◎慧慧霊や禦慧慧
CooperationandCollaborationProgramswithASEANUniversities

Thailand−JapanResearchSeminar

OnGlobalHealthandInfbctiousDiseases

Date：180ctober2013，15：00−18：00

Venues：

VODconferenceroom，Chamlongbuilding，MahidoIUniverslty

RoomE803，KobeUniversityGraduateSchoolofHealthSciences
MainLectureHa11，KobeUniversityGraduateSchoolofMedicine

CooperationandCollaboratiOnProgramswithASEANUniversities
Thailand・Japanrcsearchseminaronglob目地althandinkctiousdiseaseS
Date：180ctober2013，15：00−18：00（Japantime）
Ⅵnues：VODcon鎚renceroom，Chamlongbuilding，MahidoIUniversity

RoomE803，KobeUniversityGraduateSchoolofHealthSciences

MainLectureHall，KobeUniversityGraduateSchoolofMedicine
Program：＞

Openlmgremarks15：00
1血suhiroMinamiIⅥceDean，KobeUniversityGraduateSchoolofMedicine
l血owalarkSukthana，Dean！FacultyofTropicalMedicine，MahidoltJniversity＞

Sessionl，FacuItyofTropicalMedicine，MahidollJmiversity15‥10−15：40

1−1・ChonlatipPipattanaboon，（ClonlngandexpressionofDenguerecombinantproteinsfor
Denguevaccinedesign．）

1−2・PhanthilaSirichaiyakul，（Gambicin‥anantimicrobialpePtideasthetherapeuticoption
fortreabnentofantibiotic・reSistantbacteriaandtropicalpathogens．）＞

S鰯ion2，Mahido1−0はbCenter払rIn触伽uSDiseas飴，Osa日加ivers和15：40−16：10

2−1・Orapim Puiprom，（Characterization ofchikungunyaviruSinfbction ofa humankeratinocytecellline：Roleofmosquitosalivaryglandproteininsuppressingthehost
immuneresponse．）

2−2・Partiaporn Chaich弧g，（Sequence variation ofDengue virus2premembraneandenvelope

derived ftom patient plasma shows slgni鮎antly di飴rent biological

CharacteristicsinhumanK562cells．）＞

Session3，KobetJniversityGradtlateSchoolof丑ealthSciencesIKobetJAiversity

GraduateSchoolofMedicine16：10＿16：55

3−1・ErikoIwasaki／GraduateSchoolofHealthSciences（Bonemineraldensityandbone
tumoveramongyoungwomeninChiangMai，Thailand）
3−2・ShuheiUeda／SchoolofMedicineFacultyofHealthSciences（Detectionofanti−dengue

virusantibodiesandviralRNAinserumsamPlesderived舟omThaipatientswithfebrileillness）

3−3・ChyntiaJasirwan／GraduateSchoolofMedicine（ThehumanherpeSvirus6U21−U24
geneclusterisnotessentialforviruSgrOWth）＞

Session4，LectureSeminar17：00−18：00

4−1・Pongrama RamaSOOta／Faculty of取opical Medicine，MahidoI Uhiversity（Thailand−Japanresearchcollaborationondevelopmentoftherapheuticproductsagainst
Denguevirus）

4−2・MasanoriKameoka／KobeUniversityGraduateSchoolofHealthSciences（HIV／AⅢS
researchatoversearesearchcollaborationcenters）＞

C］osingremarks18＝00
SatoshiThkada，Dean，GraduateSchoolo柑ealthSciencesIKobelJniversity

1−lCLONmGANDEXPRESSIONOFDENGtJERECOMBmANTPROTEmSFORIDEALVACCINEDESIGN
ChonlatiDPiDattan。0nl，TadahiroSasaki2，MksuhiroNishimura2，ChayaneeSeWhapramotel，Panna血p

Pk止S頑akull，TamakiOkabayashi3，PaQiapornChaichana3，Kazuyoshirkuta2，PongramaRamasoぬI

IcenterofExcellenceforAmibodyRes脚Ch（CEAR），FacultyofTropicalMedicine，MahidoIUniversity，
Bangkok，Thailand
2ReS紺ChInstitutefor旭crobialDiseases（RIMD），OsakaUnivershy，Suha，Japan
3Mahido1−0sakaCenterR）rInfectimSDiseases，Bangkok，Thailand

