WO2024259439A1 - Nucleic acid nanocapsules for drug delivery - Google Patents

Nucleic acid nanocapsules for drug delivery Download PDF

Info

Publication number
WO2024259439A1
WO2024259439A1 PCT/US2024/034387 US2024034387W WO2024259439A1 WO 2024259439 A1 WO2024259439 A1 WO 2024259439A1 US 2024034387 W US2024034387 W US 2024034387W WO 2024259439 A1 WO2024259439 A1 WO 2024259439A1
Authority
WO
WIPO (PCT)
Prior art keywords
aou
werene
nanoarce
aen
nucec
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/US2024/034387
Other languages
French (fr)
Inventor
Jessica ROUGE
Suman PAL
Divya DIVYA
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
University of Connecticut
Original Assignee
University of Connecticut
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by University of Connecticut filed Critical University of Connecticut
Publication of WO2024259439A1 publication Critical patent/WO2024259439A1/en
Anticipated expiration legal-status Critical
Pending legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6905—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion
    • A61K47/6911—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a colloid or an emulsion the form being a liposome
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A61K31/7105—Natural ribonucleic acids, i.e. containing only riboses attached to adenine, guanine, cytosine or uracil and having 3'-5' phosphodiester links
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/02—Inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/543—Lipids, e.g. triglycerides; Polyamines, e.g. spermine or spermidine
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/549—Sugars, nucleosides, nucleotides or nucleic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6921—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
    • A61K47/6925—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a microcapsule, nanocapsule, microbubble or nanobubble
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/69—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6949—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit inclusion complexes, e.g. clathrates, cavitates or fullerenes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
    • A61K9/1271—Non-conventional liposomes, e.g. PEGylated liposomes or liposomes coated or grafted with polymers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/5115—Inorganic compounds
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/115—Aptamers, i.e. nucleic acids binding a target molecule specifically and with high affinity without hybridising therewith ; Nucleic acids binding to non-nucleic acids, e.g. aptamers
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/16—Aptamers

Definitions

  • rmu nanoarces aou suerrom oaressuesruon, non-sec aonseveoeoae escae enosomesasssosome c ceuar uae, anna o ecen ormuaons s an,n some cases, areeaooxc. ona, mannoocaus,ave aowoan caac anavesaeno oxze orroze ereore more eeecmne overa sa oeormuaon ane m caro.
  • arce comrsn non-omerc am cec acans covaen aaceo a am es are arraneowarearceens,n wceroocrous oe am es are aearce surace an areeror, anerocrous oe one or morenraceuar or exraceuar reeas cero assenns.en w some on eem oro momreensn,eres cearvae rous oe am es are crossnerou arazoe,oeer, or a oc w one or more ce en suerou n anoerav aaseec,neers.
  • ew ceane assem o a m acever vece.rs, se assem o ao_-, a mu-aere nucec m.ex, suraca cna coonncuaoessomes a sureraocrmaneafug w acre a are eenc,a wsueraeen aenonc nucreoc aoccs.aesxn,e-rmcaeaenmoaezoeo asnom 1 za 2 n ocu saer aaree ar,ereseonn neave care osome.saer sazeseosomeoreve orm a cae aroune ase m o_ senormuaenremaure ruureurn amnsraon.
  • scaears are nm.. emeraure ramn exerence mee mrou anoueeor smnuaesos oe m o_ s, ne amoun o .o. sonw s ae cna aracoveerz -ao °n o we manann sa. . ase ronamc sze o -osomes m, - moaeoao_eo-sos.
  • conu s maes o caoncosomes wncreasn suracan suracaane c cooncenraonsrom o .esusncaeaa were a suaneu sazee a snou saone a eneouo aonru mremr en scoauson oosomes,e caoncosomes consrucon ver.sus coonro.os somoews. a. ana mss oua oo asno. eme cso oescosuren saes o m oae moeosomes..
  • a. uer .oowereo a conon a wce aer soueroonaceae suseuenerae.oa m reease was vsuaze a rev e an assa run on an e.. enaurn eenrsecarannscreon m sreaes weno_-s are exoseoo eserase reease o .
  • acaoweearso a arcemno.n m-ceorrro euxneresrseae as rer we reons amonarerra reonsneac on aears as cacuaons sowneowe conocamaesnca an wemae.. nenseseween coroune an non-corouneereaeear saemese.rencee ssnnucoarnecsece wnacse caeor .. snoswsu.ore,s *ceno.ae, **., ***..
  • exermens weres reresenncea wvaes o o a ses an same ses.ereeararam o snca nera an consern- cesromerene consern.e . caeoor.. snoswon.oc, * ., **., ***.. merr exressona r a mcroscome course assaman o ea or cesn coreressuoen wne sne sruamm aese we mree reportedly an a ser °um ovreeern mea an coreaen merr exressonne nons. aernc a . an . or . an .
  • eren ses o sam –..xermens wereerormereemes w e a ° a.en seos cw a asn emao meno s an oca m ce ncues.
  • arre e wxresseona w ceens mo oae sames were mea.eere a ° or roomemeraure, e rrn serumree o_-s waseran weneverexroreusso cna o m evere souon.s resu suorse resus sow oncosomes orn erormereemes w eren sesnn.
  • sowess.caears arem. reee a roomemeraure amaenn oca memerers a enxrenscsuona wee wn m coeaseo sramresn w seerruem e or-an we mneeav.erxeresrsooun o ca monecrr mosome orenvere souoon_ even a-ers waas ere was no sncan emsson o merr exresson aer s. resus sownn.
  • ceseoosomes an m ranseceno ces usn o_ s ca werereae w eua concenraons o merroaencuae oncosomes an merr m noecamne.ames were was seen e wven a cesor.n cronoraosw,eosomes arneamraenns.eceoo mxc o soweeoox_c a-ows concenraons.ower concenraon oo_ s was sc er crossneormuaon as sucower conce reeneor aee eren.ere, unreae ces were se asnr caeo vnsa were u asne asor coneron vovro sus exeseescre rrorars aree averae oreeeren exermen rmen.
  • sucuaneous - seeeum eomoeneousever o m rouou oeumors are orrown one o mce.woerenssue secons mare as we reoxreessen weere cearecu anae meanmuaoreesc seonwnareen.s vaue os areneren areas mare areas.esus sowa m exressons reaso cae aove wenevere usn aamerunconazeonaomoeneoussrue seuenceunconazearces so _-s.n conras, scrameumor. weareerencesn exressonevesrououe .
  • eneraennc,e m sua-aer aen nanoar cocnerso“enan reoceaasseu oes” arerovea oeraren an aroone see.anocasues oesc nucec ac caroe.., m , encasuae nucec ac caro, a osure comrse aosomaaero osome, an a nucec ac na mea orancramewor aero sazee e nanocasue a nocasue oueraero, e..,nuenceearen o as we as reaen comeosveron os a nnuc meec aocs c oar usoe., ouesr, sen nancaonca asuvaes asescreeren, ever ssems.
  • s va m renesr,s moono mceoreceus,esac canncueas, waxes, seros, cos, caonc or anoncs,ervaze eres,osos, snos, eow.
  • an a meos onvenone meos noeen s or ce are conemae accornoe so consrue.eans onroucone anre o wne an-noawrncuoar moseean ss oennrouec aro,n a anns nooe exemeeren.
  • rovn meansvn, amnsern, s aso are orro or no, manuacurn, como en,srun,ranserrn s useeren,eerm “arumnacneu, ocras aecncsn.
  • aooca conon,sorer, orsease e.., arr amen”ncuenne aooca conon,sorer, orsease o esn or reucneeveomen oe conon,sorer, orsease, e rs cnca smoms or reevneaooca sease ors .., causn reresson oeaooca conon,sorer, or conon,s cornecr,a or smseaosme as.re aremaemoernae me oarns o anerw waseeene smcaom as oere a.areoeorca suecn nee ohariamens a mamma, anarcuar auman,u a, a eneeow.reamen aso meansrovn an acve comou
  • reersoe amoun o aera wcs suc or “eraeuca eecve amoun” uraon o asorer or one eno reuce or ameoraee sever anor sorer, cau or more smomsereo,n orrevene assemblemen o a roresson ose o rneer oerss moonre o s amsoomrser a,ssnocae or wrev aensoer reercurrence,eveomen, onse ormroveeroacc oreraeuc eecs ,eec asorer, or enance or eraeuc aen.n eecve amoun can r o anoererae..,roacc orerm “roaxs” reersore eure morean oneose.s useeren,e sasca
  • “reerence” or “con “reerence” are use wcs emoe as aasene o ro”eve mae areeermne vaue or rane, “onro” aso re rencmar aans wco assess a measure resu. eeremrss "o ceornaroeu exc”e orrm "enes or conro ces.
  • ncaeeren or oer eerormen a suae orer uness oerwse ex wse cear conrace conex.e use o an an a exames, or noemosaer aamnuaaoen oen.., “es succo aes o”,sennevneneon m uerneesso oeeerusraeenvenon anoes seccaon soue consrue asncan rwse came.oanuaene oenvenon as u an non-came eemen as essenaoeracce erenavee samese meaenrenn.
  • menss asoscose an an eemens cane --- meo o reeecrranonn ov ranuaes oo v eaacues seearerane a Erasmus mueereanne wneno seerve as a soranncaeeren, an eac substitutee vaues rane, uness oerwsenvua rece.or exame, ncororaenoe seccaon as were erees, ma a sze ranes sae as nmo nmor concenraons, nm ss amouns, anee,snenea vaues suc as nmo nm, nmo s s,ec ncmaoon.
  • nemse are n omno exam nme,s o or w nmaos se ncm,c eac., anree enxere,s asn e practiseraen comnaons o numerca vaueseween anncuneowes osse enumerae areoe consereoe exress vaue anees vaue urermore, wen "aou”, "aroxm saaee”n asscosure.
  • cnnroucn eemens oeresenscosure ore asecs an emomen eemens.mares, "a,”e “ aane,”c anve “ “aneo” aerre,”n wenene useo moeannroaucere are one or more oe mean one or m e an eemen,sneneo eorer easeem "ens.
  • escomnaons oese comonensn an e s useeren
  • “oona” or “oona” meansae suseuenescre saven ev oern cr ocru cmrscuamncsea mncae o or ma no occur, anaeescronncuesnsances were s useeren, “accu”rs r annsances wereoes no.
  • romo ca eerso a sra orrance canrocaron conann u are nomreono a, momes un,e ess o, ne-rwrose s,esco-ero.e, nr-esuena,ve exames o ancue, n-en,soen, neoen, n-ex, sec-u,so-u,er-u, n-e, n-oc, n-non, an n -meex,,-meen,,-meen, oer moees,en ma aso-ee ac sr.aen or anr “aance” croaun esx aamnensncrouue,euw aereen nwo meo --, - -, - -, an - -.
  • aox reerso an a rou
  • aseneeren aeneoe u are nooecmuare moo,e merooxu, e ano oxx,eron aoxom,.-eroreosexnave exames o aoxncue, exox. ,uox,er-uox,enox, an us usseuenerse anre, “naeon”e or “oaoen” reerso -, -r, - or -.
  • oruerrner,moure s,meermors cornsv,e sneceoe nno ce,o e acr.e e,n a re osc e use ae un uerareessn a oree c , no oon mexeannocaeensoe an ovnoe seccs oe suec maer,scoseerenr exame,aers.
  • muuncona nanoarcesor nanocasues,e are muaere, nanoarcesa are caae o enc nceorescre as muuncona sma moecuerus,oocs, oraessu ora ran oneu onrc moorneazeera weuc aereanesucacr
  • saliva aereanesucacr
  • se mu saeeureenc,e m,u or auncaromnaace nuaca acceae sa,rae are caae o nucec acever anaree noarces oescosure nanoarces caneareeo sec enenocown.ese muuncona ase nucec acans.or examce, c cutec renc-e, aonrsnexnn
  • roocrous oe amens,ees are arnrcanuees noowna-romeerarc acmeneroers,.en., woce , anerocrous oe am es are arraneoware ware an are crossne w one or morener arce surace exraceuar reease aenssuc aseasse c,earvoaeasee, es oenrea oser, morenraceuar or crossne mcee ,o w or easase,oorm a surace o c one or more nucec acans are covaen aaceo rromu a ara.zone so,meo eemeor, omr aenesn,e sureorouc wrou osn oe am es are crossne n some emomens,e muu e or more ceavaeners
  • oenraerncues an ac-aerou rosaene a ac ea, oroeezronreo,u sesm, ocra ar caozmone,naoonsemecreaorazone, cs-aconc rous.
  • oer ac-ae emomenns someen emero excmuenes,enerncues asuerou, wen some conons.
  • rocrous oe am cec acans are covaen aaceoe emomens, esrou aoeer or aen suerou.n some rocroue ooee aemr or aene re ssuuseromrou a re aacaconn o ae nnuec oerc a acene mansoe rocrou oe am ee.., one ammonumrou a oe one s one nuc n ao moee.., n someec e amcomanen.
  • surace ane sno,m aer eomeno,mens, nucec acans are caae o seecvenno a ce e nucec acan see ac cavreonraseo, o aroro ae nucec ac.n some emomensn wc conssn oumor-marers,nerns, ce suracen r,eceeororsens seeceromerou cannes, memraneransorroen, enzmes, anoes, a,ransmemraneroens,on emomensn wce nu n cmercroens.n some caro cec acan seecvenso a blazee,e cocarxa.ens so semeec eemorommerou conssn ocoroens, suar resues, an ac,e nucec acncuesennsn
  • -memazoe cues one or more o-memazoe, -me--erazoem, oer-m-a-zmocaarzooen,rme.azoe,enzmazoe,-me-,,-razoe, ncues-memaz n some emomens,e orancan en oe,-memazoe,-me--mazoe,mazoe, or mezmmazoazeo.en, a snomen e smomoe emmenos,me orancan comrses-memazoe, or- emomens,e meaonsn 2+ , anenes, orean orcanacnasn-mse-memamzoazoe.n ceran emomens,e orme n 2+ an-mema e, ann some ramewor- zoes aeocmaz
  • am nuceoes reace we non-omerc ncues oneea ora a mrornon e oee c aec erra c aeeuem ucs c ar nuecn re esac c a,o cnn s.
  • omanerermne emosmens ,eceroraeucm aens, a nucec ac comrsn one or more rozm nsenseonuceoes s, ereo.es, aamers, m,ce-wcn onuceoes s, or an comnaon auoroorne, s aom rae eomaoeeme nnucs,eoe nea,n ao raarocseo coomerses aanosc aen, wcncues or an eecronensea ,on,ocoero, coesero, a sero, n an asec, a an com a mea ceaor, or an comnaonereo.
  • ncun amns eo orean oranosn asease orsorersrove, escosure,n werencoe a snueercs cne naeveaeereo on aen o er meocreven armacoeunu oare nanoarce o reease aenresenne suec,us reeasn or exraceuar n some emomens, a meo oreeeraeuc aen oranosc aen.
  • smom assocae we c an orrevenn asease, asorer, or ene exres onons wc reure m anus reure anncreasennuenza orson.n ceoran c eamnceor omre an cso,men caonononerenoc.unes c communcaeseases suc assevererou arocess o vaccnao eran emomens,e meo n ceran emomens,e men,oo mr eoxaumae, o vaccne.
  • n s no morean aou , no morean aou , no more eners ceaveo mnor aen exanra aceouuar e,n nvoro mnomreen,a ann aonu some, e or no morean aou o aou , noessan aou , noessan ao momens, noessan aou , or noessan aou oenu , noessan aou , noessan n some emom ers ceaven annraceuar envronmen.
  • aers wcosso com ame nes esn,nenera are serca vesces comose o one or moreaerse.., am c mo casuae aueous souons.osomes arecaorme s veesacraeseesrnoecu eoces en a, cn suc a asuarsoe anosc o s evoemn rane esr.c or aea n uoa cnur aca o enor s,somnesa ar ceomosrsvee sa caare moecuese nucec acs, e.., an .
  • zwn- osecraoonncc,oesoncuose,au arec noonmeeo, nseeuarrao,,o-soaeoc-sno-necero csoaeaneoaamnonaemne,a,mooeoo -- o e-oososaaecanooanmene-,-maamemooomeeo-- mcoresxoane--caroxae -ma,amoosa eanoamne , n soosoeanoamne , anee, or a mxureereo.
  • n comnaon wme e omeor emxcense,n sus,core exaemre, csa maonce use aone,u arereera use ercenae oeer n aosome ma s.n some emomens,e ,reaeran,reae ereaeran,reaeran,reaeran we.n som ran,reaeran,reaeran, orreaeran aou e emomens,eeer s consueeween aou- e.., osome- m,o aeouraco-n.n, s aomoue em-om,e anosu,e-eer, aoue.-.,, an conesuee os aoeu --- rao,.
  • aarne as, caosrsm on cr asoarounse, mareeer-ams ceonmsonoase o craemmewcaor an.
  • s arenownorer surace area,unaeore szes, an exceona 3 + ourecremsaor s maea.
  • onarousneenen reresens arocaronrou conannoenzenencue aenas so omnee e omrooumse cnosn,ssen coo or —naonunconarousne orancans memazoe, anor-memazoe.
