The protection against infection by HIV probably requires the induction of both neutralizing Abs and CTL responses. Vaccination by attenuated HIV is hardly acceptable and the use of viral genes inserted in recombinant living vectors needs further development, especially with respect to safety. The peptidic vaccination is a promising approach but free peptides are usually poorly immunogenic. Because potent immune responses have been obtained in mice with modified peptides such as lipopeptides, we have designed a study to assess the immunogenicity of lipopeptides in nonhuman primates. Seven lipopeptides were synthesized, derived from known immunogenic regions of the simian immunodeficiency virus (SIV) NEF and GAG proteins. Twelve rhesus macaques, randomly chosen and not selected on their MHC basis, were immunized subcutaneously with the seven lipopeptides in IFA. An MHC class I-restricted and CD(8+)-mediated CTL response has been observed in seven macaques directed against one or two of the synthetic immunizing peptides in each case. These CTLs were able to lyse autologous target cells infected with a recombinant vaccinia virus expressing the SIV nef or gag genes, suggesting that they recognized the naturally processed peptides. These activities are detectable in peripheral blood cells for at least 10 mo after the last immunization. Abs against the immunizing peptides have also been observed in all cases. This study demonstrates that lipopeptides can generate cytotoxic and humoral immune responses in a large number of unselected animals and this approach may thus be worth considering in the vaccination against HIV.