hepatocellular carcinoma ( hcc ) is the sixth most common malignancy worldwide and the third most common cause of global cancer related mortality [ 1 , 2 ] . hcc burden disproportionately impacts developing countries and males ; as of 2008 , 85% of cases occurred in africa and asia , with worldwide male : female sex ratio of 2.4 . risk factors for the development of hcc include chronic liver inflammation from hepatitis b and c infection , autoimmune hepatitis , excessive alcohol use , nonalcoholic steatohepatitis , primary biliary cirrhosis , environmental carcinogens such as aflatoxin b , and genetic metabolic disease ( such as hemochromatosis and alpha-1 antitrypsin deficiency ) . prognostic and therapeutic options are dependent upon the severity of underlying liver disease , and median overall survival ( os ) for metastatic or locally advanced disease is estimated at 58 months . hcc is relatively refractory to cytotoxic chemotherapy , likely due to overexpression of multidrug - resistant genes , protein products such as heat shock 70   and p - glycoprotein , and p53 mutations . presently , systemic therapeutic options in the locally advanced or metastatic setting are limited to sorafenib , an oral multikinase inhibitor targeting raf kinase , vascular endothelial growth factor ( vegf ) , and platelet - derived growth factor ( pdgf ) receptor tyrosine kinase signaling .   although the transition from normal hepatocyte to hcc is not fully understood , hepatocarcinogenesis is a complex multistep process driven by accumulation of heterogeneous molecular alterations from initial hepatocyte injury to metastatic invasion . dysplastic nodules subsequently progress to early hcc through cumulative genetic alterations , while advanced hcc often involves intrahepatic metastasis and portal vein invasion . molecular alterations implicated in hcc development include mutations in oncogenes and tumor suppressor genes ( p53 and p16 ) , epigenetic alterations , chromosomal changes , and aberrant activation of signaling cascades necessary for proliferation , angiogenesis , invasion and metastasis , and survival . pathogenesis of early and advanced hcc may be modulated through different mechanisms ; for example , p53 mutations , p16 gene silencing , and aberrant akt signaling are more frequently observed in advanced hcc [ 46 ] . the molecular pathogenesis of hcc is multifactorial and is reliant upon dysregulation of multiple pathways including wnt / b - catenin , mitogen - activated protein kinase ( mapk ) , phosphatidylinositol-3 ( pi3k)/akt / mammalian target of rapamycin ( mtor ) , vegf , pdgf , insulin - like growth factor ( igf ) , epidermal growth factor ( egf ) , tgf - beta , and hepatocyte growth factor [ 6 , 7 ] . the hepatocyte growth factor ( hgf ) and its transmembrane tyrosine kinase receptor , cellular met ( cmet ) promote cell survival , proliferation , migration , and invasion via modulation of epithelial - mesenchymal interactions . hgf - cmet signaling is critical for normal processes such as embryogenesis , organogenesis , and postnatal tissue repair after acute injury . hgf - cmet axis activation is also implicated in cellular invasion and metastases through induction of increased proliferation ( mitogenesis ) , migration and mobility ( motogenesis ) , three - dimensional epithelial cell organization ( morphogenesis ) , and angiogenesis . hgf was first discovered in 1984 as a mitogenic protein for rat hepatocytes in vitro . hgf was subsequently found to be indistinguishable from scatter factor , a fibroblast - derived motility factor promoting epithelial cell dispersal   and three - dimensional branching tubulogenesis . hgf is secreted primarily by mesenchymal cells ( or by stellate and endothelial cells in the liver ) as an inactive single - chain precursor ( pro - hgf ) which is bound to heparin proteoglycans within the extracellular matrix . hgf transcription is upregulated by inflammatory modulators such as tumor necrosis factor alpha , il-1 , il-6 , tgf - beta , and vegf [ 11 , 12 ] . circulating pro - hgf undergoes proteolytic conversion via extracellular proteases including hgf activator ( hgfa ) , urokinase - type plasminogen activator , factors xii and xi , matriptase , and hepsin   into an active two - polypeptide chain heterodimeric linked by a disulfide bond . hgfa is a serine protease which is secreted primarily by the liver and circulates as pro - hgfa ; pro - hgfa is activated by thrombin in response to tissue injury and malignant transformation [ 13 , 14 ] . the active form of hgf includes an -chain containing four kringle domains ( k1 to k4 ) and an amino - terminal loop domain ( n ) , and a -chain ( c - terminal ) containing a serine protease homology ( sph ) domain . hgf is a ligand for the met receptor , also known as cellular met or cmet . the met protooncogene was first isolated in 1984 from a human osteosarcoma - derived cell line driven by a chromosomal rearrangement tpr - met , resulting from fusion of translocated promoter region located on chromosome 1q25 and met sequence located on chromosome 7q31 . the tpr - met rearrangement encodes for a prototype of the cmet receptor tyrosine kinase family . the cmet receptor is expressed predominantly on the surface of endothelial and epithelial cells of many organs , including the liver , kidney , prostate , pancreas , kidney , muscle , and bone marrow . like hgf , cmet is synthesized as an inactive single - chain precursor and undergoes proteolytic cleavage into a disulfide linked heterodimer consisting of an extracellular -chain and transmembrane -chain . the extracellular domain includes an amino - terminal semaphorin domain , terminal cysteine - rich psi domain , and four ipt repeat immunoglobulin domains . the cytoplasmic portion includes a juxtamembrane region including two phosphorylation sites , a tyrosine kinase ( tk ) domain and a carboxyl terminal region for substrate docking . under normal conditions , hgf interacts with cmet via a paracrine signaling loop and triggers cmet kinase activation within the cytoplasmic portion ( see figure 1 ) . hgf contains two cmet binding sites including a high affinity constitutively active site on the -chain ( n - terminal and first kringle domain , or n - k1 ) and a low affinity site on the -chain ( c - terminal ) [ 8 , 19 ] . the exact mechanism of cmet receptor activation and the contribution of both binding sites to receptor activation by full length hgf are not yet established , although it is evident that both - and -chain sites have distinct functions . while -chain n - k1 portion activates cmet and induces met dimerization , -chain residues bind cmet once the receptor is already occupied by hgf n - k4 and subsequently induce phosphorylation and downstream signaling . hgf binding induces autophosphorylation of tyrosine residues y1234 and y1235 within the cmet tk domain activation loop . phosphorylation also occurs at two sites within the carboxyl terminal region ( y1349 and y1356 ) , forming a multifunctional high affinity binding docking site that recruits a range of intracellular adaptors containing src homology-2 domains [ 2124 ] . recruited adaptor proteins and kinase substrates include growth factor receptor - bound protein 2 ( grb2 ) , grb2-associated binder ( gab1 ) , phospholipase c- , stat3 , shc , src , shp2 , pi3k , and ship1 [ 23 , 24 ] . these and other adaptor proteins provide scaffolding for a larger apparatus of network proteins , ultimately promoting activation of multiple signaling pathways . of these , grb2 and gab1 are critical effectors that interact directly with the receptor ; grb2 binding to the cmet docking site through y1356 results in downstream signaling via the ras / mapk pathway , while gab1 phosphorylation by met kinase activates the pi3k pathway . other signaling proteins that are activated by cmet include p38 , jnk , and nuclear factor kb . alterations in transcription induce cell cycle progression , antiapoptosis , increased cell motility , angiogenesis , and survival . the hgf - cmet pathway serves as a hub for multiple heterogeneous signaling networks and is also modulated by the activation of other receptor tk families such as the epidermal growth factor receptor ( egfr ) , human epidermal growth factor receptor 2 , insulin - like growth factor 1 receptor , raf kinase , and vegf . the hgf - cmet pathway plays an essential role in mammalian development in terms of morphogenesis , mitogenesis , and motogenesis , and angiogenesis hgf - cmet signaling is likely to be critical in many aspects of adult homeostasis including cardiac and hepatic tissue injury repair [ 26 , 27 ] , skin wound repair , and skin immunity regulation . targeted disruption of the hgf or met genes results in embryonically lethal knockouts with impaired organogenesis of the liver and placenta . preclinical models demonstrate that hgf functions as a hepatotrophic factor enhancing hepatic regeneration and suppressing hepatocyte apoptosis [ 31 , 32 ] ; expression of hgf is increased in response to liver injury , while neutralization of endogenous hgf or met knockout facilitates liver damage and fibrotic changes with delayed repair . under normal physiologic conditions , hgf - induced cmet activation is strictly controlled by paracrine ligand delivery followed by ligand activation at target cell surfaces and ligand - activated receptor internationalization / degradation . despite multiple checkpoints , hgf - cmet axis dysregulation occurs in a variety of solid tumors and hematopoietic derived malignancies and plays a key role in malignant transformation by promoting tumor cell migration , epithelial to mesenchymal transition , invasion , proliferation , and angiogenesis . dysregulated cmet signaling upregulates protease production ( plasminogen dependent