review article mtor in viral hepatitis and hepatocellular...

Review ArticlemTOR in Viral Hepatitis and Hepatocellular CarcinomaFunction and Treatment

Zhuo Wang Wei Jin Hongchuan Jin and Xian Wang

Department of Medical Oncology Institute of Clinical Science Sir Runrun Shaw Hospital School of Medicine Zhejiang UniversityHangzhou China

Correspondence should be addressed to Xian Wang wangxzju163com

Received 8 November 2013 Accepted 7 March 2014 Published 2 April 2014

Academic Editor Qing He

Copyright copy 2014 Zhuo Wang et al This is an open access article distributed under the Creative Commons Attribution Licensewhich permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

As the fifth most common cancer in men and the eighth most common cancer in women hepatocellular carcinoma (HCC) isthe leading cause of cancer-related deaths worldwide with standard chemotherapy and radiation being minimally effective inprolonging survival Virus hepatitis particularly HBV and HCV infection is the most prominent risk factor for HCC developmentMammalian target of rapamycin (mTOR) pathway is activated in viral hepatitis and HCC mTOR inhibitors have been testedsuccessfully in clinical trials for their antineoplastic potency and well tolerability Treatment with mTOR inhibitor alone or incombination with cytotoxic drugs or targeted therapy drug scan significantly reduces HCC growth and improves clinical outcomeindicating that mTOR inhibition is a promising strategy for the clinical management of HCC

1 Introduction

Hepatocellular carcinoma (HCC) is amalignant tumorwhoseincidence is increasing in many countries It is the fifth mostcommon cancer in men and the eighth most common cancerin women HCC is the leading cause of cancer-related deathsworldwide with standard chemotherapy being minimallyeffective in prolonging survival [1]

Among many factors such as environmental pollutionfatty liver and excessive alcohol consumption virus hepatitisparticularly HBV and HCV infection has been consideredas the most important high risk factor of HCC especially inAsian countries At the molecular level mammalian target ofrapamycin (mTOR) pathway was found to be associated withHCC development including chronic viral hepatitis [2 3]Inhibitors of mTORwere thus postulated to be prominent forthe clinical treatment of HCC

2 mTOR

21 Structure of mTOR Complex mTOR is a member ofPI3K-related protein kinases (PIKK)The structure of mTOR

is similar to other PIKK family members The amino ter-minus of mTOR is a cluster of HEAT (Huntingtin Elon-gation factor 3 A subunit of protein phosphatase 2A andTOR1) repeats followed by FAT (FRAP ATM and TRRAP)domain FKBP12-rapamycin binding (FRB) domain SerThrkinase catalytic domain and the carboxyl-terminal FAT(FATC) domain HEAT domain can mediate protein-proteininteractions and FRB domain is a conserved 11 kDa regionnecessary for the binding of rapamycin and regulatory-associated protein of mTOR (RAPTOR) [4]

According to different subunits mTOR can be formed astwo kinds of complexes mTORC1 and mTORC2 (Figure 1)Both mTOR complexes contain mTOR DEP domain-containingmTOR-interacting protein (DEPTOR) andmam-malian lethal with SEC13 protein 8 (mLST8) The uniquecomponents of mTORC1 are regulatory-associated protein ofmTOR (RAPTOR) and proline-rich Akt substrate of 40Kda(PRAS40) mTORC2 possesses rapamycin-insensitive com-panion of mTOR (RICTOR) protein observed with RICTOR(PROTOR) and mammalian stress-activated map kinase-interacting protein 1 (mSIN1) Among them PRAS40 is anegative regulator of mTOR and has a conserved leucine

Hindawi Publishing CorporationBioMed Research InternationalVolume 2014 Article ID 735672 9 pageshttpdxdoiorg1011552014735672

2 BioMed Research International

mTOR

RAPTORmLST8DEPTOR

PRAS40mSIN1

RICTORPROTOR

mTORmLST8DEPTOR

mTORC1 mTORC2

Figure 1 The structure of mTORC1 and mTORC2 The coremTOR machinery consists of mTOR DEPTOR and mLST8 Thecombination of core mTOR machinery with different proteinsconstitutes mTOR1 and mTORC2

charge domain (LCD) which can be phosphorylated byAKT [5 6] mLST8 can mediate protein-protein interactionswhile mSIN1 contains a Ras-binding domain (RBD) and apleckstrin homology which can interact with phospholipidCurrently the structures of RICTOR and PROTOR are stillnot clear

Rapamycin can inhibit the mTORC1 but not mTORC2because rapamycin binds with FKBP12 to disrupt the inter-action of mTOR with RAPTOR but not RICTOR [7ndash9] Therapamycin-induced dissociation of mTOR from RAPTOReventually prevents interaction of the mTOR with a numberof substrates [10 11] However long-term rapamycin treat-ment can inhibit mTORC2 [12] This effect may involve thechanges of intracellular pool of mTOR and thus reduce theassembly of mTORC2

22 Regulation of mTOR Activation mTORC1 can be acti-vated by diverse factors such as growth factors vari-ous cytokines Toll-like receptor ligands cell energy levelshypoxia and DNA damage The activation of mTORC1 playsan important role in protein synthesis ribosome biogenesisand autophagy Activated mTORC1 can phosphorylate thedownstream signalingmolecules including S6K1 or RPS6KN1(ribosomal protein S6 kinase 70 kDa polypeptide 1) andeukaryotic translation initiation factor-binding protein 1 (4E-BP1) Activation of S6K1 can promote the expression of ribo-somal protein and translation regulating protein to regulateprotein syntheses Nonphosphorylated 4E-BP1 can bind toeIF-4E to inhibit mRNA translation Once phosphorylatedby active mTOR 4E-BP1 are dissociated from eIF-4E sothat eIF-4E can bind to other translation initiation factorsto initiate protein translation [13 14] Tuberous sclerosiscomplex 1- (TSC1-) TSC2 tumor suppressor complex is anegative regulator of mTOR As a GTP activating protein(GAP) TSC2 or tuberin inactivates Ras homologue enrichedin brain (Rheb) which can directly bind to and activatemTOR TSC1 or hamartin does not have a GAP domain butit acts as a stabilizer of TSC2 by preventing it from degra-dation The activity of TSC1-TSC2 is regulated by proteinphosphorylation Activated PI3K-Akt signaling can phos-phorylate and inhibit TSC1-TSC2 while LKB1-AMPK canactivate TSC1-TSC2 by phosphorylation at different residues(Figure 2) [15 16]

The activation ofmTORC1 can be regulated by several fac-tors through signaling pathways including PI3KAktmTORLKB1AMPKmTOR andMAPK pathway Once activated byextracellular signals such as growth factors andnutrient PI3Kcan phosphorylate PIP2 to form PIP3 [17] As a result Aktand its activator phosphoinositide-dependent protein kinases1 (PDK1) translocate to the plasma membrane by binding toPIP3 When phosphorylated at Thr308 and Ser473 Akt canactivate mTOR by inhibiting TSC1-TSC2 [18] In additionAkt can suppress PRAS40 by phosphorylation to eliminateits inhibition on mTORC1

