183 Andrea Perra and Marta Anna Kowalik. / International Journal of Biological & Pharmaceutical Research. 2016; 7(4): 183-191. e- ISSN 0976 - 3651 Print ISSN 2229 - 7480 International Journal of Biological & Pharmaceutical Research Journal homepage: www.ijbpr.com IJBPR YES-ASSOCIATED PROTEIN (YAP) – A PROMISING THERAPEUTIC TARGET FOR HEPATOCELLULAR CARCINOMA Andrea Perra* and Marta Anna Kowalik Oncology and Molecular Pathology Unit, Department of Biomedical Sciences, University of Cagliari, via Porcell 4, 09124 Cagliari, Italy. ABSTRACT Hepatocellular carcinoma (HCC), the third leading cause of cancer mortality, carries a dismal prognosis and represents a major health problem. A better understanding of the molecular pathways involved in HCC development may represent an important approach for the improvement of the therapeutic strategies for this cancer. The Hippo signalling pathway, a growth- suppressive mechanism that antagonizes the transcriptional co-activator Yes-associated protein (YAP), has been recently found altered in human HCC. Moreover, using the Resistant-Hepatocyte (R-H) rat model, recently published data established that the Hippo pathway deregulation occurs already at the early stages of HCC development, making this pathway an important therapeutic target. Many strategies have been proposed to modulate the activity of the Hippo pathway, and in particular that of the final effector YAP, most of which require complex gene manipulations, useful to understand the functions of the pathway but difficult to apply on patients. One of the few drugs able to inhibit in vivo the pro-carcinogenic effect of YAP activation is verteporfin, which disrupts the formation of the complex between YAP and the TEAD transcription factors, causing a significant reduction of the number and size of preneoplastic foci induced in rats liver by R-H protocol. In this review, we give a short summary of Hippo pathway, providing the evidences of its deregulation in mouse, rat and human liver cancer and discuss the possibility to treat HCC with new drugs targeting the transcriptional co-activator YAP. Key Words: Liver, Cancer, YAP, Hippo, Therapy, Verteporfin. INTRODUCTION Liver cancer, which includes hepatocellular or cirrhosis (El-Serag et al., 2007). HCC carries a dismal carcinoma (HCC) and intrahepatic cholangiocarcinoma prognosis and only partial liver resection or liver (iCCA), represents the second cause of cancer related transplantation are potentially curative, but only a minority deaths and its incidence rate is still significantly increasing of cases is amenable to these treatments. (Bruix et al., 2015; Siegel et al., 2014). Marked variations Hepatocarcinogenesis is considered to be a very complex among geographic regions, racial and ethnic groups, and multistep process, which involves alterations in between men and women and the presence of several well- numerous signalling cascades. In particular, Wnt/β-catenin, documented environmental potentially preventable risk p53, Mitogen-Activated Protein Kinase, Ras, JAK/STAT, factors are reported as important epidemiologic features of Epidermal Growth Factor Receptor and Transforming HCC. This malignancy generally develops as a Growth Factor-β pathways have been extensively studied consequence of underlying liver diseases such as hepatitis through pathway-based approaches and functional experimental studies in order to identify potential Corresponding Author molecular targets (Aravalli et al., 2008). Recently, pioneering studies have reported that the Hippo signaling Andrea Perra pathway, an evolutionary highly conserved regulator of Email: [email protected] tissue growth, organ size and stem cell property, is 184 Andrea Perra and Marta Anna Kowalik. / International Journal of Biological & Pharmaceutical Research. 2016; 7(4): 183-191. involved in the pathogenesis of HCC (Dong J et al., 2007; resulting in their sequestration in the cytoplasm and, Zender L et al., 2010; Pan D et al., 2010; Forner A et al., finally, their ubiquitination-dependent proteosomal 2012). degradation. Therefore, the Hippo pathway acts by Many components of this signaling pathway, blocking the ability of YAP and TAZ to translocate in the which is composed of the upstream regulatory components, nucleus, where they interact with transcriptional factors of the Hippo core kinase components, and the downstream the TEAD family (TEAD1–4), leading to induction of transcriptional machinery, were initially discovered by target genes such as Axl, Birc5, CTGF, IGFBP3, ITGB2 Drosophila mosaic genetic screens (Justice RW et al., and Gli2 (Sudol M et al., 1995). 