IL-15 protects NKT cells from inhibition by tumor-associated macrophages and enhances antimetastatic activity
Daofeng Liu, … , Gianpietro Dotti, Leonid S. Metelitsa
Daofeng Liu, … , Gianpietro Dotti, Leonid S. Metelitsa
Published May 8, 2012
Citation Information: J Clin Invest. 2012;122(6):2221-2233. https://doi.org/10.1172/JCI59535.
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IL-15 protects NKT cells from inhibition by tumor-associated macrophages and enhances antimetastatic activity

Abstract

Vα24-invariant NKT cells inhibit tumor growth by targeting tumor-associated macrophages (TAMs). Tumor progression therefore requires that TAMs evade NKT cell activity through yet-unknown mechanisms. Here we report that a subset of cells in neuroblastoma (NB) cell lines and primary tumors expresses membrane-bound TNF-α (mbTNF-α). These proinflammatory tumor cells induced production of the chemokine CCL20 from TAMs via activation of the NF-κB signaling pathway, an effect that was amplified in hypoxia. Flow cytometry analyses of human primary NB tumors revealed selective accumulation of CCL20 in TAMs. Neutralization of the chemokine inhibited in vitro migration of NKT cells toward tumor-conditioned hypoxic monocytes and localization of NKT cells to NB grafts in mice. We also found that hypoxia impaired NKT cell viability and function. Thus, CCL20-producing TAMs served as a hypoxic trap for tumor-infiltrating NKT cells. IL-15 protected antigen-activated NKT cells from hypoxia, and transgenic expression of IL-15 in adoptively transferred NKT cells dramatically enhanced their antimetastatic activity in mice. Thus, tumor-induced chemokine production in hypoxic TAMs and consequent chemoattraction and inhibition of NKT cells represents a mechanism of immune escape that can be reversed by adoptive immunotherapy with IL-15–transduced NKT cells.

Authors

Daofeng Liu, Liping Song, Jie Wei, Amy N. Courtney, Xiuhua Gao, Ekaterina Marinova, Linjie Guo, Andras Heczey, Shahab Asgharzadeh, Eugene Kim, Gianpietro Dotti, Leonid S. Metelitsa

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Figure 3

mbTNF-α on NB cells induces NF-κB activation in monocytes.

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mbTNF-α on NB cells induces NF-κB activation in monocytes. (A) Cultured ...
(A) Cultured NB cells were suspended using 2% EDTA without trypsin and analyzed by FACS for cell surface expression of mbTNF-α in 2 representative NB cell lines (shaded, isotype control; open, anti–mbTNF-α). (B) Cell suspensions from freshly resected primary NB tumors were stained for surface markers. mbTNF-α expression on NB cells was analyzed after gating on CD56hiCD45– events. (C) NB cells were preincubated with 50 ng/ml anti-human TNF-α or isotype control mAb for 1 hour before addition of monocytes; NB and monocytes alone were used as controls. CCL20 concentration in the culture supernatant was determined by ELISA after 36 hours. Results are mean ± SD from 3 experiments in triplicate. ***P < 0.001, 1-way ANOVA. (D) Monocytes were cultured alone in nonadherent plates or on top of NB cell monolayer, with addition of anti–TNF-α or isotype control mAb, in normoxia or hypoxia for 16 hours followed by monocyte detachment and Western blotting for phospho-IκBα using β-actin as a loading control. Data are from a representative of 3 experiments. (E and F) The same experiment as in D was followed by intracellular staining for (E) IκBα or (F) phospho-p65 in monocytes after gating out NB cells as CD56hi events. (G) Kinetics of phospho-p65 expression in monocytes upon coculture with NB cells in normoxic and hypoxic conditions. Results are mean ± SD from 2 experiments in triplicate. **P < 0.01; ***P < 0.001.