Studies have shown that acids metabolism in mam-malian cells is adjusted by LKB1AMPKmTOR signalpathway mTOR pathway can be activated by adenosinemonophosphate-activated protein kinases (AMPKs) that inturn can be activated by LKB1 AMPKs change their con-formation in response to the intracellular AMPATP ratioWhen AMPATP ratio drops LKB1 can bind to AMPKsubunit to activate AMPKby phosphorylationThe activationof AMPK will inhibit mTOR activation reduce proteinsynthesis and inhibit cell proliferation [19]

In addition mTOR activity can be regulated in responseto the availability of amino acids [20 21] Ras-related(Rag) GTPases a family of four related small GTPase areresponsible for amino acid-regulated mTOR activity [22 23]Rag GTPases interact with the RAPTOR the subunit ofmTORC1 in an amino acid-dependent manner allowinginteraction of mTORC1 with Rheb The Rags exist as obligateheterodimers Depletion of either heterodimer partners caninhibit mTORC1 while overexpression of the heterodimerwill rescue mTORC1 from suppression by amino acid with-drawal Some specific amino acids may play distinct rolesin the regulation of mTOR activity For example glutaminein combination with leucine activates mammalian TORC1(mTORC1) by enhancing glutaminolysis and 120572-ketoglutarateproduction [24] Inhibition of glutaminolysis prevented acti-vation of RagB and mTORC1 while constitutively active Ragheterodimer activatedmTORC1 in the absence of glutaminol-ysis Conversely enhanced glutaminolysis or a cell-permeable120572-ketoglutarate analog stimulated lysosomal translocationand activation of mTORC1

3 mTOR and Viral Hepatitis

31 mTOR and HBV Infection Chronic HBV infection is acrucial factor for the development of HCC In HepG2 cellsHBV RNA transcription and subsequent DNA replicationwere inhibited by the expression of a constitutively activeAkt1 [25] This inhibition of HBV gene transcription seemsto be mediated by mTOR activation since rapamycin canabolish this inhibition Furthermore inhibitors of PI3K Aktand mTOR can increase the transcription of viral RNA aswell as the replication of HBV DNA in HBV-overexpressingcells Consistently expression of HBsAg was much higher inadjacent tissues than in tumor tissues that contain high levelof PI3K-Akt activity [26]

In contrast towild type pre-S antigenHBVpre-Smutantsare viral oncoproteins to induce endoplasmic reticulum (ER)

BioMed Research International 3

PTEN PI3K Ras LKB1

ERKAMPKIRS

AKT TSC1

TSC2Rheb mTORC1

mTORC2FOXO1 FOXO3a

S6K1

S6

4E-BP1

eIF-4E

Figure 2The regulation ofmTORThe activity ofmTOR can be regulated by PI3K-Akt and LKB1-AMPK pathway ActivatedmTOR regulatestranscriptional activity of FOXO1-FOXO3a and protein translation by pS6 and eIF-4E

stress in ground glass hepatocytes (GGHs) that have beenrecognized as the precursor lesions of HCC [27] In additionthe existence of pre-S mutants in serum of HBV carrierscan predict the development of HCC Interestingly theexpression of pre-S mutants is associated with the activationof AktmTOR signaling inHCC cells [28] Pre-Smutantsmayupregulate VEGFR-2 to activate AktmTOR which can beattenuated by VEGF-A neutralization

Intriguingly the activation of mTOR can repress HBsAgsynthesis by facilitating the interaction of histone deacetylase1 (HDAC1)with YinYang1 (YY1) a transcription factor whichbound to pre-S1 promoter [29] Such a feedback regulationof HBsAg expression during HBV-initiated tumorigenesisis that mTOR inhibitors may activate HBV replication inpatients with chronic HBV infection Indeed everolimus wasassociatedwith the risk of HBV reactivation [30] In additionthe knockdown of histone deacetylase 1 (HDAC1) can abolishthis inhibitory effect of the mTOR on pre-S transcriptionThe HDAC1 inhibitors that have been intensively underevaluation for their anticancer effect may also lead to thereactivation of HBV

HBV X (HBx) protein encoded by the HBV X geneplays a crucial role in the pathogenesis of HBV-related HCCby promoting cell cycle progression inactivates negativegrowth regulators and binds to and inhibits the expressionof p53 tumor suppressor gene and other tumor suppressorgenes and senescence-related factors [31] The expression ofmTOR and PI3KAkt in HCC cells can be increased by HBxtransfection [32] Interestingly HBx transfection increasedthe formation of autophagosomes and autolysosomes andupregulated microtubule-associated protein light chain 3beclin 1 and lysosome-associated membrane protein 2aHBx-induced autophagy was further increased by mTORinhibitor rapamycin but blocked by treatment with thePI3K-Akt inhibitor LY294002 indicating that HBx activatesautophagy through PI3K-Akt-mTOR pathway HBx activatedmTOR seems to depend on I120581B kinase 120573 (IKK120573) [33] IKK120573

inhibitor Bay 11-7082 or silencing IKK120573 expression usingsiRNA reversed HBx-induced S6K1 activation HBx upregu-lated cell proliferation and vascular endothelial growth factor(VEGF) production Similarly mTOR inhibition reducedthe growth invasion epithelial-to-mesenchymal transitionand metastasis of HBx-expressing HCC cells [34] In theHBx transgenic mouse model pIKK120573 pS6K1 and VEGFexpressions were higher in cancerous than noncancerousliver tissues Furthermore pIKK120573 levels were strongly corre-lated with pTSC1 and pS6K1 levels in HBV-associated humanhepatoma tissues and higher pIKK120573 pTSC1 and pS6K1levels were correlated with a poor prognosis in these patients

32 mTOR and HCV Infection HCV infection contributes tothe rising incidence of HCC in many developed countriessuch as Spain France Italy and Japan where the proportioncaused by the HCV ranges from 50 to 70 [35] HCV non-structural protein NS5A is a crucial factor in viral replicationand diverse cellular events NS5A can activate PI3K-mTORsignaling by directly binding to the p85 subunit of PI3K(Figure 3) [36] Inhibition of PI3K abrogated NS5A-activatedmTOR In addition NS5A can interact with FKBP38 animmunosuppressant FK506-binding protein NS5A activatedmTOR by releasing it from FKBP38 even in the absence ofserum [37] Rapamycin or NS5A knockdown can block S61 Kand 4E-BP1 phosphorylation that were increased in HCVreplicon cells and NS5A-Huh7 cell

Strikingly HCV infection can activate mTOR in anautophagy-dependent manner HCV induced autophagy byupregulating beclin 1 to activate mTOR signaling pathwaywhich in turn promoted hepatocyte growth [38] HCVinduced mTOR activated and phosphorylation of eIF-4Ewere impaired in autophagy-deficient HCC cells

Stable NS5A expression in HCC cells led to the resistanceto apoptosis that was abolished by the silence of FKBP38through RNA interference [39] Moreover NS5A can repress


mTOR

VEGFR-2

Mutants

YY1 HDAC1

HBV

eIF-4E S6K

Akt

PTEN

PAK1

HCV

NF-120581BFKBP38

Pre-s1

p85

NS5A

PI3K

X

Figure 3 Regulation and function of mTOR in viral hepatitis and HCC development In HCV infection NS5A can activate mTOR throughPI3KAkt pathway or impair the combination between mTOR and FKBP38 The complex formed by mTOR and NF-120581B can downregulatethe expression of PTEN In HBV infection pre-S1 can activate AktmTOR pathway through upregulation of VEGFR-2 YY1-HDAC1 complexcan inhibit the transcription from the pre-S1 promoter as a negative feedback HBx can increase the expression of mTOR and PI3KAkt S6Kcan activate PAK1 to regulate actin cytoskeleton and cell motility

the activation of caspase-3 and poly(ADP-ribose) poly-merase which was abrogated by rapamycin or NS5A knock-down indicating that NS5A suppresses apoptosis specificallythrough mTOR pathway

