Dendritic cells pulsed with multifunctional Wilms' tumor 1 (WT1) peptides combined with multiagent chemotherapy modulate the tumor microenvironment and enable conversion surgery in pancreatic cancer

Abstract

Background: We aimed to develop a chemoimmunotherapy regimen consisting of a novel Wilms' tumor 1 (WT1) peptide-pulsed dendritic cell (WT1-DC) vaccine and multiagent chemotherapy and to investigate the safety, clinical outcomes, and WT1-specific immune responses of patients with unresectable advanced pancreatic ductal adenocarcinoma (UR-PDAC) who received this treatment.

Methods: Patients with UR-PDAC with stage III disease (locally advanced (LA-PDAC; n=6)), stage IV disease (metastatic (M-PDAC; n=3)), or recurrent disease after surgery (n=1) were enrolled in this phase I study. The patients received one cycle of nab-paclitaxel plus gemcitabine alone followed by 15 doses of the WT1-DC vaccine independent of chemotherapy. The novel WT1 peptide cocktail was composed of a multifunctional helper peptide specific for major histocompatibility complex class II, human leukocyte antigen (HLA)-A*02:01, or HLA-A*02:06 and a killer peptide specific for HLA-A*24:02.

Results: The chemoimmunotherapy regimen was well tolerated. In the nine patients for whom a prognostic analysis was feasible, the clinical outcomes of long-term WT1 peptide-specific delayed-type hypersensitivity (WT1-DTH)-positive patients (n=4) were significantly superior to those of short-term WT1-DTH-positive patients (n=5). During chemoimmunotherapy, eight patients were deemed eligible for conversion surgery and underwent R0 resection (four patients with LA-PDAC, one patient with M-PDAC, and one recurrence) or R1 resection (one patient with M-PDAC), and one patient with LA-PDAC was determined to be unresectable. Long-term WT1-DTH positivity was observed in three of the four patients with R0-resected LA-PDAC. These three patients exhibited notable infiltration of T cells and programmed cell death protein-1+ cells within the pancreatic tumor microenvironment (TME). All patients with long-term WT1-DTH positivity were alive for at least 4.5 years after starting therapy. In patients with long-term WT1-DTH positivity, the percentage of WT1-specific circulating CD4+ or CD8+ T cells that produced IFN-γ or TNF-α was significantly greater than that in patients with short-term WT1-DTH positivity after two vaccinations. Moreover, after 12 vaccinations, the percentages of both circulating regulatory T cells and myeloid-derived suppressor cells were significantly lower in patients with long-term WT1-DTH-positive PDAC than in short-term WT1-DTH-positive patients.

Conclusions: Potent activation of WT1-specific immune responses through a combination chemoimmunotherapy regimen including the WT1-DC vaccine in patients with UR-PDAC may modulate the TME and enable conversion surgery, resulting in clinical benefits (Online supplemental file 1).

Trial registration number: jRCTc030190195.

Keywords: Dendritic; Immunotherapy; Tumor infiltrating lymphocyte - TIL; Tumor microenvironment - TME; Vaccine.

Figure 1. CE-CT and ¹⁸F-FDG PET/CT imaging. CE-CT and ¹⁸F-FDG PET/CT images of four patients with UR-PDAC (WT1-DC-01, WT1-DC-03, WT1-DC-05, and WT1-DC-09) are presented, demonstrating the changes observed before and after treatment. Yellow circles represent tumor lesions. CE-CT, contrast-enhanced CT; ¹⁸F-FDG PET/CT, positron emission tomography with 2-deoxy-2-(fluorine-18)-fluoro-D-glucose integrated with CT; UR-PDAC, unresectable pancreatic ductal adenocarcinoma; WT1-DC, Wilms’ tumor 1 peptide-pulsed dendritic cell.

