The duration of human immunodeficiency virus (HIV-1) infection prior to the development of AIDS is variable, and for most patients the exact time of infection is not known. A group of 38 HIV-1-infected subjects was tested while asymptomatic for comparative cytotoxic lymphocyte responses to the Gag and envelope antigens of HIV-1. Twenty of the 38 patients had no detectable primary cytotoxic T lymphocyte (CTL) response to Gag, and this was associated with a relative risk of 1.89 for progression to ARC or AIDS during the subsequent 3 to 40 months of observation when compared with patients who had Gag-specific CTL activity at the beginning of the observation period. In contrast, no significant association was observed between envelope-specific cytotoxic activity and disease progression. Other patient characteristics, including CD4+ T lymphocyte counts and antibody levels to the p24gag protein, measured at the start of observation, did not correlate with disease progression during the observation period. This suggests that the anti-Gag CTL response may be protective during HIV-1 infection.

By using target cells that expressed isolated env, gag, p27"'f, or p23vaf molecules introduced by recombinant vaccinia viruses containing genes encoding these polypeptides, it was possible to identify env, gag, p27"ef, and p23v1f as cytolytic target antigens for freshly isolated blood cells from human immunodeficiency virus 1 (HIV-1) seropositive patients. Most of the patients tested (95%) manifested a specific cytotoxic activity against vaccinia virus-env-infected target cells. The env-specific cytotoxic activity was not restricted by the major histocompatibility complex and was not mediated by T lymphocytes, as shown by the absence of blocking effect with an anti-CD3 monoclonal antibody and by the inefficiency of CD3+, CD8+, or CD4+ and CD8+ depletion to reduce the cytotoxic activity against the env-expressing target cells. In the same conditions, the cytotoxic activity specific for gag was abrogated and gag major histocompatibility complex-restricted cytotoxic T lymphocytes were detected in 85% of the subjects tested. Therefore, in a HIV-1 seropositive subject, distinct types of effector cells mediate the lysis of target cells expressing gag and env proteins. * Corresponding author. (100 U/ml), streptomycin (100 p.g/ml), HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) (20 mM), and 10% heat-inactivated fetal calf serum (Biological Industries, Kibbutz Haemek, Israel) (RPMIc) was used for most assays. The mouse EL4 cell line was a gift from G. Milon (Institut Pasteur, Paris, France), the mouse P815 cell line was a gift from D. Juy (Institut Pasteur, Paris, France), and the human K562 cell line was a gift from E. Gomard (Hopital Cochin, Paris, France). Monoclonal antibodies and complement. Monoclonal antibody anti-Leullb, (which recognizes the CD16 antigen) was purchased from Becton Dickinson, Grenoble, France. Monoclonal antibodies directed against the CD3, CD4, and CD8 antigens were from Ortho Diagnostics Systems, Roissy, France (OKT3, OKT4, and OKT8, respectively) or Immunotech, Marseille, France (IOT3, IOT4, and IOT8a, respectively). The low Tox H rabbit complement was purchased from Cedarlane, Tebu, France. Viruses. The different recombinant viruses used to infect the target cells have been previously described, except for the vaccinia virus (VV) Q recombinant (12, 19-21; G. Rautmann et al., in press). Briefly, the viruses used were the wild-type (WT) VV, strain Copenhagen, or various recombinants encoding either the middle T antigen of the polyomavirus (as a control) or the env (gpl60), gag (p55), p27"1f (F or 3' open reading frame) or p23"f (Q or sor) antigens of the HIV-1/BRU isolate (recombinant VV TG 1139, VV TG 1144, VV TG 1147, or VV TG 1160, respectively). The VV Q recombinant was constructed as follows: the EcoRI fragment from the plasmid pJ 19-13 (37) containing the coding sequence from the vif (Q) gene of HIV-1 was cloned in a M13 vector, and a BglII site was created upstream of the initiation codon by site-directed mutagenesis by using the oligonucleotide 5'TCCCTAAAGATCTTT3'. The resul...

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