STAT1 and STAT3 Exhibit a Crosstalk and Are Associated with Increased Inflammation in Hepatocellular Carcinoma.

Carolin Ploeger,Arianeb Mehrabi,Thomas Albrecht,Alphonse Charbel,Johannes Schreck,Stefan Pusch,Stephan Singer,Benjamin Goeppert,Peter Schirmacher,Junfang Ji,Stephanie Roessler,Thorben Huth,Kai Breuhahn,Angelika Fraas,Bruno Christian Köhler,Christoph Springfeld

doi:10.3390/cancers14051154

Abstract

Simple SummaryLiver cancer is the fourth-leading cause of cancer-related mortality worldwide and lacks effective therapies. Hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) are the two most common types of liver cancer and both are associated with underlying inflammatory diseases. Thereby, interleukin-6 (IL-6)-mediated STAT3 signaling is critically involved in early carcinogenesis and disease progression. Here, we assessed the interplay between STAT1 and STAT3 in IL-6 signaling in vitro and studied the activation of STAT1 and STAT3 in a cohort of 124 HCC and a cohort of 138 CCA patients by immunohistochemistry. We found that IL-6 induced STAT1 transcriptional activity upon STAT3 depletion, suggesting that HCC tumor cells may activate both STAT1 and STAT3 signaling under pro-inflammatory conditions. Furthermore, HCC patient tissues showed a strong positive correlation of STAT1 and STAT3 activation in distinct patient groups. These patients also exhibited a high degree of immune cell infiltration, suggesting that these tumors are immune “hot”.Liver cancers, which are mostly hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), are very aggressive tumors with poor prognosis. Therapeutic options with curative intent are largely limited to surgery and available systemic therapies show limited benefit. Signal transducer and activator of transcription 1 (STAT1) and 3 (STAT3) are key transcription factors activated by pro-inflammatory cytokines such as interferon-γ (IFN-γ) and interleukin-6 (IL-6). In this study, we combined in vitro cell culture experiments and immunohistochemical analyses of human HCC (N = 124) and CCA (N = 138) specimens. We observed that in the absence of STAT3, IL-6 induced the activation of STAT1 and its target genes suggesting that IL-6 derived from the tumor microenvironment may activate both STAT1 and STAT3 target genes in HCC tumor cells. In addition, STAT1 and STAT3 were highly activated in a subset of HCC, which exhibited a high degree of infiltrating CD8- and FOXP3-positive immune cells and PD-L1 expression. Our results demonstrate that STAT1 and STAT3 are expressed and activated in HCC and tumor infiltrating immune cells. In addition, HCC cases with high STAT1 and STAT3 expression also exhibited a high degree of immune cell infiltration, suggesting increased immunological tolerance.

Highlights

Despite growing efforts over the last two decades, liver cancer is the fourth-leading cause of cancer-related mortality worldwide [1,2]
Expression of IL-6/Signal transducer and activator of transcription 3 (STAT3) target genes Transthyretin (TTR) and Serine Peptidase Inhibitor Kazal Type 1 (SPINK1) showed that Signal transducer and activator of transcription 1 (STAT1) depleted cells treated with IFN-γ, a known inducer of the STAT1-signaling pathway, exhibited no significant induction of JAK/STAT3pathway activation at the transcriptional level in all three cell lines (Figure 1A)
As we found that IL-6 induces STAT1 transcriptional activity upon STAT3 depletion, we sought to analyze the expression of STAT1 and STAT3 regarding infiltrating immune cells in human liver cancer tissues

Summary

Introduction

Despite growing efforts over the last two decades, liver cancer is the fourth-leading cause of cancer-related mortality worldwide [1,2]. Hepatocellular carcinoma (HCC) is the most common type of liver cancer, followed by cholangiocarcinoma (CCA), which accounts for 10–15% of liver cancers [2,3] Both HCC and CCA can be caused by chronic hepatobiliary diseases such as chronic infection with hepatitis B (HBV) and C viruses (HCV) or other inflammatory liver diseases such as alcoholic and non-alcoholic steatohepatitis and primary sclerosing cholangitis [4], making HCC and CCA a paradigm for inflammation-induced carcinogenesis [5–7]. In an HCC mouse model of diethylnitrosamine (DEN)-induced tumorigenesis, ablation of IL-6 expression led to lower HCC incidence in male mice, whereas female mice did not show this effect due to estrogen-mediated inhibition of IL-6 secretion [13] Supporting these data, high serum IL-6 levels were found to be associated with rapid progression from chronic viral hepatitis to HCC in HBV- and HCV-positive patients [14,15]. It is crucial to decipher the role of IL-6 signaling in the interplay between tumor cells and the tumor-microenvironment for the development of novel treatment modalities

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