Monocytes and neutrophils as 'bad guys' for the outcome of interleukin-2 with and without histamine in metastatic renal cell carcinoma--results from a randomised phase II trial

Abstract

Histamine (HDC) inhibits formation and release of phagocyte-derived reactive oxygen species, and thereby protects natural killer (NK) and T cells against oxidative damage. Thus, the addition of histamine may potentially improve the efficacy of interleukin-2 (IL-2). We have explored this potential mechanism clinically in two randomised phase II trials in metastatic renal cell carcinoma (mRCC). In parallel with the clinical trial in Denmark (n=63), we obtained serial blood samples and tumour biopsies searching for a potential histamine effect in situ. At baseline and on-treatment weeks 3 and 8, we monitored the 'good guys' (i.e. NK and T cells) and 'bad guys' (i.e. monocytes/macrophages and neutrophils) simultaneously in blood (n=59) and tumour tissue (n=44). Patients with high number of monocytes and neutrophils in peripheral blood had very poor survival, with apparently no benefit from either IL-2 alone or IL-2/HDC treatment. Blood monocytes (r=-0.36, P=0.01) and neutrophils (r=-0.46, P=0.001) were negatively correlated with cytotoxicity, whereas blood NK cells were positively correlated with cytotoxicity (r=0.39, P=0.002). Treatment with IL-2 alone resulted in a significantly higher number of circulating monocytes (P=0.037) and intratumoral macrophages (P=0.005) compared with baseline. In contrast, IL-2/HDC resulted in an unchanged number of circulating monocytes and intratumoral macrophages, and in addition, a significantly increased number of intratumoral CD56(+) NK cells (P=0.008) and CD8(+) T cells (P=0.019) compared with baseline. The study provides evidence that circulating monocytes and neutrophils are powerful negative prognostic factors for IL-2-based immunotherapy and establishes a biological rationale for the potential use of histamine in conjunction with IL-2 in mRCC.

Figure 1

Peripheral blood monocytes, neutrophils and NK cells (CD16⁺56⁺) as prognostic factors for IL-2 based immunotherapy. High levels of monocytes and neutrophils were correlated with short survival.

Figure 2

Peripheral blood neutrophils, monocytes and NK cells as prognostic factors stratified for treatment assignment. Kaplan–Meier plots at baseline and week 8 in patients with metastatic renal cell carcinoma treated with interleukin-2 and HDC (—) or IL-2 alone (- - - -) concerning survival for (A) monocytes (B) neutrophils and (C) NK cells (CD16⁺56⁺). IL-2/HDC treatment resulted in long-term survival in patients with low numbers of monocytes and neutrophils and patients with high number of NK cells in the peripheral blood. Vertical and horizontal axes, the survival probability and months of follow-up, respectively. Tick marks (∣) indicate last date of follow-up. The hazard ratio (HR) refers to IL-2/HDC relative to IL-2 alone. CI, confidence interval.

Figure 3

Potential histamine effect on circulating monocytes during IL-2-based treatment, assessed in situ. Treatment with IL-2/HDC (white boxes) resulted in unchanged number of circulating monocytes in patients with complete response (CR), partial responses (PR) and stable disease (SD) compared with baseline values, whereas treatment with IL-2-alone (black boxes) did not prevent a significant increased number compared with baseline. All patient with progressive disease (PD) had significantly increased monocyte numbers at weeks 3 and 8 compared with baseline, irrespective of the treatment group. Vertical and horizontal axes, 10⁹ cells l⁻¹ and response to immunotherapy, respectively. The box plots represent the median (solid black line), the 25th and the 75th percentiles (boxed) and the 10th and the 90th percentiles (error bars). ‘O’ indicate outliers. ‘_^*’ indicate extremes.

Figure 4

Potential histamine effect on intratumoral NK, T-cells and macrophages during IL-2 based treatment, assessed in situ. Treatment with IL-2/HDC resulted in significantly higher numbers of intratumoral NK cells and CD8⁺T cells and unchanged numbers of intratumoral macrophages during treatment compared with baseline. IL-2 alone did not prevent a significantly higher number of intratumoral macrophages but did not result in increased intratumoral NK or T cells compared with baseline. The box plots represent the median (solid black line), the 25th and the 75th percentiles (boxed) and the 10th and the 90th percentiles (error bars). ‘O’ indicate outliers. ‘_^*’ indicate extremes. Vertical and horizontal axes, the median number of cells mm⁻² tumour tissue and therapy administered, respectively.

Figure 5

Correlation between in vitro cytotoxicity and NK cells, neutrophils and monocytes. A significantly positive correlation between blood CD16⁺CD56⁺ NK cells and cytotoxicity was observed. In contrast, there was a significantly negative correlation between blood neutrophils and cytotoxicity and also between blood monocytes and cytotoxicity.