Interestingly mTOR can influence the regulation of HCVRNA replication [40] For example mTOR downstreamkinase p21-activated kinase 1 (PAK1) has been found to takepart in antiviral signaling The suppression of PAK1 by PI3Kinhibitor or rapamycin enhanced viral RNA and proteinaffluence in established replicon HCV cell lines Similarlyknockdown of S6K abolished PAK1 phosphorylation andenhanced HCV RNA affluence while knockdown of eIF-4E increased viral RNA affluence without affecting PAK1phosphorylation

4 mTOR and HCC

As an important part of PI3KAkt pathway which is critical tocancer development mTORwas aberrantly activated in mostif not all human carcinogenesis mTOR was activated in pre-cancerous cirrhotic tissues in addition to chronic viral hepati-tis tissues [2] A small-scale immunohistochemistry staininganalysis revealed that 33 out of 73 (45) HCC patientshad increased expression of total S6k which correlated withmTOR activation as well as tumor nuclear grade and tumorsize [41] Moreover in a larger cohort of HCC patientsmTOR pathway was more remarkably altered in tumors withpoor differentiation high TNM stage vascular invasion andother poor prognostic features [42] The expression of pS6was further confirmed as an independent prognostic factorfor HCC

The activation of mTOR can confer many growth advan-tages to cancer stem or progenitor cells such as promoting cellproliferation and resistance to apoptosis induced by variousstress signals such as hypoxia and nutrient deficiency Inaddition mTOR can regulate telomerase activity in hep-atocarcinogenesis Treating HCC cells (SMMC-7721) withrapamycin significantly reduced telomerase activity by down-regulating hTERT protein level but not hTERT transcriptionindicating [43]

Neovascularization in tumor is closely associated withtumor growth Tumors that form as a result of mTORactivation are highly vascularized and inhibition of mTORby rapamycin can diminish the process of angiogenesis Theactivation ofmTORC1 can promote a variety of angiogenesis-related proteins such as hypoxia-inducible factor 120572 (HIF120572)and vascular endothelial growth factor (VEGF) Underhypoxia condition cancer cells can produce HIF120572 throughmTOR-dependent manner [43] When oxygen is sufficientactivated mTOR can promote the translation of HIF120572mRNAby the activation of 4E-BP1 or S6K1 [44] Inhibition of mTORactivity in human hepatoma cells reduced HIF120572 expressionwithout reducing its mRNA or promoting its degradation[45] In addition to affecting its expression mTOR can alsodirectly regulate the activity of HIF120572 [46] Activation ofmTORbyRheb overexpression potently enhances the activityof HIF120572 and VEGF secretion during hypoxia RAPTORdirectly interacts with HIF120572 which requires an mTOR sig-naling (TOS) motif located in the N terminus of HIF120572 Themutant of HIF120572 lacking this TOSmotif was unable to bind tothe coactivator CBPp300 and lost its transcriptional activityand proangiogenesis function


Autophagy plays a crucial role in tumor suppressionby eliminating damage cells mTOR has a regulatory rolefor autophagy and malignant cells often exhibit defects inautophagy [47] Autophagy-deficient mice such as beclin 1heterozygous mice have an increased incidence of sponta-neous tumors and also can accelerate the development ofprecancerous lesions induced by hepatitis B infection [48]Therefore mTORmay indirectly induce tumorigenesis by thesuppression of autophagy

4E-BP1 is a downstreammTORC1 key factor in regulatingcell proliferation Activated eIF4E preferably promote trans-lation of tumor-related genes such as cell cycle regulatoryproteins or antiapoptotic proteins like MCL1 [49] While therelevance of mTORC1 to human carcinogenesis has beenwell-documented whether mTORC2 is critical to humancancer development remains unknown Recently RICTORthe unique structure of mTORC2 was found to be necessaryto the tumor formation of PTEN-deficient prostate epithelialcells in nude mice indicating that mTORC2 can functionin synergy with PI3K to promote tumorigenesis [50] Inaddition inhibition of mTORC2 reduced proliferation andanchorage-independent growth of human cancer cells byinducing downregulation of cyclin D1 and cell cycle arrest atG1 phase [51]

Hepatic steatosis is a risk factor for HCC in addition tochronic viral hepatitis PTEN expression is downregulatedin the livers of rats and humans having steatosis whichwas accompanied by high plasma levels of fatty acids [52]Unsaturated fatty acids can downregulate PTEN expressionvia activation of a complex formed by mTOR and NF-kB inHepG2 cells

Aberrant lipogenesis plays a pivotal role in the develop-ment of human HCC [53]The AKT-mTORC1-S6K1 pathwayfacilitated lipogenesis via posttranscriptional and transcrip-tional mechanisms In a NASH liver activation of AKTand the mTOR pathway in turn triggered the developmentof HCC [54] mTORC1 is crucial for the activation ofthe sterol regulatory element-binding proteins (SREBPs)primary transcription factors regulating genes involved inlipid and sterol synthesis [55 56] For example fatty acidsynthase (FAS) is encoded by a target gene of SREBP and isupregulated in some human cancers [57] Overexpression ofFAS is an early phenomenon in chemically spontaneous andhormonally induced rat hepatocarcinogenesis that was foundto be associated with the activation of PI3K-Akt pathway [5859] In addition mTOR pathway can upregulate glycolysisin HCC [60] It is crucial to the transcriptional regulationof glucose transporters and various rate-limiting glycolyticenzymes such as pyruvate kinase M2 (PKM2) [61] Thelevels of PKM2 expression are upregulated in human cancercells to stimulate the transactivation of glycolytic genes [62ndash65] Transcription factors including HIF120572 and c-Myc areimportant to such transactivation effect of mTOR signaling[66 67]

5 mTOR Inhibitors for HCC Treatment

mTOR inhibitors inhibit mTOR complex 1 (mTORC1)mainly through interacting with FK506-binding protein 12

(FKBP12) At present there are 3 analogues of rapamycinwith potent biological properties and pharmacokinetics havebeen tested in clinical trials RAD001 (everolimus) CCI-779 (temsirolimus) and AP23573 (deforolimus) RAD001(everolimus) is an orally bioavailable analogue and CCI-779(temsirolimus) is a soluble ester analogue while AP23573(deforolimus) is a nonprodrug analogue of rapamycinmTORinhibitors have been tested successfully in clinical trials fortheir antineoplastic potency and well tolerability in differentmalignancies including renal cell carcinoma pancreatic neu-roendocrine tumors subependymal giant cell astrocytomasand breast cancer [68ndash72]

mTOR inhibition significantly reduces HCC growthand improves survival primarily via antiangiogenesis Afterthe treatment of mTOR inhibitor sirolimus for 4 weeksrats implanted with hepatoma cells had significantly longersurvival and developed smaller tumors fewer extrahepaticmetastases and less ascites than controls [73] Sirolimustreatment reduced intratumor microvessel density leadingto extensive necrosis Moreover vascular sprouting and tubeformation of aortic rings were also impaired

