Repeated PR1 and WT1 peptide vaccination in Montanide-adjuvant fails to induce sustained high-avidity, epitope-specific CD8+ T cells in myeloid malignancies

Abstract

Background: We previously showed that vaccination with one dose of PR1 and WT1 peptides induces transient anti-leukemia immunity. We hypothesized that maintenance of a sustained anti-leukemia response may require frequent boost injections.

Design and methods: Eight patients with myeloid malignancies were enrolled in this phase II study, and 6 completed 6 injections of PR1 and WT1 peptides in Montanide-adjuvant with GM-CSF, every two weeks.

Results: Both high- and low-avidity PR1 or WT1-specific CD8(+) T cells were detected in all evaluable patients after the first vaccine dose. Repeated vaccination led to selective deletion of high avidity PR1- and WT1-specific CD8(+) T cells and was not associated with significant reduction in WT1-expression. Additional boosting failed to increase vaccine-induced CD8(+) T-cell frequencies further and in all patients the response was lost before the 6(th) dose. PR1- or WT1-specific CD8(+) T cells were not detected in bone marrow samples, excluding their preferential localization to this site. Following a booster injection three months after the 6(th) vaccine dose, no high-avidity PR1 or WT1-specific CD8(+) T cells could be detected, whereas low-avidity T cells were readily expanded.

Conclusions: These data support the immunogenicity of PR1 and WT1 peptide vaccines. However, repeated delivery of peptides with Montanide-adjuvant and GM-CSF leads to rapid loss of high-avidity peptide-specific CD8(+) T cells. These results may offer an explanation for the lack of correlation between immune and clinical responses observed in a number of clinical trials of peptide vaccination. New approaches are needed to induce long-term high-avidity memory responses against leukemia antigens.

Trial registration: ClinicalTrials.gov NCT00499772.

Figures

Figure 1.

Kinetics of vaccine-induced PR1- and WT1-specific CD8+ T-cell response in 6 patients completing 6 courses of biweekly injections. Results in 6 individual patients who completed the vaccination program are shown. Weeks after vaccination are shown on the X-axis. PR1/HLA-A*0201+ (black diamond square and solid connecting line) and WT1/HLA-A*0201+ (black square and dashed connecting line) CD8+ T cells are expressed as absolute numbers per mL of peripheral blood (Y-axis). The vaccine-induced CD8+ T-cell responses after PR1 and WT booster injection for patients 4, 6 & 7 are presented. Abbreviations: ALC: absolute lymphocyte count, V: vaccine dose.

Figure 2.

Avidity of vaccine-induced PR1 and WT1 CD8+ T cells. (A) Avidity of PR1 and WT1 CD8+ T cells following repeated peptide vaccination (study 07-H-0159). (B) Avidity of PR1 and WT1 CD8+ T cells following a single dose of PR1 and WT1 peptide vaccination (study 06-H-0062). Stimulation of PBMC with 0.1 and 10 μM of PR1, WT1 peptides and CMV peptides determined high- and low-avidity responses to PR1 and WT1 vaccines, respectively. Results show ratios of high- to low-avidity CD8+ T-cell responses for PR1 (black bars), WT1 (gray bars) and CMV (hatched bars) in 2 CMV seropositive patients. Ratios were obtained by the following calculation: IFN-γ+ CD8+ T cell (%) with 0.1 μ peptide/IFN-γ+ CD8+ T cell (%) with 10 μM peptide. A ratio > 1.0 represents predominantly high-avidity responses, whereas a ratio of < 1.0 represents predominantly low-avidity responses. Abbreviations: V: vaccine; H/L: High-/Low-avidity ratio.

Figure 3.

Frequencies of CD4+Foxp3+ and ratio of CD4+Foxp3+/CD4+CD25+ T cells following vaccination. Frequencies are shown before each vaccine in 5 patients. (A) Following vaccination, there was a significant and transient reduction in frequencies of CD4+Foxp3+ T cells (B) and a significantly lower proportion of CD4+CD25+ T cells that were Foxp3+ve, indicating selective elimination of CD4+CD25+FOXP3+ T cells. wk: weeks post-vaccine.