Saioa Goñi

The Epidermal Growth Factor Receptor: A Link Between Inflammation and Liver Cancer

Epidemiological studies have established that many tumours occur in association with persistent inflammation. One clear example of inflammation-related cancer is hepatocellular carcinoma (HCC). HCC slowly unfolds on a background of chronic inflammation triggered by exposure to infectious agents (hepatotropic viruses), toxic compounds (ethanol), or metabolic impairment. The molecular links that connect inflammation and cancer are not completely known, but evidence gathered over the past few years is beginning to define the precise mechanisms. A central role for cytokines such as interleukin-6 (IL-6) and IL-1 (α and β) in liver cancer has been established in experimental models. Besides these inflammatory mediators, mounting evidence points to the dysregulation of specific growth and survival-related pathways in HCC development. Among them is the pathway governed by the epidermal growth factor receptor (EGFR), which can be bound and activated by a broad family of ligands. Of special relevance is the fact that the EGFR engages in extensive crosstalk with other signaling pathways, serving as a “signaling hub” for an increasing list of growth factors, cytokines, and inflammatory mediators. In this review, we summarize the most recent evidences supporting a role for the EGFR system in inflammation-related cell signaling, with special emphasis in liver inflammation and HCC. The molecular dissection of the pathways connecting the inflammatory reaction and neoplasia will facilitate the development of novel and more effective antitumor strategies.

The epidermal growth factor receptor ligand amphiregulin participates in the development of mouse liver fibrosis.

The hepatic wound‐healing response to chronic noxious stimuli may lead to liver fibrosis, a condition characterized by excessive deposition of extracellular matrix. Fibrogenic cells, including hepatic stellate cells and myofibroblasts, are activated in response to a variety of cytokines, growth factors, and inflammatory mediators. The involvement of members of the epidermal growth factor family in this process has been suggested. Amphiregulin (AR) is an epidermal growth factor receptor (EGFR) ligand specifically induced upon liver injury. Here, we have addressed the in vivo role of AR in experimental liver fibrosis. To this end, liver fibrosis was induced in AR+/+ and AR−/− mice by chronic CCl4 administration. Histological and molecular markers of hepatic fibrogenesis were measured. Additionally, the response of cultured human and mouse liver fibrogenic cells to AR was evaluated. We observed that AR was expressed in isolated Kupffer cells and liver fibrogenic cells in response to inflamatory stimuli and platelet‐derived growth factor, respectively. We demonstrate that the expression of α‐smooth muscle actin and collagen deposition were markedly reduced in AR−/− mice compared to AR+/+ animals. AR−/− mice also showed reduced expression of tissue inhibitor of metalloproteinases‐1 and connective tissue growth factor, two genes that responded to AR treatment in cultured fibrogenic cells. AR also stimulated cell proliferation and exerted a potent antiapoptotic effect on isolated fibrogenic cells. Conclusion: These results indicate that among the different EGFR ligands, AR plays a specific role in liver fibrosis. AR may contribute to the expression of fibrogenic mediators, as well as to the growth and survival of fibrogenic cells. Additionally, our data lend further support to the role of the EGFR system in hepatic fibrogenesis. (HEPATOLOGY 2008.)

Amphiregulin Induces the Alternative Splicing of p73 Into Its Oncogenic Isoform ΔEx2p73 in Human Hepatocellular Tumors

BACKGROUND & AIMS Inactivation of the product of the tumor suppressor gene TP73 does not usually occur by mutation but rather through expression of truncated isoforms that have dominant-negative effects on p73 and p53. The truncated oncogenic isoform DeltaEx2p73 is expressed in hepatocellular carcinomas (HCC) and is produced through the alternative splicing of p73 pre-messenger RNA (pre-mRNA); however, the underlying mechanisms regulating this process are unknown. METHODS We used human normal and diseased liver tissue samples, as well as human HCC cell lines, to examine the association between activation of epidermal growth factor receptor (EGFR) by its ligand amphiregulin (AR) and the alternative splicing of p73 pre-mRNA into the tumorigenic isoform DeltaEx2p73, via c-Jun N-terminal-kinase-1-mediated signaling. RESULTS DeltaEx2p73 was expressed in a subset of premalignant cirrhotic livers and in otherwise healthy livers that harbored a primary tumor, as well as in HCC tissues. DeltaEx2p73 expression was correlated with that of the EGFR ligand AR, which was previously shown to have a role in hepatocarcinogenesis. Autocrine activation of the EGFR by AR triggered c-Jun N-terminal kinase-1 activity and inhibited the expression of the splicing regulator Slu7, leading to the accumulation of DeltaEx2p73 transcripts in HCC cells. CONCLUSIONS This study provided a mechanism for the generation of protumorigenic DeltaEx2p73 during liver tumorigenesis, via activation of EGFR signaling by AR and c-Jun N-terminal kinase-1 activity, leading to inhibition of the splicing regulator Slu7.