Basically mTOR inhibitors are well tolerated To eval-uate the best dosing schedules thirty-nine patients withlocally advanced or metastatic HCC (Child-Pugh class Aor B) were enrolled in an open-label phase 1 study andrandomly assigned to daily (25ndash10mg) or weekly (20ndash70mg)everolimus in a standard 3 + 3 dose-escalation design Dose-limiting toxicities (DLTs) occurred in five of 21 patients inthe daily and two of 19 patients in the weekly cohort Dailyand weekly maximum tolerated doses (MTDs) were 75mgand 70mg respectively Grade 34 adverse events with a ge10 incidence were thrombocytopenia hypophosphatemiaand alanine transaminase (ALT) elevation In four hepatitisB surface antigen- (HBsAg-) seropositive patients grade 34ALT elevations were accompanied by significant increases inserumHBV levelsThe incidence of hepatitis flare in HBsAg-seropositive patients with andwithout detectable serumHBVDNA before treatment was 462 and 71 respectively

Another phase 1 trial of temsirolimus combined withsorafenib showed that the maximum-tolerated dose (MTD)was temsirolimus 10mg weekly plus sorafenib 200mg twicedaily [74] Grade 3 or 4 adverse events were hypophos-phatemia infection thrombocytopenia hand-foot skin reac-tion (HFSR) and fatigue There is another trial that showedthat the MTD of everolimus in combination with standard-dose sorafenib was 25mg daily [75] Most common adverseevents are diarrhea HFSR and thrombocytopenia

The randomised clinical trial to compare two everolimusdosing schedules showed 75mg better than 70mg daily inpatients with HCC Disease control rates in the daily andweekly cohorts were 714 and 444 respectively [76]Treatment of HCC patients withmTOR inhibitors can inducetemporary PR (partial response) or SD (stable disease) [77]In 21 advanced HCC patients treated with sirolimus oncedaily one had PR and five had SD at 3 months The mediansurvival was 65 months (02ndash36 months)

Interestingly the intraliver and intra-abdominal growthsof patient-derived hepatocellular carcinoma xenograftswere inhibited by bevacizumab plus rapamycin treatment to


a significantly greater degree than bevacizumab or rapamycinmonotherapy [78] Reductions in tumor growth bybevacizumab plus rapamycinwere associatedwith reductionsin tumor microvessel density as well as the expression ofVEGF cyclin D1 and cyclin B1 Eventually mouse survivalwas greatly prolonged after the combination treatment

In addition mTOR inhibitor can suppress tumor growthand sensitize tumor cell to chemotherapy or other targettherapy [79ndash82] For example RAD001 alone can repress cellproliferation but not induce apoptosis However RAD001 incombination with cisplatin can induce a remarkable increasein the number of apoptotic cells both in vitro and in vivoby downregulating the expression of prosurvival moleculessuch as Bcl-2 and survivin [79] Similarly the combinationof rapamycin with doxorubicin displayed better anticancereffect compared to either of the single agent treatments[83] Moreover mTOR inhibitor attenuated the doxorubicin-induced inhibition of endothelial cell proliferation Forexample doxorubicin can stimulate expression of p21 whichwas reversed by the addition of rapamycin FurthermoremTOR can inhibit HCC development in synergy with manyother anticancer agents such as 5-fluorouracil (5-Fu) [81]microtubule inhibitors [84] and vinblastine [82] In additionto chemotherapy drugs target therapy agents such as pro-teasome inhibitor bortezomib can synergize with rapamycinto reduce growth repress mobility and induce apoptosis ofHCC cells [85] Another phase 1 trial of temsirolimus com-bined with sorafenib showed that the maximum-tolerateddose (MTD) was temsirolimus 10mg weekly plus sorafenib200mg twice daily [74] Grade 3 or 4 adverse events werehypophosphatemia infection thrombocytopenia hand-footskin reaction (HFSR) and fatigue Two of 25 patients hada PR and 15 of 25 had SD There is another trial thatshowed that the MTD of everolimus in combination withstandard dose sorafenib was 25mg daily [75] Median timeto progression was 45 months and overall survival was 74months Most common adverse events are diarrhea HFSRand thrombocytopenia In addition to chemotherapy andtarget therapy the effect of radiation therapy can also beaugmented by mTOR inhibitors For example RAD001 atclinically relevant nanomolar concentrations enhanced theefficacy of radiation inHCCcellsThe induction of autophagymay account for this effect [86]

Although mTOR inhibitors have shown great potentialfor the treatment of HCC patients surrogate biomarkers arenecessary to identify suitable patients so as to improve clinicalefficacy and prevent drug resistance For example rapamycincan activate PI3K-Akt in an insulin-like growth factor-dependent manner by relieving S6K1-dependent inhibitoryphosphorylation of IRS-1 thus preventing IRS-1 degrada-tion and enhancing PI3K activation Rapamycin can inhibitS6K1-dependent IRS-1 serine phosphorylation increase IRS-1 protein levels and promote association of tyrosine-phosphorylated IRS-1 with PI3K [87] Such a negativefeedback regulation is important to maintain homeostasisHowever the disruption of this negative feedback by mTORinhibitors may attenuate the therapeutic effect Fortunatelythere are also some agents that havemultiple targets includingmTOR and PI3K For example NVP-BEZ235 is a dual

inhibitor of PI3K and mTOR It can decrease the levels of p-Akt and p-S6K and repress cell proliferation in HCC cell lines[88] Moreover it can repress tumor growth without loss ofbody weight Interestingly combination of everolimus withNVP-BEZ235 can synergistically suppress the proliferationof HCC cells [89] PI-103 a dual PI3KmTOR inhibitorin combination with sorafenib can effectively inhibit theproliferation of HCC cells by blocking both RasRafMAPKand PI3KAktmTOR pathways [90]

6 Conclusions and Perspectives

mTOR plays an important role in viral hepatitis and HCCdevelopment mTOR inhibitors can repress cell growth bothin vitro and in vivo Preliminary clinical trials indicated thatmTOR inhibitors alone or in combination with cytotoxicdrugs or targeted therapy drugs can improve clinical out-comes of HCC patients However no RCTs have proven thebenefits of everolimus treatment in HCC Novel biomarkersare warranted to identify suitable HCC patients who maybenefit from the treatment of mTOR inhibitors Although themost common adverse events are tolerated it is noteworthythat mTOR is associated with HBV virus replication andmTOR inhibitors may cause hepatitis B reactivation HCCcells are prone to develop multiple drug resistance due tothe heterogeneity and fragile genome Therefore the combi-nation of mTOR inhibitors with conventional chemotherapydrugs and target therapy agentsmight be a promising strategyfor the future application of mTOR inhibitors

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Authorsrsquo Contribution

Zhuo Wang and Wei Jin contributed equally to this work

References

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[2] S J Cotler N Hay H Xie et al ldquoImmunohistochemical expres-sion of components of the Akt-mTORC1 pathway is associatedwith hepatocellular carcinoma in patients with chronic liverdiseaserdquo Digestive Diseases and Sciences vol 53 no 3 pp 844ndash849 2008

[3] A Villanueva D Y Chiang P Newell et al ldquoPivotal role ofmTOR signaling in hepatocellular carcinomardquo Gastroenterol-ogy vol 135 no 6 pp 1972e1ndash1983e11 2008

