Novel hepatocellular carcinoma molecules with prognostic and therapeutic potentials

Table 1 Molecular and biochemical markers of human hepatocellular carcinoma progression and poor prognosis

Molecule	Molecular category	Biological function	Expression levels in human HCC tissue or serum	Clinical pathological indications	Target for therapy	Ref.
GPC3	Member of heparan sulfate proteoglycan membrane proteins family	Regulates the signalling pathways of several growth factors, including Wnts	Over-expression; In tissue or serum (protein)	Metastasis, overall survival decrease, recurrence, prognostic value in early diagnosis	Yes	[12,14,16-18]
DKK1	Member of dickkopf family of secreted protein with two cysteine rich regions	Cell proliferation and survival control, differentiation, and cell motility; negative regulator of Wnt/β-catenin pathway	Over-expression in tissue (mRNA and protein) and in serum (protein)	Vascular invasion and metastasis, early diagnosis in AFP-negative HCC, distinguish between malignant and non-malignant liver diseases	Potential	[22]
S100A4	Member of S100 calcium-binding protein family	Target gene of Wnt/β-catenin pathway	Over-expression in tissue	Poorly differentiated HCC, vascular invasion and recurrence, overall survival decrease, up to now limited to HBV + HCC	Potential	[24]
S100 calcium-binding protein A14	Member of S100 calcium-binding protein family	Role not yet defined	Over-expression in tissue (mRNA and protein)	HCC vascular invasion and metastasis, disease-free and overall survival decrease	No data	[25]
Sex determining region Y (SRY) related high-mobility group box 6 (SOX6)	Member of the SRY box gene superfamily of transcription factors containing a DNA-binding high mobility group (HMG) domain	Involved in cell embryogenesis, differentiation and proliferation	Decreased expression in tissue (mRNA and protein)	Overall survival decrease, advance tumor stage	No data	[27]
SUOX	Mitochondria member of molybdenum oxotransferases superfamily	Involved in oxidative phosphorylation for ATP generation via electron transport chain	Decreased expression in tissue (protein)	Overall survival decrease in serum AFP positive, with increase of AKR1B10 and CD34 early diagnosis: high grade dysplastic nodules and HCC progression	No data	[28]
xCT	Component of cystine/glutamate transporter designated as system xc–	In mammalian cells involved in redox balance by mediating cystine entry in exchange for intracellular glutamate	Over-expression in tissue (mRNA)	Overall survival decrease related to the mRNA levels	Potential	[31]
GRK6	Member of the G protein-coupled receptor kinase of the Ser/Thr protein kinase family	Phosphorylates the activated forms of G protein-coupled receptors. It is involved in migration and invasion	Over-expression in tissue (protein)	Aggressiveness related to Ki-67, overall survival decrease	No data	[34]
GPR87	Member of the family of G protein-coupled cell surface receptor related to lysophosphatidic acid receptor family	Antiapoptotic role sustaining p53-mediated pro-survival of cells, involved in cell communication	Over-expression in tissue (protein)	Aggressiveness, intrahepatic metastasis related to CD133 up-regulation	Potential	[37]
MT-1 and MT-2	Members of the cysteine-rich, low molecular weight membrane proteins, localized on Golgi apparatus	Protect from oxidative damage and regulate metals homeostasis (storage, transport and detoxification). Involved in angiogenesis, apoptosis, cell cycle progression and differentiation	Decreased expression in tissue (protein)	Agressiveness, vascular invasion, recurrence free survival time reduced and overall survival decrease	No data	[39]
RAI3	Member of G-coupled cell surface receptor	Involved in the regulation of cell proliferation.	Over-expression in tissue (mRNA and protein)	Agressiveness, vascular invasion, increased tumor recurrence, shorter overall survival	Potential	[42]
SSX2IP	Spindle protein	Helps centrosome prepare for mitosis. Essential for centrosome maturation	Over-expression in tissue (mRNA)	Larger tumor size and metastasis, overall post-operative survival time reduced	Potential	[44]
