Abstract 1080: Link between M1/M2 human macrophages and epithelial-mesenchymal status in head and neck cancer cell lines
Authors: Astorgues-Xerri Lucile , Evrard Diane , Martinet Matthieu , Raymond Eric , Faivre Sandrine , Tijeras-Raballand Annemilaï 
Published: August 2019

Abstract
Background. Head and neck carcinoma (H&N) is one of the leading causes of cancer deaths worldwide. Despite advances in diagnosis and treatment, the survival rates remain low mainly due to locoregional relapse, possibly triggered by the activation of epithelial-to-mesenchymal transition (EMT). Recently, several studies have demonstrated a positive link between macrophages, EMT and invasion in H&N cancer. The aim of this study is to analyze the interactions between human antitumoral M1/protumoral M2 macrophages and H&N human cancer cells with different EMT status, with the aim of developing new therapeutic approaches for H&N cancer patients.
Materials and Methods. M1 and M2 macrophages were obtained from THP-1 cell line (human monocyte) after 48h exposure to 25nM of PMA followed by 48h of recovery culture medium, and 72h exposure to 1ng/ml LPS + 20ng/ml IFNγ to obtain M1 phenotype or 20ng/ml IL4 + 20ng/ml IL13 to obtain M2 phenotype. Differentiation status was validated by immunofluorescence (IF) using CD14 for monocyte, CD68 for macrophage, CD80 for M1, and CD163 for M2. Eight H&N cell lines were characterized for their EMT status (E-cadherin/vimentin expression) by western blot. SQ20B (epithelial) and Hep2 (mesenchymal) cell lines were selected to study the effect of M1 and M2 conditioned medium (CM) on cell proliferation. In addition, we also analyzed the effects of CM from SQ20B and Hep2 on macrophages differentiation using IF.
Results. We confirmed the differentiation of monocytes into macrophages by a decrease of CD14 expression and an increase of CD68 expression, and the differentiation of macrophages into M1 and M2 by an increase of CD80 and CD163 expression, respectively. Among the 8 H&N cell lines, 3 cell lines showed an epithelial status (high E-cadherin expression), one a mesenchymal status (high vimentin expression), and 4 a mixed status. Based on these results, we exposed SQ20B (epithelial) and Hep2 (mesenchymal) to M1 or M2 CM. M1 CM strongly inhibited the proliferation of SQ20B cells, with moderate effect on Hep2 cells, whereas M2 CM displayed no effect on SQ20B cells and slightly increased the proliferation of Hep2 cells. Moreover, macrophages exposed to SQ20B CM displayed a M1 phenotype with an increased expression of CD80, whereas Hep2 CM induced a M2 phenotype with an increased expression of CD163.
Conclusions. In vitro, we showed that M1 and M2 macrophages displayed opposite effects on H&N cancer cells proliferation via their conditioned medium, M1 being anti-proliferative and M2 pro-proliferative. These effects were dependent on epithelial/mesenchymal status of cancer cells. In addition, we showed that factors secreted by epithelial vs mesenchymal cancer cells induced macrophages differentiation into M1 and M2, respectively. These results open up new perspectives on the role of M1/M2 macrophages in EMT-dependent H&N cancers and other tumor types such as colon, lung, and liver carcinoma.
Citation Format: Lucile Astorgues-Xerri, Diane Evrard, Matthieu Martinet, Eric Raymond, Sandrine Faivre, Annemilaï Tijeras-Raballand. Link between M1/M2 human macrophages and epithelial-mesenchymal status in head and neck cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1080.