Reduction of the CD16(-)CD56bright NK cell subset precedes NK cell dysfunction in prostate cancer

Abstract

Background: Natural cytotoxicity, mediated by natural killer (NK) cells plays an important role in the inhibition and elimination of malignant tumor cells. To investigate the immunoregulatory role of NK cells and their potential as diagnostic markers, NK cell activity (NKA) was analyzed in prostate cancer (PCa) patients with particular focus on NK cell subset distribution.

Methods: Prospective data of NKA and NK cell subset distribution patterns were measured from 51 patients initially diagnosed with PCa and 54 healthy controls. NKA was represented by IFN-γ levels after stimulation of the peripheral blood with Promoca®. To determine the distribution of NK cell subsets, PBMCs were stained with fluorochrome-conjugated monoclonal antibodies. Then, CD16(+)CD56(dim) and CD16(-)CD56(bright) cells gated on CD56(+)CD3(-) cells were analyzed using a flow-cytometer.

Results: NKA and the proportion of CD56(bright) cells were significantly lower in PCa patients compared to controls (430.9 pg/ml vs. 975.2 pg/ml and 2.3% vs. 3.8%, respectively; p<0.001). Both tended to gradually decrease according to cancer stage progression (p for trend = 0.001). A significantly higher CD56(dim)-to-CD56(bright) cell ratio was observed in PCa patients (41.8 vs. 30.3; p<0.001) along with a gradual increase according to cancer stage progression (p for trend = 0.001), implying a significant reduction of CD56(bright) cells in relation to the alteration of CD56(dim) cells. The sensitivity and the specificity of NKA regarding PCa detection were 72% and 74%, respectively (best cut-off value at 530.9 pg/ml, AUC = 0.786).

Conclusions: Reduction of CD56(bright) cells may precede NK cell dysfunction, leading to impaired cytotoxicity against PCa cells. These observations may explain one of the mechanisms behind NK cell dysfunction observed in PCa microenvironment and lend support to the development of future cancer immunotherapeutic strategies.

Conflict of interest statement

Competing Interests: An author, SMK, is an employee at ATgen who helped with the experimental process. However, this does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.

Figures

Figure 1. (A) Representative flow cytometric data of the distribution of total NK cell population represented by CD3−CD56+ cells.

(B) Representative flow cytometric data of two major NK cell subsets detected in peripheral blood. Left, upper box: CD16+CD56dim NK cell subset. Right, lower box: CD16−CD56bright NK cell subset.

Figure 2. (A) Boxplot diagrams showing flow cytometric distribution results of total NK cell population % between controls and patients, grouped according to cancer stage.

No significant differences of total NK population were noted between patients and controls, and within stage groups. (B) Boxplot diagrams showing flow cytometric distribution results of CD56dim and CD56bright subset distributions within total NK cells between controls and patients, grouped according to cancer stage. *p<0.05, **p<0.01 in relation to controls.

Figure 3. Boxplot diagram comparing NK cell activity between controls and patients grouped according to cancer stage.

**p

Figure 4. (A) ROC curves comparing the performances of NK cell activity and CD56dim-to-CD56bright ratio measurements.

(AUC = Area Under the Curve). (B) ROC curves comparing the performances of NK cell activity measurement according to PSA grouped as; 4 to 10 ng/ml and greater than 10 ng/ml. (AUC = Area Under the Curve).