[4] N Hay and N Sonenberg ldquoUpstream and downstream ofmTORrdquo Genes and Development vol 18 no 16 pp 1926ndash19452004

[5] Y Sancak C C Thoreen T R Peterson et al ldquoPRAS40 isan insulin-regulated inhibitor of the mTORC1 protein kinaserdquoMolecular Cell vol 25 no 6 pp 903ndash915 2007


[6] E V Haar S-I Lee S Bandhakavi T J Griffin and D-H KimldquoInsulin signalling tomTORmediated by theAktPKB substratePRAS40rdquo Nature Cell Biology vol 9 no 3 pp 316ndash323 2007

[7] R Loewith E Jacinto S Wullschleger et al ldquoTwo TORcomplexes only one of which is rapamycin sensitive havedistinct roles in cell growth controlrdquoMolecular Cell vol 10 no3 pp 457ndash468 2002

[8] D D Sarbassov S M Ali D-H Kim et al ldquoRictor a novelbinding partner of mTOR defines a rapamycin-insensitive andraptor-independent pathway that regulates the cytoskeletonrdquoCurrent Biology vol 14 no 14 pp 1296ndash1302 2004

[9] E Jacinto R Loewith A Schmidt et al ldquoMammalian TORcomplex 2 controls the actin cytoskeleton and is rapamycininsensitiverdquo Nature Cell Biology vol 6 no 11 pp 1122ndash11282004

[10] C K Yip K Murata T Walz D M Sabatini and S A KangldquoStructure of the human mTOR complex I and its implicationsfor rapamycin inhibitionrdquoMolecular Cell vol 38 no 5 pp 768ndash774 2010

[11] D-H Kim D D Sarbassov S M Ali et al ldquomTOR interactswith raptor to form a nutrient-sensitive complex that signalsto the cell growth machineryrdquo Cell vol 110 no 2 pp 163ndash1752002

[12] D D Sarbassov S M Ali S Sengupta et al ldquoProlongedrapamycin treatment inhibits mTORC2 assembly andAktPKBrdquoMolecular Cell vol 22 no 2 pp 159ndash168 2006

[13] K Hara K Yonezawa M T Kozlowski et al ldquoRegulation ofeIF-4E BP1 phosphorylation by mTORrdquo Journal of BiologicalChemistry vol 272 no 42 pp 26457ndash26463 1997

[14] A C Hsieh M Costa O Zollo et al ldquoGenetic dissection ofthe oncogenic mTOR pathway reveals druggable addiction totranslational control via 4EBP-eIF4E rdquo Cancer Cell vol 17 no3 pp 249ndash261 2010

[15] R J Shaw N Bardeesy B D Manning et al ldquoThe LKB1 tumorsuppressor negatively regulates mTOR signalingrdquo Cancer Cellvol 6 no 1 pp 91ndash99 2004

[16] M N Corradetti K Inoki N Bardeesy R A DePinho and K-L Guan ldquoRegulation of the TSC pathway by LKB1 evidence ofamolecular link between tuberous sclerosis complex and Peutz-Jeghers syndromerdquo Genes and Development vol 18 no 13 pp1533ndash1538 2004

[17] F Pene Y-E Claessens O Muller et al ldquoRole of the phos-phatidylinositol 3-kinaseAkt and mTORP70S6-kinase path-ways in the proliferation and apoptosis in multiple myelomardquoOncogene vol 21 no 43 pp 6587ndash6597 2002

[18] M M Hill and B A Hemmings ldquoInhibition of protein kinaseBAkt implications for cancer therapyrdquo Pharmacology andTherapeutics vol 93 no 2-3 pp 243ndash251 2002

[19] D E Martin and M N Hall ldquoThe expanding TOR signalingnetworkrdquo Current Opinion in Cell Biology vol 17 no 2 pp 158ndash166 2005

[20] K Hara K Yonezawa Q-PWeng M T Kozlowski C Belhamand J Avruch ldquoAmino acid sufficiency and mTOR regulatep70 S6 kinase and eIF-4E BP1 through a common effectormechanismrdquo Journal of Biological Chemistry vol 273 no 34pp 14484ndash14494 1998

[21] MChiu S Tardito A BarilliMG Bianchi VDallrsquoAsta andOBussolati ldquoGlutamine stimulates mTORC1 independent of thecell content of essential amino acidsrdquo Amino Acids vol 43 pp2561ndash2567 2012

[22] Y Sancak T R Peterson Y D Shaul et al ldquoThe rag GTPasesbind raptor and mediate amino acid signaling to mTORC1rdquoScience vol 320 no 5882 pp 1496ndash1501 2008

[23] E Kim P Goraksha-Hicks L Li T P Neufeld and K-L GuanldquoRegulation of TORC1 by Rag GTPases in nutrient responserdquoNature Cell Biology vol 10 no 8 pp 935ndash945 2008

[24] RVDuranWOppliger AMRobitaille et al ldquoGlutaminolysisactivates Rag-mTORC1 signalingrdquo Molecular Cell vol 47 pp349ndash358 2012

[25] H Guo T Zhou D Jiang et al ldquoRegulation of hepatitisB virus replication by the phosphatidylinositol 3-kinase-Aktsignal transduction pathwayrdquo Journal of Virology vol 81 no 18pp 10072ndash10080 2007

[26] W Wang W T London and M A Feitelson ldquoHepatitis B xantigen in hepatitis B virus carrier patients with liver cancerrdquoCancer Research vol 51 no 18 pp 4971ndash4977 1991

[27] H-C Wang H-C Wu C-F Chen N Fausto H-Y Lei and I-J Su ldquoDifferent types of ground glass hepatocytes in chronichepatitis B virus infection contain specific Pre-S mutantsthat may induce endoplasmic reticulum stressrdquo The AmericanJournal of Pathology vol 163 no 6 pp 2441ndash2449 2003

[28] J-C Yang C-F Teng H-C Wu et al ldquoEnhanced expressionof vascular endothelial growth factor-A in ground glass hepato-cytes and its implication in hepatitis B virus hepatocarcinogen-esisrdquo Hepatology vol 49 no 6 pp 1962ndash1971 2009

[29] C-F Teng H-C Wu H-W Tsai H-S Shiah W Huang andI-J Su ldquoNovel feedback inhibition of surface antigen synthesisby mammalian target of rapamycin (mTOR) signal and itsimplication for hepatitis B virus tumorigenesis and therapyrdquoHepatology vol 54 no 4 pp 1199ndash1207 2011

[30] S Sezgin Goksu S Bilal and H S Coskun ldquoHepatitis Breactivation related to everolimusrdquoWorld Journal of Hepatologyvol 5 pp 43ndash45 2013

[31] C Neuveut Y Wei and M A Buendia ldquoMechanisms of HBV-related hepatocarcinogenesisrdquo Journal of Hepatology vol 52 no4 pp 594ndash604 2010

[32] P Wang Q S Guo Z W Wang and H X Qian ldquoHBx inducesHepG-2 cells autophagy through PI3KAkt-mTOR pathwayrdquoMolecular and Cellular Biochemistry vol 372 pp 161ndash168 2013