and independent ) and increased cell dissociation via extracellular matrix degradation , facilitating tumor invasiveness and metastasis . mechanisms of pathogenic activation include aberrant paracrine or autocrine ligand production , overexpression of hgf and cmet , upregulation of genes encoding proteases for hgf / cmet processing , constitutive kinase activation independent of cmet gene amplification , and cmet gene mutations leading to ligand - independent kinase activity [ 3436 ] . the hgf - cmet axis is implicated in hepatocarcinogenesis through multiple mechanisms , many of which are still being elucidated . overexpression of hgf   and cmet [ 3741 ] is observed in 33% and 2048% of human hcc samples , respectively . the prognostic utility of cmet and hgf overexpression is uncertain ; while some studies show no correlation between cmet overexpression and tumor size or invasive behavior [ 38 , 42 ] or hgf levels and survival , others demonstrate an inverse relationship with survival . specifically , cmet overexpression was found to correlate with poorly differentiated hcc   and increased intrahepatic metastases along with decreased five - year survival . after hepatectomy , cmet overexpression in hcc tissue has been correlated with early tumor recurrence , metastasis , and shorter 5-year survival [ 41 , 45 , 46 ] . a cmet - regulated gene expression signature was found to define a subset of human hcc with poor prognosis and aggressive phenotype and correlated with increased vascular invasion , increased microvessel density , and decreased mean survival time . higher hgf levels negatively correlate with survival per biomarker analysis of the sharp trial   and prior data   and positively correlate with tumor size ; however given that hgf is secreted as an inactive precursor , overexpression alone is unlikely to guarantee pathway dysregulation . hgf has also been investigated as a potential biomarker for hcc development , but may also be a specific marker for liver cirrhosis . while gains in 7q have been reported in hcc , gene amplification has not been reported as a significant source of increased cmet . crosstalk between cmet and egfr [ 55 , 56 ] and cmet and vegf signaling pathways is also implicated in promoting tumor survival . cmet - csrc has been shown to mediate egfr phosphorylation and cell survival in the presence of egfr inhibitors . cmet is both an independent angiogenic factor and interacts with angiogenic survival signals promoted through vegf . by upregulating hypoxia - inducible factor , hypoxia results in increased hgf expression in tumor and surrounding normal interstitial cells and hgf - cmet signaling induces upregulation of tumoral vegf expression and endothelial vegfr2 expression and downregulation of endogenous inhibitors of angiogenesis [ 58 , 59 ] . dual vegf and cmet axis activity demonstrates increased capillary formation in vivo , tubulogenesis in vitro , and tumoral microvessel density . given the predominant role of dysregulated hgf - cmet signaling in hepatocarcinogenesis , pharmacologic cmet inhibition is a promising therapeutic strategy . targets for inhibition of the cmet signal transduction pathway include ligand - receptor interaction , cmet kinase activity , and cmet and adaptor protein interaction . hgf - cmet axis inhibitors can be broadly classified into biologic antagonists , c - met adaptor protein inhibitors , small - molecule downstream pathway inhibitors , and small synthetic met tyrosine kinase inhibitors ( tki ) . of these , tkis are the most common and the only ones to have completed phase 2 testing in hcc as of march 2013 . table 1 shows the selected hgf - cmet inhibitors in active clinical trials for either hcc or advanced solid malignancies ( including hcc ) as of march 2013 . a comprehensive listing of hgf - cmet inhibitors in active clinical trials for all malignancies is maintained by the bottaro nci lab and is available at https://ccrod.cancer.gov/confluence/display/ccrhgf/home . biologic antagonists inhibit cell surface interactions such as ligand - receptor binding or receptor clustering , preventing activation of downstream signaling . these include hgf competitive analogs , met decoy receptor , and anti - hgf - cmet monoclonal antibodies . hgf competitive analogs compete with ligand for receptor binding without causing cmet dimerization and activation and include nk2 [ 60 , 61 ] , nk4 [ 6264 ] , and uncleavable hgf ; preliminary safety and drug development data in humans are pending . nk2 may be the least promising hgf competitive analog due to being a potent mitogen   and promoting metastases . met decoy receptors are soluble forms of the cmet extracellular domain which compete with hgf and inhibit cmet dimerization ; in vitro and in vivo mice models demonstrate suppression of hgf - induced tumor cell migration and metastasis [ 68 , 69 ] . monoclonal antibodies targeting hgf and the extracellular domain of cmet are currently being explored in clinical trials ( see table 1 ) , but data are not yet available for hcc .   