  • memazooem,ee onz emmoazomee,nms,aezo orea annc-aamnnsoesr seeeceacr aocm,- amnen sommazoe,- e sommeo emmeonsmeen osr,anec oraans seecerom-memazoe an-memeazoe.n n some ncans-mem 2+ azoe.
  • r meoscrereeren are crossnerou am es oescosure as oerwse ners.n ceran emomens, rou.n ceran em escreeren are crossnerou arazoe cr omens, am es oescosure as oerwseescreeren are oosesrwnseeescrorue aereone areer crroosus.nen cerraoun em ano amenens s, am es oescosure as emomens,erazoe,oeer, or aen sue uerou.n ceran o ane or aene moe one crossnnrou resusrom a reacon rou an an aze oro mo rocrou oe am ee.., one ammonum rom e onener.n one exame,erazoe crossner resus ammo an
  • na anre emerovemernoms,e non-omerc am es oescosure as oerwse --- or s eren are creorocrous oe am es oescosure as oerwseescre exraceuassne w one or morenersa are ceavae one or morenraceuar or mmore enzmr reease aens.or exame,n one emomen,eners ceavae one or e es, suc as,u nomeo,eases,roeases, eserases, or easases.n some ascoosumreen ass, oeerwnseeress ccreaevaeeren a anre es ceeraavsae.en cer oanen enmracoemueanr osr, exeranceerusa or reeease cee man.n ceran em
  • ncue asuero s oescosure as oerwseescreeren excuensu ceraenr eomu uo. or amneonse,reroun ceresa ovaeescosure as oerwseescreeren eners oescosure mae seec uner reucn conons.
  • emomen no morean ao ve ceave.or exame,n one morean aou u , no morean aou , no morean aou , non an exraceuar en,v noro mnmoreen.ann a anoouer e,m oro nom meonre, noane aou oeners ceave aou , noessan aou , noessan ssan aou , noessan aou oeners ceav aou, noessan aou, or noessan exn ceran emomenes on an exraceuar envronmen.
  • escos omens,e nucec acans anaren nucec acans o m ure as oerwseescreeren are caae o seecvenn , , ano comenao,n moe , zmes, rozmes, m, s , mcro, s , escosnur ceer asas on ee emreo rwo.
  • rwosemeesncrs,ee neurceenc ar aec caaane os anenear reeuna nounc.eocr a ecxamaen,sn o s reomuea emono canmeen ss,e n,ucecz acme, ranozsm ane, marcreon nu,c ore ac acans caae oene oonuceoesncun ansense oonuceoes n oereraeuc escosnur ceer asa on eemrwosemeesncrs,ee neurceenc ca ance naan.
  • w n,eescoscu areenrosvesa one or moreeraeuc aenss encase , or encose nosome comarmen o e muuncona nanoarce.
  • raosooe,on,ocoeroo,r c exoaemsereo, a, au soerroo,o aren, e ae rcaroonaeee nuceoe, a or an comn nsea an a mea ceaor, aarmoanereo.
  • cosen envuaeae exacosae o nanocasues oescosures amnsraon are aascann vew oeaenoereae.ca,osae an enrea useorove an eecve amoun oe nanocasuesoeaen nanocasuees.
  • n o cancer an e esrove.n som n o cancere..,rosae,reas amnsern aeraeuc e emomens,ereamen o cancerncues comosoner a eecve amoun o e nanoarcesescre eren, or amnseren seuoc, aom ao suunse acnaovrn suc a canmceer a.ssa nneoceasrsacres, or comosonsereo, canencunrean, aevan, ameoran, re o aceveeesre resu as orrow o,n n e mraon o, reuevn,ean onse o,n n roresson or more smoms ore cn sever o, anor reucn
  • eoecovre aremvoeunnn cancersrove.uc meoso aea nvua.e., a suec woo o a nanoarce or comosonereo as noeenanose w can es no sa an smoms o cancer anor wo annvenvearee cer.or exame,ea nvuas ma e "mmunze" w cancer a rsnvuaarscee..,roareonsev weoomaevne a oa cmancer asnoror o on csaenc oer smaenomss ca orrn --- onoe omro mroresmen aessocc mauea wons asesvoecoaeme wn o ceveomen o canceraensavn aenec weveomen o cance anceraen
  • ra ans cae uonorrovnor asecs omsson o maeras mae useor comnaons o maeras anor eeacnseren.
  • eua oveor ae emomensa are wne scoe o euero c o anarcuar eemenorracce oeeacnserenso smarnermeseeearrse,c anve su oc amesaneorn,s m aarneuacurer, seer, user, ssem oeraor or oer enerese oeerceve accornoe sanars o consrue asearo.own examesurerusraeeresennvenonu sou noe xnam ane wamns scoe.
  • an m ormuaonsoae are neor szaaoen a oe mmer o_ -s ueoensa oeormuaonse an m su aures warmeran - °,narcuar,averoveoe uns sce o nucease.osomes, conons, areoxco ces.ereaoree, weren ce cuaaorn encv sruornamceen csa arne, usn oeern ce mran mocaon w an oerans an sncan eors are unerwa ase rouencororaon oeren earousa cana oareosomes ssues an orans.owever, even w s socazaono secc cocesorasnoc urace mocaonsor sazaon anmrove suerromme e azaono sec
  • reess merenerra a ca rne auressesemaons.. a caoncosomeormuaon, co orescenroen,soae wn caramocoeser nsrucerom--','-meamnoeane- osoeanoaorocore-oesero an,-oeo-sn-cero-- ormecne ares va sme eecrosac an roocneracons.
  • coonssomruecso wner oeo_-s,osomes werersreare as moar raon coroormoowe snresnze uner anacnner amos anere-oeseron a: e overn uner vacuum, .e mxure wasen mn oowe resersonn m- waer anurerou a eerm exnreuer o an amraorae szeer memrane.uccessu sness oosomes was .. oonuceoen s.eueenrceesesn uave nanoscae caracerzaon are sownn.
  • nma was vere - oenaecameess neave- . was oserveae m o_s surace suracan conuaeans.neres. mueoe aer ane masn oe o_ arces cane vsuazen, unee orancosoma core,e m mearamew n .
  • wou uran aceae sannueoe nanocasueaosr.,.ensa saer,eo_ was encasuae wn a nucec ac m o_.o aceevens weeman o 12 suracan moecues one o_ n orerou enecassu,aeme o 12 suracans aeo mm o maersorme,s crossnenac m o_ corearce.ncee mceencororaenoe e usn aazo eser crossner wcs covaen u cemsrarce surace usn coer caaze aze ane ccoaon m o_-.e r.esu os ses ao_ surace crossne mbibrce resennvenors’revous sewinaocee cro nsuscence ac m
  • eresence o enzmesa are eevaeurn sec e arceseraen a secc enzmee..nc csease saesueo overexresson o cros rease exressonurnnammaon an cancer.e eser o essenraseer wreasse sne wccanee esnonseomoes en oa mnoces e cneoss.omeaeraaon, asere are evesncororae ae surace oe mcee re,e eser crossner was exe m o_-s were enucraesu vaae szeo_s usnu cemsr.
  • seaex coumn.earcesa a sze o.. e nxmcu anson w a -- . m,ncane successuormaon oe c aosve suraceoena o arce surace.e sneea se aonc suracanaer asseme ae serseouaon oea enno , an zeaoena reans sues a mono anreasersereeaaree mrces a ev or seueoe n_r se ceo-. convesr weer a menunroe.reun.
  • conrmne aon oe aer ae surace oearce.e anaavess- m na sze o.. nm.ese m o_- racons wereen usenocra ae a aunraesrs ex oeremseenasrsceeesn.ca,e a s.,n ancana ameou.n o-,, co,une an wresen eenmene mau- ---aeere anar acnes.
  • mesuss ar reeease uer a .. cearan a asesonseeaeor oowe aceae conrms m reeaseses weno conon rmer an- esn areresen, raeran w s, acc an enzmaceraaon roms m reease asesnarenaeca we oan ees Kirrase- oare orne aacreaceuer mine.esusnroucen aooca ssem uns enoco e woue sae wen exoseo enosoma es sewere accaon occurs an o_- erases.oessoess, anea ces werereae w o- mea w ans c wonaonun merr exressn m n an n- an corounere-reamen.esusroms assasee.
  • ncaeansncan amouns o m were reeaserome o__--ss cnomoaere meoa aonsomes.er amouonna o m,. werneceaevsereao no cnsoesocc --- r ceomeaseex o moocas e mnvnrmonam, suesn aecrease cance oremaure m reeasen m o_ -:oenes o.e aernanroenxressonrom s n orero assessse ae sa ananrove e aer, a cereuam wenas.
  • ooso semruesm o arn m serum, wreeere m seoreao ar ° an overrnoo awneen w aaseno oeea c oers an cmeasen resreare waer souon o merr m oe same concenraon n.noearces was use as a neave conro merr cao a wasoaeoae ae same m concenraon were use as aoncosomesm oreamens are summarzen.,., an sve conro.esus oese emsson canrsneuannoe weerae ane reave ceevse.s o m.
  • a son o merr was comaraeo, ann mannsances morean, s o mae saiseerro me soraoea coenoosnosm aen as s saeen o eea mceanormuuoraesocnen –ce sanens anass. overnn areezer, wc,oreosome maave mae mes weree maaveen aece sa n serum.ne cas earcesess unorm an nuceases aonseoorransecon w e oe m, susce o serum orsme exresson.
  • m oaeo_- s aon w ea cenesnes se arnum m ae- meoeac.amesunes
  • w su vmarmanrzeosen were.nc,u wae w an ano cenese m o_-snuce no snca ere resusncae an assa.moran,o-- wasessoxca noxc asncae m-oecamnerearaons wenese a e ne m-osome an amerunc :eeceoreccuarvaeenn co oncenora_ons.-s va s oencouao oena uez nrcea aoro nen amzeeo esxoammes oaree m ever.sse case asaren nanocarrerso secc ceeess.
  • woes-s, c mesn seruom_ree m-easorunucsona mzneu weso o mr wnmozue a aamer were seccnn annernazaona ma occur va scav n non- wase an aoweo sor ener receors.e ces wereenransaenoroen.
  • a meaersaan aeern vvo smrnuseraonroueorueseneceu omo ar.amse ar s anowscanreareorenmnn c oan annexn receorsa are orurououeumor,e aamer moeor cereraeucsseever overcomeeroem oeeroenouse muaons oescosure ma s aso noaeae amouver o oonuceoeeraeucsoumors.
  • escreeren was one ore n o m aeverenearceormuaons eraureo useunres o nanor orams o mcoram.seer u mnoeuxseec weere saus,sans caom emnoanncnemeen o --- m conven soana m aearso ao wornever o orers o manueess m a aeeroass.
  • mm rossneo_- ssoven o o o -ee was souo .
  • souon was aeoo_s an 12 suracan een aeons m woe w o ms resuesor aenseneo uneerrmna ens,rraeaeoneors a meno.esnrcoourso oraeeno suer maccaene an suracee voa aroone cc reaconeweene aneea o_- 12 s useor sness wus oes resues.e - m na,earces we ere use wou anurerurcaon.
  • aamer an aamer asroows mxen a mcrocenrueue aneae ean were cona a °or mns.noer souon wasreare waenr.nereaseuer, o m , o ase an o m- secon souonoa onane arnsa so vouuon, was cooeo roomemeraure an mxe we r.
  • na concenraon s were aeo serumree o- mea a a remove an unoun oarce asn anncueae anoorer.
  • mns.nen ° ce ans were was aemosoroeur o or m exresson.es wereenmae conoc 2 eo aow croscoe a mcrosco usn aoncse eona conoca.
  • namcro scanaemrnc sazce aenrza a snuron zaec.eoena o snesze maeras were caracerze szes were aoena usnaverneaszerano-.xac cacua monea osur aever oane a szeer amnose ursnecnamae sorwaroe.oso omerae a eoowe suseuen anaze scannn eecron mcro a- s were anass waserorme on aermossco an-aracon .
  • auma -raracomeeruae,r ovaano oeran ae. e, a curren o m an a scan see o . , w a worn voae o -ase mass secrom ° mn- was useo oan aerns.- was c ercneave moeeecon oe suracan conuae ssovenonu mce a omnm anonurm araceaxeoor resuce ane amaacnsu os soa.on. sames were

Landscapes

  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Epidemiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Genetics & Genomics (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Nanotechnology (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Dispersion Chemistry (AREA)
  • Zoology (AREA)
  • General Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Physics & Mathematics (AREA)
  • Inorganic Chemistry (AREA)
  • Plant Pathology (AREA)
  • Optics & Photonics (AREA)
  • Microbiology (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The present disclosure describes a multi-layered multifunctional liposomal metal organic framework (MOF) nucleic acid nanocapsule capable of stabilizing therapeutic nucleic acid cargo, particularly mRNA. More particularly, the present disclosure relates to a multifunctional nanoparticle that functions as a nucleic acid delivery material which can both deliver and release mRNA to cells in an enzyme specific fashion; and methods of using such nanoparticles for treatment and/or diagnosis of diseases and conditions.