[33] C J Yen Y J Lin C S Yen et al ldquoHepatitis B virus X proteinupregulates mTOR signaling through IKKbeta to increase cellproliferation and VEGF production in hepatocellular carci-nomardquo PLoS ONE vol 7 Article ID e41931 2012

[34] X Xu Z Fan L Kang et al ldquoHepatitis B virus X proteinrepresses miRNA-148a to enhance tumorigenesisrdquo The Journalof Clinical Investigation vol 123 pp 630ndash645 2013

[35] C W Shepard L Finelli and M J Alter ldquoGlobal epidemiologyof hepatitis C virus infectionrdquo The Lancet Infectious Diseasesvol 5 no 9 pp 558ndash567 2005

[36] Y He H Nakao S-L Tan et al ldquoSubversion of cell signalingpathways by hepatitis C virus nonstructural 5A protein viainteraction with Grb2 and P85 phosphatidylinositol 3-kinaserdquoJournal of Virology vol 76 no 18 pp 9207ndash9217 2002

[37] L Peng D Liang W Tong J Li and Z Yuan ldquoHepatitisC virus NS5A activates the mammalian target of rapamycin(mTOR) pathway contributing to cell survival by disruptingthe interaction between FK506-binding protein 38 (FKBP38)and mTORrdquo Journal of Biological Chemistry vol 285 no 27 pp20870ndash20881 2010

[38] S Shrivastava J Bhanja Chowdhury R Steele R Ray and RB Ray ldquoHepatitis C virus upregulates Beclin1 for induction of


autophagy and activates mTOR signalingrdquo Journal of Virologyvol 86 pp 8705ndash8712 2012

[39] J Wang W Tong X Zhang et al ldquoHepatitis C virus non-structural protein NS5A interacts with FKBP38 and inhibitsapoptosis in Huh7 hepatoma cellsrdquo FEBS Letters vol 580 no18 pp 4392ndash4400 2006

[40] H Ishida K Li M Yi and S M Lemon ldquop21-activated kinase 1is activated through themammalian target of rapamycinp70 S6kinase pathway and regulates the replication of hepatitis C virusin human hepatoma cellsrdquo Journal of Biological Chemistry vol282 no 16 pp 11836ndash11848 2007

[41] F Sahin R Kannangai O Adegbola J Wang G Su and MTorbenson ldquomTOR and P70 S6 kinase expression in primaryliver neoplasmsrdquo Clinical Cancer Research vol 10 no 24 pp8421ndash8425 2004

[42] L Zhou Y Huang J Li and Z Wang ldquoThe mTOR pathwayis associated with the poor prognosis of human hepatocellularcarcinomardquoMedical Oncology vol 27 no 2 pp 255ndash261 2010

[43] B GWouters andM Koritzinsky ldquoHypoxia signalling throughmTOR and the unfolded protein response in cancerrdquo NatureReviews Cancer vol 8 no 11 pp 851ndash864 2008

[44] S Sengupta T R Peterson and D M Sabatini ldquoRegulation ofthe mTOR complex 1 pathway by nutrients growth factors andstressrdquoMolecular Cell vol 40 no 2 pp 310ndash322 2010

[45] Q Weng J Zhang J Cao et al ldquoQ39 a quinoxaline 14-Di-N-oxide derivative inhibits hypoxia-inducible factor-1120572 expres-sion and the AktmTOR4E-BP1 signaling pathway in humanhepatoma cellsrdquo Investigational New Drugs vol 29 no 6 pp1177ndash1187 2011

[46] S C Land and A R Tee ldquoHypoxia-inducible factor 1120572 isregulated by the mammalian target of rapamycin (mTOR) viaan mTOR signaling motifrdquo Journal of Biological Chemistry vol282 no 28 pp 20534ndash20543 2007

[47] B Ravikumar C Vacher Z Berger et al ldquoInhibition of mTORinduces autophagy and reduces toxicity of polyglutamineexpansions in fly and mouse models of Huntington diseaserdquoNature Genetics vol 36 no 6 pp 585ndash595 2004

[48] X Qu J Yu G Bhagat et al ldquoPromotion of tumorigenesis byheterozygous disruption of the beclin 1 autophagy generdquo Journalof Clinical Investigation vol 112 no 12 pp 1809ndash1820 2003

[49] H-GWendel R L A Silva A Malina et al ldquoDissecting eIF4Eaction in tumorigenesisrdquoGenes andDevelopment vol 21 no 24pp 3232ndash3237 2007

[50] D A Guertin D M Stevens M Saitoh et al ldquomTOR complex2 is required for the development of prostate cancer induced byPten loss in micerdquo Cancer Cell vol 15 no 2 pp 148ndash159 2009

[51] V Hietakangas and S M Cohen ldquoTOR complex 2 is neededfor cell cycle progression and anchorage-independent growthof MCF7 and PC3 tumor cellsrdquo BMC Cancer vol 8 article 2822008

[52] M Vinciguerra C Veyrat-Durebex M A Moukil L Rubbia-Brandt F Rohner-Jeanrenaud and M Foti ldquoPTEN down-regulation by unsaturated fatty acids triggers hepatic steatosisvia an NF-kappaBp65mTOR-dependent mechanismrdquo Gas-troenterology vol 134 no 1 pp 268ndash280 2008

[53] D F Calvisi C Wang C Ho et al ldquoIncreased lipogenesisinduced by AKT-mTORC1-RPS6 signaling promotes develop-ment of human hepatocellular carcinomardquo Gastroenterologyvol 140 no 3 pp 1071ndash1083 2011

[54] K Nakanishi M Sakamoto S Yamasaki S Todo and SHirohashi ldquoAkt phosphorylation is a risk factor for early disease

recurrence and poor prognosis in hepatocellular carcinomardquoCancer vol 103 no 2 pp 307ndash312 2005

[55] K Duvel J L Yecies S Menon et al ldquoActivation of a metabolicgene regulatory network downstream of mTOR complex 1rdquoMolecular Cell vol 39 no 2 pp 171ndash183 2010

[56] T Porstmann C R Santos B Griffiths et al ldquoSREBP activityis regulated by mTORC1 and contributes to Akt-dependent cellgrowthrdquo Cell Metabolism vol 8 no 3 pp 224ndash236 2008

[57] J A Menendez and R Lupu ldquoFatty acid synthase and thelipogenic phenotype in cancer pathogenesisrdquo Nature ReviewsCancer vol 7 no 10 pp 763ndash777 2007

[58] T van de Sande T Roskams E Lerut et al ldquoHigh-level expres-sion of fatty acid synthase in human prostate cancer tissues islinked to activation and nuclear localization of AktPKBrdquo TheJournal of Pathology vol 206 no 2 pp 214ndash219 2005

[59] T van de Sande E de Schrijver W Heyns G Verhoevenand J V Swinnen ldquoRole of the phosphatidylinositol 31015840-kinasePTENAkt kinase pathway in the overexpression of fattyacid synthase in LNCaP prostate cancer cellsrdquo Cancer Researchvol 62 no 3 pp 642ndash646 2002

[60] M Evert D F Calvisi K Evert et al ldquoV-AKTmurine thymomaviral oncogene homologmammalian target of rapamycin acti-vation induces a module of metabolic changes contributing togrowth in insulin-induced hepatocarcinogenesisrdquo Hepatologyvol 55 no 5 pp 1473ndash1484 2012

