1、Cancer CellReviewThe Intestinal Microbiotain Colorectal CancerHerbert Tilg,1,3,*Timon E.Adolph,1,3Romana R.Gerner,1,2,3and Alexander R.Moschen1,2,31Department of Internal Medicine I,Gastroenterology,Hepatology&Endocrinology,Medical University Innsbruck,Innsbruck,Austria2Christian Doppler Laboratory
2、of Mucosal Immunology,Medical University Innsbruck,Innsbruck,Austria3These authors contributed equally*Correspondence:herbert.tilgi-med.ac.athttps:/doi.org/10.1016/j.ccell.2018.03.004Experimental evidence from the past years highlights a key role for the intestinal microbiota in inflammatoryand mali
3、gnant gastrointestinal diseases.Diet exhibits a strong impact on microbial composition and pro-vides risk for developing colorectal carcinoma(CRC).Large metagenomic studies in human CRC associatedmicrobiome signatures with the colorectal adenoma-carcinoma sequence,suggesting a fundamental role ofthe
4、 intestinal microbiota in the evolution of gastrointestinal malignancy.Basic science established a criticalfunction for the intestinal microbiota in promoting tumorigenesis.Further studies are needed to decipher themechanisms of tumor promotion and microbial co-evolution in CRC,which may be exploite
5、d therapeuticallyin the future.IntroductionThe impact of the intestinal microbiota on health and disease isincreasingly emerging.While the microbiota evolved as a ne-glected organ in the early 2000s,our perception in 2018 hasvastly changed in that the microbiota reflects a biologicalecosystem that i
6、ntensely communicates with the host(Char-bonneau et al.,2016;Lynch and Pedersen,2016;Marchesiet al.,2016;Sonnenburg and Backhed,2016).A hype in micro-biota research in the last decade allowed to delineate thecomposition and some functions of the intestinal microbiota,which wasempowered bythe introdu
7、ctionof modern moleculartechniques.Current estimates suggest that the gastrointestinaltract contains as much bacteria as cells composing the humanbody(Sender et al.,2016).Although we can adequately depictthe enormous diversity of the human microbiota,the role ofmostbacterialspeciesinhealth anddiseas
8、eremainslargelyun-known.Key examples for a deranged interplay between thehost and the microbiota may be derived from complex diseasesincluding inflammatory bowel disease(IBD)and the tumor(mi-cro-)environment in colorectal carcinoma(CRC).Despite enor-mous efforts and substantial progress in understan
9、ding thecomposition of the human intestinal microbiota,many func-tional aspects remain unresolved.This may be partly due tothe complexity of the human intestinal microbiota,with aplethora of unpredictable host-microbe,microbe-microbe,and environmental interactions.CRC,one of the most common malignan
10、cies in the westernworld,frequently causes death and is emerging worldwide.Itis expected that CRC burden will substantially increase inthe next two decades consequent to adoption of a westernlifestyle(Arnold et al.,2017).It is well established that con-sumption of foods and nutrients affect the risk
11、 for developingCRC(Song et al.,2015).Dietary habits have substantiallychanged in the last decades especially in the western worldand might affect colorectal carcinogenesis at various steps.Diet may exhibit effects on the hosts immune response andelicit inflammation.In addition,dietary behavior treme
12、ndouslyinfluences the composition of the intestinal microbiota,whichin turn impacts on the susceptibility to intestinal diseases(Shanahan et al.,2017)(Figure 1).Exposure to antibiotics earlyin life is associated with an increased risk for colorectal ade-noma at the age of 60(Cao et al.,2017),which s
13、uggeststhat a dysbiotic microbiota is acquired and sustained over alonger period of time(Gensollen et al.,2016).The colon(i.e.,large intestine)and ileocecal valve exhibit the highest bacterialdensity along the gastrointestinal tract,which might point to-ward an important role for the microbiota in C
