Abstract 2700: AXL-positive cancer-associated fibroblasts enhance the metastatic potential of cancer cells and suppress tumor immunity in gastric cancer

Abstract Purpose: Cancer-associated fibroblasts (CAFs) are key components of the tumor microenvironment (TME). Numerous studies have demonstrated a significant correlation between increased CAFs accumulation and poor outcomes in gastric cancer (GC) patients. However, no targeting agents against CAFs have been clinically applied in cancer treatments. AXL, a receptor tyrosine kinase, has emerged as a promising target to suppress cancer cell metastasis, yet AXL inhibitors have not been approved due to limited efficacy in clinical trials. However, the role of AXL upregulated in CAFs remains unexplored. In this study, we investigated the effect of AXL-positive CAFs on GC progression and explore their potential as therapeutic targets to enhance GC treatment efficacy. Methods: We analyzed a single cell RNA-seq (scRNA-seq) dataset of human GC tissues to identify the cell types expressing AXL. Human CAFs and mouse gastric fibroblasts (MGF) expressing AXL were treated with AXL inhibitors, such as CCB3233 and 9im. Additionally, AXL knockdown (KD) was performed in CAFs and MGFs, which were subsequently co-cultured with human GC cell lines (SNU668 or MKN-1) or mouse GC cell lines (ACKPY4113) to assess the effects of AXL+ fibroblasts on GC progression. The impact of AXL-inhibited fibroblasts on T-lymphocytes or macrophages was also evaluated. Furthermore, transcriptome analysis was conducted on AXL KD and control fibroblasts. Results: The analysis using scRNA-seq dataset revealed that AXL expression was predominantly upregulated in CAFs compared to other cell types. High expression and activation of AXL in fibroblasts were confirmed across various cell lines, including GC cell lines, fibroblasts and immune cells. AXL inhibitors, such as CCB3233 and 9im, effectively reduced AXL phosphorylation in human CAFs and MGFs. Co-culture experiments demonstrated that AXL-inhibited or KD fibroblasts significantly reduced the migration or invasion of GC cells compared to control fibroblasts. Moreover, while control CAFs and MGFs suppressed the activation of T- lymphocytes or macrophages, AXL-inactivated or KD fibroblasts were unable to inhibit immune cell activation. Transcriptome analysis of AXL KD or control fibroblasts revealed reduced expression of chemokines such as CXCL9 and CXCL13, which may explain the observed experimental results. Conclusions: Our findings highlight the crucial role of AXL-positive fibroblasts in promoting the aggressive phenotype of GC cells and suppressing immune cell activity in the GC TME. Targeting AXL in CAFs holds significant promise as a novel therapeutic strategy for GC treatment. Citation Format: Tuyen Thanh Tran, Yulim Lee, In-Hye Ham, Hur Hoon. AXL-positive cancer-associated fibroblasts enhance the metastatic potential of cancer cells and suppress tumor immunity in gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2700.