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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Research Article Oncology

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 1

Contact with NB cells and hypoxia synergistically induce CCL20 in human monocytes.

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Contact with NB cells and hypoxia synergistically induce CCL20 in human ...
(A) Primary monocytes were cocultured with CHLA-255 NB cells (1:1 ratio) for 48 hours in normoxic (20% O2) or hypoxic (1% O2) conditions, and supernatants were placed in bottom chambers of dual-chambers plates with 5-μM pore membranes with or without addition of the indicated neutralizing antibodies or their isotype control. NKT cells were placed in the upper chambers and allowed to migrate for 3 hours, and the rate of NKT cell migration was quantified by FACS. Results are mean ± SD from 3 experiments in triplicate. (B) Monocytes were cocultured with or without CHLA-255 NB cells for 36 hours in normoxic or hypoxic conditions followed by mRNA isolation and quantitative real-time PCR analysis of 11 chemokine genes known to attract human NKT cells. Data are from a representative of 3 experiments in triplicate. (C) Monocytes and CHLA-255 NB cells were cultured alone or combined in hypoxic or normoxic conditions for 48 hours. CCL20 concentration was quantified in the supernatants using ELISA. Data are mean ± SD from experiments with monocytes from 6 donors in duplicate. (D) Cells were cultured as in C and analyzed for intracellular CCL20 accumulation in CD14+ monocytes and CD14– NB cells. Regions were set using corresponding isotype controls. Data are from a representative of 3 experiments in duplicate. (E) Tumor-infiltrating leukocytes (TILs) were isolated from a cell suspension of freshly resected primary NB by gradient centrifugation and cultured with GolgiStop for 4 hours followed by FACS. After gating on CD45+ events, CCL20 accumulation was examined in CD14+ TAMs and compared with the corresponding isotype control. Data are from a representative of 3 experiments. **P < 0.01; ***P < 0.001.