Induction of a primary human cytotoxic T-lymphocyte response against a novel conserved epitope in a functional sequence of HIV-1 reverse transcriptase

Abstract

To identify novel major histocompatibility complex (MHC) class I-restricted cytotoxic T-lymphocyte (CTL) epitopes conserved in HIV-1. Potential conserved CTL epitopes were selected using a predictive computer algorithm based on a human leukocyte antigen (HLA)-A*0201 peptide-binding motif and tested for actual binding to the human processing defective cell line 174.CEM T2 (T2). Hence, the amino-acid sequences of 14 full-length sequenced HIV-1 strains were analysed. An in vitro primary peptide-specific human CTL response was induced with responding lymphocytes of an HIV-1-seronegative donor. Responding T cells were cloned by limiting dilution and tested for their ability to recognize naturally processed antigen in a 51Cr-release assay using recombinant vaccinia-HIV protein-infected B-lymphoblastoid cells (B-LCL) as target cells. The analysis of peptides bearing the HLA-A*0201 motif for conservation resulted in one peptide of Env, three of Gag and 12 of Pol. Only Gag340-348, Pol83-92, Pol267-277 and Pol960-968 showed binding properties to T2 comparable with those of known CTL epitopes Gag76-84 SLYNTVATL and Pol468-476 ILKEPVHGV. A successful primary MHC class I-restricted CTL response was induced against Pol468-476 and Pol267-277 VLDVGDAYFSV, a peptide in a functional sequence of reverse transcriptase (RT). The resulting CD8+ CTL clones were peptide-specific and able to specifically lyse recombinant vaccinia-HIV-1 RT-infected HLA-A*0201-matched B-LCL. The method used to screen proteins sequences for potential CTL epitopes, test selected peptides for binding to MHC class I and induction of an in vitro primary response against optimal binding peptides resulted in the identification of at least one novel conserved CTL epitope. The novel epitope is located in an area crucial for RT activity. This study demonstrates the feasibility of identifying highly conserved HIV-1-derived peptides capable of eliciting novel anti-HIV-1 CTL responses.