DengueviruSPV）iscomposedofstruCtural（C，PrM，andE）andnonstructural（NSl，NS2A，NS2B，NS3，NS4A，NS4B，andNS5）proteins．Theenvelope（E）proteinisthe
P血cipaLtargetofIleutmlizLngantibodyandconsistsofthreedLscretedomainsPtD托andDIIf）・Antibodies againstprM，NSl，NS3，andNS5proteinsalsohavebeendetectedin
peOple．Mostly，neutralizingepltOPeShavebeenidenti鎖edandlocalizedonDIIandDIHofE

PrOtein．StudiesofepitopeSOnEproteinarenotonlynecessaryfbrunderstandingvirusー
antibodyinteractionsぅbutalsointermsofhelpingtodesignanidealdenguevaccinethatcan

PrOvidel脆−longprotection，StrOngneuhlizingactivityagainstallfburserotypeS，andlow／noenhancingactivity．

Since，136HuMAbs were successfu11y generated uslng hybridoma technique丘omperipheralbloodmononuclearcells（PBMCs）inactrteandconvalescentphasesofThai

dengue patlentS at CEAR．Characterization ofthese HuMAbsis requiredfor鮎山retherapeuticuseandvaccinedevelopment．Theaimofthestudyistoacctmtelyidentifytarget

PrOteins，antigenicdomains，andepitoperegionsofavarietyoftheseH血血生bs．Here，four

recombinantPrOteins（C，PrM，E，andNSl）andminetnmcatedEproteins鱒1−394，E1−296，

E1−192，E1−132，E1−98，E52−132，E60−121，E74−118，and E74−109）Were clonedand

expressedinHEK293TcellandE．coli．Aseriesoftheexpressedproteinswasusedto
determine binding reactivity of136HuMAbs byimmun0fluorescence assay（汀A）and

WeSkmblo血ng（WB）．

From136HuMAbs，101（74．26％），10（7．35％），and12（8．82％）HuMAbswere
determined target proteins as E，PrM，and NSI proteins，reSPeCtively・Noticeably，71（52・21％）HuMAbsrecognizingtheviralEproteinwereabletoneutralizeallfourserotypeS
OfDV・ForE−SPeCificfhiMAbs，theepitoperegionswereassignedtoE52−132（lStDH）by51

HuMAbs，E192−296（2ndDII）by2HuMAbs，andE296−394（DIH）bylHuMAb．Amongthese51HuMAbs，theminimumepltOpeSiteswerede重nedasE60−121by22HuMAbs，E74−
118by14HuMAbs，andE74−109by14HuMAbs・TheresultsimpliedthatlStDIIcontaining
thefusionloopisthepredominantlyimmunogenictargetinhumans．

Accordingtoseveralstudies，denguevaccinedeveloped録omfu11−1engthprM偲

PrOteinmightinducene血ralizingactivity，butitmayalsoinduceenhancingactivity．The
ShorttrunCatedprotein，E74−1180rE74−109，maybethecandidatetoconstruCtepltOpe−based

immunogenpreSentinthehigh1yconservedregionoffusionloop，withhighneutralizingand
lessenTlanClngaCtj、廟escomparedtofu11−1engthproteins・

KEYWORDS：DENGtJE／ENVELOPE／DOMAmII／EPITOPE／RECOMBINANT／TRtJNCATE

l

1・2

ExpreSSionandcharacterizationoftheantimicrobial■

PePtidegambicinfromCuf餅quLnqueねsc由tus

弛anthilaSirichaiyakull，RungarunSuthangkornkull，ApanChanidThepouypornl，OkabayashiTamaki2，YoshiharuMatsuura3，NaokazuTakeda4，DtmrongkietArthanl＊lDepartmentofTropicalNutritionandFoodScience，2Mahidol・OsakaCenterforInfbctious

Diseases（MOCID），FacultyofTropicalMedicine，MahidoIUniversity，Bangkok，10400
Thai1and，3DepartmentofMolecularVirology，ResearchinstituteforMicrobialdiseases，OsakaUniverslty，Osaka，5645・0871Japan，4Thailand−JapanResearchCollaborationCenter

OnEmergingandRe−emergingInfections（RCC−ERI），Nonthaburi，11000Thailand．
Tel‥＋66−2−354−9100−19ext・1粥2−5；Fax：＋66−2−644−7934
E−mail：dumrongkiet．art＠mahidol．ac．th

