Generation of tumor-specific cytotoxic T lymphocytes by stimulation with HPV type 16 E7 peptide-pulsed dendritic cells: An approach to immunotherapy of cervical cancer

Objective. The aim of this study was to generate HPV-16 E7 peptide-specific cytotoxic T lymphocytes (CTLs) in vitro for future adoptive immunotherapy of cervical cancer. Methods. Peripheral blood mononuclear cells (PBMC) were isolated from HLA-A 2+ healthy donors. The PBMCs were incubated with HPV-16 E7(11-20) peptide a nd varying cytokines in the primary culture. Restimulation was performed we ekly with peptide-pulsed, irradiated autologous PBMCs. Alternatively, the P BMCs were depleted of abundant CD4+ cells and stimulated with HPV-16 E7(11- 20) peptide-pulsed dendritic cells. Cytolytic activity was determined by a standard 4-h Cr-51-release assay. Results. After 6 weeks in culture, we were able to establish peptide-specif ic CTL lines in one of seven donors by incubating PBMCs with HPV-16 E7(11-2 0) peptide. When we employed autologous peptide-pulsed dendritic cells to s timulate CD8+ cell-enriched PBMCs, we obtained CTL lines in four of seven d onors. The primed CTLs were able to lyse the HLA-A2+ and HPV-16+ cervical c ancer cell line Caski. SiHa, an HLA-A2-, but HPV 16+, cervical cancer cell line could be lysed only after transfection with HLA-A2. In addition, a hig h cytotoxicity (>80%) was obtained against peptide-pulsed, but not unpulsed , targets such as autologous Ebstein-Barr virus-immortalized B cells or all ogeneic lipopolysaccaride-stimulated PBMCs. DCs were clearly the most poten t of ail tested antigen presenting cells to stimulate a CTL response in a p roliferation assay. Conclusion. HPV-16 E7 peptide-specific CTLs could be generated in vitro. A practical protocol to expand the CTLs to a sufficient number for an applica tion in a clinical trial is in progress. (C) 1999 Academic Press.