Abstract PO015: Intrinsic and extrinsic role of Axl in models of hepatocellular carcinoma
Authors: Breitenecker Kristina , Heiden Denise , Weber Gerhard , Mikulits Wolfgang 
Published: September 2022

The activation of the receptor tyrosine kinase Axl by binding to its high affinity ligand Gas6 is a major driver of tumor progression and metastasis in various cancer entities. In hepatocellular carcinoma (HCC), about 40% of patients show upregulation of Axl expression correlating with vascular invasion and poor survival of patients. In pre-malignant stages of HCC, Axl is essentially involved in developing hepatic fibrosis and cirrhosis by activation of hepatic stellate cells. Despite recent insights, the dichotomy of intrinsic and extrinsic Axl functions in HCC is poorly understood. Therefore, we aimed at analyzing cell-specific aspects of Axl in liver tumorigenesis using advanced mouse tumor models. To particularly address the tumor-cell specific role of Axl, we employed a transplantation-based mouse models. In order to investigate both the tumor-intrinsic and extrinsic impact of Axl, we employed an autochthonous model of diethylnitrosamine (DEN)-induced liver tumor formation in systemic Axl-deficient mice. We found that tumor-intrinsic Axl expression in HCC cells did not affect proliferation in vitro and in vivo, however increased the number of pulmonary metastatic colonies and reduced overall survival of mice. In line, Gas6/Axl signaling enhanced the invasive abilities of tumor cells in vitro as observed in 3-dimensional cell culture settings. The Axl-driven invasive phenotype of HCC cells associated with the upregulated expression of transcription factors Snai1/Snai2 and downregulation of E-Cadherin indicating changes in epithelial plasticity. Intriguingly, the tumor burden was enhanced in DEN-treated Axl−/− mice in the background of CCl4-induced inflammation by showing higher proliferative rates of malignant hepatocytes. Interestingly, these tumors were less infiltrated with cytotoxic CD8+ T cells and granzyme B+ cells. Additionally, we observed an increased infiltration of FoxP3+ regulatory T cells and F4/80+ macrophages as well as elevated expression of M2-macrophage-specific Mrc-1, Fizz-1 and TGF-β1 suggesting that Axl−/− tumors shift towards an immunosuppressive environment. Remarkably, PD-L1 levels did not vary between Axl+/+ and Axl−/− tumors, suggesting alternative immune escape mechanisms. In accordance, analysis of HCC patient datasets showed a positive correlation of Axl expression with cytotoxic immune gene signatures. From these data we conclude that tumor intrinsic Gas6/Axl signaling triggers the metastatic phenotype of HCC cells while Axl deficiency escapes immune surveillance and therefore promotes liver tumorigenesis. Together, our findings show that Axl plays a prominent role in HCC dissemination and highlight the importance of Axl expression in the tumor microenvironment.

Citation Format: Kristina Breitenecker, Denise Heiden, Gerhard Weber, Wolfgang Mikulits. Intrinsic and extrinsic role of Axl in models of hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference: Advances in the Pathogenesis and Molecular Therapies of Liver Cancer; 2022 May 5-8; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(17_Suppl):Abstract nr PO015.