IDENTIFICATION OF 2 EPITOPES ON THE DENGUE-4-VIRUS CAPSID PROTEIN RECOGNIZED BY A SEROTYPE-SPECIFIC AND A PANEL OF SEROTYPE-CROSS-REACTIVE HUMAN CD4(-LYMPHOCYTE CLONES() CYTOTOXIC T)

We analyzed the CD4(+) T-lymphocyte response of a donor who had receiv ed an experimental live-attenuated dengue 4 virus (D4V) vaccine. Bulk culture proliferative responses of peripheral blood mononuclear cells (PBMC) to noninfectious dengue virus (DV) antigens showed the highest proliferation to D-IV antigen, with lesser, cross reactive proliferati on to D2V antigen. We established CD4(+) cytotoxic T-lymphocyte clones (CTL) by stimulation with D4 antigen. Using recombinant baculovirus a ntigens, we identified seven CTL clones that recognized D4V capsid pro tein, Six of these CTL clones were cross-reactive between D2 and D4, a nd one clone was specific for D4. Using synthetic peptides. we found t hat the D4V-specific CTL clone recognized an epitope between amino aci ds (aa) 47 and 55 of the capsid protein, while the cross-reactive CTL clones each recognized epitopes in a separate location, between aa 83 and 92, which is conserved between D2V and D4V, This region of the cap sid protein induced a variety of CD4 T-cell responses, as indicated by the fact that six clones which recognized a peptide spanning this reg ion shelved heterogeneity in their recognition of truncations of this same peptide. The bulk culture response of the donor's PBMC to the epi tope peptide spanning aa 81 to 92 was also examined, Peptides containi ng this epitope induced proliferation of the donor's PBMC in bulk cult ure, but peptides not containing the entire epitope did not induce pro liferation, Also, PBMC stimulated in bulk culture with noninfectious D 4V antigen lysed autologous target cells pulsed with peptides containi ng aa 84 to 92. These results indicate that this donor exhibits memory CD4(+) T-cell responses directed against the DV capsid protein and su ggest that the response to the capsid protein is dominant not only in vitro at the clonal level but in bulk culture responses as well, Since previous studies have indicated that the CTL responses to DV infectio n seem to be directed mainly against the envelope (E) and NS3 proteins , these results are the first to indicate that the DV capsid protein i s also a target of the antiviral T-cell response.