Immunogenicity of a human immunodeficiency virus (HIV) polytope vaccine containing multiple HLA A2 HIV CD8(+) cytotoxic T-cell epitopes

Abstract

Compelling evidence now suggests that alphabeta CD8 cytotoxic T lymphocytes (CTL) have an important role in preventing human immunodeficiency virus (HIV) infection and/or slowing progression to AIDS. Here, we describe an HIV type 1 CTL polyepitope, or polytope, vaccine comprising seven contiguous minimal HLA A2-restricted CD8 CTL epitopes conjoined in a single artificial construct. Epitope-specific CTL lines derived from HIV-infected individuals were able to recognize every epitope within the construct, and HLA A2-transgenic mice immunized with a recombinant virus vaccine coding for the HIV polytope also generated CTL specific for different epitopes. Each epitope in the polytope construct was therefore processed and presented, illustrating the feasibility of the polytope approach for HIV vaccine design. By simultaneously inducing CTL specific for different epitopes, an HIV polytope vaccine might generate activity against multiple challenge isolates and/or preempt the formation of CTL escape mutants.

Figures

FIG. 1

HIV polytope insert. The first epitope and then every second CTL epitope are underlined.

FIG. 2

Agarose gel of HIV polytope PCR products from cDNA (lane B) and viral DNA (lane E) derived from rVV.HIV.pt-infected cells. Lanes A and F, 1-kb markers (Gibco BRL, Gaithersburg, Md.); lane B, RT-PCR of the HIV polytope mRNA; lane C, RT-PCR control without RT; lane D, PCR control with water as template; lane E, PCR of viral DNA. Molecular size markers indicated by arrowheads are (from the top) 394, 344, 298, and 220 bp.

FIG. 3

Epitope-specific CTL lines derived from PBMC of HIV-infected individuals (H28, H10, H19, H21, and H33) used as effectors against (i) HLA A2+ LCLs sensitized with the indicated peptide (black squares) or not sensitized (white squares) (columns headed “Peptide”), (ii) HLA A2+ LCLs infected with rVV.HIV.pt (black squares) or a control recombinant vaccinia virus (white squares) (columns headed “Polytope”), and (iii) HLA A2+ LCLs infected with rVV.nef or rVV.pol (black squares) or a control recombinant vaccinia virus (white squares) (column headed “Antigen”). The epitope listed on the left of each row was used to restimulate the bulk cultures, which were used to generate the data in that row. Bulk cultures from each individual were separately restimulated with the indicated peptide, split, and used against peptide and polytope and sometimes against whole antigen expressing target cells. A summary of patient data is shown in Table 2. The negative results, for patient H28, are shown to illustrate the specificity of the in vitro restimulation protocol.

FIG. 4

Data from HHD mice immunized with HIV polytope vaccines. (A) Mice were immunized with rVV.HIV.pt, and pooled splenocytes were restimulated in vitro with each of the indicated peptides and used as effectors against target cells sensitized with the same peptide (black squares) or not sensitized (white squares). (B) Mice were immunized with two injections, either of DNA vaccine coding for the HIV polytope followed by rVV.HIV.pt (DNA/rVV) or of a control DNA plasmid followed by rVV.HIV.pt (rVV). Splenocyte populations from each mouse were individually restimulated with each peptide and used as effectors against target cells sensitized with peptide and not sensitized. (Thus, the first six bars represent restimulation and lysis with PLTFGWCYKL.) Results were calculated as follows: percent lysis of target cells sensitized with peptide − percent lysis of target cells not sensitized (± standard error). DNA vaccination alone produced only weak CTL responses ranging from 5 to 10% (data not shown).