[61] B Chaneton and E Gottlieb ldquoRocking cell metabolism revisedfunctions of the key glycolytic regulator PKM2 in cancerrdquoTrends in Biochemical Sciences vol 37 pp 309ndash316 2012

[62] J D Dombrauckas B D Santarsiero andADMesecar ldquoStruc-tural basis for tumor pyruvate kinase M2 allosteric regulationand catalysisrdquo Biochemistry vol 44 no 27 pp 9417ndash9429 2005

[63] S Mazurek ldquoPyruvate kinase type M2 a key regulator withinthe tumour metabolome and a tool for metabolic profiling oftumoursrdquo in Oncogenes Meet Metabolism pp 99ndash124 Springer2008

[64] W Yang Y Xia D Hawke et al ldquoPKM2 phosphorylates histoneH3 and promotes gene transcription and tumorigenesisrdquo Cellvol 150 pp 685ndash696 2012

[65] W Luo H Hu R Chang et al ldquoPyruvate kinase M2 is a PHD3-stimulated coactivator for hypoxia-inducible factor 1rdquo Cell vol145 no 5 pp 732ndash744 2011

[66] J D Gordan C B Thompson and M C Simon ldquoHIF and c-Myc sibling rivals for control of cancer cell metabolism andproliferationrdquo Cancer Cell vol 12 no 2 pp 108ndash113 2007

[67] M J West M Stoneley and A E Willis ldquoTranslational induc-tion of the c-myc oncogene via activation of the FRAPTORsignalling pathwayrdquo Oncogene vol 17 no 6 pp 769ndash780 1998

[68] G R Hudes A Berkenblit J Feingold M B Atkins B I Riniand J Dutcher ldquoClinical trial experience with temsirolimusin patients with advanced renal cell carcinomardquo Seminars inOncology vol 36 supplement 3 pp S26ndashS36 2009

[69] R JMotzer B Escudier SOudard et al ldquoEfficacy of everolimusin advanced renal cell carcinoma a double-blind randomisedplacebo-controlled phase III trialrdquoTheLancet vol 372 no 9637pp 449ndash456 2008

[70] J C Yao M H Shah T Ito et al ldquoEverolimus for advancedpancreatic neuroendocrine tumorsrdquo The New England Journalof Medicine vol 364 no 6 pp 514ndash523 2011

[71] D A Krueger M M Care K Holland et al ldquoEverolimus forsubependymal giant-cell astrocytomas in tuberous sclerosisrdquoThe New England Journal of Medicine vol 363 no 19 pp 1801ndash1811 2010


[72] J Baselga M Campone M Piccart et al ldquoEverolimus inpostmenopausal hormone-receptor-positive advanced breastcancerrdquo The New England Journal of Medicine vol 366 no 6pp 520ndash529 2012

[73] D Semela A-C Piguet M Kolev et al ldquoVascular remodelingand antitumoral effects of mTOR inhibition in a rat model ofhepatocellular carcinomardquo Journal of Hepatology vol 46 no 5pp 840ndash848 2007

[74] R K Kelley H S Nimeiri P N Munster et al ldquoTemsirolimuscombined with sorafenib in hepatocellular carcinoma a phase Idose-finding trial with pharmacokinetic and biomarker corre-latesrdquo Annals of Oncology vol 24 pp 1900ndash1907 2013

[75] R S Finn R T Poon T Yau et al ldquoPhase I study investigatingeverolimus combined with sorafenib in patients with advancedhepatocellular carcinomardquo Journal of Hepatology vol 59 no 6pp 1271ndash1277 2013

[76] H S Shiah C Y Chen C Y Dai et al ldquoRandomised clin-ical trial comparison of two everolimus dosing schedules inpatients with advanced hepatocellular carcinomardquo AlimentaryPharmacology ampTherapeutics vol 37 pp 62ndash73 2013

[77] M Rizell M Andersson C Cahlin L Hafstrom M Olaussonand P Lindner ldquoEffects of the mTOR inhibitor sirolimusin patients with hepatocellular and cholangiocellular cancerrdquoInternational Journal of Clinical Oncology vol 13 no 1 pp 66ndash70 2008

[78] H Huynh P K H Chow N Palanisamy et al ldquoBevacizumaband rapamycin induce growth suppression in mouse models ofhepatocellular carcinomardquo Journal of Hepatology vol 49 no 1pp 52ndash60 2008

[79] K H Tam Z F Yang C K Lau C T Lam R W C Pang andR T P Poon ldquoInhibition of mTOR enhances chemosensitivityin hepatocellular carcinomardquo Cancer Letters vol 273 no 2 pp201ndash209 2009

[80] A-C Piguet D Semela A Keogh et al ldquoInhibition of mTORin combination with doxorubicin in an experimental model ofhepatocellular carcinomardquo Journal of Hepatology vol 49 no 1pp 78ndash87 2008

[81] X Bu C Le F Jia et al ldquoSynergistic effect of mTOR inhibitorrapamycin and fluorouracil in inducing apoptosis and cellsenescence in hepatocarcinoma cellsrdquo Cancer Biology andTher-apy vol 7 no 3 pp 392ndash396 2008

[82] Q Zhou V W Y Lui C P Y Lau et al ldquoSustained anti-tumor activity by co-targeting mTOR and the microtubulewith temsirolimusvinblastine combination in hepatocellularcarcinomardquo Biochemical Pharmacology vol 83 no 9 pp 1146ndash1158 2012


[84] D J VanderWeele R Zhou and C M Rudin ldquoAkt up-regulation increases resistance to microtubule-directedchemotherapeutic agents through mammalian target ofrapamycinrdquo Molecular Cancer Therapeutics vol 3 no 12 pp1605ndash1613 2004

[85] C Wang D Gao K Guo et al ldquoNovel synergistic antitumoreffects of rapamycin with bortezomib on hepatocellular carci-noma cells and orthotopic tumor modelrdquo BMC Cancer vol 12article 166 2012

[86] A Altmeyer E Josset J M Denis et al ldquoThe mTOR inhibitorRAD001 augments radiation-induced growth inhibition in a

hepatocellular carcinoma cell line by increasing autophagyrdquoInternational Journal of Oncology vol 41 pp 1381ndash1386 2012

[87] A Soni A Akcakanat G Singh et al ldquoeIF4E knockdowndecreases breast cancer cell growth without activating AktsignalingrdquoMolecular CancerTherapeutics vol 7 no 7 pp 1782ndash1788 2008

[88] M Masuda M Shimomura K Kobayashi S Kojima andT Nakatsura ldquoGrowth inhibition by NVP-BEZ235 a dualPI3KmTOR inhibitor in hepatocellular carcinoma cell linesrdquoOncology Reports vol 26 no 5 pp 1273ndash1279 2011

[89] H E Thomas C A Mercer L S Carnevalli et al ldquomTORinhibitors synergize on regression reversal of gene expressionand autophagy in hepatocellular carcinomardquo Science Transla-tional Medicine vol 4 Article ID 139ra84 2012

[90] R Gedaly P Angulo J Hundley et al ldquoPI-103 and sorafenibinhibit hepatocellular carcinoma cell proliferation by blockingRasRafMAPK and PI3KAKTmTOR pathwaysrdquo AnticancerResearch vol 30 no 12 pp 4951ndash4958 2010

