. DESIGN AND METHODS: We followed up 12 rhesus macaques experimentally infected with SIV. Cytotoxic T lymphocytes were detected in nine macaques, who were subdivided into a group of high responders (n = 6), with a sustained and polymorphic response directed against most SIV proteins, and a second group of weak responders (n = 3), in which the responses were only transient and directed against only a few proteins. A third group was characterized by the absence of any cytotoxic T-lymphocyte response (n = 3). Proliferative responses closely paralleled cytotoxic responses in intensity and evolution. RESULTS: Clinical profiles and CD4 cell counts were markedly linked to cytotoxic activity; five out of six macaques that responded to multiple proteins were still healthy 2 years after SIV infection, with two of them presenting a decrease in circulating CD4 cells concomitant with the disappearance of the cytotoxic T-lymphocyte response. Conversely, five non-responder or weak-responder macaques developed overt disease after 4-21 months. CONCLUSIONS: These data suggest that a cytotoxic response may predict a better clinical outcome.