Abstract 2792: Galectin 9 promotes invasion and angiogenesis in vitro and associates with an immune-suppressive microenvironment in gastric cancer patients
Authors: Hill Charlotte Nicole , Arata Maximiliano Arce , Cabrolier Camille , Luque Noymar , Gonzalez Pamela , Maita Gabriela , Vega-Letter Ana Maria , Crawford Patricia Luz , Garrido Marcelo , Owen Gareth I. 
Published: July 2021

The development of immune checkpoint inhibitors is revolutionizing gastric cancer (GC) therapy. However, the presence of an immune-suppressive tumor microenvironment and redundancy between inhibitory immune checkpoints may be responsible for why a high percentage of patients without clinical benefit. An emerging checkpoint target is Tim-3, a membrane protein that enhances immunosuppression upon binding to its ligand Galectin-9 (Gal-9). Gal-9 is overexpressed in gastrointestinal cancers and expressed in both cancer and stromal cells such as the tumor endothelium. However the role of the Gal-9/Tim-3 pathway in the GC tumor microenvironment is still undeciphered, we hypothesize that this pathway mediates immune escape though enhancement of a immunosuppressive microenvironment. We performed a bioinformatic analysis of stomach adenocarcinoma (STAD) from the TCGA database (TIMER©, Cibersort and Xcell were used). To evaluate if Gal-9 had an impact on cancer cell phenotype we transfected Gal-9 into the gastric cancer cell line AGS and the gastric epithelium cell line GES-1. To assess the potential role in CG tumor microenvironment, Gal-9 was transfected or recombinant Gal-9 (rGal-9) was administrated to Human Umbilical Vein Endothelial Cells (HUVECs). Gal-9 biological effects were assessed by migration, invasion and Matrigel tube formation assay. Western blotting assessed Tim-3 expression and PDL-1 expression. Furthermore, we used lactose to block Gal-9 binding to Tim-3 to determine if the biological effects required receptor engagement. Results from TCGA analysis demonstrated that both Gal-9 and Tim-3 are upregulated in gastric tumors. Tim-3 mRNA is significantly increased in invasive adenocarcinomas. A positive correlation was observed between Gal-9, PDL-1 and Tim-3, which was consistent with in vitro experiments, where increased expression of Gal-9 resulted in increased PDL-1 and Tim-3. In addition, Gal-9 increased cancer cell invasion and promoted HUVEC migration and tubular like structure formation in vitro. Further bioinformatic analysis revealed strong positive correlation with CD8+ T cell, Treg and macrophage/monocyte gene signatures. Tim-3 and Gal-9 were more strongly associated with these gene signatures than PD1 and PDL-1. Interestingly, Gal-9 was associated with T cell dysfunction and Treg markers, suggesting that in GC Gal-9 could promote T cell dysfunction and Treg expansion. Our results suggest an association between the Gal-9/Tim-3 pathway with a more aggressive and increased immune suppressive microenvironment and GC.

Funding: Fondecyt 118024, CONICYT-FONDAP 15130011, IMIIP09/16F, FONDECYT 1180173

Citation Format: Charlotte Nicole Hill, Maximiliano Arce Arata, Camille Cabrolier, Noymar Luque, Pamela Gonzalez, Gabriela Maita, Ana Maria Vega-Letter, Patricia Luz Crawford, Marcelo Garrido, Gareth I. Owen. Galectin 9 promotes invasion and angiogenesis in vitro and associates with an immune-suppressive microenvironment in gastric cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2792.