Elevated myeloid-derived suppressor cells in pancreatic, esophageal and gastric cancer are an independent prognostic factor and are associated with significant elevation of the Th2 cytokine interleukin-13

Abstract

We undertook a comprehensive analysis of circulating myeloid-derived suppressor cells (MDSCs) and T regulatory cells (Tregs) in pancreatic, esophageal and gastric cancer patients and investigated whether MDSCs are an independent prognostic factor for survival. We evaluated a series of plasma cytokines and in particular re-evaluated the Th2 cytokine interleukin-13 (IL-13). Peripheral blood was collected from 131 cancer patients (46 pancreatic, 60 esophageal and 25 gastric) and 54 healthy controls. PBMC were harvested with subsequent flow cytometric analysis of MDSC (HLADR(-) Lin1(low/-) CD33(+) CD11b(+)) and Treg (CD4(+) CD25(+) CD127(low/-) FoxP3(+)) percentages. Plasma IL-2, IL-4, IL-5, IL-6, IL-10, IL-12 (p70), IL-13, IL-17, G-CSF, IFN-γ, TNF-α and VEGF levels were analyzed by the Bio-Plex cytokine assay. Plasma arginase I levels were analyzed by ELISA. MDSCs and Tregs were statistically significantly elevated in pancreatic, esophageal and gastric cancer compared with controls, and MDSC numbers correlated with Treg levels. Increasing MDSC percentage was associated with increased risk of death, and in a multivariate analysis, MDSC level was an independent prognostic factor for survival. A unit increase in MDSC percentage was associated with a 22% increased risk of death (hazard ratio 1.22, 95% confidence interval 1.06-1.41). Arginase I levels were also statistically significantly elevated in upper gastrointestinal cancer patients compared with controls. There was Th2 skewing for cytokine production in all three diseases, and importantly there were significant elevations of the pivotal Th2 cytokine interleukin-13, an increase that correlated with MDSC levels.

Fig. 1

MDSC and Treg FACS and scatter plots of cancer patients and healthy controls. a live HLADR⁻ Lin1^low/− CD33⁺ CD11b⁺ MDSC FACS gating of PBMC of a pancreatic cancer patient (above) and a healthy control (below). P1, the ViViD low population represents viable cells with reduced amine staining, P2 represents the HLADR negative, lineage low/negative population and UR3, live HLADR⁻ Lin1^low/− CD33⁺ CD11b⁺ cells, calculated as a percentage of live cells in P1. b live CD4⁺ CD25⁺ CD127^low/− FoxP3⁺ Treg FACS gating of PBMC of the same pancreatic cancer patient (above) and healthy control (below). P1, the lymphocyte population. P2, viable lymphocytes. P4, CD4⁺ population. P5, CD25⁺ CD127^low/−. Histograms represent the FoxP3⁺ Treg population (FITC⁺, right) and the FoxP3⁻ non-Treg population. Tregs are calculated as a percentage of live CD4⁺ lymphocytes. c scatter plot of MDSC % in controls and cancer patients. Bar denotes median in each group. *, statistically significant difference from healthy controls. d scatter plot of Treg % in controls and cancer patients

Fig. 2

a Overall survival for MDSC % above and within normal limits. There was a significant difference between cancer patients’ survival in those with a normal MDSC% (as defined by the range seen in healthy controls, ≤2.00%) and those with an elevated MDSC% (>2.00%): ( = 16.1, P < 0.001). b Log (hazard) plot with increasing MDSC %. A unit increase in MDSC percentage was associated with an increased risk of death (hazard ratio = 1.22, 95% CI: 1.06, 1.41)