Cystathionine -Synthase in Physiology and CancerGSS GCLC SAM Betaine SAH DMG BHMT Homocyeine...

Review ArticleCystathionine 120573-Synthase in Physiology and Cancer

Haoran Zhu12 Shaun Blake12 Keefe T Chan1

Richard B Pearson 1234 and Jian Kang 1

1Division of Research Peter MacCallum Cancer Centre 305 Grattan Street Melbourne Victoria 3000 Australia2Sir Peter MacCallum Department of Oncology Australia3Department of Biochemistry and Molecular Biology University of Melbourne Parkville Victoria 3052 Australia4Department of Biochemistry and Molecular Biology Monash University Clayton Victoria 3168 Australia

Correspondence should be addressed to Richard B Pearson rickpearsonpetermacorg

Received 23 March 2018 Accepted 29 May 2018 Published 28 June 2018

Academic Editor Maria L Tornesello

Copyright copy 2018 Haoran Zhu 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

Cystathionine 120573-synthase (CBS) regulates homocysteine metabolism and contributes to hydrogen sulfide (H2S) biosynthesis

through which it plays multifunctional roles in the regulation of cellular energetics redox status DNA methylation and proteinmodification Inactivating mutations in CBS contribute to the pathogenesis of the autosomal recessive disease CBS-deficienthomocystinuria Recent studies demonstrating that CBS promotes colon and ovarian cancer growth in preclinical models highlighta newly identified oncogenic role for CBS On the contrary tumor-suppressive effects of CBS have been reported in other cancertypes suggesting context-dependent roles of CBS in tumor growth and progression Here we review the physiological functionsof CBS summarize the complexities regarding CBS research in oncology and discuss the potential of CBS and its key metabolitesincluding homocysteine and H

2S as potential biomarkers for cancer diagnosis or therapeutic targets for cancer treatment

1 Introduction

Cystathionine 120573-synthase (CBS) catalyzes the condensationof homocysteine (Hcy) with serine to form cystathioninewhich is the initial and rate-limiting step in the transsulfu-ration pathway Cystathionine is subsequently cleaved by theenzyme cystathionine gamma-lyase (CTH) to form cysteinea precursor of glutathione Besides this canonical pathwayCBS also participates in the desulfuration reactions thatcontribute to endogenous hydrogen sulfide (H

2S) production

(Figure 1) Thus CBS acting mainly through control ofHcy and H

2S metabolism exerts diverse biological functions

including mitochondrial bioenergetics redox homeostasisDNAmethylation and protein modification Deregulation ofCBS and the associated alterations in Hcy andor H

2S levels

leads to a wide range of pathological disturbances in thecardiovascular immune and central nervous systems andcontributes to disease development such as CBS-deficienthomocystinuria (CBSDH) It is now becoming clear that CBSactivity also plays an important but complex role in cancer

biology This review focuses on the current understanding ofthe functional role of CBS and the derived metabolites Hcyand H

2S in cancer pathogenesis and provides insight into

the development of novel prognosticmarkers and therapeuticapproaches for cancer patients

2 CBS Protein Structure andBiological Functions

The human CBS gene encodes a protein of 551 amino acidsThe crystal structure of the active form of human CBSformed by four of 63-kDa subunits has been fully character-ized [1 2] Each subunit consists of three structural domainsThe N-terminal domain binds to the cofactor heme whichis required for successful protein folding and assembly butnot necessary for catalytic activity [3] The catalytic domainencompasses a binding site for another cofactor pyridoxal-phosphate (PLP) [4] The C-terminal regulatory domaincontains two CBS motifs (CBS1 and CBS2) that dimerize toform a Bateman domain This domain is responsible for CBS

HindawiBioMed Research InternationalVolume 2018 Article ID 3205125 11 pageshttpsdoiorg10115520183205125

2 BioMed Research International

TranssulfurationDesulfuration

Cystathionine

Glutamylcysteine

Glutathione

3-Mercaptopyruvate

AHCY

MAT1A

MAT2A

CAT

3-MST

CBS

GSS

GCLC

SAM

SAHBetaine

DMG

BHMT

Homocysteine

Methionine

Cysteine

THF

THF

THF

5 10-Methylene

5-Methyl

Ser Gly Thr

MTHFR

SHMT

MTR

Methyl acceptors

Methylated acceptors

CTH

CBS

CTH

TransmethylationRemethylation

Homoserine + （2S

Lanthionine + （2S

（2S

Pyruvate + （2S

Cystathionine + （2S

Serine + （2S

Figure 1 Metabolic reactions catalyzed by CBS CBS catalyzes the condensation of homocysteine (Hcy) with serine to form cystathioninewhich is subsequently cleaved by cystathionine gamma-lyase (CTH) to form cysteine a precursor of glutathione CBS also catalyzes theproduction of H

2S In addition to CBS CTH and 3-mercaptopyruvate sulfurtransferase (3-MST) are also involved in the conversion

of cysteine to H2S Homocysteine is another key CBS-derived metabolite and is linked to the metabolism of methionine Methionine

is converted to homocysteine via S-adenosyl methionine (SAM) and S-adenosyl homocysteine (SAH) releasing a methyl group that isused in numerous methylation reactions SAM is an allosteric activator of CBS 3-MST 3-mercaptopyruvate sulfurtransferase AHCYadenosylhomocysteinase BHMT betaine-homocysteinemethyltransferase CAT cysteine aminotransferase CBS cystathionine 120573-synthaseCTH cystathionine gamma-lyase GCLC gamma-glutamylcysteine synthetase GSS glutathione synthetase MAT1A2A methionineadenosyltransferase 1A2AMTHFRmethylenetetrahydrofolate reductaseMTR 5-methyltetrahydrofolate-homocysteinemethyltransferaseSAM S-adenosyl methionine SAH S-adenosyl homocysteine SHMT serine hydroxymethyltransferase

subunit tetramerization and contains the binding sites for theallosteric activator S-adenosylmethionine (SAM) [1 5 6] Inthe native quaternary structure the access of substrates tothe catalytic core is occluded by the C-terminal regulatorymotifs and the binding of SAM induces a conformationalchange that improves the access of the substrates to thecatalytic site [2] The autoinhibitory function of the C-terminal regulatory domain is relieved by the C-terminaltruncation that generates a 45 kDa isoform with higher basalcatalytic activity than the full-length form [1]

CBS is predominantly expressed in the brain liverkidney and pancreas It is mainly a cytosolic enzyme butlocalization in the nucleus [7] and mitochondria [8] hadbeen detected in specific cell types CBS can be translo-cated to the mitochondria in response to hypoxia [9] ornucleolar stress [10] CBS expression is regulated at mul-tiple levels upon different stimuli For example hormonalregulation by glucocorticoids increases CBS expression atthe transcriptional level in liver cells a process that may beperturbed by insulin administration through binding to an

BioMed Research International 3

insulin-sensitive sequence localized on the CBS promoter[11] In addition testosterone can regulate CBS expressionand activity in renal tissue [12] Growthdifferentiationfactors such as EGF TGF-120572 cAMP and dexamethasoneinduced CBS protein expression in mouse astrocytes [13]Hypoxia upregulated CBS expression either via hypoxia-inducible factor- (HIF-) 1 at the transcriptional level [14]or decreased degradation of CBS protein by Lon proteasesin the mitochondria [9] Besides HIF-1 the zinc fingertranscription factor SP1 binds to the CBS gene promoterestablishing its role as a key regulator of CBS expression[15 16] Furthermore CBS activity may be enhanced viaposttranslational regulation through S-glutathionylation [17]or inhibited via epigenetic downregulation of CBS expressionthrough promoter methylation [18 19]

CBS plays a critical role in Hcy elimination Patients withCBS deficiency exhibit elevated Hcy plasma levels at excessof 200 120583M compared to 5-15 120583M in healthy adults [20] CBS-deficient homocystinuria (CBSDH) is an autosomal recessivemetabolic disease resulting from inactivating mutations inthe CBS gene CBSDH patients present multiple pathologicchanges in the eye skeleton central nervous and vascularsystems Common symptoms in CBSDH patients includethrombosis osteoporosis and impaired mental cognitivedevelopment (reviewed in [21ndash23]) Administration of highdoses of the PLP precursor pyridoxine or vitamin B