Description

--- - ,,se aunecao,n cam, wsceesnencor oora.e.rov reseorneanceaeenrenncsa enonreo.. aonansusensv oenoena wa,s an mae wo aowve n.arnmen suor uner aware e overnmenas ceran rsnenvenre eaonacenceounaon.e eecureonc comuer reaaeorm oeeuencesnse w s acaon e sumsson ansncororaenos acaon reerencens enre.e -nce_senuensce c_onsanne_ne.xme, c”re aanee onnune,,es,na svzne.ee name “- everomacerormesoeencscosureroves muaere, muuncona nanoarcesa can secca,erues, suc as m,no ceso aerene exresson.ore one or more nuceecs aecnscaonsusr aen reoaresneor mnauzea neurece, mc aucu cnacroo,na nanoarcesavn man suc nanoarce carrers, an. meos o usn s as we as meos anoss oseases an conons uc nanoarcesorreamen anor an aeneraeeoraeneuacoarsear unseere o n sucnecca anc as,e anno mn an re sseeacrccaocu,s a.so a vaccne --- nnanroasarruccuereoremuroavones u asseeooremuvear mons re vaccnesoae reure sncan suorn °o - °orann an s ure avance croreservaon conons- m an ceran nuce orae. ona,eon vra vaccneeveomen, vare oseasesncunc a ccanc mero aenesn,eanmemraaorosearseeas.roomwesvee ars, an oeraeucor a roa ae,onerms oseasee anracca access reroe more are more sae an secco cees anssues , newormuaonsa cnca areoveno,w suneerrvoemr m sasnemrsa,w sucac ass.o ar eseom neees ae on. rmu nanoarces, aou suerrom oaressuesruon, non-sec aonseveoeoae escae enosomesasssosome c ceuar uae, anna o ecen ormuaons s an,n some cases, areeaooxc. ona, mannoocaus,ave aowoan caac anavesaeno oxze orroze ereore more eeecmne overa sa oeormuaon ane m caro. m an oerraveeuecv neurce vec acces carao csa anreo nee sea.ze as we asareeever o nucec acn na ann asec,escosureroves a muuncona nanoarceancues a moreeraeucoc aaesnuse, or aa,r am maceeua ocaran accceramaeweo sra,ra,e a or soosome, an one or muuncona nanoarces,eeraeuc aen vaeereo.n suc encasene o mae a- comexs encaseneosome,eosomes n emomens,e ncues one or more o n aune- s encasene . arce comrsn non-omerc am cec acans covaen aaceo a am es are arraneowarearceens,n wceroocrous oe am es are aearce surace an areeror, anerocrous oe one or morenraceuar or exraceuar reeas cero assenns.en w some on eem oro momreensn,eres cearvae rous oe am es are crossnerou arazoe,oeer, or a oc w one or more ce en suerou n anoerav aaseec,neers. sease, asorer, or smomn asc suosuerce,rnocvuesn a c aomeasos onoen oorere maunu onrcroenvaenn a --- nanaormaarcecuesca o aeccescaoseur vee, orc ae anarmaceuca acceae saereo an aeas one sease, anso arnoer,e orr a ssmec,ome asssoccoasuo erer exrocveens. a meo orean orrevenn a ereo,e meoncun wesease orsorern a suecn a nee muuncona nan amnsern aeraeuca eecve amoun o a comosonscosoearerceenoscoese sueerce.n,n a emaormamceenusc,ae acecreae saereo, ore an oonuceoe suc as m s reease aeuc aenor exame, reease aenr one or morenraceuar or exraceuar eseese ann oneer s auseeccs, ouse rereeassennnveeneoran aeruece ascrene.n moreeaeow. meos an ceom accoosmoanns onerawscnossu arree anncu areencoorroovraee aurer unersann oe seccaon.erawnsusrae one or n an consue aar os weescron ser more emomens oescosure, anoeer . sowvse ao s exanernces an oeraon oescosure. usn m cemac reresenaon oroose mecansm o m ever eners ceuaroeure receo_or me-aes enooce cosossoo oowee ce. e mnzme reosaoensveo en_oso- escaeo successuever merr encon m oe coso.r ma noe ces ouer receors resunne enoc searces can eenn one naure oe crossner use oss oeo_-s.ex, eserases orroea ne sness oeormuaon, enosoma mxure o susresa ccaanne cornaueaees ouers,- anemaazeorse o-ear, acne a cnao renecase as.n me resu oranseaseo enn anosomearosruuconn o a mensesrr,e a c reosouocreesver o m wc resus enzmea weraeearcesroramme e cecenroen.ee o surace crossner.n eser cros mca naure oe mcee me o . cane uzesonrearr oere a ceeavea ceross eneerr esnruocsoumreas s esoewransens oorxn --- rnoeerassens vev.o.eu.eoeeer ceraosesr snereurms seanera oemeon asraereaer sa an eser .o. sows a scema meons. o_-.. ew ceane assem o a m acever vece.rs, se assem o ao_-, a mu-aere nucec m.ex, suraca cna coonncuaoessomes a sureraocrmanea wenn w acre a are eenc,a wsueraeen aenonc nucreoc aoccs.aesxn,e-rmcaeaenmoaezoeo asnom1 za2 n ocu saer aaree ar,ereseonn neave care osome.saer sazeseosomeoreve orm a cae aroune ase m o_ senormuaenremaure ruureurn amnsraon. eman o a caonc suraca wn a nucec ac nanocasue ,rs suracansearousno an aroune cae,oowe crossnne s areen suseuenu surace crossne mcee .ese m o_- ceuar uae sca nconaze aer surace w ao20 or enance exame cemca sruvcenureesr r ceacneoer es.ere a cn eesmecra co nnaaunren onee crossnersee.or reuaee reease oe m eer an eserase or r or aeenero moecue useeren encoes mer aroease, resecve.e m exressona vea c rearacueo rroenoacaeeecon oransaon an e nanosc rrezsaceonnce oo eacna ceer c ouuree aanrcne v asvo.s asse.meno am maes sown eo m_-. scaears are nm.. emeraure ramn exerence mee mrou anoueeor smnuaesos oe m o_ s, ne amoun o .o. sonw s ae cna aracoveerz -ao °n o we manann sa. . ase ronamc sze o -osomes m, - moaeoao_eo-sos. - m oaeosomesunconaze w anso maes surace anmes, - m oaeosoma s, m o_ onc, surace cr s, - m oaeosoma nucoessc ance n manocees, m o_-s, an - m oaeosoma ronamc sze cancaes oucces m o_-s, sowna canesn suraceoena oc ur a ever se o aon o a newaer.. anesn sness oe maceurraseaenersn a o enace s seur.ace.anuonwcoenranasceanraec suocncenssucaes --- eeem crensaa mna on oe m m oaeo_ . resouon nroenresen r oaeo_-s w snasor znc,osorus an scem eresennes, an m comonens.eoome sows a mcroarac os oe marceosaeen anoa_zen ... an . arenm. -s.caearsno . an . mo e su. sows caracerzaon o an anonc suracan oescosure.n orero snesrzaece u osne a ca’onoc moosomeersoeo msoroer aanmoncen w casa rreaacer, ao moe20 snesze suracan useo asseme c6 ce w ann-ouse suracan con12ua eran ormuaons oescosure.e aonoe eecorons saec wnaesr aaceonne awne aeneme neo aancveor ceareno aneoeso cmaeonn osome surace.er reacneoae we suracan a a r c suracan, was aeo a souon oe caoncosome ao o: o ure usneaex- - coumn.a s,ncuaeor anour, an was one aeo ea ss secromerncaea on averaeere srans. a recac seuraocaonrm, a ceonuueaeo we caer suer reacuansons rreoemrr neeo aorn suracan conuae . canen zeaoenarom.o -. m s a eosomeo was oserveor .s sreaowmen we mxure. conu s maes o caoncosomes wncreasn suracan suracaane c cooncenraonsrom o .esusncaeaa were a suaneu sazee a snou saone a eneouo aonru mremr en scoauson oosomes,e caoncosomes consrucon ver.sus coonro.os somoews. a. ana mss oua ooo asno. eme cso oescosuren saes o m oae moeosomes.. osomanr eoncaossomes.. crossne mceesm o_-s.e sneszeosouan surace o szesos exrusonrou a.-mc mesave aare rane o--s are reav roner wereas,osurcaon,e m oae aeo oservee oranc meaer unaso urmnenr sze..maesm aaree s oan me w uraon_ aceaoewoeveer --- caaner.e seen wou uran aceae sann . ueoenoranc surace oe merr m.-ooae. so_owe-uas re wseorensrveea neau wre o eseraos_eoo-wes. a. uer=.oowereo a conon a wce aer soueroonaceae suseuenerae.oa m reease was vsuaze a rev e an assa run on an e.. enaurn eenrsecarannscreon m sreaes weno_-s are exoseoo eserase reease o . orounenor exermensonvesaee r anow conons. roen exressonrom m o_-s.ea anoe o cee aernan corouneorevene mauraon anus a s wererereae w oea an cenesno se ccaon o enosomes.onocamannaers oneraens rum-ae or serum-ree mea sowsaencae merr exreess enonoseovmeess. acaoweearso a arcemno.n m-ceorrro euxneresrseae as rer we reons amonarerra reonsneac on aears as cacuaons sowneowe conocamaesnca an wemae.. nenseseween coroune an non-corouneereaeear saemese.rencee ssnnucoarnecsece wnacse caeor .. snoswsu.ore,s *ceno.ae, **., ***.. w cesno serum-ae -o_ an sers aeroursr oncuaon emssonaears asrer we, re um-reeo- mea.reen reresensu ons w a morearcuae caracernac an we emssonaeoarersscnenace a enm weee as reeasera w renonrso,m we a moor_eu-se casr,a acneue comareoereen emssonsromoecs sane nuce r as .o. sows conoca mcroscoma.cnae oa mrs areem. wen m ormuaons sames weree a ° annc rr exresson serum ae mea.ames werereae,,,uae wea ceorrn weremaeeen a conons sa as aer sness.reae ces amonar me. merr exresson aears asrer we reonsmannereraa ce res aonnsneaneuaocre ascnen wneensmae. c.acuaeonroocma mcerroesncoesn --- ea . an . cenes.orresonn conocamaes o sown mnerr..-err m evere o_-s resuener exresson o even aeran wase.nes m resuss sueovrereer aoueas caonc.o-so.mesm orarn o wsaeerr souon -- oun wen sames werereaenea an cesn s vaons were exermens wereerormenrcaene erumree o- mea. araram o s rerese renooca ses an same ses.eree consern nave o onera an consern- cesromerene ...e sncance was caeor .ns ., * ., **., *** exresson over.mo. sows conoca mcroscoman o merr m roomemeraureoer.meannca oe mmeesrr an exnrceussaoen w wen mea ceosrmouraons were sore a mea.. onoca mcroscomannea ces..rn se,rum ae eanuorescennens cac . . uaeromerenesnea . an m ev ceenes. ona conocamaes o are sownn.. merr m wasere o_-s resuener exresson o merran wene resus suorevere a vae caasonc.os-ome,s w orerne w smaera sro ouseornva evoenns a wer-as.s werereaenea an cesn serumre ereoun wen sames erormenrcaenerenooc e o- mea. exermens weres reresenncea wvaes o o a ses an same ses.ereeararam o snca nera an consern- cesromerene consern.e . caeoor.. snoswon.oc, * ., **., ***.. merr exressona r a mcroscome course assaman o ea or cesn coreressuoen wne sne sruamm aese we mree soare an a ser °um ovreeern mea an coreaen merr exressonne nons. aernc a . an . or . an . ces wouu seraumon o ce.s w, m..ao_-s w serum .,. an ces.onocamae mes were sore overn a °roro aononcuaon a s wereaenus aerre-mosanes,rmeanes ,.n a.erm anaore,r conorcaassmea aese wrereranecnuauso anerr-rmoes-manoessanoers,.r --- maneseoanres.ncua,on a.n ae.r aensouser sowraran msreerro esxresrsncuaonr-mos exresse asrer we reons amon onreuorescence sncan wnrorea an arerra reonsneac an wemaess sncan even ror .maes asoncaea emsson remans sames weree. ao a snoe °wrs cm ooe rnnoocans cameamnr e on moe crers.roen exresson wen m oae ae- mea.xresson o m es anncuae w cesorrn serum evererou caoncosomes vere o_-s waseran wen ere was orn souon an exressonersseor ~ wees. resus sow non snnc.an e –xress.on o. mxeremrren as werereearso.rmes rerseues conssen we ses o sames anooca ces.caears arem mes w eren sames weree. a so °ws coornnocacameamnes o an mnceurr. aero wen exress ceon woren mrn soae o- mea.xresson o merrevere osoma--s erumree s waseran wenevererou caoncosomes orn so_- exressonersseor ~ wees.ere was no sn ouon an s resus conssen w . can exresson o merr aeras. eren ses o sam –..xermens wereerormereemes w e a ° a.en seos cw a asn emao meno s an oca m ce ncues. arre e wxresseona w ceens mo oae sames were mea.eere a ° or roomemeraure, e rrn serumree o_-s waseran weneverexroreusso cna o m evere souon.s resu suorse resus sow oncosomes orn erormereemes w eren sesnn. –..xermens were . sowsman o m oe srarmreose ann exoreoca ces. weree a roomemeraureoren sson wen m oae sames serum ae- mea.xress caemes anncuae w ceorrn exresson waseran weneovne o merr m evere o_-s aeras.eres no sncan emsrseon orou m cearron excresossoomne as oerrnas s.ouso rnes euven --- s suamores an.ooc –a c..xermens wereerormereemes w eren ses o . sowess.caears arem. reee a roomemeraure amaenn oca memerers a enxrenscsuona wee wn m coeaseo sramresn w seerruem e or-an we mneeav.erxeresrsooun o ca monecrr mosome orenvere souoon_ even a-ers waas ere was no sncan emsson o merr exresson aer s. resus sownn. –..xermens wer as.s resu suorse ses o same a eerormereemes w eren .nooca ces. w sowsman o merrroen exresson wen m oae sames seerruemeree a roome mmeeara.ureeorer seamnecsa weeremees a ann °cu oar reoo wea ceorrn o merr m evere osoma--s memeraure, exresson caoncosome orn souon.s waseran wenevererou xermens wereerorme resu suorse resus o . –.. . sows resureemes w eren ses o same anooca ces. oeo_ s reavs o a ce va assa useo assesseoxcroe oecamne . ceseoosomes an m ranseceno ces usn o_ s, ca werereae w eua concenraons o merroaencuae oncosomes an merr m noecamne.ames were was seen e wven a cesor.n cronoraosw,eosomes arneamraenns.eceoo mxc o soweeoox_c a-ows concenraons.ower concenraon oo_ s was sc er crossneormuaon as sucower conce reeneor aee eren.ere, unreae ces were se asnr caeo vnsa were u asne asor coneron vovro sus exeseescre rrorars aree averae oreeeren exermen rmen. sornuc. o. sownnexn aas.merunconazaon oo_-nucnucaonaze wn c aen an sneecxnca sreecnc. a mameerrr reoearreeo aso_-- anse wearreces were eeu wence an s-c ceemascn r seerruemsenreaeo on o-ee msena anor asse mmns. o m.o aane . --- ancaumaeronun ocona-ze ceso w_ar-s.. omarson o merr exresson aer o_ e m, m oaeosomes, m oae scrame se-uencse. anes mus sowoaea aaom_erun-conaszuancononnacrezaese wse e mere aamer or a exresson, oucomene oer conons aer sorncuaon rr er re emssonexresse asrer we reon mes as evence e an wemaesore aamerunco s amonarerra reonsneac exresson as seen conoca mcroscnazeo_-s.. merr umorearn mcerosneoman omcumorssue-secons o - s snesze w an eser crcon o m oae aamerunconazeo_- ans... exer ossner.man o conro samess summarzen. same se mens wereerormereemesnerenooca ses anrere reease. o mo. sowee crossneo_-sor enzme roease reso oumorearn mce.. cemac reresenaon o merrn msvee evxerres oso mnn -ee crossneo_-s.. oserve wen ces wererea anea cesn serum ae mea was overn r,ncanreea wer ex mreoaeee crossneo_-s as ar sson was aceve usn aroease resonsve reeasenece wero mr m ever.caears arem.. -umorearn mce annexn secc aaomaeeee crossneo_- unconaze w an o_- w a srcram.e se,ueanne row an m oaeee crossne o annexn secc aamer or scr ce .,ane row. nmonecon sucuaneous -umo ame seuence wasneceno mceearn anm c rs vaa vennecon.e mce were sacrce rosnecon reasonaroosms soecenoenosus o mumor e wxerreesmsaonere un emers csonoca mcrosco.esus sowe a amonarerra reonsneac on exresse asrer we reons m severe usneee an wemaesrououeumor oserve wen ,ane row.n conras,e crossneo_- aamerormuaon . soweuorescence scrame aamerormuaon .,ane row onroumorssues caoreeceocarozme moceereaumeor werer an, emscre aeanmrear- mumoorae reons. --- rowo_ s-owesm or eemo n soc craemuaer e amsasmoenr mnoe ree caon_ne.u-rerseae. ana,sasne m exresson w umor seconreareown rsom ree aru sem useonr cuas anvsu o . s eee o cuo ca snnoceaou -unour, mes e oc. re.n,nmraoe. s ou –mnora.en.rom am uer.esus sowaeres no sncan emssonromess econ o consere as a conro exermen o .