基于scRNA-seq揭示卵巢癌外泌体相关特征并构建预后模型

中文摘要、结论在本研究中，通过一系列的生物信息学分析揭示了卵巢癌的外泌体相关特征并最终构建了预后预测模型。首先，为揭示卵巢癌的细胞组成情况，利用scRNA-seq数据，通过质控，降维聚类，共注释出7种细胞类型。通过分析不同样本中细胞类型占比，发现上皮细胞在组织中显著富集。还利用scRNA-seq与ST-seq数据结合，将注释好的scRNA-seq数据映射到ST-seq数据上，构建了卵巢癌的细胞图谱。其次，通过inferCNV分析，推断出了上皮细胞中的恶性细胞。基于流行的三种算法对ERG进行了scRNA-seq和ST-seq数据的富集评分并取均值作为最终的ERS。结果表明，恶性细胞与单核/巨噬细胞具有较高的ERS。并且通过对不同ERS评分恶性细胞进行差异分析和KEGG富集分析，揭示了恶性细胞中外泌体相关的氧化磷酸化（OXPHOS）代谢通路。接下来，基于伪时间轨迹分析，揭示了恶性细胞发育过程中ERG表达呈现上调趋势，并获取在恶性细胞发育过程中显著变化的基因。基于细胞间通讯分析，剖析了不同细胞间的通讯模式，结果表明单核/巨噬细胞与高低ERS两组恶性细胞在癌症的PD-L1表达和PD-1检查点通路上的富集存在差异。并深入挖掘了单核/巨噬细胞与恶性细胞间的L-R-TF，通过对TF调控的靶基因进行富集分析，揭示其所参与的与细胞增殖、肿瘤生长相关的信号通路。最后，基于前期所获得的三个基因集构建了PPI网络。通过衡量了网络中每个节点的点度中心性、中介中心性以及PCI，鉴定出其中的关键基因。随后，通过三种经典回归分析方法，识别出5个与卵巢癌患者生存显著相关的基因，并利用TCGA数据库中卵巢癌患者bulkRNA-seq数据构建了预后预测模型。综上所述，本研究构建了卵巢癌的细胞图谱，揭示卵巢癌外泌体的相关特征；基于不同ERS恶性细胞间的差异，揭示了恶性细胞外泌体相关的OXPHOS代谢通路；并剖析了高低ERS两组恶性细胞的动态变化及细胞通讯模式；通过上述一系列生物学分析，最终筛选出与卵巢癌患者OS呈现显著相关的5个基因，更开发出了具有临床决策支持价值的工具，为推进卵巢癌的精准医疗策略提供了新的生物标志物及理论依据。参考文献ADDINEN.REFLIST1. FilhoAM,LaversanneM,FerlayJ,ColombetM,PiñerosM,ZnaorA,ParkinDM,SoerjomataramI,BrayF.TheGlobocan2022CancerEstimates:DataSources,Methods,andaSnapshotoftheCancerBurdenWorldwide.InternationalJournalofCancer.2025;156(7):1336-1346.2. LiT,ZhangH,LianM,HeQ,LvM,ZhaiL,ZhouJ,WuK,YiM.GlobalStatusandAttributableRiskFactorsofBreast,Cervical,Ovarian,andUterineCancersfrom1990to2021.JournalofHematology&Oncology.2025;18(1):5.3. HanB,ZhengR,ZengH,WangS,SunK,ChenR,LiL,WeiW,HeJ.CancerIncidenceandMortalityinChina,2022.JournaloftheNationalCancerCenter.2024;4(1):47-53.4. CroftPK-d,SharmaS,GodboleN,RiceGE,SalomonC.Ovarian-Cancer-AssociatedExtracellularVesicles:MicroenvironmentalRegulationandPotentialClinicalApplications.Cells.2021;10(9).5. DaiJ,SuY,ZhongS,CongL,LiuB,YangJ,TaoY,HeZ,ChenC,JiangY.Exosomes:KeyPlayersinCancerandPotentialTherapeuticStrategy.SignalTransductionandTargetedTherapy.2020;5(1):145.6. MashouriL,YousefiH,ArefAR,AhadiAm,MolaeiF,AlahariSK.Exosomes:Composition,Biogenesis,andMechanismsinCancerMetastasisandDrugResistance.MolecularCancer.2019;18(1):75.7. ZhangL,YuD.ExosomesinCancerDevelopment,Metastasis,andImmunity.BiochimicaetBiophysicaActa(BBA)-ReviewsonCancer.2019;1871(2):455-468.8. ZhuJW,CharkhchiP,AkbariMR.PotentialClinicalUtilityofLiquidBiopsiesinOvarianCancer.MolecularCancer.2022;21(1):114.9. MoY,LeungLL,MakCSL,WangX,ChanW-S,HuiLMN,TangHWM,SiuMKY,SharmaR,XuD,TsuiSKW,NganHYS,YungMMH,ChanKKL,ChanDW.Tumor-SecretedExosomalMir-141ActivatesTumor-StromaInteractionsandControlsPremetastaticNicheFormationinOvarianCancerMetastasis.MolecularCancer.2023;22(1):4.10. HeL,ZhuW,ChenQ,YuanY,WangY,WangJ,WuX.OvarianCancerCell-SecretedExosomalMir-205PromotesMetastasisbyInducingAngiogenesis.Theranostics.2019;9(26):8206-8220.11. KhanM,ZhaoZ,AroojS,FuY,LiaoG.SolublePd-1:Predictive,Prognostic,andTherapeuticValueforCancerImmunotherapy.FrontiersinImmunology.2020;Volume11-2020.12. NiuM,LiuY,YiM,JiaoD,WuK.BiologicalCharacteristicsandClinicalSignificanceofSolublePd-1/Pd-L1andExosomalPd-L1inCancer.FrontiersinImmunology.2022;Volume13-2022.13. ChoKR,ShihI-M.OvarianCancer.AnnualReviewofPathology:MechanismsofDisease.2009;4(1):287-313.14. YinJ,YanXFau-YaoX,YaoXFau-ZhangY,ZhangYFau-ShanY,ShanYFau-MaoN,MaoNFau-YangY,YangYFau-PanL,PanL.SecretionofAnnexinA3fromOvarianCancerCellsandItsAssociationwithPlatinumResistanceinOvarianCancerPatients.(1582-4934(Electronic)).15. BarkleyDA-O,MoncadaR,PourM,LibermanDA,DrygI,WerbaG,WangW,BaronM,RaoAA-O,XiaBA-O,FrançaGS,WeilA,DelairDF,HajduC,LundAW,OsmanI,YanaiIA-O.CancerCellStatesRecuracrossTumorTypesandFormSpecificInteractionswiththeTumorMicroenvironment.(1546-1718(Electronic)).16. XuJA-O,FangYA-O,ChenKA-O,LiSA-O,TangSA-O,RenYA-O,CenYA-OX,FeiWA-O,ZhangB,ShenYA-O,LuWA-O.Single-CellRnaSequencingRevealstheTissueArchitectureinHumanHigh-GradeSerousOvarianCancer.(1557-3265(Electronic)).17. WangX,ChaiZ,PanG,HaoY,LiBA-O,YeT,LiYA-OX,LongF,XiaL,LiuMA-O.Exobcd:AComprehensiveDatabaseforExosomalBiomarkerDiscoveryinBreastCancer.Lid-Bbaa088[Pii]Lid-10.1093/Bib/Bbaa088[Doi].(1477-4054(Electronic)).18. HaoY,StuartTA-O,KowalskiMH,ChoudharySA-O,HoffmanPA-O,HartmanA,SrivastavaA,MollaG,MadadS,Fernandez-GrandaC,SatijaRA-O.DictionaryLearningforIntegrative,MultimodalandScalableSingle-CellAnalysis.(1546-1696(Electronic)).19. HafemeisterC,SatijaRA-O.NormalizationandVarianceStabilizationofSingle-CellRna-SeqDataUsingRegularizedNegativeBinomialRegression.