given the importance of adaptor proteins in propagating downstream cmet signaling , cmet adaptor inhibitors offer unique potential for cmet specific inhibition . as described above , cmet signaling is initiated through autophosphorylation of cytoplasmic tyrosines that form docking sites for adaptor proteins . grb2 and gab1 are critical effectors that interact directly with the cmet receptor , ultimately recruiting a larger apparatus of network proteins necessary for cmet signaling . gab1 coupling with cmet requires gab1 binding to the sh3 domain of grb2 , and cmet binding to the sh2 domain of grb2 [ 72 , 73 ] . c90 is a selective grb2 antagonist with demonstrable inhibition of gastric cancer cell motility and matrix invasion in vitro and impaired metastatic spread of human prostate cancer cells in vivo ; data in human studies have not been reported to date [ 74 , 75 ] . small - molecule downstream pathway inhibitors directed towards inhibition of stat3 phosphorylation showed preliminary tolerability in a phase i trial of advanced solid tumors , but data are not yet available for hcc . synthetic small met tkis inhibit downstream signal transduction by preventing phosphorylation , either via competitive binding of the intracellular adenosine triphosphate ( atp ) site in cmet 's tk domain , or noncompetitive binding of a cmet region outside of the atp binding site . while some of the tkis in development are cmet specific , others target multiple pathways including vegf , pdgfr , fms - related tyrosine kinase 3 ( flt3 ) , v - kit feline sarcoma viral oncogene homolog protein ( kit ) , and anaplastic lymphoma kinase ( alk ) . preclinical studies and clinical trials show tolerability and efficacy of cmet tkis across a variety of solid malignancies . as of march 2013 , promising results in the phase 2 randomized setting for hcc are available for two cmet inhibitors : tivantinib and cabozantinib . tivantinib ( arq 197 ) is an oral low - molecular - weight tki which is non - atp competitive . safety and tolerability without drug - related worsening of hepatic function were previously reported in a phase ib trial of 20 cirrhotic patients ( child - pugh a and b ) with hcc , with 2 or less prior systemic chemotherapy regimens . rimassa and colleagues reported results at asco 2012 of a phase ii trial assigning 107 patients with unresectable hcc with ecog ps 0 - 1 and child - pugh a in a 2  :  1 randomization to either second - line tivantinib or placebo with crossover allowed . although no difference in median os occurred , the primary endpoint of time to progression ( ttp ) was met and favored tivantinib versus placebo ( 6.6 versus 6.2 months , hr = 0.90 , p = 0.63 ; 6.9 versus 6 weeks , hr = 0.64 , p = 0.04 , resp . ) . patients with high met expression ( defined as 50% or more cells in the tumor specimen with 2 + or 3 + staining intensity ) versus low met expression receiving tivantinib demonstrated a significant improvement in both os and ttp ( 7.2 versus 3.8 months , hr 0.38 p = 0.01 ; 11.7 versus 6.8 weeks , hr 0.43 , p = 0.03 , resp . ) . no detrimental effect was reported in patients with low met expression , and common adverse events were neutropenia , asthenia , poor appetite , and anemia . as of november 2012 , a phase iii trial for tivantinib and hcc patients is in development . cabozantinib ( xl 184 ) is an unselective oral multikinase tki targeting cmet , kit , rearranged during transfection ( ret ) , vegfr1 and 2 and 3 , flt3 , axl receptor tyrosine kinase ( axl ) , and tie family angiopoietin 1 receptor . cabozantinib has shown efficacy in the phase 3 setting for medullary thyroid carcinoma with progression - free survival ( pfs ) improvement and phase 2 setting for advanced solid tumors ( including hcc ) . verslype and colleagues reported preliminary results of a phase 2 randomized discontinuation study of cabozantinib in 41 patients with hcc , child - pugh score a , and one prior line of systemic treatment . all patients initiated cabozantinib for a 12-week lead in time frame ; patients with partial response continued on study drug while patients with progression of disease discontinued . 