- ue secons, wc cane coreaenrom. a smowusm reorr seescenoanvree coanreoca. romu aor seusccenmaes oumor ee anumornecon.esus sowaeres sncan m euaneous -umorros werereae w m o_-s wo axressonneumorssue wen mce unconazaon.owever, merr e amer an scrame aamer eac an wema xressonexresse asrer we orra reonsn m wase es was moreomoenoussruerououeumor wene omoeneous exvere e aamerunconazearces.s resus conssen we .res ssoowns o m a oamseerrvenunc.. a sucuaneous - seeeum eomoeneousever o m rouou oeumors are orrown one o mce.woerenssue secons mare as we reoxreessen weere cearecu anae meanmuaoreesc seonwnareen.s vaue os areneren areas mare areas.esus sowa m exressons reaso cae aove wenevere usn aamerunconazeonaomoeneoussrue seuenceunconazearces so _-s.n conras, scrameumor. weareerencesn exressonevesrououe . sow a- cesneumors. re-osna onenuermesor w seacs monars seo wwnnomeoene-osuasnesrsusueon se oconmnaese w seecreo wn wans rmeaoens u onuemr coorn ceocsa. mecrosco s.annnass conrmeeuorescen oumor ces anaeeeroenenu suessomoeneoussruon mae arueoeer orescen emsson oserverououeumor a vs scrame seuence-moenceen marcee suvrearceo arnmu naoonuseoa avn aenre anceamne cre-moee ---nserresuson n.ooesee eaxaac su scaoreoe aenauosresscnenm.aes a,nu an a.ro-xm.aeee acreea asr meaar oe. caa, eneraennc,e m sua-aer aen nanoar cocnerso“enan reoceaasseu oes” arerovea oeraren an aroone see.anocasues oesc nucec ac caroe.., m , encasuae nucec ac caro, a osure comrse aosomaaero osome, an a nucec ac na mea orancramewor aero sazee e nanocasue a nocasue oueraero, e..,nuenceearen o as we as reaen comeosveron os a nnuc meec aocs c oar usoe., ouesr, sen nancaonca asuvaes asescreeren, ever ssems. naes over oer eresens ncumosuerre. onenrmssan arenrouceeow, wc are useoescreenvenon o eren,e wa ces were aecnca or scencerms no seccaene ar. vee same meann as common unersoo one o ornar sne cuuroer, e mxoamecuea,r ano noome,nmcmauunreoso use, mnc croonnoecoon, wen, anecnues o, ce anssue cemsr an rzaonescreeren ar ecs, anroen an nucec ac case o conc,eresensc e wenown an common usene ar.n an maeras ar osure,ncunenons, w conro.xemar meos es eescreeow, aou meos an maeras smar or euvaenoose erecrn.eeren cane usenracce oresn oe emomens an asecsescre carrer”, “so usosemaereena,eerms “nanocasue”, “nanoarce”, “nanocarrer”, “nanoarce ”, anee, reerancramewor-uceccanocasue”, an “o_- oerwse encomassnr oro c marurna oerne na onro mscoaree c caormo mexoeesc cuaeasoer oe encasuan wn or moreocaonsn aoe.., ce,ssue, com ver, e..,o one or comrseaersncun, e armen.anocasues oescosure can evern .., a caoncosomeaer comrsn a nucec ac caroor sur o a ce, a mea orancramewor aeraes sazeeosome, a eace cr coasse,noeo mwecee crossnaneroerm sueracaneeemaaronus o a an ca aoanecr suorrmacean aroun --- m enoancen ceeu saurr uacea oee scavenaerer re wceo arsu,nocoornma ar nouu, e.., ao20 oraer. cec ac nanocasueessan aoueerm " nnma.noaarce", as useeren, reerso anarceavn aameer o s,omers, or mxuresnoeraeroc.es csacno csoem narnsoe,a er..c, oene or more naura anor snec aameer o aounmo aou nm, w s mancue nanoarcesavn eren. urer seccearove esewere aer.eeermermo "somoseomnce"u enesco umnaamsseesa arn sr cuocmuraers,m wenc en ccoomserse a a snease one an muamear srucures, wc comrse morean one aer, "osomes" can reero unamear vesc aer.s useeren vesces anare un es an, more secca, sma unamear ooamear vesces a amear vesces ,oueaer vesces , ae o nanoa n muamear vesces .s useeren, aosomes coesero,ae-ser ormce u ". e va"m renesr,s moono mceoreceus,esac canncueas, waxes, seros, cos, caonc or anoncs,ervaze eres,osos, snos, eow. s canorm mcees, monoa s, anee, asescrenea assemenoosome ers, anaer memranes.es can se- a comosons unseo ore ores. nno,ase aermro “aramen as seurnec”, r aeeorsoe acuascanroucon onroucne comosonnoe suec,o s, or ce.n an a meos onvenone meos noeen s or ce are conemae accornoe so consrue.eans onroucone anre o wne an-noawrncuoar moseean ss oennrouec aro,n a anns nooe exemeeren. “rovn” meansvn, amnsern, s aso are orro or no, manuacurn, como en,srun,ranserrn s useeren,eerm “arumnacneu, ocras aecncsn. aresoocaoerae ano noca eae” reerso comosonsa reacon wen amnsereo a suec,reeraro auucmea ann o ar aer nocn- oru smmana sru unecow.arreera, --- a os u aseeereare onr, saeeeromve “rnmaermna ocreuscean accee.a.e”a mrmeaancso aerove a reuaor aenc armacoeaor usen anmas, an more a or oerenera reconze s useeren,eerms “rea,”a “rrecauanr,” annu “mraens. aooca conon,sorer, orsease, e.., arr amen”ncuenne aooca conon,sorer, orsease o esn or reucneeveomen oe conon,sorer, orsease, e rs cnca smoms or reevneaooca sease ors .., causn reresson oeaooca conon,sorer, or conon,s cornecr,a or smseaosme as.re aremaemoernae me oarns o anerw waseeene smcaom as oere a.areoeorca suecn nee o sucreamens a mamma, anarcuar auman,u a, a eneeow.reamen aso meansrovn an acve comou s no some as suceno measura reuce ansease smom, no aaenn an amoun reresson.eseerms a sowseaseroresson or causesease s useerens,o eenceormm "aessecervae am aonun cu"re. reersoe amoun o aera, wcs suc or “eraeuca eecve amoun” uraon o asorer or one eno reuce or ameoraee sever anor sorer, cau or more smomsereo,n orrevene avancemen o a roresson ose o rneer oerss moonre o s amsoomrser a,ssnocae or wrev aensoer reercurrence,eveomen, onse ormroveeroacc oreraeuc eecs ,eec asorer, or enance or eraeuc aen.n eecve amoun can r o anoererae..,roacc orerm “roaxs” reersore eure morean oneose.s useeren,e sasca sncanervenn or reucneroresson o asorer, eero a ecve amounese m oro aereeeecae aerson o ornar sne ar.nvua, cononoereaea, v anaroeenn uoneooca eecesrene accornoeresennvenon are secc caracerscs oe comoson comoson cane amnsereonenvua.ns resec, an suaeose oe ereae.arousener eaene..,uman, accornoee oseaseo amoun” are a conseraonsaenno accounneermnne “eecve oomannnownmaon’s:osee o sarmnacoeo arca ana asrees oescreera,e eu..,csn,m ea.n, eer aa.,m eons.,ress, --- , e aanc oem wncons’serearnmancceourocaraecences, .,acusno.,ason,a., s useeren,eerm “ reerence. or comoson,n ose”enoes anorm o an acvenreenormuaon eraeuc ecun ces,a conans an amoun sucenonae orrouce a are usenercecan wea aeearsen o.nes or us meoree aremn,nseraeromnss. “c “onorrmou,”a oron” an “comoson” erennercanea. “reerence” or “con “reerence” are use wcs emoe as aasene o ro”eve mae areeermne vaue or rane, “onro” aso re rencmar aans wco assess a measure resu. eeremrss "o ceornaroeu exc"e orrm "enes or conro ces. wenresenn an eec raeuc aen" reero a comoun or moecuea, ereo.e ve amoun,rouces aesreeraeuc eec on a suecn nee conuncon wresenenvoensomone c coonmemosaes aroa rane oeraeuc aens aner usen eer ons, asurerescreeren. acve aen, sucm as “ a coamrmaocuenu ocra sa com oosormounsa” me,a anns c aomeaoss onoens o coemr srsn aeas one a carrer.armaceuca comosons meee.. ’ usance, suc as racce sanar s oo manuacurn eesoruman or non-umanrus. comouns amrmns “cearerr.er” r “eerasrmoa aceuucean, e axccceean,e or ca vrererc”e m weans w a sucs aan acve excen,uen, or vece,as useunrearn aarmace nce, e.., enera sae, non-oxc an neerooca nor oerw uca comosonas carreras acceaeor veernar use a se unesrae, anncues a “armaceuca acceae carrer” s we asumanarmaceuca use. eerm “suec” or “ancueso one an morean one suc carrer.ncun armae en”s useereno reero an anma, suc as a mamma, a non-rmaesuc assuc a co aws a, aum,an a, c aam noen,- aumamana, armoarsee,, e a..o,a a, m ao rne, an a cmanzee, aunea, a ca, ao, a ra, a mouse, an a wae a, a see, aamser, n an emomen,e suec oraen , are.., auc or aoose, an a sar. enreae or assesseor asease,sosr aeru omr caonn suoenc, a oru amaunm aan rsaeonr a, susceas aes, asuomraenr --- ors ceoasne,osno,r aeru omr caonnavonn a asessecarse,eseorer or conon, anorumanenreaeor a ooca,s m uesecaer,e onr,n a suuaec os “enr r ne oen me. su ocrreeaammeenn”. s suucec sune ncee wouene oes no necessarresen smoms,arcuarne ca oreamenreamens. se orevenave orroaxs ercenaes are cearcceunaes an raos are cacuae we uness oerwsencae. une aease oneoa comoson uness oerwsencae.enera, ercsesn o ameorwunse o ex anrenssre seane reeerresno, "wee amo"u onr "amoun" as useeren w resecoenreen, werene raw maera maee oe raw maera comrsne acv oenreen.ereore,screereno comrseessan an uo reresene ase amoun o we ercen o an acven a comosons reecena raw maera conanne acveas use an ma or ma no one we ercenae oe acve, werenenaercenae oe acveseenen mereceno os aecscvreeneer reawn m caanera. ncaeeren or oer eerormen a suae orer uness oerwse ex wse cear conrace conex.e use o an an a exames, or noemosaer aamnuaaoen oen.., “es succo aes o”,sennevneneon m uerneesso oeeerusraeenvenon anoes seccaon soue consrue asncan rwse came.oanuaene oenvenon as u an non-came eemen as essenaoeracce erenavee samese meaenrenn. asnse csosmmeonne o uneerrwsoseo,ecn oncea o an s scennce aerrm os use scosure. s ermuaonsurn wermcore o,ne ore morsecomsuraeon esn,comasses a varaons, comnaons, an or more oes eemens, causes, anescrveermsrom one ee e cams arenrouceno anoer cam.or exame, an camas camnean osn aenoeneren ca omn caen saem meoasee caomn.cuee orene e oerm meonrse armaonsounn an oer arusrouorma, eac surou oe ee eresene asss, e..,n removeromerou. menss asoscose, an an eemens cane --- meo o reeecrranonn ov ranuaes oo v eaacues seearerane a vrea mueereanne wneno seerve as a soranncaeeren, an eac searae vaues rane, uness oerwsenvua rece.or exame, ncororaenoe seccaon as were erees, ma a sze ranes sae as nmo nmor concenraons, nm ss amouns, anee,snenea vaues suc as nmo nm, nmo s s,ec ncmaoon. nemse, are n omno exam nme,s o or w nmaos se ncm,c eac., anree enxere,s asn e anumeraen comnaons o numerca vaueseween anncuneowes osse enumerae areoe consereoe exress vaue anees vaue urermore, wen "aou", "aroxm saaee"n asscosure. escre a vaue,ss meano encom nor "susana"sare uzeo vaue.ere no sae vaue ass mnor varaonsuo- rome sae aou , srove, an eemenescre as "susana" means aeasne conex.,,,,, or more oe eemen, assoca coeren wnan ness secca saeoe conrar,or ranes sece usn "aou" examua "ee .,, "eew aeoeun a aoueso mo an ens o"e rse ecueva raenneo w "eewere sene acoue or m no. anor aouoe consrues us aes oeerne-enn,eeeerrmmss “c.eo.m, mreasnn”, “avn”, “ncun”, an “conann” are oerwse noe n “ncun,u nomeo” uness ereo,e are. cnnroucn eemens oeresenscosure ore asecs an emomen eemens.mares, "a,"e " aane,"c anve " "aneo" aerre,"n wenene useo moeannroaucere are one or more oe mean one or m e an eemen,sneneo eorer easeem "ens. uner anor," as useerenne seccaon anne cams, soue conunscooveo mreeasenn "en seorm oer coase"s a ones euenmcenves so creosnenone,.e., eemensa are se w "anor" soue consruene s n oer cases.ue eemens so conone.e ameason,.e., "one or more" oe eemens ene e "arn eeomr"e cnasu msea, w oeoenra reaee orers uennre oaeeroanosee e eeemmeennss s seeccccaa --- coennunceo.n wus, a os aen n-oenn-emannu eaxaem suce, a as re "ceormencreson " anor", wen usen onoonancun eemens oera " can reer,n one emomen,oncun eemens oeranne anonn anoer emomen,o onoonancun o er emomen,oo anoona ser u eseemeenresn ec. avee sam ne seccaon anne cams, "or" soue unersooo s, "or" or "aen meoarn" sna as "eanneorr"r aesee asneen aovnec.usovre e,xa.em.e, wen searanemsn a asoncun morean one, o a nume ,encuson o aeas one,u ems.nerms cear r ors o eemens, an, oona, aona unse wen usene cams, "cnoncsaseno o,"e w con rrearer, souce asn “counson on oe o ex”a or "exac one o," or, numer ors o eemens.nenera,eerm "or c one eemen o ancan excusve aernave " as useeren sa onenerree as excusv, s s.e., "one ore oeru noo" wenrecee erms o suc us as "eer," "one o," "on one o," or "exac one o." reerenceo ase o oenreen orn moree s eeemcencas,o sno aunne une cerasmoos,oe meraase "aeas one,"n seecerom anone or more oe eemensn n aeas one eemenncun aeas one o eac an eve es o eemens,u no necessar no excun an comnao r eemen seccase wnes o eemens an a eemens ma ns o eemensnes o eemens.senon aso aows s o eemenso w oconaeerasrees "eane oaser onea"n reeers e,e wmeenser s reecaeca or unreenaeeo wosne eemens seccaene.us, as a nonmn exame, "aeas one e euvaen, "aeas one o or," or, euvaen "aeas o an"or, one emomen,o aeas one, oona one o anor" can reer,n oonancun eemens oe ncun morean one,, w, noresenanncun morean on rann anoer emomen,o aeas one, oonane anoer em e,, w noresenan oonancun eemens oeran one, oonancoumnen m,oore aaenas on oen,e, oanon oaonnacunncu monre oaenr o eneem,en, asn e ac.eas ---nvua comeonreanses " aosn wee or m aso mre,x" ausr useeren, means aeas one, anusncues comnaon. escomnaons oese comonensn an e s useeren, “oona” or “oona” meansae suseuenescre saven ev oern cr ocru cmrscuamncsea mncae o or ma no occur, anaeescronncuesnsances were s useeren, “accu”rs r annsances wereoes no. romo ca eerso a sra orrance canrocaron conann u are nomreono a, momes un,e ess o, ne-rwrose s,esco-ero.e, nr-esuena,ve exames o ancue, n-en,soen, neoen, n-ex, sec-u,so-u,er-u, n-e, n-oc, n-non, an n -meex,,-meen,,-meen, oer moees,en ma aso-ee ac sr.aen or anr “aance” croaun esx aamnensncrouue,euw aereen nwo meo --, - -, - -, an - -. o s us2eere2n,2 22 3 2 23 2 c “aen” reerso means a sra orrance canrocaron coarnaonnnouroemono.e craerseonnsa, uvnee esxs oerwse sece, an conann aeas one caron- -roen,-me- ames o aenncue,u are nomeo, een, een,- -roen,-uen,-enen,-exen,-een,-me-- s uesceene,r aenn, “,a-nme” reoecra-,-en. conannromo ca so a sra orrance canrocaronrou eresenave exames or aon anomsn acnu ceo,nuan arne n aoemase on,eo ca arceon-cearonreon. ron,-un,-e n,-ron,- s useeren, “nn, an-un. aren m aox” reerso an a rou, aseneeren, aeneoe u are nooecmuare moo,e merooxu, e ano oxx,eron aoxom,.