A short review of the molecular events leading to the development of HCC due to different etiological factors , and its impact on future therapeutic modalities .
2. BY
Prof.Dr/ Ekbal Abo Hashem
Professor of Clinical Pathology
Mansoura University- Faculty of Medcine
3. o Introduction-Disease burden.Introduction-Disease burden.
o Pathogenesis of HCC : causal relationshipPathogenesis of HCC : causal relationship
o Molecular genetics of HCC :Molecular genetics of HCC :
-Chromosomal abnormalities-Chromosomal abnormalities
-Deregulation of signaling pathways-Deregulation of signaling pathways
o Epigenetic mechanisms :Epigenetic mechanisms :
DNA methylationDNA methylation –– Histone modificationHistone modification –– miRNAsmiRNAs
deregulationderegulation
o Targeted therapy .Targeted therapy .
o Future prospects .Future prospects .
LECTURE OUTLINELECTURE OUTLINE
4. HepatocellularHepatocellular
carcinomacarcinomaIntroduction and dieases burdenIntroduction and dieases burden
Hepatocellular carcinoma is the sixth mostHepatocellular carcinoma is the sixth most
common malignancy and the third most commoncommon malignancy and the third most common
cause of cancer deaths world wide. The majorcause of cancer deaths world wide. The major
risk factors include chronic viral infection (HBVrisk factors include chronic viral infection (HBV
and HCV), alcoholic/nonalcoholic liver disease,and HCV), alcoholic/nonalcoholic liver disease,
environmental carcinogens (i.e. aflatoxin B1environmental carcinogens (i.e. aflatoxin B1
(AFB), and inherited genetic disorders (Wilson's(AFB), and inherited genetic disorders (Wilson's
disease, hemochromatosis,disease, hemochromatosis, αα-1-antitrypsin-1-antitrypsin
deficiency, and tyrosinemia ) .deficiency, and tyrosinemia ) .
5. With a few exceptions, HCC always develops inWith a few exceptions, HCC always develops in
the setting of chronic hepatitis or cirrhosis, inthe setting of chronic hepatitis or cirrhosis, in
which there is continuous inflammation andwhich there is continuous inflammation and
regeneration of hepatocytes. The non-randomregeneration of hepatocytes. The non-random
accumulated genetic alterations oraccumulated genetic alterations or
chromosomal aberrations during the processeschromosomal aberrations during the processes
of inflammation, regeneration, and cirrhosisof inflammation, regeneration, and cirrhosis
lead to the development of HCC .lead to the development of HCC .
6. In Egypt ,HCC is the second most commonIn Egypt ,HCC is the second most common
cancer in men and the 6th most common cancercancer in men and the 6th most common cancer
in women. Disease burden is increasing with ain women. Disease burden is increasing with a
doubling of the incidence rate in the past 10doubling of the incidence rate in the past 10
years. This could be atrtributed to variousyears. This could be atrtributed to various
factors e.g biological (hepatitis B and Cfactors e.g biological (hepatitis B and C
infections ) ,environmental (aflatoxin ),endemicinfections ) ,environmental (aflatoxin ),endemic
infections (Schistosomiasis ) ,occupationalinfections (Schistosomiasis ) ,occupational
exposure to chemicals such as pesticides,exposure to chemicals such as pesticides,
besides other factors e.g obesity ,DM ,NAFLDbesides other factors e.g obesity ,DM ,NAFLD
,tobacco smoking ,etc,tobacco smoking ,etc……Estimates of the burdenEstimates of the burden
of cancer caused by these factors provide anof cancer caused by these factors provide an
opportunity for prevention .opportunity for prevention .
7. As in many developing countries , Egypt isAs in many developing countries , Egypt is
undergoing an epidemiologic transition . Theundergoing an epidemiologic transition . The
complex etiology of HCC should enable policycomplex etiology of HCC should enable policy
makers to create targeted and more efficientmakers to create targeted and more efficient
prevention and screening programs.prevention and screening programs.