14、RC.Indeed,studies in CRC patients and experimental evidence in animalslinked the intestinal microbiota with CRC,which led to theidentification of specific bacterial species that promote tumor-igenesis(Drewes et al.,2016;Louis et al.,2014;Sears andGarrett,2014).This review summarizes clinical informa
15、tionon the microbiota in CRC and aims to bridge to preclinical(i.e.,experimental)evidence that the microbiota is a drivingforce in the development of intestinal tumors.Specific Bacterial Strains Associated with CRCCRC is frequently associated with dramatic alterations in themicrobial composition of
16、the tumor and adjacent mucosa,commonly termed as dysbiosis(Feng et al.,2015;Liang et al.,2017;Nakatsu et al.,2015;Tsoi et al.,2017;Yazici et al.,2017;Yu et al.,2017a).Dysbiosis is partly characterized by the expan-sion of bacterial taxa;however,dominant species in CRC evolu-tion remain poorly define
17、d.Experimental evidence for an impor-tant role of Fusobacterium nucleatum(Fn),Escherichia coli,andBacteroides fragilis is emerging(Figure 2).In contrast,the role ofdepleted bacterial species in intestinal tumorigenesis is less wellstudied due to a lack of appropriate techniques.However,it maybe spec
18、ulated that the absence of certain(e.g.,protective orbeneficial)bacterial strains could be as important as tumor-associated bacterial overgrowth.Furthermore,we would like tohighlight that the intestinal microbiota also comprises virusesand fungi;however,their role in CRC isnot covered in this review
19、as clinical evidence is largely lacking(Collins et al.,2011).Cancer Cell 33,June 11,2018 2018 Elsevier Inc.1Please cite this article in press as:Tilg et al.,The Intestinal Microbiota in Colorectal Cancer,Cancer Cell(2018),https:/doi.org/10.1016/j.ccell.2018.03.004Fusobacterium nucleatumThe Gram-nega
20、tive,anaerobic commensal Fn was associatedwith human CRC in two initial studies by genomic analysis(Cas-tellarin et al.,2012;Kostic et al.,2012).Fusobacterium se-quences were shown to be enriched in a small number of CRCpatients and were also visualized within tumors using fluores-cence in situ hybr
21、idization(Kostic et al.,2012).Castellarin et al.(2012)linked CRC with Fn expansion as demonstrated byqPCR analysis of 99 patients,which correlated with lymphnode metastasis.Ahn et al.(2013)performed one of the first16SrRNA-basedanalysesinCRCanddemonstratedadecreased bacterial diversity in feces,depl
22、etion of Gram-posi-tive fiber-fermenting Clostridia,and an increase in oral commen-sals Fn and Porphyromonas.These data appeared particularlyinteresting in light of the pro-inflammatory role of Fn in the intes-tine and its association with human IBD(Ohkusa et al.,2003).Figure 1.Multidimensional Fram
23、ework ofCRC EvolutionMalignant degeneration of intestinal epithelialcells(IECs)and progression to CRC involves a complexinterplay from various layers of extrinsic andintrinsic factors.Together these influences resultin oncogenic mutations in Lgr5+intestinal stemcells(SCs),in altered b-catenin/Wnt si
24、gnaling,andin pro-inflammatory programs that drive CRC.(1)Extrinsic,predominantly nutritional factors candirectly damage host DNA,modulate the compo-sition and metabolic activity of the gut microbiota,interfere with gut barrier functions,affect IECmetabolism,and influence immune functions.(2)The mic
25、robiota influences intestinal tumorigenesisthrough several mechanisms.Several CRC-asso-ciated species such as Fusobacterium nucleatum,colibactin-producing Escherichia coli,and en-terotoxigenicBacteroidesfragilishavebeenimplicated in DNA damage and tumor progres-sion.Microbial metabolism of complex c