Epidermal Growth Factor Receptor (EGFR) Crosstalks in Liver Cancer

Hepatocarcinogenesis is a complex multistep process in which many different molecular pathways have been implicated. Hepatocellular carcinoma (HCC) is refractory to conventional chemotherapeutic agents, and the new targeted therapies are meeting with limited success. Interreceptor crosstalk and the positive feedback between different signaling systems are emerging as mechanisms of targeted therapy resistance. The identification of such interactions is therefore of particular relevance to improve therapeutic efficacy. Among the different signaling pathways activated in hepatocarcinogenesis the epidermal growth factor receptor (EGFR) system plays a prominent role, being recognized as a “signaling hub” where different extracellular growth and survival signals converge. EGFR can be transactivated in response to multiple heterologous ligands through the physical interaction with multiple receptors, the activity of intracellular kinases or the shedding of EGFR-ligands. In this article we review the crosstalk between the EGFR and other signaling pathways that could be relevant to liver cancer development and treatment.

Splicing regulator SLU7 is essential for maintaining liver homeostasis

A precise equilibrium between cellular differentiation and proliferation is fundamental for tissue homeostasis. Maintaining this balance is particularly important for the liver, a highly differentiated organ with systemic metabolic functions that is endowed with unparalleled regenerative potential. Carcinogenesis in the liver develops as the result of hepatocellular de-differentiation and uncontrolled proliferation. Here, we identified SLU7, which encodes a pre-mRNA splicing regulator that is inhibited in hepatocarcinoma, as a pivotal gene for hepatocellular homeostasis. SLU7 knockdown in human liver cells and mouse liver resulted in profound changes in pre-mRNA splicing and gene expression, leading to impaired glucose and lipid metabolism, refractoriness to key metabolic hormones, and reversion to a fetal-like gene expression pattern. Additionally, loss of SLU7 also increased hepatocellular proliferation and induced a switch to a tumor-like glycolytic phenotype. Slu7 governed the splicing and/or expression of multiple genes essential for hepatocellular differentiation, including serine/arginine-rich splicing factor 3 (Srsf3) and hepatocyte nuclear factor 4α (Hnf4α), and was critical for cAMP-regulated gene transcription. Together, out data indicate that SLU7 is central regulator of hepatocyte identity and quiescence.

Oral Methylthioadenosine Administration Attenuates Fibrosis and Chronic Liver Disease Progression in Mdr2−/− Mice

Background Inflammation and fibrogenesis are directly related to chronic liver disease progression, including hepatocellular carcinoma (HCC) development. Currently there are few therapeutic options available to inhibit liver fibrosis. We have evaluated the hepatoprotective and anti-fibrotic potential of orally-administered 5′-methylthioadenosine (MTA) in Mdr2−/− mice, a clinically relevant model of sclerosing cholangitis and spontaneous biliary fibrosis, followed at later stages by HCC development. Methodology MTA was administered daily by gavage to wild type and Mdr2−/− mice for three weeks. MTA anti-inflammatory and anti-fibrotic effects and potential mechanisms of action were examined in the liver of Mdr2−/− mice with ongoing fibrogenesis and in cultured liver fibrogenic cells (myofibroblasts). Principal Findings MTA treatment reduced hepatomegaly and liver injury. α-Smooth muscle actin immunoreactivity and collagen deposition were also significantly decreased. Inflammatory infiltrate, the expression of the cytokines IL6 and Mcp-1, pro-fibrogenic factors like TGFβ2 and tenascin-C, as well as pro-fibrogenic intracellular signalling pathways were reduced by MTA in vivo. MTA inhibited the activation and proliferation of isolated myofibroblasts and down-regulated cyclin D1 gene expression at the transcriptional level. The expression of JunD, a key transcription factor in liver fibrogenesis, was also reduced by MTA in activated myofibroblasts. Conclusions/Significance Oral MTA administration was well tolerated and proved its efficacy in reducing liver inflammation and fibrosis. MTA may have multiple molecular and cellular targets. These include the inhibition of inflammatory and pro-fibrogenic cytokines, as well as the attenuation of myofibroblast activation and proliferation. Downregulation of JunD and cyclin D1 expression in myofibroblasts may be important regarding the mechanism of action of MTA. This compound could be a good candidate to be tested for the treatment of (biliary) liver fibrosis.

Novel Pharmacologic Strategies to Protect the Liver from Ischemia- Reperfusion Injury

Ischemia and reperfusion (I/R) injury develops when blood flow is interrupted for a long period of time and then restarted. In the liver, this type of damage occurs in clinical settings such as liver transplantation and hepatic resection. Given the shortage of donor organs it is essential to maximize the use of sub-optimal organs, those previously rejected due to elevated risk of malfunction, and to increase split-liver transplantation interventions. Therefore, the development of strategies that preserve organ viability and promote liver regeneration is urgently needed. As observed for other organs, a brief period of ischemia followed by short reperfusion before the surgical procedure significantly increases liver resistance towards prolonged periods of ischemia. This phenomenon is known as ischemic preconditioning, and is the only protective strategy that has reached clinical practice. Recently, intensive research has improved our understanding of the mechanisms involved in I/R liver injury, and the biologic bases of ischemic preconditioning. This knowledge has generated relevant patented advances in the field, including the targeted inhibition of pro-apoptotic pathways, the interference with neutrophil activation, and the identification of cytoprotective cytokines. Here, we briefly review the mechanisms of hepatic ischemic damage, and present the most promising pharmacologic approaches against I/R injury. This article also includes recent patents on this topic.