8. Pathogenesis of HCC:Pathogenesis of HCC:
causal relationshipcausal relationship
The development of human HCCs is a multi-The development of human HCCs is a multi-
step process, from macroregen-erative nodulesstep process, from macroregen-erative nodules
(MRN)/low-grade dysplasia (LGD), to high-(MRN)/low-grade dysplasia (LGD), to high-
grade dysplasia (HGD), to early HCC . After angrade dysplasia (HGD), to early HCC . After an
initial exposure or insult by carcinogens, it mayinitial exposure or insult by carcinogens, it may
take years or decades for humans totake years or decades for humans to
accumulate the necessary genetic andaccumulate the necessary genetic and
epigenetic damages necessary forepigenetic damages necessary for
preneoplastic diseases to develop into HCC.preneoplastic diseases to develop into HCC.
9.
10. These genetic damages or changes include:These genetic damages or changes include:
The up-regulation of growth factors, inactivation of tumor suppressorThe up-regulation of growth factors, inactivation of tumor suppressor
genes, aberrant methylation, and microsatellite instability.genes, aberrant methylation, and microsatellite instability.
Up-regulation of TGF-α and IGF-2 are sequelae of degeneration dueUp-regulation of TGF-α and IGF-2 are sequelae of degeneration due
to chronic inflammation, viral trans-activation, and hepatocellularto chronic inflammation, viral trans-activation, and hepatocellular
repair and regeneration .repair and regeneration .
Aberrant hypo- or hypermethylation observed in chronic hepatitis andAberrant hypo- or hypermethylation observed in chronic hepatitis and
cirrhosis as well as HCC is due to increases in DNA methyltrans-cirrhosis as well as HCC is due to increases in DNA methyltrans-
ferases (DNMT) associated with chronic hepatitis and cirrhosisferases (DNMT) associated with chronic hepatitis and cirrhosis
Loss of heterozygosity (LOH) and microsatellite instability occur inLoss of heterozygosity (LOH) and microsatellite instability occur in
preneoplastic lesions and HCC.preneoplastic lesions and HCC.
11. Common molecular disturbances that are universal to allCommon molecular disturbances that are universal to all
liver cancers on top of the more specific mechanisms:liver cancers on top of the more specific mechanisms:
1.1. Disturbances of cell cycle regulation results fromDisturbances of cell cycle regulation results from
p53 point mutations and LOH, silencing of p16 orp53 point mutations and LOH, silencing of p16 or
Retinoblastoma genes or overexpression of cyclinRetinoblastoma genes or overexpression of cyclin
D1 .D1 .
2. Aberrant angiogenesis resulting from autocrine /2. Aberrant angiogenesis resulting from autocrine /
paracrine secretion of VEGF, PDGF or angiopoietin-2.paracrine secretion of VEGF, PDGF or angiopoietin-2.
3. Evasion of apoptosis due to de- regulation of3. Evasion of apoptosis due to de- regulation of
intrinsic or extrinsic apoptotic pathways.intrinsic or extrinsic apoptotic pathways.
4. Reactivation of TERT which ensures limitless4. Reactivation of TERT which ensures limitless
replicative potential.replicative potential.
12. Chromosomal abnormalitiesChromosomal abnormalities
Loss of heterozygosity (LOH) and microsatelliteLoss of heterozygosity (LOH) and microsatellite
instability occur in preneoplastic lesions andinstability occur in preneoplastic lesions and
HCC. The frequently deleted chromosome regionsHCC. The frequently deleted chromosome regions
by LOH in HCC contain many tumor suppressorby LOH in HCC contain many tumor suppressor
genes and some oncogenes (p53, Rb, p 16,genes and some oncogenes (p53, Rb, p 16,
PTEN, DLC1 and IGF1R ).PTEN, DLC1 and IGF1R ).
