Inhibition of EZH2 methyltransferase decreases immunoediting of mesothelioma cells by autologous macrophages through a PD-1-dependent mechanism

Our plain-English summary of Inhibition of EZH2 methyltransferase decreases immunoediting of mesothelioma cells by autologous macrophages through a PD-1-dependent mechanism, by M. Hamaidia, H. Gazon, C. Hoyos, et al (JCI Insight, 2019).

The study

• Authors: M. Hamaidia, H. Gazon, C. Hoyos, G.B. Hoffmann, R. Louis, B. Duysinx, L. Willems
• Journal: JCI Insight
• Published: 2019;4(18)
• Cited by: 49 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

The roles of macrophages in orchestrating innate immunity through phagocytosis and T lymphocyte activation have been extensively investigated. Much less understood is the unexpected role of macrophages in direct tumor regression. Tumoricidal macrophages can indeed manifest cancer immunoediting activity in the absence of adaptive immunity. We investigated direct macrophage cytotoxicity in malignant pleural mesothelioma, a lethal cancer that develops from mesothelial cells of the pleural cavity after occupational asbestos exposure. In particular, we analyzed the cytotoxic activity of mouse RAW264.7 macrophages upon cell-cell contact with autologous AB1/AB12 mesothelioma cells. We show that macrophages killed mesothelioma cells by oxeiptosis via a mechanism involving enhancer of zeste homolog 2 (EZH2), a histone H3 lysine 27-specific (H3K27-specific) methyltransferase of the polycomb repressive complex 2 (PRC2). A selective inhibitor of EZH2 indeed impaired RAW264.7-directed cytotoxicity and concomitantly stimulated the PD-1 immune checkpoint. In the immunocompetent BALB/c model, RAW264.7 macrophages pretreated with the EZH2 inhibitor failed to control tumor growth of AB1 and AB12 mesothelioma cells. Blockade of PD-1 engagement restored macrophage-dependent antitumor activity. We conclude that macrophages can be directly cytotoxic for mesothelioma cells independent of phagocytosis. Inhibition of the PRC2 EZH2 methyltransferase reduces this activity because of PD-1 overexpression. Combination of PD-1 blockade and EZH2 inhibition restores macrophage cytotoxicity.

In plain English

This laboratory and animal study examined whether mouse macrophages can kill mesothelioma cells directly. The authors used the RAW264.7 mouse macrophage line together with AB1 (sarcomatoid) and AB12 (biphasic) murine mesothelioma cell lines, which had been generated after intraperitoneal injection of BALB/c mice with crocidolite asbestos, and syngeneic BALB/c mice. Culture supernatant from macrophages caused apoptosis of AB1 cells; in the presence of LPS, blocking reactive oxygen species with apocynin or decomposing peroxynitrites with FeTTPS reduced this killing, while the NOS inhibitor L-NMMA had only a marginal effect. In direct co-culture, AB1 cells were killed without being phagocytosed, and killing fell when PGAM5 was silenced. The EZH2 inhibitor EPZ5687 (10 microM) reduced H3K27me3 and reduced both direct and supernatant-mediated killing. In mice, co-implanted macrophages almost completely abrogated AB1 and AB12 tumour development, but macrophages pretreated with the EZH2 inhibitor did not. The inhibitor increased PD-1 expression on macrophages, and anti-PD-1 antibody restored killing in culture and tumour control in mice. The authors concluded that EZH2 inhibition reduces macrophage cytotoxicity through PD-1 overexpression.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study: in vitro macrophage–mesothelioma co-culture experiments plus syngeneic immunocompetent mouse tumour experiments
• Who was studied: Mouse RAW264.7 macrophages and AB1 (sarcomatoid) and AB12 (biphasic) murine mesothelioma cell lines; male wild-type BALB/c mice aged 5-7 weeks, at least 6 mice per experimental condition
• Exposure: Mesothelioma cell lines originally derived from crocidolite asbestos-injected mice; experimental treatments were LPS, the EZH2 inhibitor EPZ5687 (10 microM), apocynin, L-NMMA, FeTTPS, and anti-PD-1 or anti-PD-L1 antibodies
• Main results:
  • Macrophage-conditioned supernatant caused apoptosis of AB1 cells; in the presence of LPS, inhibiting ROS with apocynin reduced this cytotoxicity, as did decomposition of peroxynitrites with FeTTPS
  • The NOS inhibitor L-NMMA reduced nitrites but affected supernatant cytotoxicity only marginally
  • In direct co-culture, AB1 cells were killed by LPS-stimulated macrophages without being phagocytosed
  • Silencing PGAM5 in AB1 cells significantly reduced macrophage-induced apoptosis of those cells
  • The EZH2 inhibitor EPZ5687 (10 microM) applied to RAW264.7 macrophages significantly inhibited both supernatant-mediated and direct killing of AB1 cells
  • In BALB/c mice, co-implanted RAW264.7 macrophages almost completely abrogated tumour development of AB1 or AB12 cells, but macrophages pretreated with the EZH2 inhibitor did not
  • The EZH2 inhibitor increased the proportion of PD-1-expressing RAW264.7 macrophages, with or without LPS
  • Anti-PD-1 antibody, but not isotype control, restored apoptosis of AB1 cells in culture
  • In mice, macrophages pretreated with the EZH2 inhibitor plus anti-PD-1 antibody controlled AB1 and AB12 tumour growth
  • Pretreatment of macrophages with LPS abolished their ability to control tumour growth of AB12 biphasic cells
• The authors' conclusion: The authors concluded that macrophages can be directly cytotoxic to mesothelioma cells without prior phagocytosis, that EZH2 inhibition reduces this activity by stimulating PD-1 expression, and that this supports combining EZH2 and immune checkpoint inhibitors in trials
• Limitations the authors state:
  • The authors state they could not determine whether peroxynitrites are involved in direct cytotoxicity because FeTTPS was too toxic to the macrophages
  • The authors describe findings in human macrophages pulsed with patient pleural effusion, and in spheroid cultures, as preliminary and not shown

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Mesothelioma, Macrophage, Phagocytosis, Pleural Mesothelioma, Mesothelial Cells, Pleural Cavity, Asbestos Exposure, EZH2 Inhibitor.

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