Immunoprevalence of the CD4+ T-cell response to HIV Tat and Vpr proteins is provided by clustered and disperse epitopes, respectively.

Recent studies have suggested including nonstructural proteins as Tat and Vpr in HIV vaccines. However, little is known about the CD4+ T-cell response that these small proteins induce in humans. We have therefore evaluated these responses by in vitro priming experiments of CD4+ T lymphocytes harvested in healthy donors. In the Tat protein, only one peptide primed CD4+ T cells of eight HLA unrelated healthy donors. T cells induced by this peptide recognized immature DC loaded with the native Tat protein and are restricted by multiple HLA-DR molecules, in agreement with its binding capacity. This peptide was therefore processed in an appropriate manner and was highly immunoprevalent. CD4+ T-cell response to Vpr peptides was more disperse and involved six different peptides depending on the HLA-DR molecules of the donors. Two overlapping peptides were T-cell stimulating in at least half of the donors. T-cell response to Vpr in multiple donors is the result of a combination of several CD4+ T-cell epitopes with good to moderate immunoprevalence. Altogether, our results show that the frequency of responders to HIV Tat or Vpr proteins relies on one or multiple CD4+ T-cell epitopes, respectively.

Comprehensive analysis of HLA-DR- and HLA-DP4-restricted CD4+ T cell response specific for the tumor-shared antigen survivin in healthy donors and cancer patients.

Because of the wide distribution of the survivin Ag in a variety of tumors, we have investigated the survivin-specific CD4+ T cell response in healthy donors and cancer patients. Screening of the entire sequence of survivin for HLA class II binding led to the identification of seven HLA-DR promiscuous peptides, including four HLA-DP4 peptides. All of the peptides were able to prime in vitro CD4+ T cells of eight different healthy donors. The peptide-specific T cell lines were stimulated by dendritic cells loaded with the recombinant protein or with the lysates of tumor cells. The high frequency of responders (i.e., immunoprevalence) was provided by a wide reactivity of multiple peptides. Six peptides were T cell stimulating in at least half of the donors and were close to CD8+ T cell epitopes. HLA-DR molecules were more frequently involved in T cell stimulation than were HLA-DP4 molecules, and hence immunoprevalence relies mainly on HLA-DR promiscuity in the survivin Ag. In two cancer patients a spontaneous CD4+ T cell response specific for one of these peptides was also observed. Based on these observations, the tumor-shared survivin does not appear to be the target of immune tolerance in healthy donors and cancer patients and is a relevant candidate for cancer vaccine.

Selective identification of HLA-DP4 binding T cell epitopes encoded by the MAGE-A gene family.

Because of the high frequency of HLA-DP4 in the Caucasian population, we have selectively delineated HLA-DP4 restricted T cell epitopes in the MAGE-A tumor antigens. We identified 12 good binders to HLA-DP4 and investigated the capacity of the seven best binders to induce in vitro specific CD4+ T cell lines from HLA-DP4 healthy donors. We found that the MAGE-A1 90-104 peptide exhibited a high and constant frequency of CD4+ T cell precursors in all the six tested donors. The MAGE-A1 268-282 peptide was found immunogenic in only two donors but with a high precursor frequency. The MAGE-A12 127-141 peptide was T cell stimulating in six different donors and induced fewer T cell lines. The peptide-specific T cell lines were stimulated by DC loaded with the lysates of cells transfected with MAGE-A1 or MAGE-A12, or loaded with the recombinant protein. We also show that the immunoreactivity of CD4+ T cell epitopes restricted to the same HLA II molecule may vary from one individual to another, as a result of inter-individual variations in the CD4+ T cell repertoire.

Scanning the HIV genome for CD4+ T cell epitopes restricted to HLA-DP4, the most prevalent HLA class II molecule.

HLA-DP4 alleles are carried by 75% of individuals and are the most frequent HLA II alleles worldwide. Because we have recently characterized the peptide-binding specificity of HLA-DP4 molecules, we developed a peptide-binding prediction method to identify HLA-DP4-restricted peptides in multiple Ags. CD4(+) T cell response plays a key role in the immune control of HIV infection, but few HIV-specific T cell epitopes with multi-individual specificity have been identified. They are mostly restricted to HLA-DR molecules, which are very polymorphic molecules. We therefore looked for HLA-DP4-restricted CD4(+) T cell epitopes in the whole genome of HIV. Twenty-one peptides were selected from the HXB2 HIV genome based on the prediction of binding to HLA-DP4 molecules. They were submitted to HLA-DP4-binding assays. Seventeen peptides bound to the HLA-DP401 molecule, whereas 15 peptides bound to HLA-DP402. Six peptides bound very tightly to HLA-DP401 and were investigated for their capacity to induce specific CD4(+) T cell lines in vitro using dendritic cells and CD4(+) T cells collected from eight seronegative HLA-DP4(+) donors. Four peptides from env and reverse transcriptase proteins induced in vitro-specific T cell lines restricted to HLA-DP4 molecules. Peptide-induced T cells recognized variants other than the HXB2 sequence and were stimulated by native Ags processed by immature dendritic cells. The reverse transcriptase peptide is present in 65% of the isolated HIV variants. To our knowledge, we describe the first HIV epitopes restricted to HLA-DP4 molecules.