6is

common treatment that ameliorates approximately 50 ofclinical symptoms To date 164 pathogenic genetic variantshave been identified (httpcbslf1cuniczmutationsphp) ofwhich the predominant mutations are missense mutationsc833 TgtC (pI278T) is the most frequent mutation detectedin many European populations [24] The I278T missensemutation and many of the less prevalent mutations likelyaffect the folding or stability of the CBS protein [25] whereassomemutations such as mutant D444N amissense mutationin the C-terminal regulatory domain showed an approxi-mately twofold increase in basal CBS activity but impairedresponse to SAM stimulation [2] The pathophysiology ofCBS deficiency is still not fully understood As well as theaccumulation of Hcy CBS defects lead to increased con-centrations of methionine and S-adenosyl-L-homocysteine(SAH) and depletion of cystathionine and cysteine Theseperturbations may act in concert with high Hcy to promotethe development and progression of CBSDH (reviewed in[26])

Accordingly extensive studies in the mouse models ofCBS deficiency showed mice with homozygotic CBS deletion(CBS--) died within 4 weeks after birth due to severe hepaticdysfunction and exhibited extremely high levels of circulatingHcy (reviewed in [26 27]) Wang et al showed that theneonatal lethality could be rescued by decreasing circulatingHcy levels in a transgenic mouse model with inducible CBSexpression [28] They further found that there may be athreshold effect with Hcy meaning that moderately loweringhomocysteinemia can improve mouse viability during theneonatal period [29] In support of the Hcy threshold effectCBS+- heterozygote mice were fully viable with a 3-foldincrease of Hcy levels compared to the 8-fold increase inhomozygous mice [30]

3 Homocysteine and H2S the Major CBS-Derived Metabolites

31 Homocysteine Hcy is a sulfur-containing nonproteino-genic amino acid linked to the metabolism of methionineand cysteine Methionine is converted to Hcy via S-adenosylmethionine (SAM) and SAH releasing a methyl group thatis used in numerous methylation reactions Hcy can reformMet by the remethylation pathway either via 5-methyltetra-hydrofolate-homocysteine methyltransferase (MTR 5-meth-yltetrahydrofolate as the methyl group donor) or betaine-homocysteine methyltransferase (BHMT betaine as themethyl group donor) (Figure 1)Hcy is also irreversiblymetab-olized by CBS to cystathionine that subsequently converts tocysteine via CTH in the transsulfuration pathway (Figure 1)Hcy metabolism mainly occurs in the liver and conversion tocystathionine by CBS is amajor elimination route of Hcy [31]

Hyperhomocysteinemia (HHcy) is recognized as an inde-pendent risk factor for atherosclerotic vascular disease [32]HHcymay result frommutations in genes encoding enzymesof Hcy biosynthesis and metabolism or deficiencies of vita-min cofactors including vitamin B

12and B

6[33] The molec-

ular mechanisms underlying HHcy-induced atherosclerosisare complex and multifactorial (Figure 2) Elevated Hcyconcentration reduces nitric oxide (NO) bioavailability andcauses oxidative stress HHcy also leads to formation ofHcy thiolactone as a result of error-prone editing by themethionyl-tRNA synthase [34] This Hcy derivative cancause protein N-homocysteinylation in which the thioestergroup of thiolactone binds to the lysine residues in pro-teins consequently impairing protein function resulting inunfolded protein response and endoplasmic reticulum stress(reviewed in [35 36]) Moreover an elevated Hcy levelcould lead to accumulation of SAH a competitive inhibitorof most methyltransferases consequently inducing DNAhypomethylation [37] Through this epigenetic mechanismHcy has been reported to inhibit endothelial cell growth bydecreasing the expression of cyclin A [38] fibroblast growthfactor 2 [39] and hTERT expression [40] and by upregulationof platelet-derived growth factors and P66shC [41]

HHcy has also been implicated in the pathogenesis ofcancer Increased release of Hcy by tumor cells is relatedto their rapid proliferation rate [42] Hcy accumulationresults from defects in methionine synthesis leading to amethionine-dependent malignant phenotype [43] A meta-analysis revealed the association of elevated circulating Hcylevels with increased overall risk of cancer [44] A higherHcy plasma level has been detected in the patients withhepatocellular carcinoma (HCC) [44] and head and necksquamous cell carcinoma [45] Although the mechanismsunderlying this association between elevated Hcy levelsand malignant transformation are unclear a recent studyproposed a mechanism linking Hcy to lipid metabolismand HCC [46] It demonstrated that Hcy transcriptionallyupregulated CYP2J2 a cytochrome P450 (CYP) epoxygenaseby stimulating DNA demethylation and increasing SP1AP1activity on the promoter of CYP2J2 which promotes epoxye-icosatrienoic acid synthesis and hepatocellular tumorigene-sis


CBS

Hyperhomocysteinemia

Homocysteinethiolactone

Nrf2 activation+

Increased glutathioneproduction

Increased antioxidants

Protein sulfhydration

Modulation of protein activity

Vasorelaxation

MitochondrialETC

Unfolded proteinresponse

ER stress

DNA hypomethylationROS production

Normal CBSactivity

High CBSactivity

Low CBSactivity

SAM

SAH

I

II

III

IV

Oxidative stress

Homocysteine

HIGH

LOW

Biol

ogic

al eff

ect

Glutathione

＋+

４０ channel

[（2S]

Excess （2S

（2S

（2S

Figure 2 Potential mechanisms underlying CBS deregulation with alterations of homocysteine and H2S levels in cancer pathogenesis CBS

deficiency causes hyperhomocysteinemia Elevated Hcy concentration can increase reactive oxygen species (ROS) production and induceoxidative stress Hyperhomocysteinemia also leads to formation of homocysteine thiolactone as a result of error-prone editing by themethionyl-tRNA synthase This homocysteine derivative can cause protein N-homocysteinylation that impairs protein function resultingin an unfolded protein response and endoplasmic reticulum (ER) stress The elevated Hcy level can lead to accumulation of S-adenosylhomocysteine (SAH) a competitive inhibitor of most methyltransferases consequently inducing DNA hypomethylation and affectinggene transcription CBS-driven endogenous H

2S production maintains mitochondrial respiration and ATP synthesis promotes antioxidant

production by enhancing Nrf2 activation and increasing glutathione production and modulates protein activity via protein sulfhydrationSecreted H

2S can cause vasodilation via activation of ATP-sensitive K+ channels

32 H2S Like nitric oxide and carbon monoxide H

2S is

a diffusible gaseous transmitter in the human body and ismainly synthesized during cysteine metabolism and excretedas urinary sulfates by the kidney (reviewed in [47]) CBScatalyzes the production of H

2S via at least three path-

ways including (i) converting cysteine to serine and H2S

(ii) condensing cysteine and Hcy to yield cystathionineand H

2S and (iii) condensing two cysteine molecules to

lanthionine and H2S (Figure 1) In addition to CBS CTH

and 3-mercaptopyruvate sulfurtransferase (3-MST) are alsoinvolved in the conversion of cysteine to H

2S (Figure 1)

While H2S has diverse biological functions in the ner-

vous cardiovascular and immune systems the pathologicalrole of H

2S in cancer biology has attracted substantial

attention in recent years CBS-driven endogenous H2S pro-

duction has been reported to support tumor growth by (i)maintaining mitochondrial respiration and ATP synthesis(ii) stimulating cell proliferation and survival (iii) redoxbalance and (iv) vasodilation (Figure 2) H

2S modulates

mitochondrial functions and cellular bioenergetics in a

concentration-dependent manner At low concentrationsH

2S acts as a mitochondrial electron donor to mitochondrial

complex II resulting in bioenergetic stimulation [48 49] Athigher concentrations H

2S acts as a mitochondrial poison

via the inhibition of cytochrome c oxidase in mitochondrialcomplex IV [50] H

2S stimulates cell proliferation through

activation of specific kinase pathways (eg MAPK andPI3KAkt) and inhibition of selective phosphatases such asPTEN and PTP1B [51ndash53] Modulation of protein activityby H

2S either occurs via protein sulfhydration (reviewed

in [54]) or intracellular formation of polysulfides by H2S

followed by oxidative inactivation of proteins [55 56] Thesulfhydration of nuclear factor kappa B (NF-120581B) by H

2S has

also been shown to inhibit apoptosis andmay be of particularrelevance to cancer cell survival [57] The protective effectof H

2S from oxidative stress has been extensively studied

in endothelial cells and neurons [58ndash62] Studies showedH

2S inhibited H

2O

2-mediated mitochondrial dysfunction

by preserving the protein expression levels and activity ofkey antioxidant enzymes inhibiting reactive oxygen species