Gain of chromosome 10q is unique to HCV-Gain of chromosome 10q is unique to HCV-
related-HCC, while loss of 4q and 16q and gain ofrelated-HCC, while loss of 4q and 16q and gain of
11q are seen in HBV positive cases.11q are seen in HBV positive cases.
13.
14.
15. Molecular basis of pathogenesisMolecular basis of pathogenesis
HBV-associated HCC may occur in the liver withoutHBV-associated HCC may occur in the liver without
cirrhosis and is distinct from HCC related to HCV andcirrhosis and is distinct from HCC related to HCV and
other etiologies. Although it is still under debate, HBVother etiologies. Although it is still under debate, HBV
may have both direct and indirect oncogenic effects onmay have both direct and indirect oncogenic effects on
hepatocytes:hepatocytes:
One direct effect is that viral DNA may be integrated intoOne direct effect is that viral DNA may be integrated into
the hepatocyte genome, causing disruption ofthe hepatocyte genome, causing disruption of
chromosomal stability or tumor suppressor genes andchromosomal stability or tumor suppressor genes and
activation of proto-oncogene.activation of proto-oncogene.
Another direct oncogenic effect may be attributable to theAnother direct oncogenic effect may be attributable to the
154-amino acid (16.5-kDa) viral protein HBx, which may154-amino acid (16.5-kDa) viral protein HBx, which may
transactivate or up-regulate a variety of viral and cellulartransactivate or up-regulate a variety of viral and cellular
genes .genes .
16.
17. HBx protein amplifies TGF-HBx protein amplifies TGF- ββ signaling throughsignaling through
direct interaction with Smad4, and both directdirect interaction with Smad4, and both direct
and indirect interactions with DNA repair proteinand indirect interactions with DNA repair protein
UVDDB, tumor suppressor proteins (p53 andUVDDB, tumor suppressor proteins (p53 and APCAPC
gene product), cell cycle regulators, growthgene product), cell cycle regulators, growth
factors and receptor genes, cytokines, genesfactors and receptor genes, cytokines, genes
involved in apoptosisinvolved in apoptosis ,, proteasome subunitsproteasome subunits ,, andand
NF-kB .NF-kB . .Indirectly, HBV infection causes liver.Indirectly, HBV infection causes liver
cell injury mediated by cellular immunecell injury mediated by cellular immune
responses, resulting in carcinogenesis byresponses, resulting in carcinogenesis by
promoting cell death, proliferation, and geneticpromoting cell death, proliferation, and genetic
mutations.mutations.
18. In chronic HBV infection, non-random DNAIn chronic HBV infection, non-random DNA
integration of HBV leads to promoter activationintegration of HBV leads to promoter activation
of oncogenes, DNA rearrangement andof oncogenes, DNA rearrangement and
chromosomal instability. Chronic inflammationchromosomal instability. Chronic inflammation
leads to oxidative stress, with genomic andleads to oxidative stress, with genomic and
DNA damage.DNA damage.
19.
20. The carcinogenesis of HCV infection-associatedThe carcinogenesis of HCV infection-associated
HCC differs from that related to HBV infection,HCC differs from that related to HBV infection,
and is mainly due to the indirect effects of viraland is mainly due to the indirect effects of viral
infection. HCV viral RNA is never integratedinfection. HCV viral RNA is never integrated
into the genome of hepatocytesinto the genome of hepatocytes but HCVbut HCV
infection may cause the accumulation ofinfection may cause the accumulation of
genetic abnormalities during the degeneration-genetic abnormalities during the degeneration-
regeneration process.regeneration process.
21. Viral proteins, viral core protein in particular,Viral proteins, viral core protein in particular,
may interfere with intracellular signalingmay interfere with intracellular signaling
pathways (activating TNF-pathways (activating TNF- αα receptor, Raf-1receptor, Raf-1
kinase, and NF-kkinase, and NF-k ββ pathways, resulting inpathways, resulting in
inhibition of TNF-inhibition of TNF- αα and Fas-and Fas-
mediatedmediated apoptosis) and interact with the hostapoptosis) and interact with the host
immune system .immune system .
