Identification of a new hTERT-derived HLA-A*0201 restricted, naturally processed CTL epitope

By the use of a neural network capable of performing quantitative predictions of peptides binding to HLA-A*0201 molecules, we identified a number of nonamer peptides derived from the catalytic subunit of telomerase, human telomerase reverse transcriptase (hTERT). Five nonimmunogenic peptides with measured binding affnities for HLA- A* 0201 ranging from 155 to 1,298 nM were modified at the P1, P2 and P9 positions, respectively, to achieve stronger HLA- A*0201 binding. One peptide, mp30 - 38 (mp30), with an L to V substitution at position 9 was subsequently found to be immunogenic in mp30 immunized HLA- A* 0201/ H2K(b) or HHD transgenic mice. The T cell reactivity obtained was directed against both the mp30 and against the unmodified p30. Anti-mp30 specific T cells generated in HLA- A* 0201 transgenic mice were dependent on TCR- CD8/ MHC- I alpha 3 binding and therefore not capable of recognizing mp30- pulsed human HLAA* 0201(+) cells or murine HLA-A*0201 transfectants. In order to show reactivity against naturally processed peptide in human tumor cells, an hTERT positive HLA- A*0201 negative colon carcinoma cell line (CCL220) was transfected with an HLA-A*0201/ H2Kb cDNA construct and used as target in ELISPOT and cytotoxicity assays. The data show that T cells from mp30 immunized HHD transgenic mice react specifically against the CCL220 transfectant indicating that p30 is naturally processed. In conclusion, we have identified a new CTL HLA-A*0201 restricted hTERT epitope, which is now, included in an ongoing phase 2 vaccine trial of patients with disseminated cancer.