Phase 2 trial of a multivalent WT1 peptide vaccine (galinpepimut-S) in acute myeloid leukemia

Abstract

A National Cancer Institute consensus study on prioritization of cancer antigens ranked the Wilms tumor 1 (WT1) protein as the top immunotherapy target in cancer. We previously reported a pilot study of a multivalent WT1 peptide vaccine (galinpepimut-S) in acute myeloid leukemia (AML) patients. We have now conducted a phase 2 study investigating this vaccine in adults with AML in first complete remission (CR1). Patients received 6 vaccinations administered over 10 weeks with the potential to receive 6 additional monthly doses if they remained in CR1. Immune responses (IRs) were evaluated after the 6th and 12th vaccinations by CD4⁺ T-cell proliferation, CD8⁺ T-cell interferon-γ secretion (enzyme-linked immunospot), or the CD8-relevant WT1 peptide major histocompatibility complex tetramer assay (HLA-A*02 patients only). Twenty-two patients (7 males; median age, 64 years) were treated. Fourteen patients (64%) completed ≥6 vaccinations, and 9 (41%) received all 12 vaccine doses. Fifteen patients (68%) relapsed, and 10 (46%) died. The vaccine was well tolerated, with the most common toxicities being grade 1/2 injection site reactions (46%), fatigue (32%), and skin induration (32%). Median disease-free survival from CR1 was 16.9 months, whereas the overall survival from diagnosis has not yet been reached but is estimated to be ≥67.6 months. Nine of 14 tested patients (64%) had an IR in ≥1 assay (CD4 or CD8). These results indicated that the WT1 vaccine was well tolerated, stimulated a specific IR, and was associated with survival in excess of 5 years in this cohort of patients. This trial was registered at www.clinicaltrials.gov as #NCT01266083.

Graphical abstract

Figure 1.

Survival curves for vaccinated patients. (A) DFS (from time of CR) for entire cohort. (B) OS (from time of diagnosis) for entire cohort. (C) EFS (from time of first vaccination). (D) OS (from time of first vaccination). (E) DFS (from time of CR) for patients <60 years. (F) OS (from time of diagnosis) for patients <60 years. (G) DFS (from time of CR) for patients ≥60 years. (H) OS (from time of diagnosis) for patients ≥60 years.

Figure 2.

CD4⁺T cell proliferation. CD4⁺ T cells from prevaccination (time 0), post-6 GPS vaccinations (time 6), and post-12 GPS vaccinations (time 12) from patient 4 were incubated with indicated peptides at 20 µg/mL (“20”) or 50 µg/mL (“50”) for 5 days, and 1 µCi [³H]-thymidine was added to the cultures for 20 hours. The cell proliferation was determined by [³H]-thymidine incorporation. Data are mean ± SD from quadruplicate cultures. Negative controls were also used (incubation with no peptide present and with irrelevant peptides [B2A2 long fragment of BCR-ABL]). After 6 vaccinations, cell proliferation increased sixfold to 331, eightfold to 427, 11-fold to 122A1, and 13-fold to 122A (P = .008) There was no significant dose dependency of the peptides, and the CD4⁺ T-cell response was sustained through the period of vaccination, although the degree varied among the individual peptides.

Figure 3.

IFN-γ secretion by CD8⁺cells. CD3⁺ T cells from patient 1 (Pt. 1) 1 and patient 14 (Pt. 14) were stimulated with WT1-A (native) peptide. IFN-γ–secreting T cells were measured by ELISPOT assay after challenge with indicated peptides. Controls were no peptide (only CD14⁺ antigen-presenting cells) or irrelevant Ewing sarcoma–derived peptide (EW). Data are mean ± SD from quadruplicate cultures from before GPS administration (time 0), after 6 GPS vaccinations (time 6), and after 12 GPS vaccinations (time 12). A positive response was defined as meeting all of the following criteria: (1) a twofold increase in the IFN-γ–secreting cells and in frequencies of CD8⁺ WT1-A tetramer–positive cells over the controls, (2) P < .05, and (3) >30 spots per 10⁵ cells. Results indicate that a WT1-A–specific response can be generated by challenge to both native and heteroclitic peptide, suggesting processing and presentation of the antigen. In patient 1, a stronger response was seen after 6 vaccinations and then faded. In patient 14, a stronger response was seen after 12 vaccinations.

Figure 4.

Tetramer assays. CD3⁺ T cells from patient 1 (A) and patient 14 (B) were stained with WT1-A/HLA-A*02:01 tetramer with CD8⁺ and other T-cell markers. Percentage of tetramer-positive CD8⁺ T cells (number shown in the top right of each histogram and highlighted in bold print) were gated on CD3⁺ events after establishing a lymphocyte gate. Cells from before vaccine (time 0 [T0]), after 6 GPS vaccinations (T6), and after 12 GPS vaccinations (T12) are shown. (A) CD8⁺ cells only from patient 1 are shown. Cultures were preincubated with native (WT1-A) and heteroclitic (WT1-A1) peptides. The data are controls (no peptide [−]) on the left followed by representative staining from triplicate cultures following antigen exposure (+) on the right. Before vaccination, there were low percentages of tetramer-positive cells in the CD8⁺ population. (B) Both CD8⁺ and CD8⁻ cells from patient 14 are shown. Cultures are incubated with WT1-A peptides. Bottom row of histograms show concurrent testing with irrelevant tetramers. After vaccination, a large increase in the percentage of WT1-A–specific CD8⁺ T cells was noted in primed cultures indicating that vaccination with the heteroclitic peptide can induce T cells that recognize the native sequence.

Figure 5.

Survival curves according to immunologic response. (A) DFS (from time of CR; median not reached vs 15.6 months, P = .11). (B) OS (from time of diagnosis; median not reached vs 35.8 months, P = .08).