22. Chronic HCV infection causes an increase ofChronic HCV infection causes an increase of
TGF-alpha and IGF-2 leading to acceleratedTGF-alpha and IGF-2 leading to accelerated
proliferation. HCV core protein acts on Wntproliferation. HCV core protein acts on Wnt
ligand, transactivates Ras signaling andligand, transactivates Ras signaling and
inactivates p53.inactivates p53.
23.
24. Exposure to food contaminated by Aflatoxin B1Exposure to food contaminated by Aflatoxin B1
(AFB1), a fungal metabolite produced by(AFB1), a fungal metabolite produced by
Asperigillus flavus and related fungi, isAsperigillus flavus and related fungi, is
associated with an increased incidence ofassociated with an increased incidence of
HCC .HCC .
AFB1 and its metabolite may result in a highAFB1 and its metabolite may result in a high
frequency of mutations affecting 249serfrequency of mutations affecting 249ser inin
thethe p53p53 tumor suppressor gene . Moreover,tumor suppressor gene . Moreover,
there is a dose-dependent relationshipthere is a dose-dependent relationship
betweenbetween p53p53 249ser249ser mutation load in cells andmutation load in cells and
the intake of AFB1 in non-tumorous liverthe intake of AFB1 in non-tumorous liver
tissue .tissue .
25. The molecular mechanism for carcinogenesisThe molecular mechanism for carcinogenesis
associated with Wilson's disease, primaryassociated with Wilson's disease, primary
hemochromatosis, and other genetic diseaseshemochromatosis, and other genetic diseases
affecting the liver is also related to poorlyaffecting the liver is also related to poorly
controlled immune responses to copper , iron ,controlled immune responses to copper , iron ,
or other metabolite accumulation.or other metabolite accumulation.
This immune response results in inflammationThis immune response results in inflammation
and generates oxidative free radicals thatand generates oxidative free radicals that
damage human DNA and cause genomicdamage human DNA and cause genomic
alterations in hepatocyte genes association withalterations in hepatocyte genes association with
tumor suppression, cell cycle regulation, DNAtumor suppression, cell cycle regulation, DNA
repair, and apoptosis .repair, and apoptosis .
26.
27. Though specific genetic alterations depend onThough specific genetic alterations depend on
HCC etiology, the main proteins affected includeHCC etiology, the main proteins affected include
cell membrane receptors (in particular tyrosinecell membrane receptors (in particular tyrosine
kinase receptors) as well as proteins involved inkinase receptors) as well as proteins involved in
cell signaling (specifically Wnt/beta-catenin,cell signaling (specifically Wnt/beta-catenin,
Ras/Raf/MEK/ERK and PI3K/Akt/mTORRas/Raf/MEK/ERK and PI3K/Akt/mTOR
pathways), cell cycle regulation (i.e. p53,pathways), cell cycle regulation (i.e. p53,
p16/INK4, cyclin/cdk complex), invasivenessp16/INK4, cyclin/cdk complex), invasiveness
(EMT, TGF-(EMT, TGF-β)β) and DNA metabolism.and DNA metabolism.
28.
29.
30. EPIGENETICS AND
HCC
Epigenetics is defined as heritable states of
gene expression without altering DNA
sequences. Epigenetic mechanisms
encompass genomic DNA modifications
(methylation of DNA cytosine bases),
chemical modifications of histone tails,
and non-coding miRNA regulation.
31.
32.
33. During cell division, these epigenetic modifications are
passed down faithfully to daughter cells to maintain
“cellular memory”. DNA methyltransferases (DNMTs)
catalyze the addition of methyl groups (CH3) to the 5’
cytosine nucleotides.
DNA Methylation
Mechanistically, DNA methylation leads to transcriptional
gene silencing in two ways:
First, methylation at CpG sites sterically hinders
accessibility of transcription factors to their cognate binding
sites on respective gene promoters .