-eroreosexnave exames o aoxncue, exox. ,uox,er-uox,enox, an us usseuenerse anre, “naeon”e or “oaoen” reerso -, -r, - or -. oron oe name uness aasncea reeas o “eerwose r.o”r ex.ea.m,ee, a1a-c6m aenoxs vcaaroenas e sameun1c6 ona smar ara an –aar-nc-- an - -ancaee sameunc ae morean one so son eua o a. r asco,e uen oreresro ooea s,ens c oerr aan meosescreerenancue e orern wce ses or acs oe m cs oe meos no necessarmeo oarercwusaer. oruerrner,moure s,meermors cornsv,e sneceoe nno ce,o e acr.e e,n a re osc e use ae un uerareessn a oree c , no oon mexeannocaeensoe an ovnoe seccs oe suec maer,scoseerenr exame,aers. muuncona nanoarcesor nanocasues,e are muaere, nanoarcesa are caae o enc nceorescre as muuncona sma moecuerus,oocs, oraessu ora ran oneu onrc moorneazeera weuc aereanesucacr nousc seucc a acs ora sonvsaseucer aeso a.n azmoen, or se mu saeeureenc,e m,u or auncaromnaace nuaca acceae sa,rae are caae o nucec acever anaree noarces oescosure nanoarces caneareeo sec enenocown.ese muuncona ase nucec acans.or examce, c meuc renc-e, aonrsnexnn-co,rraonrsaenrrn an n ourceon a recaemers canearee usneruse aamer seuences.n aon,ear ors e an reease oenernazeru caro canerora eereaown o seccroeases.e., caesns, marx meao mmeoreraaon e neeornraceuar or exra roeases,or exame eenn on . ceuar reease, uzne unue crossneresn oe aress a numeenes oe muaere, muuncona nanoarces oescosure a o easeru onc counrraenze ane smuroarcaen o arurueveerverur veesceresoernenraeeu arc, suc ase acaons,oeraa, ane caacor comnaonera, as orarenoae w one or morerus, ane surace moe w a s eneror cane omoecue earaeeraeuc nanocasuen. one as, aec m,e aa m ouraunnccona nanoarcesrove,ncun a nucec ac aens, or aarmaceuca accearaem seaw,orrae or, s aovaoesomeer,eo an,n on we ocr moreeeerraaeeuucc --- a coemnesx en acnaseene-osomseeosomes encasene o mae a- n some emom encasene . roocrous oe amens,ees are arnrcanuees noowna-romeerarc acmeneroers,.en., woce , anerocrous oe am es are arraneoware ware an are crossne w one or morener arce surace exraceuar reease aenssuc aseasse c,earvoaeasee, es oenrea oser, morenraceuar or crossne mcee ,o w or easase,oorm a surace o c one or more nucec acans are covaen aaceo rromu a ara.zone so,meo eemeor, omr aenesn,e sureorouc wrou osn oe am es are crossne n some emomens,e muu e or more ceavaeners. nucec aca ncona nanoarceurerncues aaren s, smanneree.rn., aamesrss,erase,u mcc oroonucsemoes, asn, rsenosze oonuceoes ee nucec acs s,oce nucec acs s, mes, zmes, ereo aa anee, or comnaons nceoe exeror oe . ero scomroeu ems oomee anms,erea aznoe,eoneeerr, re osru asernom s auerou crossnn aene moe onerocrou oe am ee. reacon o ane or aze oro moe onener. ., one ammonumrou an an rrocrou oe am ens so omnee o ermwoo omenros,--en a onre oror a--eenne mo onee o ornwoe o eo a-m-n, oer osner orrowo- o-n-ro---aemnno, a onrrna somaem emomens,erocrou o ease orn sroomeaes eem,no wmecns,eeeneersn acueaess an woeo.eor exame, ceavae w a amno acnos acon, or aeas amno acson, or aeasree ve am our amno acson oreeeseweenwo anwen- :s,on.n some emomens,e:eenerncues ,a-,:a,-s,a-r, ore-:s., , ::, a ,e-eu,a- --- ceavae wn o ane ers emerasoe,m wencs,,ne sonmeer emncouemse onnes, orn mcourees o eeseororwounsrooru exame,
Figure imgf000024_0001
. orm meonres, oenraerncues an ac-aerou rosaene a ac ea, oroeezronreo,u sesm, ocra ar caozmone,naoonsemecreaorazone, cs-aconc rous. , or oer ac-ae emomenns, someen emero excmuenes,enerncues asuerou, wen some conons. ssuerou or oerrou ceavae uner reucn reease aenn, s aonmen em ooer emmenos,meeners ceavae onenraceuar or exraceuar exr ns,eners ceavaewo or morenraceuar or rouacse eaucar c reeaevaase aeen ase.e.,r werenener comrseswo or moreeren cemcaeasworous seecerom ane ens reer,easer aazeonne,. senm socme emomens,enerncues a aconc ame, oroeser, acea,ea,oeer,suaer, aanzon ae,eosee,m wcaerreanzone, cs- aeaswo amno acson, or aeasree amno a eeeson oreeeseween a cson, or aeasour amno acswen-ve amno acson, aoouuwwoooowern a ammnnoo a accssoonn,, aoro euxame, aouwo an acson, aouwooen amno acson, o wooeen amno n some emomens,e nu r aouwoove amno acson. rocrous oe am cec acans are covaen aaceoe emomens, esrou aoeer or aen suerou.n some rocroue ooee aemr or aene re ssuuseromrou a re aacaconn o ae nnuec oerc a acene mansoe rocrou oe am ee.., one ammonumrou a oe one s one nuc n ao moee.., n someec e amcomanen. am essro--n orrso,-e-e an,n oer orr ao-en-en mo,e or oron-e-enr,o annc soromues oe --- emn-o-ammenns,e ane or aene moe onerocrou oe am esro-- no or aamno. surace ane sno,m aer eomeno,mens, nucec acans are caae o seecvenno a ce e nucec acan see ac cavreonraseo, o aroro ae nucec ac.n some emomensn wc conssn oumor-marers,nerns, ce suracen r,eceeororsens seeceromerou cannes, memraneransorroen, enzmes, anoes, a,ransmemraneroens,on emomensn wce nu n cmercroens.n some caro cec acan seecvenso a carorae,e cocarxa.ens so semeec eemorommerou conssn ocoroens, suar resues, an ac,e nucec acncuesennsnn wc,e nuc, meco acean, se meocveens,o a nzucec rozmes, m, s , mcro, s , an comna mes, n some emomens,e nucec aca onsereo. werene nucec acs s , zme, rozmne, ms ccaroae o, ore one reeruearaone, an oonuceoe.n some emomens,e nuce uc o a ce receor, c acans caae o seecvenn eveomena s oreco ac c cee suurrnace a a snecencseveomena saee.., saeavn umor ceaasumor-assocae anens or orumoo ar- csen a seccsease saee.., a emomens,e nanoarcesareeo a secc ceec recce anor, nenosn.musn,n e some suc receorsncun nuceon,ranserrn, annexn an mucn- receors.xames o earn non-omerc am es,n some emomens,e am es are soemrveerom
Figure imgf000025_0001
saoen, ann s
Figure imgf000025_0002
--- aso aaoen.n some emomens,e non-omerc am es
Figure imgf000026_0001
em menesa,oens an onrcaunes a ans are coornaeoorm aorous,ree-men c e2+,4+,nr4 s sonaramewor arou +,ome4+ e,mo2+,me+ns,4+e m3+ea2−onncu3+es one or more on2+n,e2+,oas2o+m,ee.3+, onsn2+,2+,a2+,e3+, o,re2,+, an,n s,o amne emo, wmeenns some emomens2,+e mea n some emomens,e orancann ,e meaonsn. -memazoe,- cues one or more o-memazoe, -me--erazoem, oer-m-a-zmocaarzooen,rme.azoe,enzmazoe,-me-,,-razoe, ncues-memaz n some emomens,e orancan en oe,-memazoe,-me--mazoe,mazoe, or mezmmazoazeo.en, a snomen e smomoe emmenos,me orancan comrses-memazoe, or- emomens,e meaonsn2+, anenes, orean orcanacnasn-mse-memamzoazoe.n ceran emomens,e orme n2+ an-mema e, ann some ramewor- zoes aeocmazoae caonces aanr- an onee. or moosorem neo,nn-c saomonec emeomens,eosome comrses one or more more caoncss a coesero-ase , wecrn sosm.en e smomoe emmenosmsen-s,-e o',ne or meamnoeane-caramocoeserorocore '- n some emomens,e one or more non-cao-o. as neounrca or zweronc asooc nceer sncues a non-ca a, ann some emomens,e one or more n soemeer emosm coemns,rsees na,n-oaorecoe o-sn-ecero--osoeanoamne . uracanonuae ncororaenoeosomescaoesru,r ee.u.,reorsonmceuesa aer.n some --- emo-cmoenesse,roe c noannouaare,ce o-oecosceorsureurerncues -suracan conuae, a nc o conuae, or an comnaonereo aorso arnaoencn.on soeme eomsomoemaeenr.s,n seome emncoumesen a ns,on-eo nanmoerarc acmencuees a accoraner saoen an aoae
Figure imgf000027_0001
am e seecerom
Figure imgf000027_0002
s neunceo, aeosunen an, a nouuceo anesn nuceoesnen, aou an nuceoesn an nu en, aou an nuceoesnen, aou nen,c aeooues annen nu,c aeou an nuceoesnen, aou an nuceoes some emomens,eoaeo oesonnuceneo, or aou an nuceoesnen, ann e vn some emomens,eoae oesoo20. am nuceoes reace we non-omerc ncues oneea ora a mrornon e oee c aec erra c aeeuem ucs c ar nuecn re esac c a,o cnn s. ,o ame ermooomcen smsae moeeraceuc aen an an-cancer aen, an anoc, an anvra, an anarasc aen, ueruncun anaesc aen, an anesec aen, anon canneoenaor an ancoauan, an nammaor, a meaoru, an an comnaonereo., anon cannenor, an an- aunorucnn, s vomncer esmneo,macenasx,e,e neoecraarzeunosca aenn, canccueaems ccamn, coseacnn,, caorxooruacnn,, --- o mxeaoarexna, sea, vranaasn,n, vconcrasn,n,urooecan, annamcn,oceaxe,amoxen, erucn, armaceuca ac oecan,rensone,rensoone, a-, or a n soceae sa,rae or sovaeereo, or an comnaonereo. omanerermne emosmens,,eceroraeucm aens, a nucec ac comrsn one or more rozm nsenseonuceoes s, ereo.es, aamers, m,ce-wcn onuceoes s, or an comnaon auoroorne, s aom rae eomaoeeme nnucs,eoe nea,n ao raarocseo coomerses aanosc aen, wcncues or an eecronensea ,on,ocoero, coesero, a sero, n an asec, a an com a mea ceaor, or an comnaonereo. smomn a s osonorrean orrevenn asease, asorer, or escosure our aecarsmarocevueca,e ac ccoemaoseo sannceureon an a aeasea osne on oe nanoarces o acceae vece earmaceuca n an as anec,o ar e mxcen. ncun amns eo orean oranosn asease orsorersrove, escosure,n werncoe a snueercs cne naeveaeereo on aen o er meocreven armacoeunu oare nanoarce o reease aenresenne suec,us reeasn or exraceuar n some emomens, a meo oreeeraeuc aen oranosc aen. smom assocae we c an orrevenn asease, asorer, or ene exres onons wc reure m anus reure anncreasennuenza orson.n ceoran c eamnceor omre an cso,men caonononerenoc.unes c communcaeseases suc assevererou arocess o vaccnao eran emomens,e meo n ceran emomens,e men,oo mr eoxaumae, o vaccne. mouanemmune resonse o esemmune resonse o a suec n ceran emomens,ee m suuecunc aernene exresson. escreeren, arearcua ona nanoarce oescosure asn conuncon werar acaeoeever orooc sma moecuerus s , ansense oonuceueoc oes,o mnuccroeoe,s asoammeer nso,nmznm eexs,amoezs amree ms, ec.,a are --- u msueuunocrnonraace nanuoararenceesn oocown an aernroen exressoneves.or exame,eoosomerasenor an cancererusc,o asuorne a wses acrezemreen seecvecra camesoneecn,o aarenea cmemavaaoen oawas- mna.now-nsrse aamranscr anonac coeraeraens aanomn.oreaanv roen m can resunoceroen ae o n some emomen ransaon. an aou , n s, no morean aou , no morean aou , no more eners ceaveo mnor aen exanra aceouuar e,n nvoro mnomreen,a ann aonu some, e or no morean aou o aou , noessan aou , noessan ao momens, noessan aou , or noessan aou oenu , noessan aou , noessan n some emom ers ceaven annraceuar envronmen. c ens,e reease mecansmss an enzme exresse umor ceaves,o aane ane snome emomens,e reease aens asosome aen, enosome aen, anor o an.oarces oes romon auceeaose.ans s aome ℃ emoo ameaec snoss,ureea sem saonsra a ℃ oe en .n seo na man encoe ema sra ocems o enssc,oeseever seareen cane nanoarcesscoseeren canerom aeasas a room a oe a roomemeraureor examee sa emeraureo aeas as as.n an emomen,e sa cane aeasas,as,as, or aeas as,as,as,as,a oe nanoarces cane aeasas,as, emomen,e sa oe nanoas,as,as, or aeas as a ℃.n an or aeas as a roome rces cane aeasas,as,as,as, o meraure. aers wcosso com ame nes esn,nenera, are serca vesces comose o one or moreaerse.., am c mo casuae aueous souons.osomes arecaorme s veesacraeseesrnoecu eoces en a, cn suc a asuarsoe anosc o s evoemn rane esr.c or aea n uoa cnur aca o enor s,somnesa ar ceomosrsvee sa caare moecuese nucec acs, e.., an . arcuar neave care --- eren,enr saosmee “c eamooncmens”, reeorssomoe asn co omrse one or more caoncs.s use osve care a a seece, su a numer o secesaave a ne more caonc c assooca.n some emomens,e one or -','-mesa mmanoee canoee-scaerroa-maosec.o seusearoe coroecseoror-ease caoncss- n some emomen,e one or more caoncs ma or “-o”. oeoxro-,,-rmea e--,- car mmonum core or “ ”,- carooxxamseromec-ne,-ocmaeec-am-roe oarna “mnum”, or, “-oeo”x,-,--oseoerm-n-e- “mea”m.monum-roane or “ ”,,-oeo--rmeammonum-roane or amnoroane oro “na exem”,ar,- caooencox-s, a-somnecue-,-searox-,-me-- noeox-,-me--amnoroan -amnoroane or “ ”,,- -amnoroane or “en ” e or “n ”,,-noenox-,-me- ,-sear-,-meam,-oe-,-meammonum core or “ ”, ,-me--roxemonumrome or “ ”,-,-mrsoxro--- -en--ea ammonumrome or “ ”,-meamno--coes- coes--e-no-x-ueaa-no-x-o-x′--ox-acesn,cosx-,---ocmaeeca-enoxroane or “n ”,-′- ocaecaenoxroane or “n ”,,-me-cs,cs-′,-′- “ ”,,-,′-oecaram--meam-,-oeoxenzamne or noeoox-,-meroamne or “ noroane or “car ”,,- meamnoroane or “ncar ”, n”,,-,′-noecaram-- or “n ”,,-noe-- ,-noeocaram--meamnoroane ,-noe--meamn meamnome-,-oxoane or “n-- ”, ,-ocaeca-,-eoe-,-oxoane or “n-- - ”, an-,- ,,-noe--mn---,-oxoan---,-meeanamnen-- earacona-,,,- eamnoe-,-oxoane or “”, ,,,- ,-me- eraen---meamnouanoae, or “”,a,s,a— amne, or “,--”,,,-,oc-aresca--ox,o---eunneceraamnoro-roa-c-co,ena-un,eco-xo-- --- ,ro,an,e,- oerr “aazaexa”,e ocarn me-x,ure-samereoe, or “ -”,,-oe--meammonum e.,,n c-er,an- emmeom-e-ns,,e-oocs.oameecsa c-om,r-seen a-nonzae caonc, suc as, ,,-,-me--,-ocaeca-eracosa-,-en--amneren--amne , an ,-,-me -,-en--eracosa-,,- eracosa-,,-ren--amn --,-ocaeca-,-en-- n some emomense,e caonc, or ms cxouresereo. n some emomens,eercenae o camonrcse-n ao. an,reaeran,reaeran,reae osome maereaer ,reaeran, orreaera ran,reaeran,reaeran s consueeween ao n we.n some emomens,e caonc aou-, aou-, aun-eee. o., aou-, aou-, aou-, emomens,e caonc e. eosome moeracon.n some aou , or aou oeo.s,-o consues aou , aou , aou , s.s usne someree enm,oemreanss,oosmoemes c moomer raseo o.ne or more non-caonc“eer” o e “non-caonc” reerso an neura or zweronc. zwn- osecraoonncc,oesoncuose,au arec noonmeeo, nseeuarrao,,o-soaeoc-sno-necero csoaeaneoaamnonaemne,a,mooeoo -- o e-oososaaecanooanmene-,-maamemooomeeo-- mcoresxoane--caroxae -ma,amoosa eanoamne , n soosoeanoamne , anee, or a mxureereo. n comnaon wme e omeor emxcense,n sus,core exaemre, csa maonce use aone,u arereera use ercenae oeer n aosome ma s.n some emomens,e ,reaeran,reae ereaeran,reaeran,reaeran we.n som ran,reaeran,reaeran, orreaeran aou e emomens,eeer s consueeween aou- e.., osome- m,o aeouraco-n.n, s aomoue em-om,e anosu,e-eer, aoue.-.,, an conesuee os aoeu --- rao,. aou , aou , aou , aou , or aou oeosome moe e meaons orea ca o us or era rsan cnc ocr orraam nme aew ewoorrs os s orancs. aarne as, caosrsm on cr asoarounse, mareeer-ams ceonmsonoase o craemmewcaor an. s arenownorer surace area,unaeore szes, an exceona 3+ ourecremsaor s maea. ea 3s,+ on2−e,2+ o3+,e 3rr2o+u,sa2 c+o,nscs3+o ,nns 3 o m +a ,n4+2,+e, a 3 m +e ,2+a,2− sa ,a2+,3 a +n ,e3+or a a,3+,e2 m+ea r,3+,4+ o,x o3r+4e ,+.,3+e ,4+a,on n3+2,+s,nc n+2u +,e ,4+ a, e+,e+,u2++,u2+2+,s33 ++,3s+2−,3+o3+,4+o2+,2−2+,4++,2+r2+,r4− +,2 2+,+,2+,u2+,u+, s+,,u5+,,3+ a,n,+.an s,omne, emo,me,ns,e mee,aeon,snncu,en a+,eas4+, one2+,s5+,s3−, con 2s +s 2+ 2+ 2+ 3+ 2+ 4+ 4+ 4+ 2+ + 4+ 3+ 2 o−ero3u+s we,,nn a2 s+o o ,men e3 e+m, , anom, e2e+na,e,e,,r,,,,,,, an , .s,n soem mee eamoonsmnecnus,e ae meaesa onoen os arer2o+us conssn on2+, n varous asecs an emomensescreeren,ee orna. oranc moeesa conan one or more coor ncans reero srucure.n one o em naonunconarousns moecuar e v omens,e coornaonunconarousne orancans maeasa ornoeus ounecoronuasr coounsss cana oe — oorm,n — coor, —nae,o —ns w, — mea, —ons,ncun a memazoe,-memazoe,-me2--m2azoe,maz 3, —,- me-,,-razoe,-me--erazoe,-m oe,enzmazoe,- s4, —, — 2, —, — 3, —, — 22,, —azocaron — 23 a,r n —e, —42, — 2,, — w —s3, —3, —neunc2 3 22 23 2 3eren eac rns. onarousneenen reresens arocaronrou conannoenzenencue aenas so omnee e omrooumse cnosn,ssen coo or —naonunconarousne orancans memazoe, anor-memazoe.2,enzmazoe,mazoe,- --- are exemee oreaonwc.oran exasm conea,nen oraen acovea-nmesnnconuee coeornaonunconarous -nroereac ac,-roxereac ac,,- reac ac,rmesc ac, amnoereac ac, monosoum-suoereaa roxereac ac,- nrosoac ac,-roxsoac ac, monose,-amnosoac ac,- ,′-encaroxc ac,,′-nro oum-suosoaae,enzoc ac, encaroxc ac,,′- r-,′-encaroxc ac,,′-amno-,′- eneracaroxc ac,mercox-,′-encaroxc ac,,′,,′- ,-naaene aosuccnc ac,,,,-naaeneeracaroxc ac, - caroxc ac, naaene-,-caroxc ac,-oso--reonne, mercaoso-o-,,--rroasznoe,e,ar,u-mm-ercaososou-cc-ncc aecra,e,-mercaoroonc ac,-amno-- me--suoerazoe,mazo -meox--mercaoenzmazoe,- caronmazoe,- e,enzmazoe,-mercaoenzmazoe,,- memazoe,-mmemazoe,-e--memazoe,,-memazoe,- memazoe,meazomazoe,-nromazoe,-canomazoe,,- roxme- e-,-caroxc ac,-amno--mazoecaroxame,- cano -enzmazoe,-meenzmazoe,,-meenzmazoe,,-sorommazaozoe,e,en-zemnza-zo-me-e-carmoxazco aec,-,nro-mmaazzooee-,--aomrmnoc aecr,- monorae,erazoe acec ac,,,-razoe,razoe-- azoerazoe,-n caroxc ac,-amno--,,- nr so-,,-razoe. memazooem,ee onz emmoazomee,nms,aezo orea annc-aamnnsoesr seeeceacr aocm,- amnen sommazoe,- e sommeo emmeonsmeen osr,anec oraans seecerom-memazoe an-memeazoe.n n some ncans-mem2+azoe. memazoe.n e cmomens,e meaonsn, ane or2+ancans- orms aeocmaezroaan emomens,e orme n an-memazoe esraee o comerrsaeme ow zorn- - arouneosome.