26、arbohy-drates,bile acids,and luminal iron,including hemeiron,are important for barrier function and immunehomeostasis as are bacterial signals integratedby surface and intracellular pattern recognitionreceptors such as Toll-like receptors,NOD-likereceptors,and inflammasomes expressed byvarious cells
27、 types.(3)Chronic inflammation rep-resents an important intrinsic factor that promotescarcinogenesis by inducing DNA damage,andreactive oxygen and nitrogen species,by modu-lation of IEC polarization and the tumor microen-vironment,by activating transcriptional programssuch as nuclear factor kB and S
28、TAT3 in IECs,andby hampering anti-tumoral immunity.Furthermore,Fn expansion is associatedwith periodontal disease which is linkedto an increased CRC risk(Momen-Heraviet al.,2017).One study not only revealedincreased presence of Fn both in high-grade dysplasia and established CRCbutalsoacorrelationwi
29、thpatientoutcome(Flanagan et al.,2014).Enrich-ment of Fn in cancer tissue was alsoassociatedwithshortersurvivalinanother study and may therefore act aspotential prognostic marker(Mima et al.,2016b).Fn expression was related to mi-crosatellite instability(MSI)independent from CpG island meth-ylator p
30、henotype(CIMP)and BRAF mutation status(Mima et al.,2016b).Fnpresence alsocorrelated withCIMP-highlesions,andFnwas increasingly detected inmalignant(CRC56%)comparedwithbenignlesions(hyperplastic polyp24%and sessileserratedadenomas 35%)(Ito et al.,2015).As Fn abundance increasesfrom rectum(2.5%)to cec
31、um(11%),this bacterial strain maybe particularly relevant in specific colonic locations(e.g.,right-sided tumors)(Mima et al.,2016a).A large patient cohort fromJapan demonstrated Fn colonization in 8.6%of CRC subjectsand confirmed the association with MSI(Nosho et al.,2016).Inanother clinical study,F
32、n was detected in CRC tissue in 76(13%)of 598 cases,and was inversely associated with the den-sityofCD3+Tcellssupportingtheideathatdiseasemechanismsinvolve regulation of immune responses(Mima et al.,2015).Fnabundance was not only associated with promotion of CRC2Cancer Cell 33,June 11,2018Cancer Cel
33、lReviewPlease cite this article in press as:Tilg et al.,The Intestinal Microbiota in Colorectal Cancer,Cancer Cell(2018),https:/doi.org/10.1016/j.ccell.2018.03.004but furthermore conferred chemoresistance and recurrence inCRC patients by interfering with TLR4 and MyD88 signaling(Yuetal.,2017b).Fnwas
34、showntotargetspecific microRNAsre-sulting in activation of the autophagy pathway,thereby alteringthe chemotherapeutic response(Yu et al.,2017b).Remarkably,the CRC-associated microbiota including Fn can be detectedin metastases(Bullman etal.,2017).Mouse xenografts of humanprimary CRC retained viable
35、Fn,and antibiotic treatmentreduced Fn load,tumor cell proliferation,and growth(Bullmanet al.,2017).As such,clinical evidence is accumulating that Fnmight promote colonic neoplasia.This notion is also supported by experimental evidence.Fn isabletoinhibitnaturalkiller(NK)cell killingofvarioustumors,an
36、dthis effect is mediated by the human(but not mouse)T cellimmunoglobulin and ITIM domain(TIGIT)(Gur et al.,2015).The inhibitory receptor TIGIT is present both on human NK cellsand T cells,and this inhibitory effect has been demonstrated tobe dependent on the Fap2 protein of Fn.These data indicatetha
37、t Fn-derived factors are able to regulate tumor-immuneevasion.Furthermore,infection of CRC cells with Fn increasestheir proliferation rate,invasive activity,and potential to inducexenograft tumors in mice(Yangetal.,2017).Fn hasbeenshownto modulate the tumor-immune microenvironment and theFigure 2.Mi
38、crobial MechanismsInfluencing Cancer Development andProgressionThe tumor-associated luminal environment repre-sents a niche characterized by an impaired barrierfunction and a cluster of commensal bacteria thathave been implicated in tumor initiation and pro-gression.(1)Fusobacteriumnucleatums(Fn)Fad