The second mechanism involves direct binding of methyl
CpG binding domain (MBD)-containing proteins to the
methylated DNA, causing transcription repression
34. HISTONE MODIFICATIONS IN HCCHISTONE MODIFICATIONS IN HCC
Within the chromosome, DNA is packaged intoWithin the chromosome, DNA is packaged into
chromatin where the DNA coils around anchromatin where the DNA coils around an
octamer of histones. One hundred and forty-fiveoctamer of histones. One hundred and forty-five
base pairs of DNA are wrapped around thebase pairs of DNA are wrapped around the
histone octamer, comprising H2A, H2B, H3 andhistone octamer, comprising H2A, H2B, H3 and
H4, forming the repeating unit of chromatin, theH4, forming the repeating unit of chromatin, the
nucleosome. Histone tails protruding out of thenucleosome. Histone tails protruding out of the
nucleosome are targets of post-translationalnucleosome are targets of post-translational
modifications, including acetylation andmodifications, including acetylation and
methylation of lysine (K) and arginine (R)methylation of lysine (K) and arginine (R)
residues, phosphorylation of serine (S) andresidues, phosphorylation of serine (S) and
threonine (T) residues, and ubiquitination ofthreonine (T) residues, and ubiquitination of
lysine residues.lysine residues.
35. These modifications can turn transcription ofThese modifications can turn transcription of
genes on or off, and are therefore key playersgenes on or off, and are therefore key players
in establishing the gene expression patterns ofin establishing the gene expression patterns of
cells by adjusting the tightness of DNA boundcells by adjusting the tightness of DNA bound
to histones, thereby affecting accessibility ofto histones, thereby affecting accessibility of
transcription factors.transcription factors.
36.
37.
38.
39. Histone acetylation is controlled by two families of enzymes: histone
acetyltransferases (HATs) that “write” the acetyl mark. Acetylation
counteracts the positive charge of histones, thereby loosening the tight
interaction between histones and DNA.
Conversely, histone deacetylases (HDACs) “erase” the acetyl group,
resulting in tight coiling of DNA around the histones, leading to the
transcriptionally inactive or closed chromatin state.
In contrast, histone methylation is associated with either
transcriptionally active or closed chromatin, depending on which
histone or which lysine residue is modified.
40. Roles of histone modifications in the regulation of gene
transcription.
41. miRNAs IN HCCmiRNAs IN HCC
MicroRNAs (miRNAs) are non-coding small RNA
(ncRNA) molecules that are 20-23 nucleotides in length.
They play important regulatory roles in plants and
animals by targeting mRNAs for cleavage or translational
repression.
More than 1000 miRNAs have been identified to date.
Through their roles in post-transcriptional gene
regulation, miRNAs regulate diverse cellular functions
including proliferation, differentiation, apoptosis, cell fate,
and plasticity.
48. Targeted therapy, or molecularly targeted therapy is oneTargeted therapy, or molecularly targeted therapy is one
of the major modalities of medical treatment for cancer. Itof the major modalities of medical treatment for cancer. It
blocks the growth of cancer cells by interfering with allblocks the growth of cancer cells by interfering with all
rapidly dividing cells through molecular pathways ofrapidly dividing cells through molecular pathways of
carcinogenesis, without affecting normal cells. They arecarcinogenesis, without affecting normal cells. They are
a cornerstone of (precision medicine) that usesa cornerstone of (precision medicine) that uses
information about a personinformation about a person’’s genes and proteins tos genes and proteins to
prevent, diagnose, and treat disease.prevent, diagnose, and treat disease.