-s a aree-menso consn²⁺ coornaeomazoae anons,ormnooo,.e., anoaroursam seruwcourre w w aaoreo coav reesse cmonnence zeoe nsa,r wrow w a snooawse.- s --- saave un.ueroeres,ncun surace area anoros,erma sa an cemca use comrnsese anre rsenscosure,e amoun o or rao oe mea sourceso oranc ac mo, aou. maoa resusneesre rouc an mae aou. moo aou. meosooo mo, aou. moo aou mo, aou. moo aou mo, mo, w r aou mo, aou. moo aou. mo, or aou. moo aou. uceceo mo oe orancans. sc o ace nanocasues s roocrous oscosurencue non-omerc am es,n wce orneracn wee am, anees arero arranceroouwsa oree amarceneesr aorre o.ree.,noewaorw,ao earce surace, crossne w one or moreners ceav r or exraceuar reease ae ae one or morenraceuar more nucec acansne.s,.,ooormae a surace craonsssn aree c movcaeene aac,eo woocrm on ae or n some emomens,eroocrous oe am es are crossn .razoe,oeer, or aen suerou w one or more c erou a ace.., anes sazeearc eavaeners.e ouer nucec acae seecvenno a c en waer anuer an canemseves unconazee.., va enzmaec suraaocne a wne an,rooenan o nru ccaerco acraea,n orsee.u.r,aer nucec acsor ce secc ren maeraaon-s noom aerc am uaees..s useeren,eerm “non-omerc” means a escosureaveoromeorc.e.,r aou mos aercruane ceomowosaer oe reaerac uennse.ror,e am es o rous are aearce surace.n ceran emomens an roc am e as oerwseescreerenncue- a,eroocrous oe rou, eac oona susue wa 622 ,6-22 aen, or6-22 an - a.n oer emomenso,1-4 a,2-4 aen,1-6 aox, or – unsu1su22e- a,- aen,, oeroocrous oe am encue roocro6u2s2 622 r6-22 anrou.n oer emomens,e 20 a or unsus ouee am6-18 aen ocru unesu unssuuseue6-165- a22 a or u onrsu unssuuseue6-162- a or unsusue6-10 a or unsuussuuee10--22 a a o orr u unnssuussuue- e10--- a or unsusue- a or uns -20 a or unsusue10-18 a or unsusue10-15 a or uns 1222 a.n oer emome1n2s,20eroocrous1218 usue12-15 susue a.n oer emomens,ero ooe acmrouencue oona unsusue10 a.n ceran emomens,erocrs oe am encue oerwseescr10eerenncue an ammonum ous oe am e as arazoe,eo aemer, or aese on seusceosruorue as w oe orr nwou es oe. r meoscrereeren are crossnerou am es oescosure as oerwse ners.n ceran emomens, rou.n ceran em escreeren are crossnerou arazoe cr omens, am es oescosure as oerwseescreeren are oosesrwnseeescrorue aereone areer crroosus.nen cerraoun em ano amenens s, am es oescosure as emomens,erazoe,oeer, or aen sue uerou.n ceran o ane or aene moe one crossnnrou resusrom a reacon rou an an aze oro mo rocrou oe am ee.., one ammonum rom e onener.n one exame,erazoe crossner resus ammo an reacon o ane moe onerocrou oe am ee.., one cr umrou w an aze moe onener.n one exame,e aen sue eo.s.s, onnere re asmumsonroumm ar roeuaco wn o a aone m mooee o onneenerro.n once eroxuam oe am e crossner resusrom a reacon o aene moe on e,eoeer e.., one ammonumrou w ao moe oneenerrocrou oe am e ane or aene moe onerocrou oe a .n ceran emomens,e oerwseescreerens one orwo oro- m e oescosure as n, or one orwo oro--en -n orro--en one orwo oro-- n--amno orraam .n ceran emomens,erocrousrro-- reacon oro--n.n cneor.ann c eemraon emmenos,meen crso,ssen cronssnroun resruous resusrom a oaam rom a reacon escrene cn reo er. na anre emerovemernoms,e non-omerc am es oescosure as oerwse --- or
Figure imgf000036_0001
Figure imgf000036_0002
s eren are creorocrous oe am es oescosure as oerwseescre exraceuassne w one or morenersa are ceavae one or morenraceuar or mmore enzmr reease aens.or exame,n one emomen,eners ceavae one or e es, suc as,u nomeo,eases,roeases, eserases, or easases.n some ascoosumreen ass, oeerwnseeress ccreaevaeeren a anre es ceeraavsae.en cer oanen enmracoemueanr osr, exeranceerusa or reeease cee man.n ceran emomens,eners oescosure as oerwseescreeren are covarseeswo or morenraceuar or exraceuar reease aense.., werenener nw ceor or moreeren cemcarous eac ceavae aeren reease aen. ncue aeaeno emr exoammeens,eners oescosure as oerwseescreeren easwo amno acson.n, c cereaavna eme w aease orroease, wereneees a n ceran emomens, aeasree amnoo acmesnso,eees aeaswo amno acson. amno acson.n ceran emomens,eene.sne cwereaen emomens, aeasour acson oreweenree anwen-ve amno acson onrweow aenenwoen-ve amno amno acson.n some em ur anwen-ve a-,a-:s,, omen a-r, ore-s:s, .n,eeener comrses som :,e-eu,a-a, :, e emome:ns,, oereener comrses --- :.n some em:o,mens,eeeners:,e-eu,a-a :, a-r, o ,a-,a-s, : re-s.n some emomens,eeeners ee,ner :, or :.n some emomens,e or exame, as oerwseescreeren comrsesorurer comrseswosrous uoeer or a a eaecn en su oeeroeu crener, sucae suur oneeener maes ener.n s ossnnerocrou oe am e an :, ome emomens,eeener comrses :,s-:a-,- :, :, :, or :s.n s :, :, :, ome emome:ns,, oener comrses :.n some emome r :,ns,eners : :, :,s-a--s em:o, , :, :, or :.n some mens,eners:, n c:er,an emomens,en:ers, o orescosure as oe :. ncue one or more o eserrousor exame, ceavae w anrwseescreeren emomen,eners oescosure as oerwseescreer eesnernacseu.en one
Figure imgf000037_0001
rou. ncue one or more or eners oescosure as oerwseescreeren oroeser, ac azone, semcarazone,osemcarazone, cs-aconc ame, a areroea,ea, oroeerrous, or a comnaonereo, or oer ac-aerousncue ane caesrzaa wno eemn rouoe sm seesno ecss,om eee. romn aenrs e oser,esrcazoosnuere, s aesm ocaerrwazsoenee,screeren osemcarazone, cs-aconc ame, oroeser, acea,ea,oeer,sue, an a --- oer aeea,s woeurer anmneo aecseosn a oeraseweo amenos acson, or aeasree amno acson,on.n ceran emomen eweenwo anwen-ve amno acs exrace s,eners ceavaewo or morenraceuar or semcaruazaorn ree,easoes aemencasr anazonncue aeasworous seecerom an eser,razone, sue, a e, cs-aconc ame, oroeser, acea,ea,oeer, nn ce araen emeo asmeescnrs,eeerneenr. ncue asuero s oescosure as oerwseescreeren excuensu ceraenr eomu uo. or amneonse,reroun ceresa ovaeescosure as oerwseescreeren eners oescosure mae seec uner reucn conons. emomen, no morean ao ve ceave.or exame,n one morean aou u , no morean aou , no morean aou , non an exraceuar en,v noro mnmoreen.ann a anoouer e,m oro nom meonre, noane aou oeners ceave aou , noessan aou , noessan ssan aou , noessan aou oeners ceav aou, noessan aou, or noessan exn ceran emomenes on an exraceuar envronmen. enzme e meos oescosure,e reease mecansms an sosome anr ce ees nrs ,ae enn eomu soomo memr c aeenss. oe meos oescosure,e reease aens a ucec acanse ann, anaroern cav neuocaee aen. acanss corovvaene a aovaec,ee moueuanrccoena a ncan aocarcaenss oescosurencue nucec emomensurernc s oerwseescreeren, ann some acans.or examue,ena sormeen em nuocemcen acs,ea nnucseca ac arenaneo one or more nucec acans oescosure are caae o see s anaren nucec aamer. cvenno a ce surace anen scosuren ar seom caea emeo o smeeencs,vee nuncenc aco aroanens a onr a caarreon nrauce.ecn a scomeans oe --- e cmaaoem oen ses,ecev neucenc acno aanros aenn, wareerenn neucrec acans oescosure are conssn oumor-marers,ner oens seeceromerou c ns, ce surace receors,ransmemraneroens,on emanonems,e mnesm,rean neucraencs aocrroaenns, a ennzmaes, anoes, an cmercroens.n some caae o seecvenno ren nucec acans oescosure are rou cons a carorae, werene caroraes seecerome ns cenra on emcoroens, suar resues, ancocax. escos omens,e nucec acans anaren nucec acans o m ure as oerwseescreeren are caae o seecvenn , , ano comenao,n moe , zmes, rozmes, m, s , mcro, s , escosnur ceer asas on ee emreo rwo. semeesncrs,ee neurceenc ar aec cans anaren nucec acans o secceveomena saee.., aae o seecvenno a ceurn a oro a cen a saeavneveomena secc ce surace anensumor-secc a snececsease saee.., aumor ceaasumor-assocae anens or escosnur ceer asa on eemns. rwosemeesncrs,ee neurceenc ar aec caaane os anenear reeuna nounc.eocr a ecxamaen,sn o s reomuea emono canmeen ss,e n,ucecz acme, ranozsm ane, marcreon nu,c ore ac acans caae oene oonuceoesncun ansense oonuceoes n oereraeuc escosnur ceer asa on eemrwosemeesncrs,ee neurceenc ca ance naan. ves a onaren nucec acans o osoroaeacones, an’rmeroece ronurc moe,ncun eer or nanoarce oveeno, sr mso coasvuee ec acs, or cane an aamer, en re re cc o aor u nnnm razneo sorae,. en n cuecreacn a ecmoamnens,e muuncona oun aarcuareor,seeveae nucec ac oe coso. ouen a o reace cosoueos covaen aacmen an mae asssensoearce’sroocrou oe am es.oe am es.e.,s reaons --- erosrcovroues a oovee, ame nuceces a.cn cearnans o emoescosure are covaen aaceo moecues are aaceoerocr mens, uowo nucec ac n ceran emomens ous oe am ese.., uowoer ane. am es.n ceran, e omneo nucmeencs a,cwo m nouecceuce asc a macoeecuoes aererocrous oe rous oe am e aaceoerocoeroe nuccerc acs.ans,or examen ceran emomens, are covaen aace oeer or aen souus oeroeu ams ma reseusrrooum a r aeacooen oer o ar aneen or a sueerou.uc rocrou oe am ee.., one ammonumrou ne moe one s one nucec acan.n ceran emome an ao moee.., rocrous oe am essro ns,e ane or aene moe one en ore ane or --n orro--en, orro--n, orro-- amno or a aene moe onerocrou oe am esro--n-- erenarec meuamn szuo en. s,ceonean nanno, ea.r.,ces asescreeren caneroven a vare o em.or exame one am es an crossners useor man as aarce sze w,n cneraen ra enmeo om aeonus, nem muo auonuconma nnanoarce asescreeren nm, or nmo nm, or nmo nm, or n ameer, e.., nmoo nm, or nmom, or nmo nmm,o nm, or nmo nm, or nm nmo nm, or nmo nm, or nm or nmo nm, or nmo nm, or nm, or nmo nm, or nmo nm,o nm, or nmo nm, or nmo nmo nm, or nmo nm, or or n nmo nm, or nmo nm, or emomens,e muuncona na mo nmnameer.n ceran rane o a noarce asescreerenas aarce sze wne muuou nmo aou nmnameer, ann some emomens,e nmnnacmoenear, n aan aosa mreacseureas aarc.e szee wersonn oe or rannaer o aou nmo aou e maeras an meosescreeren sne ar can,n vew o nanoarce. ,rove aesrearce szeo a muuncona - -- screeanr ccerea snze em ano armee mnosn,oes muuncona nanoarces asescre erenave a n some emomens,e zeearseor s.ue.r, unorm. m.or exame,e zeaor ace cares aou - mvom o aou m, m, m, m, m su,race cares aou - m, - m, - m, - m, eraeu m, m, m, or aou m. w n,eescoscu areenrosvesa one or moreeraeuc aenss encase , or encose nosome comarmen o e muuncona nanoarce. ru, suc a cs,eruan no emmoemeno,s a,n aen-ecraanceeur ac aen comrses a rooc sma moecue aen, an ancoauan, an anaesc aen, an aneesn,e anc a aneno, anc,o ann a cnanvra, an anarascon cannen or, an an-nammaor, a neoenaor, an exame,n cera meaoru, an an comnaonereo.or oxorucn,aunnor eumcon,m venncrs,snee,earcaeauxec, a neeoncars seecerom camoecn, caroan, oxa an, saraan,coan,uroecanz, annonsaamnc, cna,coceeaamxec,n,a cmsoan, erucn, meorexae, vnasn, vncrsn,ooec xen, an comnaoner an,rensone,rensoone, a- , or manenre sormne emeoo. smens,,ecreoraeucm aens, a nnucseensce ac comrsn one or more o rozmes, aamers, m , crcuarze m , onuceoes s, sneue s ce-wcn onuocreoaresn s,s or a ,asms, ereo. n comnaon eanons cecr aanen em maome,ens,e muuncona nanoarcencues aanosc aen. raosooe,on,ocoeroo,r c exoaemsereo, a, au soerroo,o aren, e ae rcaroonaeee nuceoe, a or an comn nsea an a mea ceaor, aarmoanereo. acceae carureruc sne ocuoc ona rma a noarnmou armaaro accn ees uscasc coomseosereonn, m acacoren como anneo wer asecarm oaceeuca --- onvenuon c.arrercsanc,ueessooooccaa sa acncee,aose caarreerus aerer, nroesennuorm.xames ma, waer,uere waer,. sane,. cne,cormauere sane- sa e.., aumn,oroen,oun, ec., a roensorove enance carrers mae cosen, e..,ase o nee.ncesooca acceaeocaon oearessue, ne roue o amnsraon asescreeow,e ee,ere are a eruenevere,eme course oever oeru, an emo we vare o suaeormuaons oarmaceuca comosonsor e.,me.ns oeresenscosureee, e..,emnon'sarmaceucacences, an meansnoewnarnmaece aur,canc cuomnos oraon asn oaresnnevreanon cane amnsereo aaen arenera roues areesrae snc roues.n ceran emomensne amenar cana. ee avo conac weesve enzmesa areoun amnsere n ccorno suc emomens,nvenve comosons maenecon, reca,e vcaonnae.,.,oncraamusacsuar,nowraveerns,o curse,a smus,cu oaneous,nraeroneanaaonas nmens, orros, or ormuae ancceca sreasre.araons,or exame, serenecae aueous susensons mae s suusseennsonn an aeonr nos.n noxeo c see arreeno nnw eren ac aar u acesn cere s aaureaaone ue mnase or ars ssoon vee or n a w , se oernen aeecnas aen souon or ,-uaneo.mone acceae veces an s r exame, as a souonn waer,ner's souon,..., ansoo ovensa mae emoencue ormuaons cane serze,or nc soum core souon.enecaencororaen serz exame, raonrou aacera-reanner, or orsers n aensneorm o sere so comosons wc canessove wn seree a wareecr or oer serenecae meumroro use. cosen envuaeae exacosae o nanocasues oescosures amnsraon are aascann vew oeaenoereae.ca,osae an enrea useorove an eecve amoun oe nanocasuesoeaen nanocasuees. mas w vare aeenrecnae on sucosaec oor osr asnare sesrenoso arc,a een eeocnv,e aemoun o --- exearmaeue,ces eoecevee avmeoreun, oe naarnoecasssuuees, coen raounen o a aamrncseraon, anee.or cancereraeuc, as we as a seconor mor moeras aso an an- amouna resusn a reuconnumoeeraeucne core reon, m ee me. onaacors wc mae cr sze aesre amoun over aesreero o we anener oeaenenonserencuee sever oesease sae ae, comnaons reacon sen reaee,me anreuenc o amnsraonru e nanocasuses ovese anscoosuerreanceresonseoera. amnsraon an unorm oos maeormuaen unosaeormor ease oo ascasc ae.e exresson "unosaeorm" as useeren reers unersoo, ree un o nanocasue aroraeoreaenoereae. we eeceowever,aeoaa usae oe comosons oeresenscosure w nanocasue oe aenn scan wne scoe o soun mecaumen.or ann r nanocasue