(ROS) production and lipid peroxidation [60] Additionallythese effects may be associated with sulfhydration of Keap1and activation of Nrf2 [61] or increasing the production ofthe antioxidant glutathione Vasorelaxation is one of the firstrecognized biological effects of H

2SThemechanisms ofH

2S-

mediated vasodilation include the activation of ATP-sensitiveK+ channels inhibition of phosphodiesterases and a synergywith NO (reviewed in [63])

H2S-donating compounds deliver H

2S exogenously

including fast H2S donors such as sulfate salts (eg NaHS

and Na2S) and naturally occurring compounds (eg the

garlic constituent diallyl trisulfide sulforaphane erucinand iberin) and slow H

2S-releasing synthetic moieties such

as GYY4137 (reviewed in [64]) The cellular response toexogenous H

2S released by the donors has been considered

as a biphasic response in which low H2S concentrations

(or low H2S production rates) showed enhancement of cell

proliferation rates and cell viability whereas high H2S caused

deleteriousadverse effects in cells [50 65] This biphasiccellular response is consistent with the special action modelof H

2S on mitochondrial respiration described above that

is stimulation of mitochondrial respiration at low levels andinhibition at high levels This bell-shape pharmacology ofH

2S may at least in part explain the inconsistent results of

the effect of exogenous H2S in colon cancer cell line HCT116

reported by different groups including a growth inhibitoryeffect (using NaHS at 400 120583M and 800 120583M) by the Deng lab[66] and a growth stimulatory effect (using NaHS at 30-300120583M) by the Szabo lab [49 65 67]

4 CBS and Cancer

41 Promoting Tumor Growth by Activation of CBS Elevatedexpression of CBS in tumor tissues or cell lines has beenreported in colon [49 68] ovarian [8] prostate [69] andbreast cancer [70] compared to adjacent normal tissue ornontransformed cells A series of studies from the Hellmichgroup characterized the oncogenic role of CBS in coloncancer [49 68 71] Through modification of CBS expres-sion (overexpression or RNAi knockdown) or CBS activity(allosteric activator SAM or the inhibitor aminooxyacetate)in the HCT116 colon cancer cell line they demonstrated thatCBS promoted cancer cell proliferation The antiproliferativeeffect observed by silencing or inhibiting CBS was recapit-ulated in the xenograft mouse models and patient-derivedtumor xenografts [49] CBS not only promotes tumor growthand progression but also initiates tumor formation [68]Overexpression of CBS in adenoma-like colonic epithelialcell line NCM356 enhanced cell proliferative anchorage-independent growth and invasive capability in vitro andtumorigenicity in vivo Mice heterozygous for CBS showedfewer numbers of mutagen-induced aberrant crypt focithan wild-type controls Through a similar approach Bhat-tacharyya et al [8] reported that CBS knockdown inhib-ited cell proliferation and suppressed tumor growth in anorthotopicmousemodel of cisplatin-resistant ovarian cancerInterestingly in breast cancer silencing CBS did not affectcell proliferation in culture but significantly attenuated tumorgrowth in a xenograft mouse model [70]

The protumorigenic effect of CBS occurs through anautocrine mechanism by regulation of bioenergetics antioxi-dant capacity and apoptosis-related pathways Targeting CBSgenetically or pharmacologically impairs cellular bioener-getics through inhibiting mitochondrial electron transportoxidative phosphorylation and glycolysis H

2S was identi-

fied to be responsible for such metabolic and bioenergeticrewiring in colon cancer cells as CBS expression and activitycorrelated with H

2S production and exogenous H

2S stim-

ulated cell proliferation and bioenergetics [49] Systematicmetabolomic analysis of CBS-overexpressing NCM356 cellsuncovered an anabolic metabolic phenotype with signifi-cantly enhanced glycolysis nucleotide synthesis and lipo-genesis which is thought to promote malignant transfor-mation [68] CBS may also promote tumor cell survival byincreasing cell intrinsic antioxidant capacity Ovarian cancercells depleted of CBS showed enhanced ROS productionAntioxidant glutathione but not H

2S fully rescued viability

of CBS-depleted cells suggesting that the effect of CBS inovarian cancer cells is mediated through regulation of ROSproduction by glutathione [8] Similarly reduced glutathioneabundance was observed in breast cancer cells upon CBSsilencing and was accompanied by decreased Nrf2 expression[72] CBS downregulation reduced antioxidant capacity andenhanced the sensitivity of cancer cells to chemotherapeuticdrugsThe cytoprotective effect of CBS is also associated withregulation of NF-120581B and p53 apoptosis-related signaling [8]A recent study further suggested CBS is involved in nucleolarstress-induced apoptosis [10]The authors demonstrated thattreatment of p53-- colon cancer cells with 5-fluorouracilcaused nucleolar stress which led to accumulation of theribosome-free form of ribosomal protein L3 (rpL3) rpL3decreased CBS protein abundance through suppression ofSP1-mediated CBS gene transcription and increase of CBSprotein degradation by translocation of CBS into mitochon-dria Decreased CBS abundance and in turn reductionof H

2S production have been suggested to contribute to

mitochondrial cytochrome C release and induction of theintrinsic cell death pathway [10]

In addition to autocrine regulation CBS acts via aparacrine mechanism to modulate the tumor microenviron-ment including stimulating angiogenesis and vasodilationvia H

2S production and release as reported in colon and

ovarian cancer xenografts [8 49] and regulating macrophageactivation in breast cancer xenograft mouse models [70]

42 CBS Associated Oncogenesis Is Tumor Type-SpecificUnlike in colon ovarian and breast cancer CBS does notappear to have a functional role in melanoma [73] CBSexpression is absent in dysplastic nevi detected in only25 of primary melanoma samples and unregulated in fourof five melanoma cell lines examined More importantlymodulation of CBS expression had a minimal effect onmelanoma cell proliferation [73]

Downregulation of CBS through promoter methylationhas been observed in multiple gastric cancer cell lines andfour colon cancer cell lines (including HCT116) [74] How-ever the biological consequence of CBS epigenetic silencingin gastric cancer has not been determined Evidence from


Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS

Figure 3 CBS associated oncogenesis is tumor type-specific Activation of CBS promotes tumor growth in colon ovarian and breast cancerbut suppresses tumor growth in gliomaThe role of CBS in liver cancer gastric cancer and melanoma is still conflicting and inconclusive

glioma supports a tumor-suppressive role for CBS [75] CBSdeficiency in U87-MG glioma cells did not affect cell prolif-eration in 2D culture but increased colony formation in softagar indicative of enhanced anchorage-independent growthConsistently CBS knockdown decreased tumor latency inU87-MGxenografts and increased tumor volume in an ortho-topic model Enhanced glioma tumorigenicity upon CBS losswas associated with upregulation of HIF-2120572 protein level andHIF-2120572-dependent transcriptional activation of angiopoietinlike 4 (ANGPTL4) and vascular endothelial growth factorA (VEGFA) The lack of function or suppression of tumorgrowth by CBS in certain tumor types indicates that CBSassociated oncogenesis is tumor-specific (Figure 3)

43 Conflicting Role of CBS in Hepatocellular CarcinomaClinical evidence from patient samples strongly supports anegative regulatory role for CBS in hepatocellular carcinoma(HCC) Downregulation of CBS expression and activitycontributes to the pathogenesis of multiple liver diseases(Reviewed in [76]) Analysis of 120 HCC specimens foundthat CBS mRNA was markedly lower in tumor tissuesthan surrounding noncancerous liver [77] Reduced CBSexpression was significantly correlated with the poor clinicpathological parameters including tumor stage Edmondsongrade alpha-fetoprotein (AFP) level and overall survivalFurther data analysis suggested that the expression levelof CBS mRNA could be used as a prognostic marker foroverall survival especially in patients with low AFP levels[77] Diminished CBS levels were also detected in the tumortissues from the mouse model of HCC [78ndash80] Furthersupporting the tumor-suppressive role for CBS exogenousH

2S induced autophagy and apoptosis in HCC cells through

the PI3KAktmTOR pathway [81]

Intriguingly distinct from this clinical data a recentstudy showed that several HCC cell lines exhibited higherCBS expression than normal liver cells HL-7702 and QSG-7701 [82] Both genetic (by siRNA) and pharmacological (byAOAA) inhibition of CBS in the SMMC-7721 HCC cell linewith reduced H