32 patients with stable disease were blindly randomized 1  :  1 to continue cabozantinib or receive placebo . the primary endpoint was overall response rate ( rr ) during the lead in time phase and pfs for patients entering the randomization phase . independent of prior sorafenib use , median os was 15.1 months , median pfs was 4.4 months , and median time on study was 6 months . common grade 3 adverse effects were hand - foot syndrome , diarrhea , and thrombocytopenia . met expression was not evaluated prospectively , and given the broad activity of cabozantinib , it is unclear how much activity is attributable to met inhibition alone . in fact , the inhibition of both vegf and met concurrently may be particularly effective . vegf inhibition leads to increased met signaling , either from resultant hypoxia or direct interactions between vegfr2 and met [ 81 , 82 ] . preliminary evidence of efficacy against hcc was seen in a phase i combination study of tivantinib with sorafenib including a complete response and another prolonged partial response lasting greater than one year . inc280 is a selective met tki in phase i testing for early hcc , currently accruing . foretinib ( gsk 1363089 ) is a small - molecule tki targeting both met and vegf in phase i / ii testing for advanced hcc . based on preclinical trials and phase 2 data for tivantinib and cabozantinib , inhibition of cmet signaling is a promising therapeutic strategy in hcc . although it is unclear which genetic and molecular abnormalities implicated in hgf - cmet dysregulation are predictive of sensitivity to cmet targeted therapy , ongoing trials must address the challenge of identifying patients most likely to achieve maximal benefit and minimal toxicity . two current strategies for patient selection based on tumor biomarkers are quantification of tumor cmet content via immunohistochemical ( ihc ) and immunoassay tissue analysis and assessment of met sequence status . pharmacodynamic serum markers are under evaluation in ongoing trials as a strategy to assess clinical response . tumoral cmet overexpression as measured by commercially available ihc kits appears to correlate with efficacy of cmet inhibitors in hcc and other solid tumors , although more prospective data are necessary for validation . identification of cmet phosphorylation status is a potentially powerful target for targeted antibodies ; two site immunoassays of flash - frozen tissue samples yielding precise measurements of met content and phosphorylation activation are currently under development . another promising stratification strategy is assessment of met sequence status including met mutations , met amplification , and chromosome 7 polysomy . preclinical studies of cmet targeted agents demonstrate variable efficacy based on met mutation location ; for example , pf-2341066/4217903 is a selective inhibitor with increased activity against certain cmet atp binding site mutations in comparison to met kinase domain activation loop mutations . in vitro studies of su11274 , a small - molecule tk competitive atp binding site inhibitor , show selective inhibition of two out of four identified met mutations . amplification of cmet is associated with increased clinical response to foretinib ( xl 880 ) in phase 2 gastric cancer data , and cmet copy number correlates with increased clinical response to tivantinib in addition to erlotinib in advanced nsclc . ancillary pharmacodynamic and pharmacokinetic marker studies in a variety of solid tumors show variable correlation with clinical response . preclinical data with crizotinib in gastric cancer cell lines demonstrate variable biomarker modulation of cmet inhibition with cmet dependent gastric cancer cell lines versus cmet - independent lines [ 88 , 89 ] . clinical data shows changes in plasma concentrations of hgf , vegf , soluble met , soluble vegfr2 , pigf , and epo during treatment with various tkis [ 90 , 91 ] . the predictive utility of these biomarkers in patients with cmet - dependent hcc is unclear ; further analyses and prospective validation are necessary . hgf - cmet is an intriguing target for the development of companion diagnostic tools as an adjunct tool for patient selection and stratification for cmet therapy . in vitro and in vivo animal studies suggest that radiolabelled dyes containing cmet binding peptide successfully targets cmet receptors with higher imaged based tumor uptake . a variant of su11274 with radiomethylation modification is being utilized as a pet visualization agent to quantify cmet receptor in xenograft models . inhibition of cmet is a promising therapeutic strategy in hcc . given the heterogeneous mechanisms underlying cmet dysregulation , there is an urgent and unmet need for the development of predictive biomarkers to identify which subsets of cmet - dependent hcc tumors are most likely to benefit from specific classes of inhibitors . as more agents move into phase 2 and 3 trials for hcc , one important consideration is the emergence of acquired and primary resistance mechanisms from de novo or preexisting mutations . these may be overcome by rational combination therapy directed against multiple pathways , different levels of ligand - receptor - tki interaction , and the presence of cmet addiction . innovative clinical trial designs ( such as the discontinuation cabozantinib design ) with incorporation of enriched patient cohorts , biomarker analyses , pharmacodynamic markers , and companion diagnostics are essential in moving forward .