Consumption of soy isoflavone enriched bread in men with prostate cancer is associated with reduced proinflammatory cytokines and immunosuppressive cells

Abstract

We hypothesized that soy phytochemicals may have immunomodulatory properties that may affect prostate carcinogenesis and progression. A randomized, phase II trial was conducted in 32 patients with prostate cancer with asymptomatic biochemical recurrence but no measurable disease on standard staging studies. Patients were randomized to two slices of soy bread (34 mg isoflavones/slice) or soy bread containing almond powder daily as a source of β-glucosidase. Flow cytometry and bioplex assays were used to measure cytokines or immune cell phenotype in blood at baseline (day 0) and following intervention (day 56). Adequate blood samples were available at enrollment and day 56 and evaluated. Multiple plasma cytokines and chemokines were significantly decreased on day 56 versus baseline. Subgroup analysis indicated reduced TH1 (P = 0.028) and myeloid-derived suppressor cell (MDSC)-associated cytokines (P = 0.035). TH2 and TH17 cytokines were not significantly altered. Phenotypic analysis revealed no change in CD8(+) or CD4(+) T cells but showed increased CD56(+) natural killer (NK) cells (P = 0.038). The percentage of cells with a T regulatory cell phenotype (CD4(+)CD25(+)FoxP3(+)) was significantly decreased after 56 days of soy bread (P = 0.0136). Significantly decreased monocytic (CD33(+)HLADR(neg)CD14(+)) MDSC were observed in patients consuming soy bread (P = 0.0056). These data suggest that soy bread modulates systemic soluble and cellular biomarkers consistent with limiting inflammation and suppression of MDSCs. Additional studies to elucidate impact on the carcinogenic process or as a complement to immune-based therapy are required.

Figure 1. Soy bread consumption is associated with modulation of cytokines in plasma from patients with prostate cancer

Plasma from patients obtained on Day 0 and Day 56 were analyzed via bioplex analysis for the expression of 54 soluble cytokine and chemokine mediators. (A) Data are presented as the mean fold change in expression for each factor across the 23 patients who were evaluable on study with adequate samples for analysis. Significantly reduced levels of both canonical pro-inflammatory cytokines (IL-1β, TGF-β, TNF-α, IFN-γ; shown in blue) and MDSC-associated cytokines (G-CSF, IL-6, GM-CSF, M-CSF, IL-10, IL-13, VEGF); shown in red) were observed following dietary soy intervention. (B) Representative raw data for plasma IL-6 (in pg/mL) from patients prior to (Day 0) and following soy intervention (Day 56).

Figure 1. Soy bread consumption is associated with modulation of cytokines in plasma from patients with prostate cancer

Plasma from patients obtained on Day 0 and Day 56 were analyzed via bioplex analysis for the expression of 54 soluble cytokine and chemokine mediators. (A) Data are presented as the mean fold change in expression for each factor across the 23 patients who were evaluable on study with adequate samples for analysis. Significantly reduced levels of both canonical pro-inflammatory cytokines (IL-1β, TGF-β, TNF-α, IFN-γ; shown in blue) and MDSC-associated cytokines (G-CSF, IL-6, GM-CSF, M-CSF, IL-10, IL-13, VEGF); shown in red) were observed following dietary soy intervention. (B) Representative raw data for plasma IL-6 (in pg/mL) from patients prior to (Day 0) and following soy intervention (Day 56).

Figure 2. Soy bread consumption and phenotypic changes in circulating NK and T cell populations in patients with prostate cancer

(A) Percentages of circulating CD4⁺, CD8⁺ and CD56⁺ lymphocytes or (B) CD4⁺CD25⁺FoxP3⁺ T regulatory cells were evaluated at Day 0 and Day 56 by flow cytometry. Data are presented as the mean % positive cells for each phenotypic subset across the 25 patients who were evaluable on study. (C) The ratio of CD4⁺CD25⁺FoxP3⁺ T regulatory cells to CD8⁺ cells was significantly reduced at Day 56 after soy bread intervention as compared to Day 0. Data are presented as the mean ratio of T reg:CD8⁺ cells across the 25 evaluable patients.

Figure 3. Soy bread consumption and phenotypic changes in circulating MDSC populations in patients with prostate cancer

Peripheral blood mononuclear cells (PBMCs) were obtained from the 25 evaluable cancer patients on Day 0 and Day 56, and stained for MDSC using flurochrome-labeled antibodies targeting CD33, HLADR and CD14 or the appropriate isotype controls. MDSC levels were evaluated by flow cytometry based on a minimum of 20,000 live events and presented as the percentage of total cells. Data were analyzed and presented as the total percentage of (A) CD33⁺HLA-DRnegCD14⁺ monocytic MDSC. Each symbol represents data from an individual patient while the horizontal lines indicate the mean within each group. (B) Representative flow cytometric dot plots validating effective depletion of MDSC from patient peripheral blood samples. Depletion of CD33⁺ cells results in enhanced proliferation following a 72 hour stimulation with CD3/CD28 beads in CFSE-labeled (C) CD4⁺ and (D) CD8⁺ T cells from peripheral blood of patients. Data are presented as the percentage of T cell proliferation from blood taken on Day 0 from five representative patients on study. Cryopreserved cells were thawed, CFSE-labeled, stimulated, stained for CD4 or CD8 markers and analyzed by flow cytometry.