Immunol Lett. 2016 Aug;176:1-7.

Indomethacin sensitizes resistant transformed cells to macrophage cytotoxicity.

Totary-Jain H1Sionov RV2Gallily R3.

1Department of Molecular Pharmacology and Physiology, Morsani College of Medicine, University of South Florida, Tampa, FL, USA. Electronic address: totaryjainh@health.usf.edu.

2Department of Developmental Biology and Cancer Research, IMRIC, The Hadassah Medical School-Hebrew University of Jerusalem, Israel. Electronic address: ronit.sionov@mail.huji.ac.il.

3The Lautenberg Center for General and Tumor Immunology, The Hadassah Medical School-Hebrew University of Jerusalem, Israel. Electronic address: ruthg@ekmd.huji.ac.il.

 

Abstract

Activated macrophages are well known to exhibit anti-tumor properties. However, certain cell types show intrinsic resistance. Searching for a mechanism that could explain this phenomenon, we observed that the supernatant of resistant cells could confer resistance to otherwise sensitive tumor cells, suggesting the presence of a secreted suppressor factor. The effect was abolished upon dialysis, indicating that the suppressor factor has a low molecular weight. Further studies showed that prostaglandin E2 (PGE2) is secreted by the resistant tumor cells and that inhibition of PGE2 production by indomethacin, a cyclooxygenase (COX) inhibitor, eliminated the macrophage suppression factor from the supernatant, and sensitized the resistant tumor cells to macrophage cytotoxicity. This study emphasizes the important role of tumor-secreted PGE2 in escaping macrophage surveillance and justifies the use of COX inhibitors as an adjuvant for improving tumor immunotherapy.

Keywords: Indomethacin; Macrophage cytotoxicity; PGE2; TNFα; Tumor cell resistance

 

Supplementary

Macrophages are innate immune cells differentiated from the blood white cells monocytes and reside in the tissues throughout the body where they play important roles in eliminating foreign invaders and in stimulating adaptive immune responses. In addition, they are involved in tissue repair, matrix remodeling and angiogenesis. In the context of cancer, macrophages have been attributed both anti- and pro-tumor functions, which during the years have been recognized to be effectuated by different subpopulations of macrophages. This observation has led to the terminology of pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages1, which represent extremes of a whole spectrum of differential activation statuses. M1 macrophages have been considered to have anti-tumor functions, while M2 macrophages being pro-tumor. A major aim in cancer immunology is to switch the macrophage phenotype from being pro-tumor to possessing anti-tumor properties. In the study conducted by Totary-Jain et al.2, we observed that while some tumor cells are killed by activated macrophages, others are seemingly resistant. A major question was whether the resistant tumor cells were intrinsic resistant to the macrophage-mediated anti-tumor mechanisms, or somehow they escape killing by preventing the proper activation of macrophages. The data showed that the resistant tumor cells do undergo cell death when exposed to the supernatant of activated macrophages or purified TNFα, indicating that the seemingly resistant tumor cells are sensitive to macrophage-mediated anti-tumor activities. However, when the macrophages were incubated in the conditioned medium from the resistant tumor cells, macrophage activation, as measured by TNFα production, was prevented. In a search for the component in the conditioned medium that inhibits macrophage activation, we found that the small bioactive molecule prostaglandin E2 (PGE2), highly produced by the tumor cells (1-2 nM in the conditioned medium), was the component that prevented macrophage activation. Dialysis of the conditioned medium restored macrophage activation and even enhanced TNFα secretion upon activation by lipopolysaccharide (LPS), and exogenously added PGE2 prevented macrophage activation. Intriguingly, the PGE2 effect on macrophage activation was dose-dependent where concentrations above 1 nM inhibited, while concentrations below 10 pM even enhanced TNFα secretion (Fig. 1A). The latter may explain the enhanced TNFα production observed when dialyzed conditioned medium was used where the remaining PGE2 concentration is very low. Of clinical importance is the observation that the non-steroid anti-inflammatory drug (NSAID) indomethacin could overcome the tumor-mediated suppression of macrophage activation and sensitizes the tumor cells to macrophage-mediated cytotoxicity (Fig. 1B). Our data suggest a potential use of indomethacin as an adjuvant agent in cancer immunotherapy (Fig. 2).

 fig1

Fig 1. A. Dose-dependent effect of PGE2 on TNFα-secretion by LPS-activated macrophages. At concentrations above 1 nM, PGE2 inhibited, while concentrations below 10 pM, PGE2 even enhanced TNFα secretion. B. Indomethacin sensitizes tumor cells to macrophage-mediated cytotoxicity. Resistance to macrophage cytotoxicity can be overcome by the NSAID drug indomethacin.

fig2

Fig 2. Reactivation of macrophage tumor cytotoxicity by Indomethacin. Classical activation of macrophages by LPS leads to TNFα secretion and tumor cell killing. Secretion of PGE2 by tumor cells leads to immunosuppression, a condition where macrophage activation by LPS is inhibited, and consequently there is no killing of tumor cells. However, the application of indomethacin, which prevents PGE2 production, enables macrophage activation and restores macrophage cytotoxicity towards tumor cells.

References

1. Martinez FOGordon S. The M1 and M2 paradigm of macrophage activation:  Time for reassessment. F1000Prime Rep. 2014 Mar 3;6:13.

2.  Totary-Jain H, Sionov RV, Gallily R. Indomethacin sensitizes resistant transformed cells to macrophage cytotoxicity. Immunol Lett. 2016 Aug;176:1-7.

 

 

 

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