comoson,eeraeuca eecveose cane esmae anmaa m eoeersn a ceso u csueuroe a acssaevse o arne asnramae c moonceens,ra usuoan ran mece, ras,os, ors.e ucnormaon canene useoeermne use an roue o amnsraon. umans.eraeuc ecac anoxc uoses an rouesor amnsraonn armaceucaroceuresn o nanocasues caneeermne sanar eraeuca eec ce cuures or exermena anmas, e..,50eoses ouaon.eosev reano oox ocoeoeruaon an50eoseseao oe exresse ase rao, .armaeuc eecsseeraeucnex, an canences mae useun s50 50 aceuca comosons wc ex areeraeuc anma sues ca ome emomens.eaa oanerom ce cuure assas an n somne eem usoemnenorsmuan a rane oosaeoruman use. oe varous emo , a comosonsrovencun aura o nanocasues carrer, aarmaceumcaens acecsecraee eexrcen,en an, o orne oarrm maocreeu oca aa arcmceaceauecau acceae comosons canurer comrse an oer comonens as aro en.uc nanocasue raeor annene use oe nanocasuen c s so aom smeos sc emon oseo.mereenns, an a co am soousoonn su suaaeeoorrrereezeznn, e.s.r ao svucreos,en scouuonns - -- aeearocees nraonmoc aasrueea sunse unsoonn.reezen su.croosre ma ac, e.., as a croroecanoreven ormuaon comrsn aura exame,rove erens a nanocasue nanocas o scose nanocasues, sucrose an waer werene -uessucrosewaers aou- - - www or aou- - ncun an amnew ansoww seec o. rn orea oemen a susceocsurne, n meeeoesre oo arena enec aves aeamsoe orsorer arerove, nanoarce o escosure, weren one or mor un o e muunconanraceuar or exraceuar re enerss ceavae one or more aen oranos ease aenresenne suec,us reeasn eeraeuc acera, vorra e,x oamc a rarea,en sn. sco,mea enm, asommae,nns,amesease orsorers cancer,necone.., zemer’ssease,arnson’ssease, ec..mna cerona,n n eemurooomceans,seaese osreasseo orrersoer..,s asma,nammaone.., asma-nucenammaon or cronc er sease -nucenammaon, ornecone..,o osrucveumonar n some emomens, a meoorereamewer resraornecons. cancer, an e esrove.n som n o cancere..,rosae,reas amnsern aeraeuc e emomens,ereamen o cancerncues comosoner a eecve amoun o e nanoarcesescre eren, or amnseren seuoc, aom ao suunse acnaovrn suc a canmceer a.ssa nneoceasrsacres, or comosonsereo, canencunrean, aevan, ameoran, re o aceveeesre resu as orrow o,n n e mraon o, reuevn,ean onse o,n n roresson or more smoms ore cn sever o, anor reucnncence o onencue amnn sosmeren ema aour ermes aeen o us c,a c an a mceer. eoecovre aremvoeunnn cancersrove.uc meoso aea nvua.e., a suec woo o a nanoarce or comosonereo as noeenanose w can es no sa an smoms o cancer anor wo annvenvearee cer.or exame,ea nvuas ma e "mmunze" w cancer a rsnvuaarscee..,roareonsev weoomaevne a oa cmancer asnoror o on csaenc oer smaenomss ca orrn --- onoe omro mroresmen aessocc mauea wons asesvoecoaeme wn o ceveomen o canceraensavn aenec weveomen o cance anceraensnece a vrus assocae cancer ec raens was anoreses assocae weveomen o smoms. o c caanereaeroro or susana conemoraneous we onse o as we as sec snace cer em. xaemnsesereenre recnrnces, asecs, an emomens oescosure, euvaen o, areneneo encomasso srucura anuncona nown eusvaeerenos. as weo ansa eu,vasennseneveeoae sunce euuvuaree,n.es.,n acnueo curren aerorme sameuncon, rearess o sruc eemenseveoe oeeacnarsouesre on.ero crom exaomnene,s a maoenanc muur aeee. ra ans, cae uonorrovnor asecs omsson o maeras mae useor comnaons o maeras anor eeacnseren.eua oveor ae emomensa are wne scoe o euero c o anarcuar eemenorracce oeeacnserenso smarnermeseeearrse,c anve su oc amesaneorn,s m aarneuacurer, seer, user, ssem oeraor or oer enerese oeerceve accornoe sanars o consrue asearo.own examesurerusraeeresennvenonu sou noe xnam ane wamns scoe. acevesonneermos soawnza exames, a muaer, muuncona nanocasuesescrea noweeere are currenon no an o eenrz mme secevce rreeoarsme oua caornso maav.eoeeneensvenors’ ever m n an enzme secc manner.n aon, we sown c neo muaereo_- nanocasue oescosure c e cuure suesae exressono a ce va aamerunconaza an eecvearee ms secc ce receor,ns on wena ceasreaer exresson o a reure anener case annexn.eo_- was eecve sowno enca om ever comonenunosesn.e caoncosoma core reeasseuaroese,e an m a coanrao, en anosomea escaaee craraoves aueeo sea-me, am resaozonseve --- wean ass an r eenezasmese w-aeeneeraraaeso.n meecnaans omuera enaaerroves maxma sa as m ans suseuen exressonn es enzme secc reease oe enaess ouer oonuceo ces.aserorammae naure oe cane useoare cancer cees oanversexoreessnae un wue c aea rmeceers,o arnsn vs sroo awnnerenaesen vvo, a moreomoeneous exresson o m s o a, wen e conroe reease o m,aren c serve,eeernoeumor. muaereo_- nanocasue oeasaes, anroone see oe oorunesoreev cosure, canrovemoranransaona :ern oesersa aenucaara recevan nucec ac caro. an m ormuaonsoae are neor szaaoen a oe mmer o_ -s ueoensa oeormuaonse an m su aures warmeran - °,narcuar,averoveoe uns sce o nucease.osomes, conons, areoxco ces.ereaoree, wern ce cuaaorn encv sruornamceen csa arne, usn oeern ce mran mocaon w an oerans an sncan eors are unerwa ase rouencororaon oeren earousa cana oareosomes ssues an orans.owever, even w s socazaono secc cocesorasnoc urace mocaonsor sazaon anmrove suerromme e azaono secc ses,e maor o ormuaons s sowsowee msnn ao ,rs eoo samca e nesscea os ece xr. reess merenerra, a ca rne auressesemaons.. a caoncosomeormuaon, co orescenroen,soae wn caramocoeser nsrucerom--','-meamnoeane- osoeanoaorocore-oesero an,-oeo-sn-cero-- ormecne ares va sme eecrosac an roocneracons. coue enca seosome sa ssue, weer a caonc ormuaonnves se wn a mea orancramewor , raeranen ae, was uae. wasnvesaeoeermne weer cou sazeeosomeanmsazooeen coauraovoenenee scaa eec os osn m enosom caar eosca seeco ann, wre,e wrese-rme cou --- nmanoacra cosuneroaeeraaon anarenroeres wen encasuae wn a nucec ac oows: momen. coonssomruecso wner oeo_-s,osomes werersreare as moar raon coroormoowe snresnze uner anacnner amos anere-oeseron a: e overn uner vacuum, .e mxure wasen mn oowe resersonn m- waer anurerou a eerm exnreuer o an amraorae szeer memrane.uccessu sness oosomes was .. oonuceoen s.eueenrceesesn uave nanoscae caracerzaon are sownn. – aeras aneos secon,eow.seo a wsasesm useeoeeaerrmcense arerovenae arces werene sarn ro e averae sze o a corroor namc sze oeosomess.. nm, as o.a me.e snazme acn sur saccae ceranre a.e caoncosomes sowe a zeaoena eo er successu sness o eac aonaaer o armeua:ounmsm saurmm oarzeneae. am aeonr ove eac sz veaer c anearc me oua zeeons aon_ zea-oensa measuremens aer eamoena osom
Figure imgf000047_0001
es surace care was neee, ase caonc comonens o,e e wouencomae we caonc c - snessrooco se are oeosomes as sneszenerevous use.n o reraecrno m aooaee care w oeosome exeror, suracan conuaes were conuae mocaon oeosome a wna as suncecesrsesuen an suracan .. suracan ne surace carerom caonc. mo anonc-c.e mve an re.sue.n w casa onsnervee --- suaracean sz ceo annu saaeso oosomoseosm.enss ose,enen su ornacaen co cnocnenraon rao oe were vareromo reaveo axe conce uae concenraons anass waserormeoeermneees sunraon oosome mm. concenraon rao anouna a surracan conuaeoosome mm oosoma souon,e sae an s acan conuae an aroxmae enouoe uze as a cor ze oeosome was mos unorm an sma er successu senoresse oescre s mu-aere aroac .. sze excuson cromaor uracan conuae moeosomes, souon.e s a urcaon waserormeo remove a excesssn . nm, anzee oeosomes aer mocaonncreaserom..o. encas surace zeaoenaecame more neave-. m.ex,e rowu oaoen sne-m wasazoereorameer uosrmne a a mroounee- s sunraecsasn-rmooocoeo coonsroome amoun o znc anmazoe ae, aneme aoweor mneraz es.e mananarce szene nanomeer sze rane aon was vareo mue wasesneonze waerrou cen.ese m o_s wereure anass,earce was sownoaruaon aneen oearces.sn ase ronamc szeeermnaonve a sze o.. nma was vere - oenaecameess neave- . was oserveae m o_s surace suracan conuaeans.neres. mueoe aer ane masn oe o_ arces cane vsuazen, unee orancosoma core,e m mearamew n . wou uran aceae sannueoe nanocasueaosr.,.ensa saer,eo_, was encasuae wn a nucec ac m o_.o aceevens weeman o12 suracan moecues one o_ n orerou enecassu,aeme o12 suracans aeo mm o maersorme,s crossnenac m o_ corearce.ncee mceencororaenoe e usn aazo eser crossner wcs covaen u cemsrarce surace usn coer caaze aze ane ccoaon m o_-.e r.esu os ses ao_ surace crossne mceearce resennvenors’revous seuvreaocee cro nsuscence ac mc neaneocasuese sruecsurna. co uren ouee --- asremeca oureeeraaoens anns reeeas cero oss nnucnec s ace w carco sserveso sazee mceerevenn ever.e crossners a versae com uraceansurn ceuar arce’seraaon eren enzmaconaen oe esn ass mouare cemca naure oe crossne res can asoeroramme mone conann cr r.or mos oe worescreeren, an esereeser crossner waoss ussen.er aos w weer aes u as mearnx or meeraooersoeena esnezme sec rceso onsveeeee roease seccee crossners werencororaeoave sn. eresence o enzmesa are eevaeurn sec e arceseraen a secc enzmee..nc csease saesueo overexresson o cros rease exressonurnnammaon an cancer.e eser o essenraseer wreasse sne wccanee esnonseomoes en oa mnoces e cneoss.omeaeraaon, asere are evesncororae ae surace oe mcee re,e eser crossner was exe m o_-s were enucraesu vaae szeo_s usnu cemsr. seaex coumn.earcesa a sze o.. e nxmcu anson w a -- . m,ncane successuormaon oe c aosve suraceoena o arce surace.e sneea se aonc suracanaer asseme ae serseouaon oea enno , an zeaoena reans sues a mono anreasersereeaaree mrces a ev or seueoe n_r se ceo-. convesr weer a menunroe.reun. nconaze aoae s.e ouer anes sazeearcen waer anuae mr an canoe_- unconaze waren oonuceoesor ce secc uae va en urer oae sncororae va aone reacon usn an azomacaon.e arcesormem o_-s areenurerom excessreeona uor. excuson cromaora usn -- seaexracons an wer sn sze er unconazaon,e zeaoena oearces caneo ne coece. conrmne aon oe aer ae surace oearce.e anaavess- m na sze o.. nm.ese m o_- racons wereen usenocra ae a aunraesrs ex oeremseenasrsceeesn.ca,e a s.,n ancana ameou.n o-,, co,une an wresen eenmene mau- ---aeere anar acnes. ase wen a ans, aore aerouseooes ann sa m an a-nmem ouearzoaee oe ower-ran orerracoonnvesae anae resssu wnas cerrsoarmen, an oe wasaer wnr.earce, oservencae a crsaneaernas smar ounaeeas .eereenecaroeres o a -eoaer w-,ou s eren . .e.aeeraaon caaes an s ere evennearceesnnvesae. azaon oe m anosome wasen o assesse s:anzzmne arnoerese osoensveo_eease o m ranscrase assa waseve -s, ann vro reverse ormuaonn resonseo secc enzomaeco an assesseee anmeonun s om mu.e r aesesaasero.menucuer,erea.n, eseera mse enzmoae,eo eos_erase a-n acs wc co ener acc cononsaceae reease m was evauae re ons, or as a conro.oa vsuaze va en verseranscronoowe amcaon an sruommmarezeon_.a -,u wrn se wreo eneacr ee ra aem rsue cess se aro e ewec reaaro e weoress eseser aamseoun o. mesuss ar reeease uer a .. cearan a asesonseeaeor oowe aceae conrms m reeaseses weno conon rmer an- esn areresen, raeran w s, acc an enzmaceraaon roms m reease asesnarenaeca we oan ees meurase- oare orne aacreaceuer searae.esusnroucen aooca ssem uns enoco e woue sae wen exoseo enosoma es sewere accaon occurs an o_- erases.oessoess, anea ces werereae w o- mea w ans c wonaonun merr exressn m n an n- an corounere-reamen.esusroms assasee. o . ncaeansncan amouns o m were reeaserome o__--ss cnomoaere meoa aonsomes.er amouonna o m,. werneceaevsereao no cnsoesocc --- r ceomeaseex o moocas e mnvnrmonam, suesn aecrease cance oremaure m reeasen m o_ -:oenes o.e aernanroenxressonrom s n orero assessse ae sa ananrove e aer, a cereuam wenas. conouroceunen wsncow cnooro eune was amnsereo cesroro nanoarce orevoewnen ace mcaon oo_e eno-someec.ve s so.reve w snnnccae ane msauara ow ceeon r ms ore ean ereo r wsome exre csoroune wenreae w ourarces wou coroune.s ssonan enosoma accaons resu suorse concusona necessar resonseoreraaon oe aer – a seas aso conrmorsee r aeeaese s oae m oea enaoees marcererseroen exresson.s exermen reease o m. sn an rue ou nonsecc a :o_ -scevenance m xresson an escosureo, e av saeuraese oe eo_cenc- oe m ceu sauerees w neurcee cacrr ace oue avsesres nanocasues o reuorescence oserve as a res sne amoun o merr m wn ces.u oe exresson oe m roen encoe uorescen as aroeo assesss aen suncceasseus u,aoe_ oe-arcse w wereours moae. we rueoe-roescenoa aeno_ a-es w-ere mnceuaa.e we resus o cess exorermrenno serum summarzen..e aearance o eves oreen are rououe coasm oe cess ev emsson ouorescen ever veces caae o remann saeen e evveennnneereresseennccee o o meeraa,n suesne serum. enzmesn ese suesee e exceennc o o m m encoevner m vaeo_-s wasen assesse.or ever was assesse as auncon ome onceurraorene wrreoere an as meae crers m.e ce cse --- wnererem,a me usnoa coenocao_ mcros-co ws, ao enxc waon o aer con nmro an sam emesssomn anar nm. ooso semruesm o arn m serum, wreeere m seoreao ar ° an overrnoo awneen w aaseno oeea c oers an cmeasen resreare waer souon o merr m oe same concenraon n.noearces was use as a neave conro merr cao a wasoaeoae ae same m concenraon were use as aoncosomesm oreamens are summarzen.,., an sve conro.esus oese emsson canrsneuannoe weerae ane reave ceevse.s o m. .er- rrea e. mxernesss woenren concauecee re serum ae an serumree mea.ames werencuaeornca no conoca mcrosco.ncan exresson o merr wa emes anmae ea ces an aerr oncuaonor ces s even aerr oncuaonor asence o serum was aroro n serum. merr exressonneonermeon cenes.o sncan enancemenn re emsson aer omeo sncaear anror eac resecve cenes a sucen amoun oservaorn evraomuans e mxerm eexnr weasssonan m suse euxenre esxsoenr wmaesn ss.nnoenesrmoran wc suessaeres aerssen an connua reea e even aerr, e m over se o m anor sazaon o e r seeavevraours wne ce. m ever caeo enccenocsom oe ms a ever eo_-s comareo cear sowea merr exresnare m was assesse.e resus. m wasevere vaeo_son-nos c oenes was sncan er wene eosomes or asare m,rre escosure, raeran wenevere was noeae exres secve oe mea cononsserum ae or no. a son o merr was comaraeo, ann mannsances morean, s o mae meueerro me soraoea coenoosnosm aen as s saeen o eea mceanormuuoraesocnen –ce sanens anass. overnn areezer, wc,oreosome maave mae mes weree maaveen aece sa n serum.ne cas earcesess unorm an nuceases aonseoorransecon w e oe m, susce o serum orsme exresson. oue a o aransecon aen areee reason --- me course souur wears e cvaarrueae oue a rseseassvne s raeave o mee mrr exr wessonneeveo_-s, a ormuaon wase eerne rereraor ° or a roomemer s aere me an comaree resusoe merr exre aureoronuraons o osomes, orare m as a neave c ssoneves aceve eerarewoeren mea cononsw or wonro.woeren cenes anea an eenenc ano asses ou serum were useo assess an cene revousesc sroaer aca oe resuse same mea conons as mcroscoosrreea wmeerne u wzea.r mcese arrer ex,res,s aonnevesa wseroer asseesasean c,ono,ca sorae anmae usn conoca mcrosco.esus sownn. an orncaea m exressonserno cenes wenevere usneo_.