ABSTRACT

A血microbialpeptides（AMPs），Ⅴ血ichcanbefoundinmanyorganisms，aregenerally
low molecular weightpePtides contaimingless thanlOO amino acid residues．Due toine飴ctive conventionalantibiotic treatments causlng numerOuS mOrtalities ofinfbcted

Patientsinthe world，AMPs areaninterestlngapPrOaChtothedevelopmentofnovel
ther叩euticsagalnStPathogens，Particularlyantibiotic−reSistantbacteria，becausetheyaresafb，

non−tOXictomammalS，nObacterialresistanceandshowlngbroad−SPeCtrumantimicrobialactivity・Interestingly，anantimicrobialpePtidegambicinisolatedftomAn・gambiaecelllines

POSSeSSantimicrobialactivityagainstgram−POSitiveand−negativebacteria，BlamerltOuSfungi，andPhLymOdium speCleS・However，theisolationofCulexgambicinandexpressionof
recombinant mOSqultO gambicins remain unreported・This study aimed to express

recombinantCulexgambicininPichiapastorisand／OrbaculoviruSeXPreSSionsystem，and

SuCCeSSfullyclonedandexpressed・SDS−PAGEofCulexgambicininPichiapastorisrevealed

anobviousproteinbandwithamolecularweightof7．OkDa，indicatinghighproductionof
recombinantCulexgambicin・TestingofantimicrobialactivityagainstEcoliDH5αStrainby

agar−WelldifRISionassayexhibitednoactivity・Arefbldingapproachtotherecoveryof
antimicrobialpeptidesisrequiredforfurtherstudies，Whiletheexpressionofrecombinantgambicininthebaculovirusexpressionsystemisontheprocess血ichwillbealsodiscussed．

22−1
CharaCtCrizationofchikungunyavirusin騰CtiOnofahumankeratinocyte
CCllline：RoleofmosquitosalivaryglaTLdproteininsuppresslngthehost
immunerespon＄e

完霊慧恕慧訝荒…慧芸慧雲丹慧霊恕労賃
悪霊等慧慧監慧慧認諾認諾琵琶票慧慧S。。ial
窓慧芸芸憲琵器霊慧宅謡霊慧篭急設霊慧器怒。触・MicrobialDiseases，OsakaUniversity，Suita，Osaka，JapanThe chikungunyaviruS（CHIKV）is a mosquito−bome Virus that has recently re−emergedinseveralcountries．Oninfbction，thenrstvertebratecellstocomeintocontactwith

CHIKVareskincells；mOSqultOeSinoculatethevirustogetherwithsalivaryglandprotein
intohostskinwhleprobingandfbedingonblood．However，thereislittleknownaboutthe

SuSCeptibilityofhumanskincellstoCHIKVin免ction．Toclarifythis，WeinvestigatedthekineticsofCHrKVinthehumankeratinocytecellline，HaCaT．CHIKVactivelyreplicatedin

HaCaTcells，withviruStitersinthesupematantincreasingto2．8×10（4）plaque−forming
units（PFU）ml（−1）24

h postinfbction．CHIKVinfbction suppressed production ofinterleukin，8（IL−8）inHaCaTcells．ThefunctionofIL−8istorecmitimmunecellstovirus−
infbctedsites，aPrOCeSSknownaschemotaxis．FurthermOre，WeaSSeSSedtheroleofmosquito

Salivary gland proteinin CHIKVinfbctions by eomparlng thelevels of CHIKV gene

expressionandchemokineproductioninHaCaTcellswithandwithoutsalivaryglandextract（SGE）．SGEenhancedboththeexpressionoftheCHIKVgeneandthesuppressionef臨ctof
CHⅨVonIL−8production．OurdatasuggestthattheHaCaTcelllinerepresentsane飴ctive

toolforinvestigatingthemechanismofCHIKVtranSmissionandspreadinskincells．Atthe

mosquitobitesite，CHIKVworkstogetherwithSGEtoensuretheviruSrePlicatesinskin
CellsandescapeSthehostimmunesystembysuppressionofIL−8production．