Targeted therapy
49. Recently introduced molecular targetedRecently introduced molecular targeted
therapies are specific for groups of HCCs withtherapies are specific for groups of HCCs with
similar genetics. The targeted therapy aims tosimilar genetics. The targeted therapy aims to
inactivate activated oncogenes, recover tumorinactivate activated oncogenes, recover tumor
suppressor genes, or repair other genes andsuppressor genes, or repair other genes and
molecules related to HCC development, therebymolecules related to HCC development, thereby
correcting abnormal genes or functions as wellcorrecting abnormal genes or functions as well
as biological behavior.as biological behavior.
50. Some of these targeted therapies, such as monoclonalSome of these targeted therapies, such as monoclonal
antibodies, small molecules and antisense drugs, haveantibodies, small molecules and antisense drugs, have
reached phase II and III clinical trials for therapeutic use,reached phase II and III clinical trials for therapeutic use,
and many have been shown to be effective. Sorafenib,and many have been shown to be effective. Sorafenib,
an oral multikinase inhibitor of vascular endothelialan oral multikinase inhibitor of vascular endothelial
growth factor receptor (VEGFR) and Ras kinase, hasgrowth factor receptor (VEGFR) and Ras kinase, has
been approved by the FDA as a molecularly targetedbeen approved by the FDA as a molecularly targeted
anticancer agent. Some other agents targeting similaranticancer agent. Some other agents targeting similar
genes or molecules are being tested in preclinical andgenes or molecules are being tested in preclinical and
clinical trials for HCC.clinical trials for HCC.
51. 1.1. Anti-Epidermal growth factorAnti-Epidermal growth factor
receptor (anti-EGFR) therapy.receptor (anti-EGFR) therapy.
2.2. Anti-vascular endothelial growthAnti-vascular endothelial growth
factor (antiangiogenesis)factor (antiangiogenesis)
3.3. Multikinase targetsMultikinase targets
Main lines of drugs include
52. Epigenetic drugsEpigenetic drugs
In contrast to conventional or molecularly-targetedIn contrast to conventional or molecularly-targeted
therapies for inhibiting dysregulated genes ortherapies for inhibiting dysregulated genes or
signaling pathways, epigenetic drugs provide ansignaling pathways, epigenetic drugs provide an
alternative approach by reversing the methylationalternative approach by reversing the methylation
status and histone modifications of aberrantlystatus and histone modifications of aberrantly
expressed genes. There are currently four FDA-expressed genes. There are currently four FDA-
approved epigenetic drugs, including two DNMTapproved epigenetic drugs, including two DNMT
inhibitors, 5-azacytidine and decitabine, and twoinhibitors, 5-azacytidine and decitabine, and two
HDAC inhibitors, vorinostat and valporic acid. TheseHDAC inhibitors, vorinostat and valporic acid. These
drugs have been also successful in treatingdrugs have been also successful in treating
hematological cancers, specifically myelodysplastichematological cancers, specifically myelodysplastic
syndrome,syndrome,
53. FUTURE PROSPECTS:FUTURE PROSPECTS:
Current data of the molecularCurrent data of the molecular
pathogenesis of HCC are still verypathogenesis of HCC are still very
elementary and represent only aelementary and represent only a
minor part of the key hits requiredminor part of the key hits required
for HCC initiation andfor HCC initiation and
progression.progression.
Novel high-throughputNovel high-throughput
technologies are needed totechnologies are needed to
identify other relevant geneticidentify other relevant genetic
pathways and provide a molecularpathways and provide a molecular
54. Experimental models and cancer HCCExperimental models and cancer HCC
stem cell lines should be developed to teststem cell lines should be developed to test
new drugs.new drugs.
Biomarker approaches should beBiomarker approaches should be
developed to identify treatmentdeveloped to identify treatment
responders to specific drugs, the first stepresponders to specific drugs, the first step
towards personalized medicine.towards personalized medicine.
55. Molecular approaches, such as geneMolecular approaches, such as gene
expression microarray and SNP array,expression microarray and SNP array,
should be used to develop a newshould be used to develop a new
classification system for HCCs that betterclassification system for HCCs that better
predicts clinical outcome and facilitatespredicts clinical outcome and facilitates
targeted molecular therapy.targeted molecular therapy.