Submit your manuscripts athttpwwwhindawicom

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014


MEDIATORSINFLAMMATION

of


Behavioural Neurology

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Immunology ResearchHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary and Alternative Medicine

Volume 2014Hindawi Publishing Corporationhttpwwwhindawicom


mTOR

RAPTORmLST8DEPTOR

PRAS40mSIN1

RICTORPROTOR

mTORmLST8DEPTOR

mTORC1 mTORC2

Figure 1 The structure of mTORC1 and mTORC2 The coremTOR machinery consists of mTOR DEPTOR and mLST8 Thecombination of core mTOR machinery with different proteinsconstitutes mTOR1 and mTORC2

charge domain (LCD) which can be phosphorylated byAKT [5 6] mLST8 can mediate protein-protein interactionswhile mSIN1 contains a Ras-binding domain (RBD) and apleckstrin homology which can interact with phospholipidCurrently the structures of RICTOR and PROTOR are stillnot clear

Rapamycin can inhibit the mTORC1 but not mTORC2because rapamycin binds with FKBP12 to disrupt the inter-action of mTOR with RAPTOR but not RICTOR [7ndash9] Therapamycin-induced dissociation of mTOR from RAPTOReventually prevents interaction of the mTOR with a numberof substrates [10 11] However long-term rapamycin treat-ment can inhibit mTORC2 [12] This effect may involve thechanges of intracellular pool of mTOR and thus reduce theassembly of mTORC2

22 Regulation of mTOR Activation mTORC1 can be acti-vated by diverse factors such as growth factors vari-ous cytokines Toll-like receptor ligands cell energy levelshypoxia and DNA damage The activation of mTORC1 playsan important role in protein synthesis ribosome biogenesisand autophagy Activated mTORC1 can phosphorylate thedownstream signalingmolecules including S6K1 or RPS6KN1(ribosomal protein S6 kinase 70 kDa polypeptide 1) andeukaryotic translation initiation factor-binding protein 1 (4E-BP1) Activation of S6K1 can promote the expression of ribo-somal protein and translation regulating protein to regulateprotein syntheses Nonphosphorylated 4E-BP1 can bind toeIF-4E to inhibit mRNA translation Once phosphorylatedby active mTOR 4E-BP1 are dissociated from eIF-4E sothat eIF-4E can bind to other translation initiation factorsto initiate protein translation [13 14] Tuberous sclerosiscomplex 1- (TSC1-) TSC2 tumor suppressor complex is anegative regulator of mTOR As a GTP activating protein(GAP) TSC2 or tuberin inactivates Ras homologue enrichedin brain (Rheb) which can directly bind to and activatemTOR TSC1 or hamartin does not have a GAP domain butit acts as a stabilizer of TSC2 by preventing it from degra-dation The activity of TSC1-TSC2 is regulated by proteinphosphorylation Activated PI3K-Akt signaling can phos-phorylate and inhibit TSC1-TSC2 while LKB1-AMPK canactivate TSC1-TSC2 by phosphorylation at different residues(Figure 2) [15 16]

The activation ofmTORC1 can be regulated by several fac-tors through signaling pathways including PI3KAktmTORLKB1AMPKmTOR andMAPK pathway Once activated byextracellular signals such as growth factors andnutrient PI3Kcan phosphorylate PIP2 to form PIP3 [17] As a result Aktand its activator phosphoinositide-dependent protein kinases1 (PDK1) translocate to the plasma membrane by binding toPIP3 When phosphorylated at Thr308 and Ser473 Akt canactivate mTOR by inhibiting TSC1-TSC2 [18] In additionAkt can suppress PRAS40 by phosphorylation to eliminateits inhibition on mTORC1

Studies have shown that acids metabolism in mam-malian cells is adjusted by LKB1AMPKmTOR signalpathway mTOR pathway can be activated by adenosinemonophosphate-activated protein kinases (AMPKs) that inturn can be activated by LKB1 AMPKs change their con-formation in response to the intracellular AMPATP ratioWhen AMPATP ratio drops LKB1 can bind to AMPKsubunit to activate AMPKby phosphorylationThe activationof AMPK will inhibit mTOR activation reduce proteinsynthesis and inhibit cell proliferation [19]

In addition mTOR activity can be regulated in responseto the availability of amino acids [20 21] Ras-related(Rag) GTPases a family of four related small GTPase areresponsible for amino acid-regulated mTOR activity [22 23]Rag GTPases interact with the RAPTOR the subunit ofmTORC1 in an amino acid-dependent manner allowinginteraction of mTORC1 with Rheb The Rags exist as obligateheterodimers Depletion of either heterodimer partners caninhibit mTORC1 while overexpression of the heterodimerwill rescue mTORC1 from suppression by amino acid with-drawal Some specific amino acids may play distinct rolesin the regulation of mTOR activity For example glutaminein combination with leucine activates mammalian TORC1(mTORC1) by enhancing glutaminolysis and 120572-ketoglutarateproduction [24] Inhibition of glutaminolysis prevented acti-vation of RagB and mTORC1 while constitutively active Ragheterodimer activatedmTORC1 in the absence of glutaminol-ysis Conversely enhanced glutaminolysis or a cell-permeable120572-ketoglutarate analog stimulated lysosomal translocationand activation of mTORC1

3 mTOR and Viral Hepatitis

31 mTOR and HBV Infection Chronic HBV infection is acrucial factor for the development of HCC In HepG2 cellsHBV RNA transcription and subsequent DNA replicationwere inhibited by the expression of a constitutively activeAkt1 [25] This inhibition of HBV gene transcription seemsto be mediated by mTOR activation since rapamycin canabolish this inhibition Furthermore inhibitors of PI3K Aktand mTOR can increase the transcription of viral RNA aswell as the replication of HBV DNA in HBV-overexpressingcells Consistently expression of HBsAg was much higher inadjacent tissues than in tumor tissues that contain high levelof PI3K-Akt activity [26]

In contrast towild type pre-S antigenHBVpre-Smutantsare viral oncoproteins to induce endoplasmic reticulum (ER)


PTEN PI3K Ras LKB1

ERKAMPKIRS

AKT TSC1

TSC2Rheb mTORC1

mTORC2FOXO1 FOXO3a

S6K1

S6

4E-BP1

eIF-4E










mTOR

VEGFR-2

Mutants

YY1 HDAC1

HBV

eIF-4E S6K

Akt

PTEN

PAK1

HCV

NF-120581BFKBP38

Pre-s1

p85

NS5A

PI3K

X




4 mTOR and HCC




























References






































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















PTEN PI3K Ras LKB1

ERKAMPKIRS

AKT TSC1

TSC2Rheb mTORC1

mTORC2FOXO1 FOXO3a

S6K1

S6

4E-BP1

eIF-4E










mTOR

VEGFR-2

Mutants

YY1 HDAC1

HBV

eIF-4E S6K

Akt

PTEN

PAK1

HCV

NF-120581BFKBP38

Pre-s1

p85

NS5A

PI3K

X




4 mTOR and HCC




























References






































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of

















mTOR

VEGFR-2

Mutants

YY1 HDAC1

HBV

eIF-4E S6K

Akt

PTEN

PAK1

HCV

NF-120581BFKBP38

Pre-s1

p85

NS5A

PI3K

X




4 mTOR and HCC




























References






































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of








































References






































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















































































































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















review article mtor in viral hepatitis and hepatocellular...

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