3

2−2

SEQUENCEVARIATIONOFI）ENGUEVIRUS2PREMEM旧RANEANDENVELOPEDERIVEDFROMPATIENTPLASMASHOWSSIGNIFICANTI，YDIFFERENTBIOLOGICALCHARACTERISTICSINHUMANK562CELLS
P．Chaichanal，T．Kurosu2，0．fhiproml，K．kuta2，P．Ramasoota3，T・Okal）ayaShil′′l／ ′／肌仙り、〟血（、州JぐJ・、仙り叫h仙Jハ〃ハ仙ぐ．ヾ．八日小川／什叩′＝〟抽 J′州ぐ‥l山／削Mと加vem砂，βα乃卸た7協α血〝dご／）岬〟川JJ州Jr）川川ん脚＼J〜ハぐ 肌カ／′日日仙一声げ仙げ〃h′日加し・〟．、ぐ．、JJ．、〟hJ仙ぐハ申．∫J′血
0ざ血565−087ノ，J甲α乃

十古川．八九叫1・吋／叩，Jぐ（〟〝肌〟川町仙／仇hH厄日，打、町、仇川か√止〃MJん仇／BACKGROUND：Denguevirus（DV），likemostRNAviruSeS，hasbeenshowntoexistasa
populationofcloselyrelatedgenome，SO−CalledquasISPeCieswithinindividuals．However，thebiologiCalsignincance ofquasl坤eCiesinviruSinftction andimmunopathogenesis remains
unclear．

OBJECTIVE：Inthis study，Weaimedto determinethe biologicalsignificance ofsequence

VariationofDV2struCturalregion（prM−E）derived舟ompatientsinThailand．

METHODS：TheprM−EreglOnOfdenguevirus舟om4clinicalsampleswereampli鮎dbyRT−
PCR，Clonedintoplasmid．Thesequencesofthesegenesweredeterminedandtheirvariations

WereanalyzedbyABIautomatedsequencerandMEGA5，reSPeCtively．Foranalyzeaneffbctof
SequenCeVariationofprM−EreglOnS，theprM−EreglOnOfDENV−2infbctiouscDNAplasmid
CloneRO5−624wasreplacedwiththerepresentativeprM−Egene駐omeachvariantSderivedfrom
OneDHfpatientD30．Virusreplicationkinetics，neutralizingandinfection−e血anClngef艶ctof
thevari弧tSWereanalyzed．

RESULTS：TheextentofsequencediversltyOfprM−Ereg10nreVealedthatDV2hm4padents
demonstratedsequencediversltywiththemeanPalrwisedistanceranglng丘omO・0．046％．For
hI也erinvestlgation，WeSelected薫verepresentadvevariantS免・OmDHFpatientD30toconstructinftcdouscDNAclonesbyreplacingprM−Ereg10n．VariantsdlOWed signi鮎antlydi飴rent
replicationrateinhumanleukemiaK562cells，butnotinC6β60rVerocells．Itisnoteworthy

thatthevariantCOntaimingsubstitutionataminoacidposition365indomainIIIofEprotein
ShowedstrongenhanClngef托ct，andlowFRm50titerbymouseanti−Emonoclonalantibody（MAb）4G2comparedwithothervariants．
CONCLUSIONS：OurresultsuggestedthatsubsdtutionsofDV2prM−EreglOnderived録om
Patientplasmaa蝕ctviruSPrOductioninhumancelHineeitherinthepresenceorabsenceof
antibodies．

KEYWORDS：dengueviruS，quaSISPeCies

4

3・l

Bone血er出血n8iけandbonetWnOVeramOngyOⅦngWOnenhCl血皿gM姐m鋤皿d

ErikoIwasaki，NuntanaMorakote，SomsakChaovitsaree，andHiroyaMatsuo

Preseder；ErikoIwasaki（Ed姫Un柁rSity；Gra血ateS血0lofHealthScienαS，D印artme血OfI血emationalHea址軸

ThepresentstudywascamiedollttOinvestigatetheinmlenCeOflife＄tyleonbonemineraldensity（BMD）andbonetumoveramongyoungwomeninChiangMai，Thailand．Atotalof177youngwomenamliatedwithChiang

MaiUniversityhospitalwereenrolled．QlleStionnairesabouttheirli鈷StyleandtheOsteoporosisKnowledgeTbst（0KT）wereexamined，ThemeasurementofBMDwasasSeSSedbyQuantitativeUltrasound（QUS）．Basedonthe

measurementofBMD，the＄ubjectsweredividedinto2groups，aLewBMDgroup（Lgroup：lessthanYAM・1．OSD）

andaNormalBMDgroup（Ngroup：morethanYAM・1．OSI））．Lgroup（n＝23）andNgroup（n＝23）wereexaminedusingOC，NTxanducOCasboneturnOVermarkers，andserumCa，1，25・（OH）2VitaminD，ⅥtaminKlandⅥtamin