56. Further ReadingFurther Reading
Li MaLi Ma,, Mei-Sze ChuaMei-Sze Chua,, Ourania AndrisaniOurania Andrisani, and, and Samuel SoSamuel So, Epigenetics in, Epigenetics in
hepatocellular carcinoma: An update and future therapy perspectives. World Jhepatocellular carcinoma: An update and future therapy perspectives. World J
Gastroenterol. 2014 Jan 14; 20(2): 333Gastroenterol. 2014 Jan 14; 20(2): 333 ––345.345.
El-Serag HB. Hepatocellular carcinoma.El-Serag HB. Hepatocellular carcinoma. N Engl J Med.N Engl J Med. 2011;365:11182011;365:1118––1127.1127.
Egger G, Liang G, Aparicio A, Jones PA. Epigenetics in human disease andEgger G, Liang G, Aparicio A, Jones PA. Epigenetics in human disease and
prospects for epigenetic therapy.Nature.prospects for epigenetic therapy.Nature. 2004;429:4572004;429:457––463.463.
Herceg Z, Paliwal A. Epigenetic mechanisms in hepatocellular carcinoma: howHerceg Z, Paliwal A. Epigenetic mechanisms in hepatocellular carcinoma: how
environmental factors influence the epigenome.environmental factors influence the epigenome. Mutat Res.Mutat Res. 2011;727:552011;727:55––61.61.
Wang XW, Heegaard NH, Orum H. MicroRNAs in liverWang XW, Heegaard NH, Orum H. MicroRNAs in liver
disease.disease. Gastroenterology.Gastroenterology. 2012;142:14312012;142:1431––1443.1443.
Yang H, Fang F, Chang R, Yang L. MicroRNA-140-5p suppresses tumor growthYang H, Fang F, Chang R, Yang L. MicroRNA-140-5p suppresses tumor growth
and metastasis by targeting transforming growth factor β receptor 1 and fibroblastand metastasis by targeting transforming growth factor β receptor 1 and fibroblast
growth factor 9 in hepatocellular carcinoma.growth factor 9 in hepatocellular carcinoma. Hepatology.2013;58:205Hepatology.2013;58:205 ––217.217.
Chan SL, Yeo W. Targeted therapy of hepatocellular carcinoma: present andChan SL, Yeo W. Targeted therapy of hepatocellular carcinoma: present and
future.future. J Gastroenterol Hepatol.J Gastroenterol Hepatol. 2012;27:8622012;27:862––872.872.
Lachenmayer A, Toffanin S, Cabellos L, Alsinet C, Hoshida Y, Villanueva A,Lachenmayer A, Toffanin S, Cabellos L, Alsinet C, Hoshida Y, Villanueva A,
Minguez B, Tsai HW, Ward SC, Thung S, et al. Combination therapy forMinguez B, Tsai HW, Ward SC, Thung S, et al. Combination therapy for
hepatocellular carcinoma: additive preclinical efficacy of the HDAC inhibitorhepatocellular carcinoma: additive preclinical efficacy of the HDAC inhibitor
panobinostat with sorafenib.panobinostat with sorafenib. J Hepatol.J Hepatol. 2012;56:13432012;56:1343––1350.1350.
Ozen C, Yildiz G, Dagcan AT, Cevik D, Ors A, Keles U, Topel H, Ozturk M.Ozen C, Yildiz G, Dagcan AT, Cevik D, Ors A, Keles U, Topel H, Ozturk M.
Genetics and epigenetics of liver cancer.Genetics and epigenetics of liver cancer. N Biotechnol.N Biotechnol. 2013;30:3812013;30:381––384.384.
Jain SH, Singhal SH, Lee P, Xu R, Molecualr genetics of hepatocellularJain SH, Singhal SH, Lee P, Xu R, Molecualr genetics of hepatocellular
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