Protein Phosphatase magnesium- Ppm1d	Member of PP2C family of Ser/Thr protein phosphatases	Inhibits p53 signalling, down-regulates p38 mitogen-activated protein kinase and thus is a putative oncogenetic protein	Over-expression in tissue (mRNA and protein), correlates with high AFP level	Larger tumor size, metastasis, shorter overall survival	No data	[46]
BCL9	Protein associated with B-cell acute lymphoblastic leukemia	Involved in signal transduction through Wnt pathway, promotes β-catenin’s transcription activity	Over-expression in tissue (protein)	Aggressiveness, vascular invasion and metastasis. Shorter disease free status after hepatectomy	No data	[47]
IRF-1 and IRF-2	Belongs to transcription factors, member of the interferon regulatory factor family	IRF-1 and its functional antagonist IRF-2 are involved in the modulation of IFNs	IRF-1 decreased expression or IRF-2 over-expression in tissue (protein)	Aggressiveness, shorter overall survival and early recurrence	No data	[49]
CDK4	Member of the Ser/Thr kinase family	Controls the G1-S phase by phosphorylation of retinoblastoma protein	Over-expressionin tissue (mRNA and protein)	Larger tumor size, HBV and shorter median survival	potential	[50]
LASP-1	Not yet elucidated	Involved in actin assembly	Positive cytosolic expression in tissue (protein)	Poorer survival	No data	[51]
PTP4A3	Member of PRL subgroup of protein tyrosine phosphatases	Involved in signaling pathways modulating cell proliferation, growth and motility	Over-expression in tissue (protein)	Overall survival decrease and shorter recurrence free survival	Potential	[52]
Steroyl-CoA-desaturase	Endoplasmic reticulum enzyme that catalyzed the biosynthesis of monounsaturated fatty acids	Catalyzed the D9-cis desaturation of a range of fatty acyl-CoA substrates such as palmitoyl- and stearoyl-CoA (converted into palmitoleoyl-and oleyl-CoA, respectively)	Over-expression in tissue (mRNA)	Distinguishes between high and low risk HCC, positively related to AFP levels, tumor size and staging	Yes	[54]
PAK5	Member of the p21-activated kinase (Pak) family that serves as target for GTP-binding proteins involved in modulation of cell shape, movement, proliferation and survival	A mediator of filipodia formation	Over-expression in tissue (mRNA)	Early diagnosis and possibly indicator of HCC progression	Potential	[56]
hnRNP-L	Member of the hnRNPs that are involved in the processing of pre-mRNAs	Regulator of alternative splicing with both activator and repressor functions (retention and skipping of the exons)	Over-expressed (serum and tissue)	Progression in HBV-HCC patients, tumor size increase and overall survival decrease that correlates with high serum levels	Potential	[57]
CYLD	Protease, displaying endodeubiquitinase activity. Specifically cleaves 'Lys-63'-linked polyubiquitin chains.	Involved in the regulation of cell survival, proliferation, and differentiation. Plays an important role in pathways activating NF-κB and is a negative Wnt regulator. Involved in modulation of cell polarization, migration and angiogenesis	Downregulation at mRNA and protein levels (tissue)	Relates to resistance to chemotherapeutic agents	Yes	[58]
MAGE-D4	Component of the MAGE family protein	Expressed during development and involved in regulation of cell survival, cell cycle progression and apoptosis	Over-expression at mRNA and protein levels (tissue)	Relates to HCC poor differentiation and invasion. Marks early recurrence and decrease overall survival	No data	[59]
EphA3	Member of the receptor of tyrosine kinase family	Involved in development, particularly of the nervous system. Binds to ephrin-A ligands (especially EFNA5). Regulates cell-cell adhesion, cytoskeletal organization, cell differentiation and migration	Over-expression at protein level (tissue)	Decrease in overall survival	potential	[60]
Flot-1	Member of the Flotillin subfamily	Acts as a scaffolding protein within caveolar membranes and mediates cell signalings. Involves in the formation of caveolae and caveolae-like vesicles to regulate membrane trafficking. Participates to cytoskeleton organization, cell adhesion and invasion	Over-expressed at mRNA and protein levels (tissue)	High levels relates to increase tumor size, invasion and short disease-free time and overall survival	No data	[61]