2S production decreased cell viability and

enhanced ROS production in vitro Another study showingthat the PI3KAKT pathway regulated the CTHH

2S to

promote HCC proliferation also supports the oncogenic roleof H

2S in HCC [53] Clearly the biological function of CBS

in liver cancer is complex and requires further investigation

5 CBS in Cancer Therapy

Consistent with the complex roles of CBS in cancer biologydescribed above it is also becoming evident that both theactivators and inhibitors of CBS have antitumor activity indifferent cancer models This genetic context dependencedetermines different types of cancer will display distinct effi-cacy and toxicity profiles in response to CBS-based targetedtherapies

51 CBS Inhibitors Aminooxyacetate (AOAA) is currentlyconsidered as the most potent CBS inhibitor compared withthe other drugs such as trifluoroalanine and hydroxylamine[65] It has shown antitumor actions in the mouse xenograftmodels of colon [49] and breast cancer [83] and patient-derived colon cancer xenografts [49] Decreased H

2S level

in plasma was detected in a colon xenograft mouse modeltreated with AOAA while the drug effect on circulating Hcylevel was not investigated While these antitumor responsesare encouraging the therapeutic effect of CBS inhibitionrequires further investigation as AOAA is actually not


selective for CBS [65 84] The pharmacological action ofAOAA is not limited to suppression of the CBS H

2S axis

It binds irreversibly to the cofactor PLP and therefore inaddition to CBS it inhibits other PLP-dependent enzymessuch as CTH 3-MST and glutamate oxaloacetate transam-inase 1 (GOT1) AOAA has been reported to target CTHpreferentially overCBS (IC50 852120583MforCBS versus 109120583Mfor CTH) [85] Furthermore inhibition of GOT1 by AOAAdisrupted the malateaspartate shuttle decreased glucose-derived carbon flux into mitochondrial tricarboxylic acidcycle and ATP synthesis [83]

To identify new CBS inhibitors two groups performedsmall-molecule screening [86 87] The Barrios group [87]and the Wu group [86] used recombinant CBS enzymes andemployed fluorescent H

2S readouts to screen a composite

library of 1900 compounds and a chemical library consist-ing of 20000 compounds respectively Several compoundsshowed some selectivity for CBS compared with CTH withIC50 20-50 120583MHowever as the studies did not use AOAA asa reference in the screen whether these drugs are superior toAOAA in terms of potency and selectivity remains unknown

52 CBS Activator S-Adenosyl-L-Methionine (SAM) SAM isa vital molecule for transmethylation and transsulfurationreactions It is the principle methyl-donor for DNA aminoacid protein and lipid methyltransferase and a key precursorfor glutathione and polyamine synthesis (reviewed by [88])It is synthesized from methionine and ATP by methionineadenosyltransferase (MAT Figure 1) SAM as an allostericactivator modulates CBS activity by inducing a conforma-tional change in the C-terminus of CBS that facilitates theentrance of substrates into the catalytic site of the enzyme [1]Although SAM has been used for treatment of osteoarthritis[89] depression [90] and liver diseases [88] the clinicalevidence for its efficacy in these diseases is still inconclusiveRecent data support the concept of using SAMas a chemopre-ventive agent in HCC and colon cancer consistent with theproposed tumor-suppressive role of CBS in HCCTheMat1aknockout mice spontaneously develop HCC supporting thefact that hepatic SAM deficiency predisposes to HCC [91]In several rodent models of HCC administration of SAMis effective in preventing liver carcinogenesis [92 93] Onephase II clinical trial is evaluating SAM as a potentialchemoprevention agent in patients with hepatitis C cirrhosis[94] SAM also showed a similar chemoprevention effect inan inflammation induced colon cancer mouse model [95] Inaddition to chemoprevention SAM exerted a proapoptoticeffect in liver (at 02mMover 5 days) [96] gastric (10120583Mover7 days) [97] and colon cancer cells (ranging from 025 to 5mMfor 24 hours) [98] Interestingly similar to the conflictingdata regarding CBS function and effects of H

2S donors

in colon cancer the Szabo group [71] reported a biphasicresponse to SAM in colon cancer cells At low concentrationsfor the short-time period (01-1 mM for 12 hours or 01 mMfor 24 hours) SAM induced a stimulatory effect on CBS acti-vation H

2S production and cell proliferation while at higher

concentrations or chronic exposure (01-5mMafter 24 hours)the inhibitory effects became more prominent and were notattenuated by CBS silencing suggesting nonspecificity or

toxicity [71] Therefore more work in multiple experimentmodels is required to better define the role of SAMCBS axisin cancer pathogenesis

6 CBS in Cancer Prognosis

With the identification of the pathogenic role of CBS in can-cer the use of CBS as a prognostic and predictive biomarkeris becoming attractive As described above the negativecorrelation of CBS expression with the pathologic parametersin HCC indicates its potential as a prognostic marker in HCC[77] Modulation of CBS activity can be indicated by thechanges of Hcy andor H

2S levels The potential prognostic

values of Hcy in cancer have been extensively studied [99ndash101] However the biological sources of Hcy were not definedin these studies and thus the link between the levels ofHcy and CBS function remains unknown Neverthelesssignificant progress in the detection and quantitation of Hcyfrompatient samples has been made in recent years Methodsof measuring plasma Hcy have evolved from ion-exchangechromatography to high-performance liquid chromatogra-phy (HPLC) gas-chromatography mass spectrometry liq-uid chromatography-electrospray tandemmass spectrometry(LC-MSMS) and fluorescence polarization immunoassay(FPIA) [102] In terms of H

2S elevated H

2S in exhaled

breath or its degraded form in urine in cancer patientsprovides support for the clinical utility of H

2S as a marker of

cancer [101] However in order to determine the prognosticand predictive values of H

2S in cancer development of

the methods that can accurately measure H2S levels in the

circulation or in the targeted organs is imperative

7 Summary and Future Directions

A functional role for CBS in tumor biology is supported by(i) clinical evidence of altered CBS expression level and CBS-derived Hcy and H

2S levels in cancer patients (ii) preclinical

studies showing dysregulation of CBS function and activityin cancer cell culture and animal models (iii) mechanisticinvestigations linking CBS to cancer-related cellular andmolecular changes and signaling pathways The distinctbiological effects of CBS alterations in different cancermodelsreveal the complexity of CBS signaling in cancer pathogene-sisThe contradictory role of CBS in cancer biology (Figure 3)is possibly due to the existence of alternative Hcy and H

2S

metabolic pathways and multiple modes of regulation ofCBS expression and activity by hormones growth factorsand other metabolites Therefore the functional role ofCBS is determined by the distinct metabolic and geneticprofiles in different types of cancer and is context-dependentFurthermore the current conflicting data adds an additionallayer of complexity indicating that multiple experimentaland analytical approaches as well as in-depth mechanisticinvestigations are required to clarify the role of CBS in cancerbiology

Increased understanding of the role of theCBS-controllednetwork in cancer biology will greatly promote the devel-opment of pharmacological reagents targeting CBS and theidentification of appropriate patient populations CBS acts


through two main metabolites Hcy and H2S which have

important physiological roles in specific tissues such as theliver brain and blood vessels Given its central metabolicrole it is possible that CBS-based targeted therapy may causeside effects due to accumulation of unfavorable metabolitesFor example CBS inhibitors may elevate Hcy levels withpotential risk for developing HHcyTherefore further studieswill be required to define the therapeutic windows of thenovel CBS targeting agents Additional investigations areclearly required to better elucidate the complex role of CBSin malignant transformation including (i) characterizing therole of CBS-related metabolic signaling in cancer pathogene-sis including but not limited toCBSHcy H

2S and the related

enzymes (ii) determining the interaction of tumor cell-derived CBS and its metabolites with the microenvironment(iii) identifying biomarkers of CBS-based therapies in clinicalsamples and cancer models Certainly a greater appreciationfor the complexity of CBS in cancer biology will give rise tonew prospective biomarkers or targets for cancer

Conflicts of Interest

The authors declare that there are no conflicts of interestregarding the publication of this paper

Authorsrsquo Contributions

Richard B Pearson and Jian Kang contributed equally to thiswork

Acknowledgments

The authors apologize to the authors of many primaryresearch papers that were not cited here due to spaceconstraints but whose work influenced their understandingsubstantially This work was supported by the NationalHealth andMedical ResearchCouncil (NHMRC) ofAustraliaproject and program grants and Cancer Council VictoriaResearchers were funded by NHMRC Fellowships (RichardB Pearson) a Melbourne International Research Scholarship(Haoran Zhu) and Research Training Program Scholarship(the University of Melbourne) (Shaun Blake)