(1474-760X(Electronic)).20. KorsunskyIA-O,MillardN,FanJA-O,SlowikowskiK,ZhangFA-O,WeiK,BaglaenkoYA-O,BrennerM,LohPA-O,RaychaudhuriSA-O.Fast,SensitiveandAccurateIntegrationofSingle-CellDatawithHarmony.(1548-7105(Electronic)).21. AndreattaM,CarmonaSJ.Ucell:RobustandScalableSingle-CellGeneSignatureScoring.(2001-0370(Print)).22. QiuXA-O,MaoQ,TangY,WangL,ChawlaR,PlinerHA,TrapnellCA-O.ReversedGraphEmbeddingResolvesComplexSingle-CellTrajectories.(1548-7105(Electronic)).23. ZhangY,LiuT,HuX,WangM,WangJ,ZouB,TanP,CuiT,DouY,NingL,HuangY,RaoS,WangDA-O,ZhaoX.Cellcall:IntegratingPairedLigand-ReceptorandTranscriptionFactorActivitiesforCell-CellCommunication.(1362-4962(Electronic)).24. HuC,LiT,XuY,ZhangX,LiF,BaiJ,ChenJ,JiangW,YangK,OuQ,LiXA-O,WangPA-O,ZhangY.Cellmarker2.0:AnUpdatedDatabaseofManuallyCuratedCellMarkersinHuman/MouseandWebToolsBasedonScrna-SeqData.(1362-4962(Electronic)).25. MaY,ZhouX.SpatiallyInformedCell-TypeDeconvolutionforSpatialTranscriptomics.NatureBiotechnology.2022;40(9):1349-1359.26. 魏思亮.外泌体介导环状rna调节肺腺癌细胞pc9对吉非替尼耐药的相关研究.硕士,2020.27. BarkleyD,MoncadaR,PourM,LibermanDA,DrygI,WerbaG,WangW,BaronM,RaoA,XiaB,FrançaGS,WeilA,DelairDF,HajduC,LundAW,OsmanI,YanaiI.CancerCellStatesRecuracrossTumorTypesandFormSpecificInteractionswiththeTumorMicroenvironment.NatureGenetics.2022;54(8):1192-1201.28. KanehisaM,GotoS.Kegg:KyotoEncyclopediaofGenesandGenomes.(0305-1048(Print)).29. 裴夏明,李鸿,梁宇,谢宇.生物标志物炎症因子、细胞焦亡因子和铁死亡因子在膀胱癌治疗与预后中的价值.重庆医科大学学报.2023;48(08):863-868.30. BergersG,BenjaminLE.TumorigenesisandtheAngiogenicSwitch.(1474-175X(Print)).31. Pàez-RibesM,AllenEFau-HudockJ,HudockJFau-TakedaT,TakedaTFau-OkuyamaH,OkuyamaHFau-ViñalsF,ViñalsFFau-InoueM,InoueMFau-BergersG,BergersGFau-HanahanD,HanahanDFau-CasanovasO,CasanovasO.AntiangiogenicTherapyElicitsMalignantProgressionofTumorstoIncreasedLocalInvasionandDistantMetastasis.(1878-3686(Electronic)).32. WuY,HooperAtFau-ZhongZ,ZhongZFau-WitteL,WitteLFau-BohlenP,BohlenPFau-RafiiS,RafiiSFau-HicklinDJ,HicklinDJ.TheVascularEndothelialGrowthFactorReceptor(Vegfr-1)SupportsGrowthandSurvivalofHumanBreastCarcinoma.(0020-7136(Print)).33. QiuX,LiY,ZhangZ.CrosstalkbetweenOxidativePhosphorylationandImmuneEscapeinCancer:ANewConceptofTherapeuticTargetsSelection.CellularOncology.2023;46(4):847-865.34. WuY,YangS,MaJ,ChenZA-O,SongG,RaoD,ChengY,HuangS,LiuY,JiangS,LiuJ,HuangX,WangX,QiuS,XuJA-O,XiR,BaiF,ZhouJ,FanJ,ZhangX,GaoQ.SpatiotemporalImmuneLandscapeofColorectalCancerLiverMetastasisatSingle-CellLevel.