K2（MK▲4）asboneturnoverrelatedhctors．ThepercentageofIJOWBMDgroupwas23．2％．Concer血gli触tyle

andBMD，theBMDofthelowcheeseirrtakegroupwas99．7土17．0andtheBMDofthehighcheeseintakeonewasllO．0土23．3b＜0．05）．TheBMDofthefractureexperiencegroupwas82．5ill．6andtheBMDofno・魚・aCturegpOuP

WaSlO3．3j＝19．6（p＜0．05）．Thereweresigni丘cantdi飴rencesinucOCandl，25・（OH）2Vitaminl）betweenLandN

groupsb＜0．05）．ItwassuggestedthatBMI，foodand蝕・aCtureeXperiencemighta飴ctBMDlevelandSuppreSSion

Ofboneformatio11mightbemntributedtothelowBMDinyoungwomeninChiangMai，Thailand．

53−2
Detectionofanti−denguevinlSantibodiesandviralRNAinserumsampIesderived

fromThipatientswithfbbrileitlness

UedaShuhei

KobeUniversitySchoolofMedicineFacultyofHealthScience

Abstract
InsoutheastAsia，thereareseveralmosquito−borneviralinfbctiousdiseaseswhich

are prevalentintropicaland subtropicalreglOnS．Amongthem，dengue fbveris an
importantonethatneedstobepaidattention．DengueviruSistransmittedbytwostrains

OfmosqultOeS，Aedasa唱汐tiandAedbsalbq，ictus．DengueviruSisclassi鮎dinto4
SerOtypeS，andcausesdenguefbverin50−100millioninfbctionperyear．2．5billion
PeOPleliveindengue，endemictropICalandsubtropICalareas．Inthis study，dengue

SuSPeCtedpatientswithftbrileillnessinNakhonsawan，Thailandwerediagnosedby

detectingviralRNAbyRrlPCRandbydetectlngantiEdengueantibodiesbyELISA・In

addition，ViralserotypingwasalsocarriedoutbyRトPCR．Twenty−Sixof28samples
Were POSitivefor dengue viralRNA．In addition，the most epidemic serotype wasdenguetype3；however，thepadentsinfbctedwithdenguevirustype1，2and4werealsopresent，indicatingtheco−Circulationofal14serotypesoftheviruSeS．TheELISA
Ofpaired，aCute−COnValescentserumsamplesrevealedthatanti−dengueantibodies，IgG

and／orI岳M，WerePOSitiveatacutephaseinallsamPles．Inaddition，thehighertiterof
theantibodywasdetectedatconvalescentthanacutephaseinmostsamples．Inapaired
SamPles，IgGwasnegativeatacutephase，Whileitwaspositiveatconvalescentphase，

indicatlngthis patient was prlmaryin丘ctedwithdengue viruS．Two samples were

POSitiveforanti−dengueantibodies，butwerenegativebyPCR，SuggeStlngthatthosePatieTrtSWerein氏ctedwithotherviruSintheFlaviviridae，SuChasJapaneseencephalitis

ViruS（JEV）andZikaviruS．Thesuperinftctionofdi飴rentserotypesofdengueviruSeS
POtentiallycauses severeilluneSS；therefore，mOreePidemiologicalstudiesincluding

Viralserotypingisrequiredindengueendemic訂ea．

6

3＿3

Thehumanherpesvirus6U21IoU24geneclusterisnoIessendalbrviruSgrOWth

ChyntiaJasirw弧，HuaminThg，MayukoHayashi，M喝umi0也andYasukoMori
DivisionofClinicalVirology，CenterforhfectiousDiseases，KobeUmiversityGraduateSchoolofMedicine．

Batkgr仙nd

HtmanherpeSViruS6（HHV−6）isaTlbrmphotrophicviruSbelongingtothegenusRoseolovirus
Within betaherpesvirus sub偽mily．TheU20」U24g孤e Clusteris tLnlque tO Roseoloviruses；

however，itisnotknOwnhowtheyfunctionorwhethertheyareessendalforvirusgrow血．h
addidon，We蝕undthatCD6血ichisoneoftheimmtmologicalsyn叩SeCOmPOnentSinvoIved
inlymphocyteactivationanddi飴re鵬iationprocesses，WaSSPeCifiCallydownregulated拉omthe
CellsurぬceofHHVN6−infectedeells．Thereforetheaimsofourstudyaretoanalyzethefunction