References

[1] J Ereno-Orbea T Majtan I Oyenarte J P Kraus and L AMartınez-Cruza ldquoStructural basis of regulation and oligomer-ization of human cystathionine 120573-synthase the central enzymeof transsulfurationrdquo Proceedings of the National Acadamy ofSciences of the United States of America vol 110 no 40 ppE3790ndashE3799 2013

[2] J Ereno-Orbea T Majtan I Oyenarte J P Kraus and L AMartinez-Cruz ldquoStructural insight into the molecular mecha-nism of allosteric activation of human cystathionine 120573-synthaseby S-adenosylmethioninerdquoProceedings of theNational Acadamyof Sciences of the United States of America vol 111 no 37 ppE3845ndashE3852 2014

[3] T Majtan L R Singh L Wang W D Kruger and J P KrausldquoActive cystathionine 120573-synthase can be expressed in heme-freesystems in the presence of metal-substituted porphyrins or a

chemical chaperonerdquo The Journal of Biological Chemistry vol283 no 50 pp 34588ndash34595 2008

[4] M Meier M Janosik V Kery J P Kraus and P BurkhardldquoStructure of human cystathionine 120573-synthase a unique pyri-doxal 51015840-phosphate-dependent heme proteinrdquo EMBO Journalvol 20 no 15 pp 3910ndash3916 2001

[5] V Kery L Poneleit and J P Kraus ldquoTrypsin cleavage ofhuman cystathionine 120573-synthase into an evolutionarily con-served active core Structural and functional consequencesrdquoArchives of Biochemistry and Biophysics vol 355 no 2 pp 222ndash232 1998

[6] K-H Jhee P McPhie and EWMiles ldquoDomain architectureofthe heme-independent yeast cystathionine 120573-synthase providesinsights intomechanisms of catalysis and regulationrdquo Biochem-istry vol 39 no 34 pp 10548ndash10556 2000

[7] O Kabil Y Zhou and R Banerjee ldquoHuman cystathionine 120573-synthase is a target for sumoylationrdquo Biochemistry vol 45 no45 pp 13528ndash13536 2006

[8] S Bhattacharyya S Saha K Giri et al ldquoCystathionine Beta-Synthase (CBS) Contributes to Advanced Ovarian CancerProgression and Drug Resistancerdquo PLoS ONE vol 8 no 11Article ID e79167 2013

[9] H Teng B Wu K Zhao G Yang L Wu and R WangldquoOxygen-sensitive mitochondrial accumulation of cystathion-ine 120573-synthase mediated by Lon proteaserdquo Proceedings of theNational Acadamy of Sciences of the United States of Americavol 110 no 31 pp 12679ndash12684 2013

[10] V Pagliara A Saide E Mitidieri et al ldquo5-FU targets rpL3 toinduce mitochondrial apoptosis via cystathionine-120573-synthasein colon cancer cells lacking p53rdquo Oncotarget vol 7 no 31 pp50333ndash50348 2016

[11] S Ratnam K N Maclean R L Jacobs M E Brosnan J PKraus and J T Brosnan ldquoHormonal regulation of cystathionine120573-synthase expression in liverrdquoThe Journal of Biological Chem-istry vol 277 no 45 pp 42912ndash42918 2002

[12] V Vitvitsky A Prudova S Stabler S Dayal S R Lentz andR Banerjee ldquoTestosterone regulation of renal cystathionine 120573-synthase Implications for sex-dependent differences in plasmahomocysteine levelsrdquo American Journal of Physiology-RenalPhysiology vol 293 no 2 pp F594ndashF600 2007

[13] Y Enokido E Suzuki K Iwasawa K Namekata H Okazawaand H Kimura ldquoCystathionine 120573-synthase a key enzyme forhomocysteine metabolism is preferentially expressed in theradial gliaastrocyte lineage of developing mouse CNSrdquo TheFASEB Journal vol 19 no 13 pp 1854ndash1856 2005

[14] N Takano Y-J Peng G K Kumar et al ldquoHypoxia-induciblefactors regulate human and rat cystathionine beta-synthasegene expressionrdquo Biochemical Journal vol 458 no 2 pp 203ndash211 2014

[15] K N Maclean E Kraus and J P Kraus ldquoThe Dominant Roleof Sp1 in Regulating the Cystathionine 120573-Synthase -1a and -1b Promoters Facilitates Potential Tissue-specific Regulation byKruppel-like FactorsrdquoThe Journal of Biological Chemistry vol279 no 10 pp 8558ndash8566 2004

[16] Y Ge M A Konrad L H Matherly and J W Taub ldquoTran-scriptional regulation of the human cystathionine 120573-synthase -1b basal promoter Synergistic transactivation by transcriptionfactors NF-Y and Sp1Sp3rdquo Biochemical Journal vol 357 no 1pp 97ndash105 2001

[17] W-N Niu P K Yadav J Adamec and R Banerjee ldquoS-glutathionylation enhances human cystathionine 120573-synthase


activity under oxidative stress conditionsrdquo Antioxidants ampRedox Signaling vol 22 no 5 pp 350ndash361 2015

[18] F Qi Y Zhou Y Xiao et al ldquoPromoter demethylation ofcystathionine-120573-synthetase gene contributes to inflammatorypain in ratsrdquo PAIN vol 154 no 1 pp 34ndash45 2013

[19] H-H Zhang J Hu Y-L Zhou et al ldquoPromoted interac-tion of nuclear factor-120581B with demethylated cystathionine-120573-synthetase gene contributes to gastric hypersensitivity indiabetic ratsrdquo The Journal of Neuroscience vol 33 no 21 pp9028ndash9038 2013

[20] W D Kruger L Wang K H Jhee R H Singh and LJ Elsas II ldquoCystathionine 120573-Synthase Deficiency in Georgia(USA) Correlation of Clinical andBiochemical PhenotypewithGenotyperdquoHuman Mutation vol 22 no 6 pp 434ndash441 2003

[21] M Meier J Oliveriusova J P Kraus and P Burkhard ldquoStruc-tural insights into mutations of cystathionine 120573-synthaserdquoBiochimica et Biophysica Acta (BBA) - Proteins and Proteomicsvol 1647 no 1-2 pp 206ndash213 2003

[22] M Yamanishi O Kabil S Sen and R Banerjee ldquoStruc-tural insights into pathogenic mutations in heme-dependentcystathionine-120573-synthaserdquo Journal of Inorganic Biochemistryvol 100 no 12 pp 1988ndash1995 2006

[23] A A M Morris V Kozich S Santra et al ldquoGuidelines forthe diagnosis and management of cystathionine beta-synthasedeficiencyrdquo Journal of Inherited Metabolic Disease vol 40 no 1pp 49ndash74 2017

[24] V E Shih J M Fringer R Mandell et al ldquoA missense muta-tion (I278T) in the cystathionine 120573-synthase gene prevalentin pyridoxine-responsive homocystinuria and associated withmild clinical phenotyperdquo American Journal of Human Geneticsvol 57 no 1 pp 34ndash39 1995

[25] E W Miles and J P Kraus ldquoCystathionine 120573-synthase Struc-ture function regulation and location of homocystinuria-causingmutationsrdquoThe Journal of Biological Chemistry vol 279no 29 pp 29871ndash29874 2004

[26] W D Kruger ldquoCystathionine 120573-synthase deficiency Of miceand menrdquo Molecular Genetics and Metabolism vol 121 no 3pp 199ndash205 2017

[27] M Watanabe J Osada Y Aratani et al ldquoMice deficient incystathionine 120573-synthase animal models for mild and severehomocyst(e)inemiardquo Proceedings of the National Acadamy ofSciences of the United States of America vol 92 no 5 pp 1585ndash1589 1995

[28] L Wang K-H Jhee X Hua P M DiBello D W Jacobsenand W D Kruger ldquoModulation of cystathionine 120573-synthaselevel regulates total serum homocysteine in micerdquo CirculationResearch vol 94 no 10 pp 1318ndash1324 2004

[29] S Gupta J Kuhnisch A Mustafa et al ldquoMouse models ofcystathionine 120573-synthase deficiency reveal significant thresholdeffects of hyperhomocysteinemiardquoThe FASEB Journal vol 23no 3 pp 883ndash893 2009