(2159-8290(Electronic)).35. ZhaoS,WangQ,NiK,ZhangP,LiuY,XieJ,JiW,ChengC,ZhouQ.CombiningSingle-CellSequencingandSpatialTranscriptomeSequencingtoIdentifyExosome-RelatedFeaturesofGlioblastomaandConstructingaPrognosticModeltoIdentifyBard1asaPotentialTherapeuticTargetforGbmPatients.FrontiersinImmunology.2023;14.36. 张圣林,邱法波,吴力群.Th1/Th2偏移与恶性肿瘤关系的研究进展.细胞与分子免疫学杂志.2005;21(z1):113-114,117.37. KharkevitchDD,SeitoD,BalchGC,MaedaT,BalchCM,ItohK.CharacterizationofAutologousTumor‐SpecificT‐Helper2CellsinTumor‐InfiltratingLymphocytesfromaPatientwithMetastaticMelanoma.InternationalJournalofCancer.2006;58(3):317-323.38. 任中华,刘风玲.Pd-1/Pd-L1在结直肠癌中的研究进展.中国肿瘤生物治疗杂志.2017;24(10):1134-1138.39. BieF,TianH,SunN,ZangR,ZhangM,SongP,LiuL,PengY,BaiG,ZhouB,GaoS.ResearchProgressofAnti-Pd-1/Pd-L1ImmunotherapyRelatedMechanismsandPredictiveBiomarkersinNsclc.(2234-943X(Print)).40. HanH,ChoJW,LeeS,YunA,KimH,BaeD,YangS,KimCY,LeeM,KimE,LeeS,KangB,JeongD,KimY,JeonHN,JungH,NamS,ChungM,KimJH,LeeI.TrrustV2:AnExpandedReferenceDatabaseofHumanandMouseTranscriptionalRegulatoryInteractions.(1362-4962(Electronic)).41. VogelsteinB,LaneD,LevineAJ.SurfingtheP53Network.Nature.2000;408(6810):307-310.42. ColicE,PatelPU,KentOA-O.AberrantMapkSignalingOffersTherapeuticPotentialforTreatmentofOvarianCarcinoma.(1178-6930(Print)).43. HuX,liJ,FuM,ZhaoX,WangW.TheJak/StatSignalingPathway:FromBenchtoClinic.SignalTransductionandTargetedTherapy.2021;6(1):402.44. NguyenVHL,HoughR,BernaudoS,PengCA-O.Wnt/Β-CateninSignallinginOvarianCancer:InsightsintoItsHyperactivationandFunctioninTumorigenesis.(1757-2215(Electronic)).45. TherachiyilL,AnandA,AzmiA,BhatA,KorashyHM,UddinSA-O.RoleofRasSignalinginOvarianCancer.(2046-1402(Electronic)).46. SzklarczykD,GableAL,NastouKC,LyonD,KirschR,PyysaloS,DonchevaNT,LegeayM,FangT,BorkP,JensenLJ,vonMeringC.TheStringDatabasein2021:CustomizableProtein-ProteinNetworks,andFunctionalCharacterizationofUser-UploadedGene/MeasurementSets.(1362-4962(Electronic)).47. CsardiG,NepuszT.TheIgraphSoftwarePackageforComplexNetworkResearch.InterJournal.2005;ComplexSystems:1695.