39、A antigen binds E-cadherin on IECs to activateb-catenin.This leads to uncontrolled cell growth,acquisition of a stem cell-like phenotype,loss of cellpolarization,and possibly MSI instability.By mech-anisms including TLR4 and MyD88,Fn has pro-in-flammatory effects on the tumor microenvironment.Furthe
40、rmore,Fn has been shown to modulate auto-phagy in IECs by activating regulatory microRNAs.(2)E.coli strains harboring the polyketide synthases(pks)island encoding the genotoxin colibactin arefrequently observed in human colorectal tumors.Besides the genotoxic activity of colibactin,cellgrowth is sus
41、tained by cellular senescence associ-atedwiththeexpressionofhepatocytegrowthfactor(HGF).(3)EnterotoxigenicBacteroidesfragilis(ETBF)causes inflammation and tumors in animal models.ETBF induces spermine oxidase(SPO)generatingreactive oxygen species(ROS),thereby inducingDNA damage.ETBF is associated wi
42、th Th17 re-sponses.(4)Certain sulfate-reducing bacteria suchas Bilophila wadsworthia or Alistipes spp.producehydrogen sulfide(H2S)capable of inducing geno-toxic insults.Both strains promote inflammation insusceptible animals.E-cadherin/b-catenin signaling pathway.Rubinsteinandcolleaguesdemon-strated
43、 that Fn,through FadA adhesion,binds to E-cadherin thereby activatingtheb-cateninsignalingpathwayresultingin induction of oncogenic and inflamma-tory responses(Rubinstein et al.,2013).Importantly,FadA gene expression inhuman CRC tissue is extraordinarily high compared withhealthy controls,and inhibi
44、tion of this pathway protectedagainst pro-oncogenic activity.Fn was able to increase tumormultiplicity in the adenomatous polyposis coli/multiple intesti-nal neoplasia(ApcMin/+)model of intestinal tumorigenesis(Kos-tic et al.,2013).Increased tumor load was paralleled byincreased tumor infiltration o
45、f myeloid cells and pro-inflamma-tory cytokine expression.Notably,Fn did not deteriorate colitis,enteritis,or inflammation-associated intestinal carcinogenesis,demonstratingthatFnparticularlycontrolstumorigenesisinthismodel.These studies support the idea that Fn plays a role inhuman CRC.The mechanis
46、ms deserve further attention butmay involve immune evasion and/or the promotion of an inflam-matory tumor microenvironment.Escherichia coliWhile Fn is among the most prevalent bacterial strains in CRCtissues and its relevance in CRC is supported by both clinicaland pre-clinical studies,the evidence
47、for E.coli is mainly basedon preclinical investigations.E.coli are gut commensals,although certain strains have acquired the ability to promote in-testinal inflammation and to produce toxins such as colibactinwith oncogenic potential(Denizot et al.,2015).A western diet af-fects microbial composition
48、 and enhances susceptibility towardCancer Cell 33,June 11,20183Cancer CellReviewPlease cite this article in press as:Tilg et al.,The Intestinal Microbiota in Colorectal Cancer,Cancer Cell(2018),https:/doi.org/10.1016/j.ccell.2018.03.004the pathogenic potential of adherent invasive E.coli(Agus et al.
49、,2016).Mucosa-associated E.coli are significantly more preva-lent in CRC tissue and correlate with tumor stage and prognosis(Bonnet et al.,2014).Interestingly,pathogenic colibactin-ex-pressing E.coli strains were more prevalent in advanced dis-ease,andcolonizationofacoloncancer-associatedE.colistrai
50、ninto ApcMin/+mice resulted in a marked increase in number ofpolyps,suggesting that certain E.coli strains might indeed pro-mote tumorigenesis.Studies using bioluminescent inflammationprobes and fluorescence optical imaging also showed a correla-tion of E.coli with pro-inflammatory infiltrates which