1995). Successively, genetic and biochemical studies In mammals the most conserved component of demonstrated that the Hippo pathway is highly this pathway is YAP, which is considered to be the evolutionary and functionally conserved in mammals principal and less tissue-specific nuclear effector of the (Zhao B et al., 2010). Recently, increasing number of Hippo pathway. It owes the name to its ability to interact studies confirmed that Hippo pathway perturbation can with Yes protein; however, the biological relevance of this trigger tumorigenesis in mice and altered expression of interaction remains elusive (Harvey KF et al., 2013). genes targeted by this pathway have been observed in Originally sequenced in chicken, human and mouse various human cancers (Steinhardt AA et al., 2008; Harvey homologues of YAP were identified by using cDNA to KF et al., 2013). Indeed, the Hippo pathway results probe cDNA libraries (Sudol M et al., 1995). Initially, it deregulated in a broad variety of human cancers, including was demonstrated that variations of YAP levels can hepatocellular carcinoma (Zhou D et al. 2009; Xu MZ et overcome growth arrest at confluence, in vitro, and organ al., 2009; Li H et al., 2012). In recent times, an important size control in vivo. YAP effects on liver size is goal in cancer therapy is to develop drugs able to target astonishing. Indeed, overexpression of YAP in transgenic selectively a pathway or molecule demonstrated to be mice causes an increase of the liver weight which can be crucial for tumor progression. In this regard, Hippo from 5% to 25% of body weight, largely due to increased pathway emerged as an important candidate, as critical for cell number. This effect is reversible as the enlarged liver the growth of cancer cells, but with minimal role in adult reverts to almost normal size once overexpression of YAP tissue homeostasis (Stanger BZ, 2012). In this review, we is ceased (Camargo FD et al., 2007). Moreover, sustained provide a short summary of Hippo pathway components induction of YAP expression leads to formation of and discuss the role its effector, the transcriptional co- hepatocellular adenomas and carcinomas (Dong J et al., activator YAP (Yes-associated protein), in hepato 2007). YAP activation is also associated with liver carcinogenesis and the new therapeutic strategies targeting hyperplasia, increased liver size and HCC also in non- the Hippo downstream transcriptional machinery. transgenic mice, following treatment with TCPOBOP (1,4- Bis-[2-(3,5-dichloropyridyloxy)]benzene), an agonist of The Hippo pathways and its effectors in mammals the constitutive androstane receptor (CAR) (Kowalik MA The Hippo pathway is conserved signaling et al., 2011). cascades that regulates organ size by controlling cell This intense and reversible effect on liver size proliferation and apoptosis, and participate to caused by YAP activation, places the Hippo pathway as a organogenesis and cell differentiation. These features make major mechanism inducing liver hyperplasia in mammals. the study of the alterations of this pathway extremely Furthermore, in two different animals models of YAP relevant to several human diseases, including hypertrophic activation, transgenic over-expression and drug-induced, cardiopathy, alteration of the inflammatory response, delay primary, liver tumors were observed, strengthening the in wound healing, and, recently, in primary tumors of the concept of YAP as a relevant oncogene in liver (Perra A et al., 2014). Basically, the Hippo pathway hepatocarcinogenesis, as already demonstrated for other can be divided into three parts, the upstream regulatory carcinomas (Harvey KF et al., 2013). components, the Hippo core kinase components, and the downstream transcriptional machinery, each one YAP structure and functions constituted by several proteins (figure 1). In mammals, the The human YAP gene (also known as YAP1) is upstream components are still poorly understood, while the located at 11q13 and can be transcribed in a least 8 core kinase that controls the transcriptional machinery is isoforms, that are derived by differential splicing, that are well characterized. It contains four proteins, two of which classified into two main groups, YAP-1 (or YAP1-1) and are kinases, Mst1/2 and Lats1/2, the other two are the YAP-2 (or YAP1-2) (Gaffney CJ et al., 2012). The most adaptor/activator proteins WW45 and Mob. Upstream studied and cited isoform belongs to the YAP-2 group and regulators activate, by phosphorylation, the Mst1/2-WW45 is that containing 488 residues. The analysis of the protein complex, which, in turn, can phosphorylate and activate sequence allowed the identification of several domains, the LATS1/2-Mob complex. The major target of this common to all isoforms, that are crucial for the modulation cascade is the phosphorylation of the effectors YAP and of the Hippo pathway (figure 2). The so called Lats TAZ, that