Figure 2. K-M estimates of PFS and OS for patients with UR-PDAC. (A) PFS (left panel) or OS (right panel) of patients with UR-PDAC (n=9) who were treated with WT1-DC vaccines combined with chemotherapy. (B) PFS (left panel) or OS (right panel) for patients with LA-PDAC (n=5), patients with M-PDAC (n=3), and one patient with solitary lung recurrence (n=1). (C) PFS (left panel) or OS (right panel) in patients with UR-PDAC with long-term WT1-DTH positivity (n=4) or short-term WT1-DTH positivity (n=5). (D) PFS (left panel) and OS (right panel) of patients with LA-PDAC with long-term WT1-DTH positivity (n=3), patients with LA-PDAC with short-term WT1-DTH positivity (n=2), patients with M-PDAC with short-term WT1-DTH positivity (n=3), and one recurrent patient with long-term WT1-DTH positivity (n=1). K-M, Kaplan-Meier; LA-PDAC, locally advanced pancreatic ductal adenocarcinoma; M-PDAC, metastatic pancreatic ductal adenocarcinoma; OS, overall survival; PFS, progression-free survival; UR-PDAC, unresectable pancreatic ductal adenocarcinoma; WT1-DC, Wilms’ tumor 1 peptide-pulsed dendritic cell; WT1-DTH, Wilms’ tumor 1 peptide-specific delayed-type hypersensitivity.

Figure 3. Immunohistochemical analysis of the pancreatic TME in patients with PDAC who underwent conversion surgery. The mean cell counts/mm² tumor area for CD3+, CD4+, CD8+ (A) or PD-1+ (B) cells were analyzed in the pancreatic TME obtained from six surgically resected PDAC samples (four with LA-PDAC (WT1-DC-01, WT1-DC-03, WT1-DC-04, and WT1-DC-09) and two with M-PDAC (WT1-DC-02 and WT1-DC-05)). The mean percentage of Foxp3+/CD4+ cells (C) and the CD68+/CD163+ratio (D, E) in the pancreatic TME. Foxp3, forkhead box p3; IgG, immunoglobulin G; LA-PDAC, locally advanced pancreatic ductal adenocarcinoma; M-PDAC, metastatic pancreatic ductal adenocarcinoma; PD-1, programmed cell death protein-1; TME, tumor microenvironment; WT1-DC, Wilms’ tumor 1 peptide-pulsed dendritic cell.

Figure 4. IFN-γ-producing or TNF-α-producing WT1-specific CD8+ T cells restricted to HLA-A*24:02. (A) IFN-γ (left panel) or TNF-α (right panel) levels in one WT1-DC-07 patient before and after the 4 vaccines are shown. (B) The left panel shows HLA-A*24:02-restricted WT1-tetramer+IFN-γ+ CD8+ T cells among total CD8+ T cells (red bar), whereas the right panel shows HLA-A*24:02-restricted WT1-tetramer+TNF-α+ CD8+ T cells among total CD8+ T cells (red bar). (C) Changes in IFN-γ-producing (upper panel) or TNF-α-producing (lower panel) WT1-CTLs among total CD8+ T cells restricted to HLA-A*24:02 during vaccination (0, 2, 4, 6, 8, and 10 vaccinations) are shown. Three patients (WT1-DC-01, WT1-DC-07, and WT1-DC-09) presented long-term DTH (blue line), whereas four patients (WT1-DC-02, WT1-DC-04, WT1-DC-05, and WT1-DC-08) presented short-term WT1-DTH positivity (red line). (D) CD8+ T cells from before and after vaccination (0, 2, 4, 6, 8, and 10 vaccinations) were analyzed via HLA-A*24:02-restricted WT1-tetramers and IFN-γ (upper panel) or TNF-α (lower panel) production on stimulation with WT1 peptides in vitro. Patients with UR-PDAC with long-term WT1-DTH positivity (blue bar) or short-term WT1-DTH positivity (red bar) are shown. Each data point is presented in scatter plots with mean bar graphs. (E) PFS (upper panel) or OS (lower panel) for seven patients with UR-PDAC with HLA-A*24:02. These patients were divided into two groups: Those with a sustained high percentage of functional WT1-CTLs producing IFN-γ or TNF-α (>median percentage: WT1-DC-01, WT1-DC-07, and WT1-DC-09) (blue line) and those with relatively low percentages of these WT1-CTLs (≤median percentage: WT1-DC-02, WT1-DC-04, WT1-DC-05, and WT1-DC-08) (red line). *p<0.05, **p<0.01. HLA, human leukocyte antigen; OS, overall survival; PFS, progression-free survival; UR-PDAC, unresectable pancreatic ductal adenocarcinoma; WT1, Wilms’ tumor 1; WT1-CTLs, WT1-specific cytotoxic T lymphocytes; WT1-DC, WT1 peptide-pulsed dendritic cell; WT1-DTH, WT1 peptide-specific delayed-type hypersensitivity.