Immunogenicity for CD8+ and CD4+ T cells of 2 formulations of an incomplete freund's adjuvant for multipeptide melanoma vaccines

Abstract

An incomplete Freund's adjuvant (IFA) commonly used in experimental cancer vaccines has recently been reformulated. Oleic acid used in the surfactant was purified from a vegetable source (olives, IFA-VG) rather than an animal source (beef tallow, IFA-AN). To provide an insight into the adjuvant properties of the new formulation, we reviewed T-cell responses, by enzyme-linked immunospot assay, to multipeptide vaccines in 2 sequential clinical trials that spanned this transition of adjuvants. Analyses included 194 patients who received either IFA-AN or IFA-VG for all vaccines, and a subset of 93 patients best matched by study arm for vaccine antigens (12 melanoma peptides restricted by major histocompatibility complex class I, 12MP; plus a tetanus helper peptide, tet) administered with IFA but without granulocyte macrophage-colony stimulating factor. Inflammation was observed at vaccine sites clinically for almost all patients, even including ulceration in a subset with each IFA formulation. CD8 T-cell response rates to the 12 melanoma peptides were 53% [95% confidence interval (CI), 44%, 61%)] for IFA-AN and 46% [95% CI, 32%, 59%)] for IFA-VG. In the 93 patient subset, these rates were 73% [95% CI, 61%, 83%)] and 70% [95% CI, 47%, 87%)], respectively. CD4 T-cell responses to tetanus helper peptide were identified in 94% [95% CI, 86%, 98%)] and 96% [95% CI, 78%, 100%)], respectively. Responses to individual human leukocyte antigen (HLA)-A1, A2, and DR associated peptides were largely preserved, but reactivity trended lower for some HLA-A3 associated peptides. Despite the necessarily retrospective nature of the analysis and limitations of multiple comparisons, our summary data support the use of IFA-VG as an adjuvant with multipeptide vaccines in melanoma patients.

Figure 1. CD8 T cell responses to 12MP

Immune response rates, by study, to one or more of 12 MHC Class I restricted melanoma peptides were assayed by Direct ElIspot assay and are shown for patients vaccinated with peptides in Montanide ISA-51 AN (white bars) and in Montanide ISA-51 VG (black bars). The data in (A) represent 137 patients receiving Montanide ISA-51 AN (118 on Mel43, and 19 on Mel44) and 57 patients receiving Montanide ISA-51-VG on the Mel44 trial. The data in (B) represent the subset of 93 patients receiving 12MP + tetanus peptide, in Montanide ISA-51, without GM-CSF (Mel43 arms A+C and Mel44 arms A+B).

Figure 2. CD8 and CD4 T cell responses by HLA allele

The percentage of patients with a T cell response to any of the peptides restricted by HLA-A1, A2, or A3 are shown in panels A and B, where the data in (A) represent Mel43 patients receiving Montanide ISA-51 AN (vertical hatched bars), Mel44 trial receiving Montanide ISA-51 AN (white bars), and Mel44 patients receiving Montanide ISA-51 VG (black bars). The data in (B) represent the subset of 93 patients vaccinated with 12MP + tetanus peptide, without GM-CSF (Mel43 arms A+C and Mel44 arms A+B), for patients receiving Montanide ISA-51 AN (white bars) and Montanide ISA-51 VG (black bars). Note that the for the IFA-VG group, the HLA-A1 data represent just 6 patients. In (C), the percentage of patients with a CD4+ T cell response to the tetanus peptide, also detected by direct IFN-gamma ELIspot assay, is shown for the subset of patients vaccinated with 12MP + tetanus peptide, without GM-CSF (Mel43 arms A+C and Mel44 arms A+B), for patients receiving Montanide ISA-51 AN (white bars) and Montanide ISA-51 VG (black bars).

Figure 3. The absolute magnitude of T cell responses

The percentage of CD4⁺ or CD8⁺ T cells producing IFN-gamma in response to each of the 12 peptides in 12MP (abbreviated with 3–4 letter abbreviations), or to the tetanus helper peptide (TET), are shown. For each peptide, the maximum value across multiple time points was used for each patient. These are shown for the subset of patients vaccinated with the 12MP + tetanus peptides, without GM-CSF, with IFA-AN (A) or IFA-VG (B). In this box plot format, the box represents the 25^th to 75^th percentile values, the horizontal line inside the box is the median, the symbol in the box is the mean, and the tails of the stems represent the minimum and maximum values.