[30] N Tyagi N Qipshidze U SenW Rodriguez A Ovechkin andS C Tyagi ldquoCystathionine beta synthase gene dose dependentvascular remodeling in murine model of hyperhomocysteine-miardquo International Journal of Physiology Pathophysiology andPharmacology vol 3 no 3 pp 210ndash222 2011

[31] K Robert J Nehme E Bourdon et al ldquoCystathionine 120573synthase deficiency promotes oxidative stress fibrosis andsteatosis in mice liverrdquo Gastroenterology vol 128 no 5 pp1405ndash1415 2005

[32] L M Graham L E Daly H M Refsum et al ldquoPlasmahomocysteine as a risk factor for vascular diseaseTheEuropean

Concerted Action Projectrdquo Journal of the American MedicalAssociation vol 277 no 22 pp 1775ndash1781 1997

[33] S Brustolin R Giugliani and T M Felix ldquoGenetics ofhomocysteine metabolism and associated disordersrdquo BrazilianJournal of Medical and Biological Research vol 43 no 1 pp 1ndash72010

[34] H Jakubowski ldquoProofreading in vivo Editing of homocysteinebymethionyl-tRNA synthetase in Escherichia colirdquo Proceedingsof the National Acadamy of Sciences of the United States ofAmerica vol 87 no 12 pp 4504ndash4508 1990

[35] WK C Lai andMY Kan ldquoHomocysteine-induced endothelialdysfunctionrdquo Annals of Nutrition andMetabolism vol 67 no 1pp 1ndash12 2015

[36] X C Wang W T Sun C M Yu et al ldquoER stress mediateshomocysteine-induced endothelial dysfunction modulation ofIKCa and SKCa channelsrdquo Atherosclerosis vol 242 no 1 pp191ndash198 2015

[37] S Zhou Z Zhang and G Xu ldquoNotable epigenetic role ofhyperhomocysteinemia in atherogenesisrdquo Lipids in Health andDisease vol 13 no 1 article no 134 2014

[38] M S Jamaluddin I Chen F Yang et al ldquoHomocysteine inhibitsendothelial cell growth via DNA hypomethylation of the cyclinA generdquo Blood vol 110 no 10 pp 3648ndash3655 2007

[39] P-Y Chang S-C Lu C-M Lee et al ldquoHomocysteine inhibitsarterial endothelial cell growth through transcriptional down-regulation of fibroblast growth factor-2 involving G protein andDNAmethylationrdquoCirculationResearch vol 102 no 8 pp 933ndash941 2008

[40] D Zhang X Sun J Liu X Xie W Cui and Y Zhu ldquoHomo-cysteine accelerates senescence of endothelial cells via DNAhypomethylation of human telomerase reverse transcriptaserdquoArteriosclerosis Thrombosis and Vascular Biology vol 35 no1 pp 71ndash78 2015

[41] C-S Kim Y-R Kim A Naqvi et al ldquoHomocysteine promoteshuman endothelial cell dysfunction via site-specific epigeneticregulation of p66shcrdquoCardiovascular Research vol 92 no 3 pp466ndash475 2011

[42] C-F Sun T R Haven T-L Wu K-C Tsao and J T WuldquoSerum total homocysteine increases with the rapid prolifera-tion rate of tumor cells and decline upon cell death A potentialnew tumor markerrdquo Clinica Chimica Acta vol 321 no 1-2 pp55ndash62 2002

[43] P Cavuoto and M F Fenech ldquoA review of methionine depen-dency and the role of methionine restriction in cancer growthcontrol and life-span extensionrdquoCancer Treatment Reviews vol38 no 6 pp 726ndash736 2012

[44] D Zhang XWenWWu Y Guo andW Cui ldquoElevated homo-cysteine level and folate deficiency associated with increasedoverall risk of carcinogenesis Meta-analysis of 83 case-controlstudies involving 35758 individualsrdquo PLoS ONE vol 10 no 5Article ID e0123423 2015

[45] GAlmadori F Bussu J Galli et al ldquoSerum folate andhomocys-teine levels in head and neck squamous cell carcinomardquoCancervol 94 no 4 pp 1006ndash1011 2002

[46] D Zhang J Lou X Zhang et al ldquoHyperhomocysteine-mia results from and promotes hepatocellular carcinoma viaCYP450metabolism by CYP2J2 DNAmethylationrdquoOncotarget vol 8 no 9 pp 15377ndash15392 2017

[47] C Szabo ldquoGasotransmitters in cancer From pathophysiologyto experimental therapyrdquo Nature Reviews Drug Discovery vol15 no 3 pp 185ndash203 2016


[48] E Lagoutte S Mimoun M Andriamihaja C ChaumontetF Blachier and F Bouillaud ldquoOxidation of hydrogen sulfideremains a priority in mammalian cells and causes reverseelectron transfer in colonocytesrdquo Biochimica et Biophysica Acta(BBA) - Bioenergetics vol 1797 no 8 pp 1500ndash1511 2010

[49] C Szabo C Coletta C Chao et al ldquoTumor-derived hydro-gen sulfide produced by cystathionine-120573-synthase stimulatesbioenergetics cell proliferation and angiogenesis in coloncancerrdquo Proceedings of the National Acadamy of Sciences of theUnited States of America vol 110 no 30 pp 12474ndash12479 2013

[50] C Szabo C Ransy K Modis et al ldquoRegulation of mito-chondrial bioenergetic function by hydrogen sulfide Part IBiochemical and physiological mechanismsrdquo British Journal ofPharmacology vol 171 no 8 pp 2099ndash2122 2014

[51] W-J Cai M-J Wang P K Moore H-M Jin T Yao and Y-C Zhu ldquoThe novel proangiogenic effect of hydrogen sulfide isdependent on Akt phosphorylationrdquo Cardiovascular Researchvol 76 no 1 pp 29ndash40 2007

[52] P Manna and S K Jain ldquoHydrogen sulfide and L-cysteineincrease phosphatidylinositol 345-trisphosphate (PIP3) andglucose utilization by inhibiting phosphatase and tensinhomolog (PTEN) protein and activating phosphoinositide 3-kinase (PI3K)serinethreonine protein kinase (AKT)proteinkinase Czetalambda (PKCzetalambda) in 3T3l1 adipocytesrdquoThe Journal of Biological Chemistry vol 286 no 46 pp 39848ndash39859 2011

[53] P Yin C Zhao Z Li et al ldquoSp1 is involved in regulation ofcystathionine 120574-lyase gene expression and biological functionby PI3KAkt pathway in human hepatocellular carcinoma celllinesrdquo Cellular Signalling vol 24 no 6 pp 1229ndash1240 2012

[54] B D Paul and S H Snyder ldquoH 2S signalling through proteinsulfhydration and beyondrdquo Nature Reviews Molecular CellBiology vol 13 no 8 pp 499ndash507 2012

[55] R Greiner Z Palinkas K Basell et al ldquoPolysulfides link H2S to

protein thiol oxidationrdquoAntioxidants amp Redox Signaling vol 19no 15 pp 1749ndash1765 2013

[56] Y Kimura Y Mikami K Osumi M Tsugane J-I Oka andH Kimura ldquoPolysulfides are possible H

2S-derived signaling

molecules in rat brainrdquo The FASEB Journal vol 27 no 6 pp2451ndash2457 2013

[57] N Sen B D Paul M M Gadalla et al ldquoHydrogen sulfide-linked sulfhydration of NF-120581B mediates its antiapoptoticactionsrdquoMolecular Cell vol 45 no 1 pp 13ndash24 2012

[58] P B L Pun J Lu E M Kan and S Moochhala ldquoGases in themitochondriardquoMitochondrion vol 10 no 2 pp 83ndash93 2010

[59] K Suzuki G Olah K Modis et al ldquoHydrogen sulfide replace-ment therapy protects the vascular endothelium in hyper-glycemia by preserving mitochondrial functionrdquo Proceedings ofthe National Acadamyof Sciences of the United States of Americavol 108 no 33 pp 13829ndash13834 2011

[60] Y-D Wen H Wang S-H Kho et al ldquoHydrogen sulfideprotects HUVECs against hydrogen peroxide induced mito-chondrial dysfunction and oxidative stressrdquo PLoS ONE vol 8no 2 Article ID e53147 2013

[61] G Yang K Zhao and Y Ju ldquoHydrogen sulfide protects againstcellular senescence via S-sulfhydration of keap1 and activationof Nrf2rdquo Antioxidants amp Redox Signaling vol 18 no 15 pp1906ndash1919 2013

