Induction of Survivin-Specific, HLA-A24 Restricted Cytotoxic T Lymphocytes Using an Immunodominant Peptide; In-Vitro Feasibility Study and a Clinical Trial of Survivin Peptide Vaccination.

Survivin, a member of the inhibitors of the apoptosis family, is overexpressed frequently in a variety of cancers and hematological malignancies, but not in normal tissues. Murine in vivo and human in vitro studies have suggested that immunotherapy of cancer patients using survivin peptide might be feasible. In the present study, we examined whether HLA-A24 restricted cytotoxic T lymphocytes (CTL) which recognize survivin peptide can be generated from peripheral blood of lymphoma patients. HLA-A24 positive four lymphoma patients and two healthy volunteers were enrolled. Three immunodominant 9-mer candidate peptides (2B, 3A, 3B) were selected on the basis of anchoring motif of peptide binding to HLA-A24 molecule. CD8 T cells from the patients and healthy volunteers were stimulated several times with autologous monocyte-derived dendritic cells pulsed with survivin or control HIV peptides and tested for peptide-specific cytotoxicity by an LDH-release assay. CTL generated with survivin 2B peptide lysed autologous monocytes pulsed with a relevant peptide. However, other survivin peptides did not elicit CTL response. Non-pulsed or HIV peptide-pulsed monocytes were not lysed. On the other hand, CTL generated with HIV peptide only lysed HIV peptide-pulsed monocytes. CTL did not lyse allogeneic monocytes regardless of the peptide pulse. Cytotoxic activity was inhibited by the pretreatment of target cells by anti-HLA class I, not by anti-HLA-DR monoclonal antibody, indicating that the lysis was HLA class I (A24) restricted. These cells did not lyse Daudi and K562, excluding the involvement of LAK or NK activity. Importantly, these survivin peptide-specific CTL showed cytotoxicity to the patient’s lymphoma cells and HLA-A24 positive lymphoma cells. Based on these preclinical data, we have just started a pilot clinical study to examine the safety and the efficacy of peptide vaccination to relapsed, chemotherapy-resistant malignant lymphoma patients who are HLA-A24 and survivin positive. A 46-year old male patient with diffuse large B-cell lymphoma has just completed two courses of four vaccinations at two-week intervals with survivin 2B peptide (1 μg subcutaneously) in an incomplete Freund’s adjuvant (Montanide ISA-51, SEPPIC Co. France). We observed a marked decrease in the size of extra-nodular surface and cervical lymphnodes following vaccinations without serious adverse events. Immunological evaluations using HLA-tetramer and T cell receptor clonality assays revealed an increase in survivin-specific CTL frequency after vaccinations. The in-vitro feasibility study and pilot clinical trial indicate that a vaccination with a survivin peptide is safe and might be a promising novel strategy for the treatment of lymphoma patients.