Of血is gene clusterin HHV−6in飴cdonand to elucidate whetherit plays rolesin thedownregulationofCD6分omthece11surface．

Me仙odsWe used two step Red Reco血bination procedure to generate

the Bacterial

Arti負cial

Chromosome（BAC）mdreconstitutetherecombinantviruSWithadeletionin蝕lllengthofU21

toU24geneandalsotherevertant．WenamedtheresultantBAC，HHV−6ABACAU21−U24，and

HHV・6ABACAU21・U24rev．WeisolatedtheBACDNAandcon丘medthedigestionpattern
withBamHlenzyme．ThenwereconstitutedtheinfeCtiousviruSbytranS良ctionintotheJJHAN
Cellsandcoculturingto CBMCs．ThetitreofviruSWaSmeaStWeduslngTCID50methodin

JJHANce11S．ThegrowthcurVeOftheseviruSeSWaSdeterminedbyreal−timePCR，and was
COmPared．In addition，the CD6doⅧegulationininfectedcellswasexamineduslngthe
ⅥmSeS．

Results

HerewesuccessfullygeneratedrecombinantviruSeSOfaU211J24deletionmutantandthe托Ⅵ汀払出8AC．罰le柁Ⅵ切5a Sim血r pmpO血0乃OfGFPや0扇ve ce地雨血加地erHHV」6ABAC−andrHHV16ABACAU21−U24−infectedCBMCpopulations．Thiswas
COnfirmedbytheexpressionofgQlinthecelllysates．Theviralgrowthcurvesshowed

thatthere was no signi鮎ant di飴renceinthe growth ofthe viruses（wild−tyPe，recombinantandits revertant），SuggeStingthatU2トU24is notrequired fbrviruS
PrOPagationin vitro・The

flOw cytometric analysis showed tha CD6was not

dovvTrregulated録omthecellsurfhceinHHV−6AU21−U24deletionmutantvirus−infbctedMolt3

CenS，indicatingthatU21・U24reglOnmayCOn扇butetodleCD6、sdownregulahon倉omdle

Cdl飢鵬ce．

7

4＿1

Thailand−JapanresearchcollaborationondevelopmentofTherapeuticHtlnan

MonoclonalAntibodiesagainstfbtlrSerYltyPeOfDeJlgtleVirusM

物eg助成呼托抑0お′，Cゐ胡お鍋申物故即ぬ0邦7，肋α研娩少乃ね柳Jムそ7地鋸和肋かj，
肋砂のカブ肋血タ，Po乃g和研α月α椚α∫00細り＊∫α醸rげ肋β肋乃Ceカγ血舶0㊥励館α打力榊，凡慮か沼殉ぬ鋼廠如如牒肋血相油肌吻

助輌7勉血刀db軍畑脚柏悦血柏励肋如椚細山血崗喝助成舛押桓如摘誠励再励磁舶加珊瑚
及明が戚∴相加触感
jRe∫朗化あお融虚かA館cmみねJp加がβ，伽血肋fvg相和頻用

DengueviruSin鈷ctionare−emerglngvirus，COnStitutethelargeStVeCtOrJbomedis飽SeviruS，with50−100

millioncasesreportedeveryyear，hasbeenverylmPOrtantPtiblichealthprobleminTropicalandshb−
tropicalcountries．TもailandisoneofDenguea飽ctedcountry．Recendy，倉omJanuarytillSepte血ber

2013，therearebigoutbreakofDengueinthenordlemandsouthempr？VincesofThailand，Whichleadt0

120，000caseswith110deadcases・Even血oughseveralDengueYaCClneandtherapeuticproductshave
beendevelopedandtested．Butuntilpresent，therehavenoe飴ctlVeVaCCineandspeCi重ctreatmentfor

dengueviruS（DENV），eSpeCiallyagainsta114serotype．

Ffomtheyears2009−2013，withthesupported丘ⅦmJapanScienceandTbchnologyAgency（JST）amd
JapanI血emat10nalCooperationAgency（JICA），ProfbssorKazuyoshiktrta丘omResearchhstitutefbr

MicrobialDiseases（RIMD），OsakaUmiversityandThaiResearChersgroups倉omDeparbnentofMedical

Sciences，MinistryofPublicHealth，tOgetherwithFacultyof廿opicalMedicine，Mal1idoIUniversityhad
StartedtheresearchcollaborationprqJeCtentitle