[62] S Koike Y Ogasawara N Shibuya H Kimura and K IshiildquoPolysulfide exerts a protective effect against cytotoxicity causedby t-buthylhydroperoxide through Nrf2 signaling in neuroblas-toma cellsrdquo FEBS Letters vol 587 no 21 pp 3548ndash3555 2013

[63] R Wang ldquoPhysiological implications of hydrogen sulfide awhiff exploration that blossomedrdquo Physiological Reviews vol92 no 2 pp 791ndash896 2012

[64] K Kashfi and K R Olson ldquoBiology and therapeutic potentialof hydrogen sulfide and hydrogen sulfide-releasing chimerasrdquoBiochemical Pharmacology vol 85 no 5 pp 689ndash703 2013

[65] M R Hellmich C Coletta C Chao and C Szabo ldquoThetherapeutic potential of cystathionine 120573-synthetasehydrogensulfide inhibition in cancerrdquo Antioxidants amp Redox Signalingvol 22 no 5 pp 424ndash448 2015

[66] Z W Lee J Zhou C-S Chen et al ldquoThe slow-releasingHydrogen Sulfide donor GYY4137 exhibits novel anti-cancereffects in vitro and in vivordquo PLoS ONE vol 6 no 6 Article IDe21077 2011

[67] W-J Cai M-J Wang L-H Ju C Wang and Y-C ZhuldquoHydrogen sulfide induces human colon cancer cell prolifera-tion Role of Akt ERK and p21rdquo Cell Biology International vol34 no 6 pp 565ndash572 2010

[68] CM Phillips J R ZatarainM E Nicholls et al ldquoUpregulationof cystathionine-120573-synthase in colonic epithelia reprogramsmetabolism and promotes carcinogenesisrdquo Cancer Researchvol 77 no 21 pp 5741ndash5754 2017

[69] H Guo J-W Gai Y Wang H-F Jin J-B Du and J Jin ldquoChar-acterization of hydrogen sulfide and its synthases cystathionine120573-synthase and cystathionine 120574-lyase in human prostatic tissueand cellsrdquo Urology vol 79 no 2 pp 483e1ndash483e5 2012

[70] S Sen B Kawahara D Gupta et al ldquoRole of cystathionine120573-synthase in human breast Cancerrdquo Free Radical Biology ampMedicine vol 86 pp 228ndash238 2015

[71] K Modis C Coletta A Asimakopoulou et al ldquoEffect ofS-adenosyl-l-methionine (SAM) an allosteric activator ofcystathionine-120573-synthase (CBS) on colorectal cancer cell pro-liferation and bioenergetics in vitrordquo Nitric Oxide Biology andChemistry vol 41 pp 146ndash156 2014

[72] B Kawahara T Moller K Hu-Moore et al ldquoAttenuation ofAntioxidant Capacity in Human Breast Cancer Cells by CarbonMonoxide through Inhibition of Cystathionine 120573-SynthaseActivity Implications in Chemotherapeutic Drug SensitivityrdquoJournal of Medicinal Chemistry vol 60 no 19 pp 8000ndash80102017

[73] E Panza P De Cicco C Armogida et al ldquoRole of the cystathio-nine 120574 lyasehydrogen sulfide pathway in human melanomaprogressionrdquo Pigment Cell amp Melanoma Research vol 28 no1 pp 61ndash72 2015

[74] H Zhao Q Li J Wang et al ldquoFrequent epigenetic silencingof the folate-metabolising gene cystathionine-beta-synthase ingastrointestinal Cancerrdquo PLoS ONE vol 7 no 11 Article IDe49683 2012

[75] N Takano Y Sarfraz D M Gilkes et al ldquoDecreased expressionof cystathionine 120573-synthase promotes glioma tumorigenesisrdquoMolecular Cancer Research vol 12 no 10 pp 1398ndash1406 2014

[76] L K Sarna Y L Siow and O Karmin ldquoThe CBSCSE systemA potential therapeutic target in NAFLDrdquo Canadian Journal ofPhysiology and Pharmacology vol 93 no 1 pp 1ndash11 2015

[77] J Kim S J Hong J H Park et al ldquoExpression of cystathionine120573-synthase is downregulated in hepatocellular carcinoma andassociated with poor prognosisrdquo Oncology Reports vol 21 no6 pp 1449ndash1454 2009

[78] MAAvila C Berasain L Torres et al ldquoReducedmRNAabun-dance of themain enzymes involved inmethioninemetabolismin human liver cirrhosis and hepatocellular carcinomardquo Journalof Hepatology vol 33 no 6 pp 907ndash914 2000


[79] A Prudova Z Bauman A Braun V Vitvitsky S C Lu andR Banerjee ldquoS-adenosylmethionine stabilizes cystathionine 120573-synthase and modulates redox capacityrdquo Proceedings of theNational Acadamy of Sciences of the United States of Americavol 103 no 17 pp 6489ndash6494 2006

[80] D F Calvisi M M Simile S Ladu et al ldquoAltered methioninemetabolism and global DNA methylation in liver cancer rela-tionship with genomic instability and prognosisrdquo InternationalJournal of Cancer vol 121 no 11 pp 2410ndash2420 2007

[81] S S Wang Y H Chen N Chen et al ldquoHydrogen sulfidepromotes autophagy of hepatocellular carcinoma cells throughthe PI3KAktmTOR signaling pathwayrdquo Cell Death amp Diseasevol 8 no 3 Article ID e2688 2017

[82] H Jia J Ye J You X Shi W Kang and T Wang ldquoRole of thecystathionine 120573-synthaseH2S system in liver cancer cells andthe inhibitory effect of quinolone-indolone conjugate QIC2 onthe systemrdquoOncology Reports vol 37 no 5 pp 3001ndash3009 2017

[83] J M Thornburg K K Nelson B F Clem et al ldquoTargetingaspartate aminotransferase in breast cancerrdquo Breast CancerResearch vol 10 no 5 article no R84 2008

[84] K Modis E M Bos E Calzia et al ldquoRegulation of mito-chondrial bioenergetic function by hydrogen sulfide Part IIPathophysiological and therapeutic aspectsrdquo British Journal ofPharmacology vol 171 no 8 pp 2123ndash2146 2014

[85] A Asimakopoulou P Panopoulos C T Chasapis et alldquoSelectivity of commonly used pharmacological inhibitors forcystathionine 120573 synthase (CBS) and cystathionine 120574 lyase(CSE)rdquo British Journal of Pharmacology vol 169 no 4 pp 922ndash932 2013

[86] Y Zhou J Yu X Lei et al ldquoHigh-throughput tandem-microwell assay identifies inhibitors of the hydrogen sulfidesignaling pathwayrdquo Chemical Communications vol 49 no 100pp 11782ndash11784 2013

[87] M K Thorson T Majtan J P Kraus and A M BarriosldquoIdentification of Cystathionine 120573-Synthase Inhibitors Using aHydrogen Sulfide Selective Proberdquo Angewandte Chemie Inter-national Edition vol 52 no 17 pp 4641ndash4644 2013

[88] S C Lu and J M Mato ldquoS-adenosylmethionine in liver healthinjury and cancerrdquoPhysiological Reviews vol 92 no 4 pp 1515ndash1542 2012

[89] A W Rutjes E Nuesch S Reichenbach and P Juni ldquoS-Adenosylmethionine for osteoarthritis of the knee or hiprdquoCochrane Database of Systematic Reviews (Online) no 4 pCD007321 2009

[90] I Galizia L Oldani K Macritchie et al ldquoS-adenosyl methio-nine (SAMe) for depression in adultsrdquo Cochrane Database ofSystematic Reviews vol 2016 no 10 Article ID CD011286 2016

[91] M L Martınez-Chantar F J Corrales L A Martınez-Cruzet al ldquoSpontaneous oxidative stress and liver tumors in micelacking methionine adenosyltransferase 1Ardquo The FASEB Jour-nal vol 16 no 10 pp 1292ndash1294 2002

[92] R M Pascale M M Simile M R D Miglio et al ldquoChemopre-vention by s-adenosyl-l-methionine of rat liver carcinogenesisinitiated by 12-dimethylhydrazine and promoted by oroticacidrdquo Carcinogenesis vol 16 no 2 pp 427ndash430 1995

[93] S C Lu K Ramani X Ou et al ldquoS-adenosylmethionine in thechemoprevention and treatment of hepatocellular carcinoma ina rat modelrdquoHepatology vol 50 no 2 pp 462ndash471 2009