GPC3: Glypican-3; DKK1: Dickkopf-1; S100A4: S100 calcium-binding protein A4; SUOX: Sulfite oxidase; xCT: Cystine/glutamate transporter; GRK6: G protein-coupled receptor kinase 6; GPR87: G protein-coupled receptor 87; MT-1: Metallothionein 1; MT-2: Metallothionein 2; RAI3: Retinoic acid-induced protein 3; SSX2IP: Synovial sarcoma X break point 2 interacting protein; Ppm1d: Protein phosphatase magnesium-dependent 1d; BCL9: B-cell CLL/lymphoma 9 protein; IRF-1: Interferon regulatory factor-1; CDK4: Cyclin-dependent kinase 4; LASP-1: LIM and SH3 protein 1; PTP4A3: Protein tyrosine phosphatase type IVA member 3; PAK5: P21-activated kinase 5; hnRNP-L: Heterogeneous nuclear ribonucleo protein L; CYLD: Cylindromatosis (turban tumor syndrome); MAGE-D4: Melanoma-associated antigen D4; EphA3: Ephrin receptor A3; Flot-1: Flotillin-1.

Table 2 Micro-interfering RNA, miRBir and epigenetic markers of human hepatocellular carcinoma progression and poor prognosis

Molecule	Molecular category	Biological function	Expression levels in human HCC tissue	Clinical pathological indications	Target for therapy	Ref.
miRNA-18b	Belongs to Oncomir-1 or miR17-92 cluster	Involved in embryogenesis and spermatogenesis	Over-expressed	Poor differentiated HCC, high levels positively correlate with lower survival time	Potential	[70]
miRNA-372	Oncomir	Not well defined: it can act as an oncomir or as an anti-oncomir depending on the malignancy	Over-expressed	High levels positively correlate with advance TNM stage, shorter recurrence-free survival and overall survival	Potential	[74]
miRNA-650	Oncomir	Targets protein involved in cell proliferation and survival	Over-expressed	Higher levels in less than 60-year-old patients; levels positively correlate with HCC differentiation status	Potential	[75,76]
miRNA-100	Tumor suppressor	Regulates cell differentiation and survival	Down-regulated	Levels negatively correlate with tumor grade, metastasis and recurrence	Potential	[77]
miRNA-132	Tumor suppressor	Involved in the repression and control of inflammation and in neuronal morphogenesis	In HBV-HCC down-modulated by HBx protein causing promoter methylation	Levels of miR-132 inversely correlate with those of HBx in HBV-HCC patients. Early diagnosis of HBV-HCC	Potential	[80]
Ago2	Belongs to eIF2C/AGO subfamily	One of the effectors of RNA interference either at initiation or elongation phases, endowed with PIWI domain (endonuclease activity)	Overexpressed (mRNA and protein levels) and gene amplification	Can signed metastasis	No data	[83]
Dicer	Belongs to the RNaseIII family	Cleaves double-stranded RNA and pre-miRNA in the short (20-25 nt) active miRNA	Down-regulated	Shorter recurrence-free survival time	No data	[84]
TFPI2 gene	Member of the Kunitz-type serine protease inhibitors gene	The encoded protein inhibits plasmin-and trypsin-mediated activation of matrix metalloproteinases.	Up-regulation of gene promoter methylation (HCC tissues and serum)	TFPI2 methylation levels positively correlate with high grade TNM, improved detection of HCC in patients with AFP levels less than 400 μg/L	No data	[87]
Ten-eleven translocation protein 1 gene	Member of the TET family of methylcytosine dioxygenase enzymes gene	The encoded protein convert 5-methylcytosine to 5-hydroxymethylcytosine (5hmC) in various embryonic and adult tissues	Up-regulation of promoter methylation	Low levels of TET 1 and 5hmC relates to shorter overall survival	No data	[89]
MAT1A, MAT2A genes	Member of the SAM synthesizing isozymes genes	The encoded enzymes are essential to SAM biosynthesis. MAT1A is expressed in mature liver, whereas MAT2A is expressed by fetal liver, extrahepatic tissues or in HCC.	Up-regulation of MAT1A promoter methylation and down-regulation of MAT2A promoter methylation	Based on in cell culture and animal models studies, decreased MAT1A/MAT2A expression ratio has been proposed as s poor prognostic factor in HCC patients	No data	[90]

TFPI2: Tissue factor pathway inhibitor-2; Ago2: Argonaute-2; MAT1A: Methionine adenosyltransferase; HCC: Hepatocellular carcinoma; HBx: Hepatitis B virus x; SAM: S-adenosylmethionine.

