A phase I/II study of a MUC1 peptide pulsed autologous dendritic cell vaccine as adjuvant therapy in patients with resected pancreatic and biliary tumors

Abstract

Pancreatic and biliary cancers are relatively resistant to chemotherapy and radiation and may therefore provide an opportunity for testing the potential of immunotherapy. MUC1 is an epithelial cell glycoprotein that is highly overexpressed and aberrantly glycosylated in many adenocarcinomas, including pancreatic tumors, providing a tumor specific antigen and target. We performed a Phase I/II clinical trial of a MUC1 peptide-loaded DC vaccine in 12 pancreatic and biliary cancer patients following resection of their primary tumors. The primary endpoints were vaccine toxicity and immunogenicity and the secondary endpoint was clinical outcome. The vaccine was well tolerated and no toxicity was observed. Three patients had pre-existing MUC1 antibody responses that remained stable post vaccination. MUC1-specific T cell responses were difficult to evaluate due to increases in activity of all CD8 and CD4 T cells following each vaccination. Prior to vaccination, patients entered onto this trial had a significantly higher percentage of FoxP3+CD4+ T cells compared to age matched healthy controls. The percentage of these cells also increased transiently following each injection, returning to baseline or below before the next injection. Vaccinated patients have been followed for over four years and four of the twelve patients are alive, all without evidence of recurrence. Study of the immune parameters in long-term survivors several years after vaccination may yield the sought after immune correlates of clinical responses that analysis of immune responses shortly after vaccination has not revealed.

Figure 1

MUC-1 vaccination transiently increases the percentages of functional CD4 and CD8 T cells in both survivors and non-survivors. Pre and post vaccination PBMCs were thawed, counted, and 10⁶ cells were aliquotted into 96 well plates. The cells were stimulated with PMA and ionomycin for 6 hours at 37°C in the presence of Golgistop and then stained for CD3, CD4, CD8 and intracellular IFN-γ and TNF-α. Percentage of T cells producing IFN is marked by closed circles and the percentage producing TNF is shown in the open squares. V1, V2 and V3 correspond to three consecutive vaccine administrations.

Figure 2

MUC-1 vaccination transiently increases the percentages of CD8 T cells expressing the effector molecules perforin and Granzyme B. Pre and post vaccination PBMCs were thawed, counted, and 10⁶ cells were aliquotted into 96 well plates. The cells were surface stained for CD3, CD4, and CD8, permeabilized and stained with anti-perforin and anti-granzyme B antibodies. Percentage of cells positive for Granzyme B is marked by a solid line and the percentage positive for perforin is marked by a dashed line. V1, V2 and V3 correspond to three consecutive vaccine administrations.

Figure 3

Pancreatic cancer patients have a higher percentage of regulatory T cells compared to age-matched healthy controls. PBMCs were thawed, counted and 10⁶ cells were aliquotted into 96 well plates. The cells were surface stained for CD3, CD4, CD8, permeabilized and then incubated with either an isotype control antibody (A) or with an anti-FoxP3 antibody (B). Representative dot plots are shown in A and B and a summary graph of all patients analyzed (normals n=9 and cancer patients n=7) is shown in C. Statistical analysis by a Wilcoxon ranked sum test resulted in a p-value of 0.012.

Figure 4

MUC-1 vaccination transiently increases the percentages of regulatory T cells in cancer patients. Pre and post vaccination PBMCs were thawed, counted, and 10⁶ cells were aliquotted into 96 well plates. The cells were surface stained for CD3, CD4, and CD8, permeabilized and then stained with an antibody against FoxP3. V1, V2 and V3 correspond to three consecutive vaccine administrations.