[94] T R Morgan ldquoChemoprevention of hepatocellular carcinomain chronic hepatitis Crdquo Recent Results in Cancer Research vol188 pp 85ndash99 2011

[95] T W H Li H Yang H Peng M Xia J M Mato and S C LuldquoEffects of S-adenosylmethionine and methylthioadenosine oninflammation-induced colon cancer in micerdquo Carcinogenesisvol 33 no 2 pp 427ndash435 2012

[96] Y Wang Z Sun and M Szyf ldquoS-adenosyl-methionine (SAM)alters the transcriptome and methylome and specifically blocksgrowth and invasiveness of liver cancer cellsrdquo Oncotarget vol8 no 67 pp 111866ndash111881 2017

[97] J Luo Y-N Li F Wang W-M Zhang and X Geng ldquoS-adenosylmethionine inhibits the growth of cancer cells byreversing the hypomethylation status of c-myc and H-ras inhuman gastric cancer and colon cancerrdquo International Journalof Biological Sciences vol 6 no 7 pp 784ndash795 2010

[98] T W H Li Q Zhang P Oh et al ldquoS-adenosylmethionineand methylthioadenosine inhibit cellular FLICE inhibitoryprotein expression and induce apoptosis in colon cancer cellsrdquoMolecular Pharmacology vol 76 no 1 pp 192ndash200 2009

[99] L L Wu and J T Wu ldquoHyperhomocysteinemia is a risk factorfor cancer and a new potential tumor markerrdquo Clinica ChimicaActa vol 322 no 1-2 pp 21ndash28 2002

[100] Y Ozkan S Yardim-Akaydin H Firat E Caliskan-Can SArdic and B Simsek ldquoUsefulness of homocysteine as a cancermarker Total thiol compounds and folate levels in untreatedlung cancer patientsrdquoAnticancer Reseach vol 27 no 2 pp 1185ndash1189 2007

[101] M R Hellmich and C Szabo ldquoHydrogen sulfide and cancerrdquoHandbook of Experimental Pharmacology vol 230 pp 233ndash2412015

[102] H J Powers and S J Moat ldquoDevelopments in the measurementof plasma total homocysteinerdquo Current Opinion in ClinicalNutrition amp Metabolic Care vol 3 no 5 pp 391ndash397 2000

Stem Cells International

Hindawiwwwhindawicom Volume 2018


MEDIATORSINFLAMMATION

of

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

Hindawi Publishing Corporation httpwwwhindawicom Volume 2013Hindawiwwwhindawicom

The Scientific World Journal

Volume 2018

Immunology ResearchHindawiwwwhindawicom Volume 2018

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine


Behavioural Neurology

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary andAlternative Medicine

Volume 2018Hindawiwwwhindawicom

Submit your manuscripts atwwwhindawicom


TranssulfurationDesulfuration

Cystathionine

Glutamylcysteine

Glutathione

3-Mercaptopyruvate

AHCY

MAT1A

MAT2A

CAT

3-MST

CBS

GSS

GCLC

SAM

SAHBetaine

DMG

BHMT

Homocysteine

Methionine

Cysteine

THF

THF

THF

5 10-Methylene

5-Methyl

Ser Gly Thr

MTHFR

SHMT

MTR

Methyl acceptors

Methylated acceptors

CTH

CBS

CTH

TransmethylationRemethylation

Homoserine + （2S

Lanthionine + （2S

（2S

Pyruvate + （2S

Cystathionine + （2S

Serine + （2S

Figure 1 Metabolic reactions catalyzed by CBS CBS catalyzes the condensation of homocysteine (Hcy) with serine to form cystathioninewhich is subsequently cleaved by cystathionine gamma-lyase (CTH) to form cysteine a precursor of glutathione CBS also catalyzes theproduction of H

2S In addition to CBS CTH and 3-mercaptopyruvate sulfurtransferase (3-MST) are also involved in the conversion

of cysteine to H2S Homocysteine is another key CBS-derived metabolite and is linked to the metabolism of methionine Methionine

is converted to homocysteine via S-adenosyl methionine (SAM) and S-adenosyl homocysteine (SAH) releasing a methyl group that isused in numerous methylation reactions SAM is an allosteric activator of CBS 3-MST 3-mercaptopyruvate sulfurtransferase AHCYadenosylhomocysteinase BHMT betaine-homocysteinemethyltransferase CAT cysteine aminotransferase CBS cystathionine 120573-synthaseCTH cystathionine gamma-lyase GCLC gamma-glutamylcysteine synthetase GSS glutathione synthetase MAT1A2A methionineadenosyltransferase 1A2AMTHFRmethylenetetrahydrofolate reductaseMTR 5-methyltetrahydrofolate-homocysteinemethyltransferaseSAM S-adenosyl methionine SAH S-adenosyl homocysteine SHMT serine hydroxymethyltransferase

subunit tetramerization and contains the binding sites for theallosteric activator S-adenosylmethionine (SAM) [1 5 6] Inthe native quaternary structure the access of substrates tothe catalytic core is occluded by the C-terminal regulatorymotifs and the binding of SAM induces a conformationalchange that improves the access of the substrates to thecatalytic site [2] The autoinhibitory function of the C-terminal regulatory domain is relieved by the C-terminaltruncation that generates a 45 kDa isoform with higher basalcatalytic activity than the full-length form [1]

CBS is predominantly expressed in the brain liverkidney and pancreas It is mainly a cytosolic enzyme butlocalization in the nucleus [7] and mitochondria [8] hadbeen detected in specific cell types CBS can be translo-cated to the mitochondria in response to hypoxia [9] ornucleolar stress [10] CBS expression is regulated at mul-tiple levels upon different stimuli For example hormonalregulation by glucocorticoids increases CBS expression atthe transcriptional level in liver cells a process that may beperturbed by insulin administration through binding to an




6is






12and B

6[33] The molec-




CBS



Nrf2 activation+





Vasorelaxation

MitochondrialETC


ER stress


Normal CBSactivity

High CBSactivity

Low CBSactivity

SAM

SAH

I

II

III

IV

Oxidative stress

Homocysteine

HIGH

LOW

Biol

ogic

al eff

ect

Glutathione

＋+

４０ channel

[（2S]

Excess （2S

（2S

（2S







2S is








2S (Figure 1)






2S modulates












2S has




2S inhibited H

2O





2SThemechanisms ofH

2S-



2S exogenously
















4 CBS and Cancer



2S was identi-



2S stim-












Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS









2S to







2S level




2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of






















6is






12and B

6[33] The molec-




CBS



Nrf2 activation+





Vasorelaxation

MitochondrialETC


ER stress


Normal CBSactivity

High CBSactivity

Low CBSactivity

SAM

SAH

I

II

III

IV

Oxidative stress

Homocysteine

HIGH

LOW

Biol

ogic

al eff

ect

Glutathione

＋+

４０ channel

[（2S]

Excess （2S

（2S

（2S







2S is








2S (Figure 1)






2S modulates












2S has




2S inhibited H

2O





2SThemechanisms ofH

2S-



2S exogenously
















4 CBS and Cancer



2S was identi-



2S stim-












Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS









2S to







2S level




2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of




















CBS



Nrf2 activation+





Vasorelaxation

MitochondrialETC


ER stress


Normal CBSactivity

High CBSactivity

Low CBSactivity

SAM

SAH

I

II

III

IV

Oxidative stress

Homocysteine

HIGH

LOW

Biol

ogic

al eff

ect

Glutathione

＋+

４０ channel

[（2S]

Excess （2S

（2S

（2S







2S is








2S (Figure 1)






2S modulates












2S has




2S inhibited H

2O





2SThemechanisms ofH

2S-



2S exogenously
















4 CBS and Cancer



2S was identi-



2S stim-












Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS









2S to







2S level




2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of





















2SThemechanisms ofH

2S-



2S exogenously
















4 CBS and Cancer



2S was identi-



2S stim-












Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS









2S to







2S level




2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of




















Glioma

Ovarian Cancer

Breast Cancer

Colon Cancer

Liver Cancer

Stomach Cancer

Melanoma

CBS









2S to







2S level




2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of





















2S axis






2S donors









2S elevated H

2S in exhaled


2S as a marker of









2S






2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of






















2S and the related






Acknowledgments


References




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of




















































































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















































































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of
















































of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of























of




Disease Markers



OncologyJournal of





PPAR Research



Volume 2018


Journal of

ObesityJournal of



















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