DevelopmentofTherapeuticproductsagamstDe鱒gue

virus

Oneofthe o句ecdveof也isresearchprqectwasto developthetherapeutichmanmonoclonal
antibodies（HuMAbs）withstrongneutralizingactivkyagainstallfourserotypesofDENVusingnovel

myelomafusionpartnernamelySPYMEG．Peripheralbloodmononuclearcells（PBMC）obtained丘om

acuteandconvalescentdengue patientsthatwereusedforfusionwithSPYMEG cells．Then，fusedhybridomacellswerecultured，SCreeningofDENVspeci負cantibodyusingImmuno恥orescenceAssay（IFA），followedbylimkingdilution，Finally，CdturefluidscontainingHuMAbsweretestedbyIFAand
Viralneutralization（NT）．Total136hybridomas producing anti−denguelmMAbs were success嵐山ygenerated・hterestingly，high1yefncacyinnumberofhybridomaclonesproduchgspeCinchuMAbwas
Obtained倉omPBMCsofacutephase（121HuMAbs）thanthose倉omconvalescentphase（15HuMAbs）．
AftercharacterizationuslngIFandNTassay，200utOf136HuMAbspotentiallyshowed85−100％NT
activityagainstal14serotypesofDENVThreecandidates愈・Omthese20HuMAbswerefurthertestedand

foundthatitcouldne血alized20clinicalDENVisolated（5isolatespereachserotype）withhigherNT
thanthoseoflaboratoryisolates．DENVinoculatedsucklingmousethattreatedwidlthesethreecandidate

NhuMAbs could survivelongerthanun廿eated group slgnincantly，andalso DENV were clearly
eliminatedwithin2daysamongNhuMAbb・eatedmamosetgroups，incontrarywithcontrolgroup（using

nondenguehumanIgGandPBS）thatDENVcopystillveryhighwithin2−3days．These3candidate
NhuMAbsarenowongoingfbrlaTgeSCaleIgGstableexpressioninChineseHamsterOvary（CHO）cell，WhichaimingforuseinhumanClinicaltrialinthenear鮎ture‥＊PresentinvideoeonfbrenceseminarofKobeUniversityandMahidoIUniversityon180ctober，2013at

FacultyOfTropicalMedicine，Mal1idoIUniversity．

8

4−2
ⅡⅣ仏IDSresearchatoversearesearchcollaborationcenters

Masan0riKameOka，Division ofInfbctious Diseases，Department ofInternational

Health，KobeUmiversityGraduateSchoolofHealthSciences

Humanimmunode貢ciencyvirustypel（HIV−1）isam毎Orcausativeagentofacquired

immunode鮎iency syndrome（AIDS）．HIVllis prevalentalloverthe world，and
according to the reportfromtheJoint United Nations Programme on HIV／AIDS（UNAIDS），34millionindividualsarelivingwithHIV−1，While2．5millionsarenewiyin鈷ctedwithHIVlandl．7millionsdied舟omAIDSduring2011．AmongSoutheast
Asiancountries，theanrrualincidentrateofHIV・1infectionhasdeclinedincou止ries
includingCambodia，MalayslaandThailand，Whereasithascontinuouslyincreasedin

COtmtriessuchasIndonesia，BangladeshandthePhilipplneS．HIVisablood−borne

virusthatspreadsthroughcontaminatedbloodandotherbody丑山d．Antiretroviraltherapyuslngmultipleanti−HIVdrugSiscurrentlyavailable，Whilethedevelopmentof

VaCCineshasnotbeensucceeded．As acollaborativeresearchwiththeMimistryof
PdblicHealthofThailand，WeStudiedtheimmunologicalchamcteristicsofThaistrains

OfHIVl，andfbundthemechamismofhowthevirusisescaped丘omtheinhibitorye飴ctofaneutralizingmonoclonalantibodyonviralreplication．Inaddition，aSa
COllaborationwiththeInstituteofTropicalDisease，AirlanggaUniversityofIndonesia，

SerOandmoleclllarepidemi010gyofHIVhavebeencarriedoutinIndonesia，andthe
highprevalenceofHIVamongcommercialsexworkersinSurabayawasrevealed．h

this talk，Iwill present the overview of global HIV／AmS endemic and fⅡV・1
Pathogenesis，aSWellastheseresearchoutcomesproducedatresearchcollaboration
centersinThailandandIndonesia．

9