Table 3 Some secreted factors from cellular components of tumor microenvironment and their contribution to hepatocellular carcinoma prognosis

Molecule	Molecular category	Biological function	Expression in human HCC	Clinico-pathological indications	Target for therapy	Ref.
Interleukin-4	Cytokine	Alternative activation of macrophages into M2 cells	Over-expressed (elevated level in serum)	Associated with extrahepatic metastasis	Not presented	[108]
TGF-β1	Multifunctional cytokine	Proto-oncogene/tumor suppressor	Over-expressed (elevated level in serum)	Decreased overall survival, increased risk of metastasis and recurrence, positively correlated with vascular invasion, higher PVT, higher overall invasiveness	Potential	[123]
FGF19	Growth factor	Activation of mitogen-activated protein kinase; activation of Wnt/β-catenin pathway	Over-expressed in tissue	Decreased disease-free and overall survival, increased risk of metastasis, positively correlated with vascular invasion	Potential	[117]
Fox C1	FOX transcription factor family member	Induction of EMT, promotion of epithelial cell migration via inhibition of E-Cadherin transcription through transactivation of SNAI1 expression	Over-expressed in tissue	Increased risk of metastasis and recurrence, increased tumor size and number, poorer tumor differentiation, poorer TNM stage, higher vascular invasion	Potential	[130,131]
Lcn2	Member of lipocalin protein family	Inhibition of epithelial-to-mesenchymal transition; suppression of JNK and PI3K/Akt signaling pathways, down regulation of Twist1	Over-expressed in tissue	Positively correlated with poor prognosis and high risk HCC, decreased overall survival, however, elevated levels of Lcn2 is correlated with the inhibition of EMT in vitro and in vivo	Not presented	[134,135]
uPA	Serine protease	Plasminogen activation, extracellular proteolytic activity	Over-expressed in tissue	Decreased disease-free and overall survival, increased HCC invasion, poorer pathological grade	Potential	[144,146]
uPAR	Urokinase receptor	ECM degradation, migration, invasion and metastasis	Over-expressed in tissue	Decreased disease-free and overall survival, higher risk of HCC invasion, portal cancer embolus and tumor metastasis, poorer tumor cell differentiation	Potential	[145,146]
PAI-1	Protease inhibitor	Regulating extracellular matrix degradation	Over-expressed in tissue	Decreased disease-free and overall survival, increased risk of metastasis and recurrence, increased regional invasion, poorer pathological grade	Not presented	[144,146]
VEGF	Growth factor	Regulation of vascularization and angiogenesis	Over-expressed in tissue, elevated level in serum	Poorer overall survival, increased risk of metastasis and recurrence, higher vascular invasion	Potential	[156]
HIF-1α	Subunit of HIF-1 transcription factor	Activating the transcription of a variety of genes involved in angiogenesis, survival pathways, immune evasion, invasion, and metastasis	Over-expressed in tissue	Decreased disease-free (and overall) survival, increased risk of metastasis, higher vascular invasion	Potential	[162,163]

PVT: Portal vein thrombosis; EMT: Epithelial-to-mesenchymal transition; TNM: Tumor node metastasis; FGF19: Fibroblast growth factor 19; Fox C1: Forkhead box C1; Lcn2: Lipocalin-2; uPA: Urokinase plasminogen activator; uPAR: Urokinase plasminogen activator receptor; PAI-1: Plasminogen activator inhibitor type-1; VEGF: Vascular endothelial growth factor; HIF-1α: Hypoxia-inducible factor-1α.