8、附录附表1外泌体基因集SupplementaryTable1ExosomegenesetGenesymbleEnsembleChromosome/scaffoldnameGenestart(bp)Geneend(bp)StrandCDC42ENSG00000070831122052627220929461CPENSG000000474573149162410149222055-1CD47ENSG000001967763108043094108091862-1EEF1A1ENSG0000015650867351575073523797-1EPCAMENSG00000119888247345158473876011ERBB2ENSG000001417361739687914397304261FGFR2ENSG0000006646810121478334121598458-1CHEK2ENSG00000183765222868774328742422-1BARD1ENSG000001383762214725646214809711-1TSC2ENSG0000010319716204746520894871CASP7ENSG00000165806101136791621137309071SFNENSG00000175793126863138268644571TYRENSG000000774981189177452892957591CCKBRENSG0000011014811625973662721271CYP11A1ENSG00000140459157433775974367740-1GATA4ENSG00000136574811676959117600021ABCB11ENSG000000737342168922938169031322-1IVLENSG0000016320711529085451529118861BIDENSG00000015475221773413817774770-1SDHAENSG0000007357852182412570821USF1ENSG000001587731161039251161045977-1RHOENSG0000016391431295286401295351691TAC1ENSG00000006128797731908977404721HLA-DQA1ENSG00000196735632628179326470621RIPK1ENSG000001372756306399131151871CUL4AENSG00000139842131132081931132671081ABCB5ENSG00000004846720615207207770381ADCYAP1ENSG00000141433189049439121721FZD5ENSG000001632512207762586207769563-1RANBP1ENSG000000999012220115938201273571CASC3ENSG000001083491740140318401721711GPBAR1ENSG0000017992122182594962182638591PSMC5ENSG000000871911763827152638320261POLR2KENSG0000014766981001505841001540021续附表1外泌体基因集GenesymbleEnsembleChromosome/scaffoldnameGenestart(bp)Geneend(bp)StrandPLCB4ENSG0000010133320906876394812421DOK1ENSG00000115325274549026745575541SF3B4ENSG000001433681149923317149928344-1GTF2IENSG00000263001774657667747606921NOC2LENSG000001889761944204959309-1MEF2AENSG000000683051599565417997164661CYC1ENSG0000017909181440950271440975251MRPL21ENSG00000197345116889127668903835-1MRPL4ENSG000001053641910251901102600451BDKRB2ENSG000001683981496204679962441661PIK3R5ENSG000001415061788789118965712-1PRPF3ENSG0000011736011503214761503531951PSMD2ENSG0000017516631842987091843090541SNRPA1ENSG0000013187615101281510101295282-1RPS10ENSG0000012461463441745434426125-1ADRA1BENSG0000017021451598650801599725441TRIOBPENSG000001001062237696988377765561RPL26L1ENSG0000003724151729587291729697711FAM162AENSG0000011402331223841761224123341SF3A2ENSG0000010489719223650422486791HER2ENSG000001417361739687914397304261CD82ENSG000000851171144564427446203581CD24ENSG000002723986106969831106975627-1Del-1ENSG0000016417658394055484384793-1DicerENSG00000100697149508622895158010-1POSTNENSG00000133110133756258337598844-1FGFR3ENSG000000680784179329318088721FIBPENSG00000172500116588374165888539-1FRS2ENSG000001662251269470349695797891FTH1ENSG00000167996116195971861967660-1FTLENSG000000870861948965301489668781GSTP1ENSG000000842071167583595675866561HOXC6ENSG000001977571253990624540308231HTR7ENSG00000148680109074082390857698-1IGF1RENSG000001404431598648971989645301IL1RAPENSG0000019608331905140511906597501ITGB1ENSG00000150093103290031933005792-1续附表1外泌体基因集GenesymbleEnsembleChromosome/scaffoldnameGenestart(bp)Geneend(bp)StrandKISS1RENSG00000116014199172879210151KRT14ENSG00000186847174158227941586921-1LDHAENSG000001343331118394388184084251MTDHENSG00000147649897644179977287701NANOGENSG0000011170412778779477991411NEUROD1ENSG000001629922181673088181680876-1PlauENSG000001228611073909177739174971RAB13ENSG000001435451153981617153986358-1RPL28ENSG000001081071955385345554032501S1PR2ENSG00000267534191022143310231331-1TGFBR1ENSG00000106799999104038991541921TRPC5ENSG00000072315X111774315112082776-1TSG101ENSG00000074319111846833618527232-1UCHL1ENSG