Leukemia-associated antigen-specific T-cell responses following combined PR1 and WT1 peptide vaccination in patients with myeloid malignancies

Abstract

We describe the safety and immunogenicity of a combined vaccine of 2 leukemia-associated antigenic peptides, PR1 and WT1. Eight patients with myeloid malignancies received one subcutaneous dose each of PR1 and WT1 vaccines in Montanide adjuvant, with granulocyte-macrophage colony-stimulating factor. Patients were reviewed weekly for 4 weeks to monitor toxicity and immunologic responses. Toxicity was limited to grades 1 to 2. Using peptide/HLA-A 0201 tetramers and intracellular interferon-gamma staining, CD8(+) T cells against PR1 or WT1 were detected in 8 of 8 patients after a single vaccination. To monitor the kinetics of vaccine-induced CD8(+) T-cell responses and disease regression after vaccination, absolute PR1 and WT1(+)CD8(+) T-cell numbers and WT1 expression were studied weekly after vaccination. Responses occurred as early as 1 week after vaccination. After vaccination, the emergence of PR1 or WT1(+)CD8(+) T cells was associated with a decrease in WT1 mRNA expression as a marker of minimal residual disease, suggesting a vaccine-driven antileukemia effect. Conversely, loss of response was associated with reappearance of WT1 transcripts (P < .01). This is the first demonstration that a combined PR1 and WT1 vaccine is immunogenic. These results support further studies of combination immunization strategies in leukemia patients.

Trial registration: ClinicalTrials.gov NCT00313638.

Figure 1

CD8⁺ T-cell response to PR1 and WT1 vaccination by peptide/HLA-A2 tetramer analysis. Tetramer analysis of PBMCs was performed by 5-color flow cytometry. Longitudinal data from patient 8 are presented. PR1/HLA-A*0201⁺ and WT1/HLA-A*0201⁺ CD8⁺ T cells were gated on CD3⁺ events after passing through a small lymphocyte gate; HLA-A2–null tetramer was used similarly as a negative control.

Figure 2

CD8⁺ T-cell responses to PR1 and WT1 vaccination by intracellular IFN-γ assay. (A-C) Longitudinal data on IFN-γ production by CD8⁺ T cells in PBMC samples from patient 8, cultured for 6 hours with PR1 (A), with WT1 (B), or without peptide (negative control [C]) are presented. Results are expressed as percentages of total CD8⁺ T cells. (D,E) Frequencies of PR1- and WT1-specific CD8⁺ T cells by tetramer analysis and IFN-γ production were compared and found to be highly correlated; P < .001.

Figure 3

PR1- and WT1-specific CD8⁺ T-cell responses in peripheral blood in relation to disease response as measured by WT1/ABL and BCR-ABL/ABL gene expression. Results in 8 individual patients are shown. Weeks after vaccination are shown on the x-axis. PR1/HLA-A*0201⁺ (black square and connecting line) and WT1/HLA-A*0201⁺ (dark blue diamonds and connecting line) CD8⁺ T cells are expressed as absolute numbers per milliliter of peripheral blood (left y-axis); the shaded area represents absolute numbers of CMVpp65₄₉₅/HLA-A*0201⁺ CD8⁺ T cells. The absolute lymphocyte count (gray triangle and connecting line) is expressed as absolute number per liter of peripheral blood (left y-axis); WT1 and BCR-ABL gene expression in peripheral blood is expressed as the ratio of WT1/ABL (red circles and connecting line) and BCR-ABL/ABL (red circles and dashed connecting line) (right y-axis).