-- ms o, weerescoesuore n aos e cxomresasroen oo me merrer wras e oxsreesrvson oserveorosomes, oror aso comareo m evere usnoec e aaermeons.esus were merr exresson was a amne as aosve conro,n wc sncan we as non-seccor cearene a,n as exs weeceocuomweevneer,. we areseooeunnca snoocancao rxecc as were carre ou resus soweaeo_- oescosure can s aes or uo aeas asas seen merr exresson, wen sore a aze memeraure conons. ° or room m :o_ -sceveoomemeraurereservaon o oen a - ° m,aoorrrreaswervaaco on o curreen sr mucureormuaonsse neeorrozen sorae, exresson sues were conuce oeormuaon ane m.ere, m s, m oaeosomes an a mer sora ae o ro ae samesm oaeo_-n anea ces.eoa m exressomemeraure,oowe ncuanem an as aererearao on was assesse conoca mcrosco,, exresson waseroreon_s were- maesee.,. -.. merr m.urrsn,e m oaeo_orm-uaon canora oeosome orree merrroenor uo wees, wc wa onnueo suore exresson o sore a roomemeraure, an m s noso arnou usnex unescaee re asu su cconsemerernaurees saomr suecs are --- sumeer scoar sesa.zsa reosnu on mcaes weer meueaaeerresoo_res-s oescosure oere enaes eecveever, wc resuene ervee sa oe m an roen. rssen ~-a sa an exresson o vauaon o:ea esero eoena ceuaroxc oeo_- nanocarrer was m-rooso mmovnnon vvo sues. m oaeo_- s, aon w ea cenesnes se arnum m ae- meoeac.amesunes, w are su vmarmanrzeosen were.nc,u wae w an ano cenese m o_-snuce no snca ere resusncae an assa.moran,o-- wasessoxca noxc asncae m-oecamnerearaons wenese a e ne m-osome an amerunc :eeceoreccuarvaeenn co oncenora_ons.-s va s oencouao oena uez nrcea aoro nen amzeeo esxoammes oaree m ever.sse case asaren nanocarrerso secc ceeess. woraarrecreso omaceromsocoecsuures suc as anoesor nucec acsan , wc caneunconaz e,e oueraers a e.., oonuceoes.o a e w one or morearen moees was mo ssess ce secc uae,e ouer mosaer oeo_- unco e w an aamer..ecca,e nanocarrers were receonraazes w oen an ov aer-aemxerressae cann caare ann excusveo annexn, a ce surace surace oe m ncer ces.e aamer wasunconazeoe s annexnos_ overex-resses vna enzmacaon. ceneor assess - ces,s cene was cosen as a moe scosure ne a oaree exresson oe m everaorm oencuae. woes-s, c mesn seruom_ree m-easorunucsona mzneu weso o mr wnmozue a aamer were seccnn annernazaona ma occur va scav n non- wase an aoweo sor ener receors.e ces wereenransaenoroen. m a-nooe_r.-ro asowun acno mnaze wnern aa sczreamneo see cueensceoee --- samoe s anze anare c memsr w aesre aes aocnvceu aaeam wer, as c we asosomesoae w m m .rncuaono serve asre esors sorme mnsoowereae w c comarsons.erre ces were wase anenncaeaoee acsam searnu anncomnaaeze un naneroc caornroercae mcrosco ..esus cearos ra receor-seccanne vere merr wn mn,eue conraso a xnnn annernazaon a.s resusn sar asence o re eme osseorn saemxrees csoen aso _n_s_ensea,aace an acvearen moe weree no ecenransaenese c c an wemaesncaea m wasnece noo- mce-aas suunce cuos an. naeozues w- aaummeorrs orre scrame seuence werenecon r,e mce were euanze, a own oner s.os- sucrose,rozen an secone an assaeorn meerurm eoxrsrexsseonnsorman, croreserven uorescenceexresse as ___nac an eeeos.e scramerearaon wemaes was seenno aamer an exresse. sncana m wasevereoumorsno cases an ca :roeaseesonsveever o m ossesseraeucncororaeon coomnoe_ rece-orarse snn wesze en wzme a sreoceacseever, merr m was surace, raerane eser cro secc crossner as evau ssnerescre aove.eee seuence coseno ea eex wracenuaer seoc_ree e- esn :saree sneszeo evauae m nzme .ere, an secco_- was comare w a scrame seueencveer con anrosn exres-so cnens. vveos.usesearces wereen sowr re emssonexresse as __ , summarzen., ormuaons werencu _nac an wemaes weno oese osome an m onae w ces overn r eo nouorescence waseecen :rneaevo comnraon ce asn.aren --- wnearoc_e a-sunconaze wo seccee crossnaes e ne annexn secc aamer ons surace were evauaeo6r m cveroumors aernecon.ce weremane sucuaneous wx - aens.nceeumors were sucenare,o_-soae w merr macnunc monaerrze m wem- aam aerso or w scram eeer a seaumeenrce o,r o srcr saamnee or seuoe-ncec-s weernenneecceon.noe mec mec we onros, v ere.en nm euoanze aamrero osrn.encon an meumooars wn vaa rocesseor conocauorescence mcroscoseeeo eremaes oumor ee anumor coreore s.. sows reresenave eae conoca mcroscomawnoe rserous ane conrorou. omoeneousrououeu veae merr exressonoe reasona unconazeo_ mor ee an core wen m severe usne aamer usne scrame se-s., row.owever,n conras, m evere reons o merr exrueenceo_-s., row sowe morescree wneu sson oenocaze aeumor ee as we as someme areas vas mor—oenane vcn onra-umora sacesa m reresen acuoenc mmcrsee.,., an ..eseerencesn exresson acevrnes a wnnexnne auammoerr a veresaurs socraemeeo seeue snceec acnn ocnaeernazaon mecansms e a o caneesruon o m exresson wneaume aamer maave eeroenous exressono a moreomoeneou orrom a more anernazaone on verooa_aere-nnc vaae annas e exnex - areas nmsoern nsae .ern. a meaersaan aeern vvo smrnuseraonroueorueseneceu omo ar.amse ar s anowscanreareorenmnn c oan annexn receorsa are orurououeumor,e aamer moeor cereraeucsseever overcomeeroem oeeroenouse muaons oescosure ma s aso noaeae amouver o oonuceoeeraeucsoumors. escreeren was one ore n o m aeverenearceormuaons eraureo useunres o nanor orams o mcoram.seer u mnoeuxseec weere saus,sans caom emnoanncnemeen o --- m conven soana m aearso aoworeever o orers o manueess m a aeeroass. ane meamnaa oeoonn acca ne-on csaoo rmaeo mss o woomerceess oreesaero reo sarn rnersoms. commerca avaae--','- sn-cero--os ocore-oesero an,-oeo- mm soc o coeoroanoormam anne on a: voroumme ra mom soc on c-corooeoserorom ese souons were ae uner aronureo rm. coroorm w c anceao sowowne evaoraon o m aeoomo onsaen sasrr,n. wasere comn ae vaec euvaoraon oe coroormeavn an rem mannsno coworeoorm s.onceaonnorm an waosu rer.susnaenuem , wnesccaor as o overn woa reerm aonve srereor assne susensonrou ererou a mn exruer ocaronae memraneer r aesecm,ve, nm- anmeens eanca.e as nmreoarree-sze souon wase a osoma m oa °eorurer use. m o-ness.-uracanonuae oeano-osome rroarooreecareammons,u amoroameeo20 su oraconaunceoe was reace w,,- re suracan, oae12 . vaone cc reacon.n roxeox1-2-merooeno o20 , an -rox--- - n aaone reacor we snrernor m-nso.oenanwaore w,ere mxeo asnom eexsoseooae w commerca avaae merr m omm so were oecnooesncuan a roomemeraureo c,rn was aeoe m oae nanooso r mn. o snesze s ermosaer a mesman ana voume anace on a aeaex- °- an coum rnm, a.ne resunrac mons w-ereo co-ecaer.ces wereure usn --- an m onaeess oo asomnecmazoe-aseearancramewor roun ou aroecvemaer o mo_ encasn an m caro, o an asereaa orrean mcrameowaoerosom aero suonuone waosome w om uer an o mema sue m u zoe souonrearen exara er was ae. e srrns souon, o znc nrae annsan aen so ouseornvareeuarren.merr o surrner, w mas aeoaerowse wc resuen cenrue a rmor mn ane suernaan wa o_ were meo,e were wasewce w sscare.oowne same re uner vacuum waer.na,e m oaeo_s were ness an o me a -oa °eoronosomerma sorae. m o_ - -uceccanocasues soncaonor m mnm- . sous.o on o m o_s wasreare vorexnoowe conane. m. mmo a on as sreuraarcean so.uoen so wuaso an weaso so anncaeenorue wc crossne mceesoae w1o2soma-s, or mn.o aon w o m soum asc aazo eser crossner. mmo even orer.e souo orae, an o mu- were aenemerau n wasen a soncaonaor we man surea ure usren o aeeaae rexma-ns co.- ceo eusmern. croossrneaere me -o_ c-s wereen s,.m. mm rossneo_- ssoven o o o -ee : was souo .see souon was aeoo_s an12 suracan een aeons m woe w o ms resuesor aenseneo uneerrmna ens,rraeaeoneors a meno.esnrcoourso oraeeno suer maccaene an suracee voa aroone cc reaconeweene aneea o_-12s useor sness wus oes resues.e - m na,earces we ere use wou anurerurcaon. surace o asreare m o_reunc-onasze a wn a an oonaonucoeone ce sceu reeancceo ane --- esw seeen, a aoae souo mno oec auser aenaree anse weearous oe suracan moecues.or oonuceoen re reace w ooaeo20 as,e m-eor_esence- os wereure us unnereaerraaonor mn. everseranscron x- - coumn. nzme an resonsve m eeasessa. m o_-sm o r-eease_ o ms wer wearse aanea wze.wo o sammeso o ver eserase orema an on rcne ae- mea e eac wasreae w - waer o merr m wasncuae o wr o ouemr.n es aeraarsae.e sa sme, o nm o or mns on aermosaer a ° an a rm.er mnso,uons o were m man sane aceaeuer was aeo one samereae worcnever eserase, e soum reae w ae- mea, an earae m ame manan an enosoma o err m sameo same conons.e oer., an reurneo aermosaeror an aona mn ae useuen,ouan eu aes connueo sae wou anurersuranceor mn. usn a sanar r moun o m reease, reverseranscron was conuce unreae m,ooowoeco w-uer ocnr a nvroen on ae sames aon w an sem.rmer seuences useor amcaono area onnecea-me eecon ae:onuceoeeuencerso avnennear seeu.ences ame euence
Figure imgf000059_0001
r --- a aan e mce o or r r -
Figure imgf000060_0001
ne --- nerr
Figure imgf000061_0001
e varousreamense sames were runn an oa er assa unerenaurn conons urea an rucramee eecrooress sane w o, exce a nm an emsson wans vosruaz men a a .ee was eaveexresson o merr m. nm. o examne m ever ecenc oo_-s, noeaaev cea coonnrcos, mosaomneasnmne sameo m were us ceon,c aesn wraeo anssrnee a m casee m srr.merr m as - ce were use as moe cenes.ames werereaeneres ,ea, an serum an werencuaeorror an - ces orrorence or asence o were cosenueoe oserve exressonroes sow ea.esem4es cesm wereae an cuure overnn serumn connan..roxmae× .ea was caneo serum ae or serumree mearnro mreoa a ° an e2rrorr oncuaon we sames, wase ces werem samereamen. cse croscoeon conoca.er sanne nuceusa we usonec aon exresson wasmae nmaser excaon an emsson a nm.ms, merr rocesse mae soware usne same correconsor am aes were exermens,e m wasevere a a concenraon o nma.es.n an vro --- oe eevramunaaeone e onana o m. escr ce sa oe m caro aore nanocasues soreeneeorennz,e s wnaeesr aze m ° or roomo_emera-ur, m o anure m were erorme asescre aove. e sames wereen.cueauear w exresson a snues weren serum ae an serumree mea, ea ces,o nuc anncuaeore corresonnmesoowe anear sanans a aner smaan usn conoca mcrosco.ereamens wereerorme,, n were snesze anes .-smerearaon. ean o enmexcan an snamne eex srnec ze ss anc a' s-craammer.e seuencecr oe same sze aamer an'- -' :or ' :ore s - rm crame seuence were ame cces o usn enenrs : an :.eoue srane roucs were eecro vorroesrsanscre. we′- aame arn useureeren cenaurnoacramee aaveeen sownomrov onanswo muaons rme es anos receorare.n aon, usne- e ar,am aer- annuceoe naeno sacers aeoe' en oe aamerorovesanceeween m-o_ea nmaneor -carraar esr, sounce rao. c me a waasmu- enrco sco_ rnaamze- e ws. aamer an aamer asroows: mxen a mcrocenrueue aneae ean were cona a °or mns.noer souon wasreare waenr.nereaseuer, o m , o ase an o m- secon souonoa onane arnsa so vouuon, was cooeo roomemeraure an mxe we r. wasueo anumre oerou.s s-ou coonum wans.reancnocnusa-e w aere °or --- coreeacree s aanm coenscenraeo exac concenraon aowemeraure see vacuum meo. onoc aaonm wan o oer conamroer samoese were useor ceman sues. - ces×4 cesm we m o_ -s. amer moe m o_-reaen an we conoca se overn. na concenraon s were aeo serumree o- mea a a remove an unoun oarce asn anncueae anoorer. mns.nen ° ce ans were was aemosoroeur o or m exresson.es wereenmae conoc 2 eo aow croscoe a mcrosco usn aoncse eona conoca. manuacurer’sonosxrcuco wnassss .ea ×s.e4 c uesnsm a co wnevreenona assaromeaer ormuaons an conr aen weae overn.e varousncuaonor a o sames werencuae w cesorroowe a weae rneoaeerrorenrenseenrce o.n rreeaaeen c.ess woreraence a nm was measuren was oane usn a souon o reaen.reesnc sam usee re as ccoanoronss w anere a usaeseonre aaaons. nmamae erx mermens woe_re conuce acrceoarmnenonumuorexnerse assnnce. envers oonneccueaenernsuonanma arove eoree sar oe su.ve mon - cesaare anseommee e were mxe w mareornn , maareenurcaseromnece sucuaneousnoe s o amc nue mce e, ces an enumors were- mm3ao urcaseromaresver. ormuaons or sane uree wees, mce werenece w ue .ce were euanze raer.es wumors werexe w sreormanor ~r a °,en wase w mnwas anaceno wucrosen mn an wa ssoerse. aum °ors. wereer r ~emoaves,es wumors were remove, wase casseesn cromarx an ra rozenn ansoenarnoemrecee,a cu.nearo aznenaoccesn weore --- e mn ccrroonssec suoneec,e covoer csonoecauorescecneron w anen secons oumor aorrs mcrosco,o a ess an aonnosccaesear. namcro scanaemrnc sazce aenrza a snuron zaec.eoena o snesze maeras were caracerze szes were aoena usnaverneaszerano-.xac cacua monea osur aever oane a szeer amnose ursnecnamae sorwaroe.oso omerae a eoowe suseuen anaze scannn eecron mcro a- s were anass waserorme on aermossco an-aracon . auma -raracomeeruae,r =ovaano oeran ae. e, a curren o m an a scan see o . , w a worn voae o -ase mass secrom ° mn- was useo oan aerns.- was c ercneave moeeecon oe suracan conuae ssovenonu mce a omnm anonurm araceaxeoor resuce ane amaacnsu os soa.on. sames were

Claims

--- as cames: o.r muuncona nanoarce comrsn: a nucec ac nanocasue , a mea aarnmcaceruamceawor acce, aosome, an one or moreeraeuc aens, or a wereneerae sa,rae or sovaeereo, wereneoaseuc aens encaseneosome werene-omes esn ecnacsaesennee o. mae a- comex an n.ucec ace muanusn ccovoanaen nan aoaacrcee oo a caarmc,e w coerene comrses one or more werenroocrous oe am esm arer asrnran noen-oowamrerce amarceneesr,or, we a crn reonssnreorocurou asn oer we am es are aearce surace an are nraceuar or exraceuar reeas oen ae oern ms.oreners ceavae one or more ac.ane mauamuenrc aonaace nanooare ecxee oro cra ome or,.urer comrsn aaren nucec .e ae muuncona nanoarce o eer o cams or, werenerocrous o moremners cesea avre crossnerou arazoe,oeer, or aen suerou w one or reacon o anae one or morenraceuar or exraceuar reease aens resusrom a ammonumroue o arn a aenn aez moee or ono meoero onecnrouer. oe am ee.., one .roe mucruonuc oonae na anmoarce o cam, werene ane or aene moe one ro--n, or one orwo oro-e-esn one. orwo oro--n orro--en one orwo os.rero mu--unn-c-onaam nnanoo orarrace oam cnaom., werenerocrou oe am e --- co.mres mesu anu ennzcmoneas nuacno asarece oase a,nr ooneea ose, c esaemrasse-, o,r w eaesreasne.e reease aen .eee muoruncona nanoarce o an one o cams-, werenener comrses a amno acs eoxame, ceavae w aease orroease, wereneees aeaswo eesewnee,n or aeasree amno acson, or aeasour amno acson ore s resues or azweo m aoneewse onn e-acve e anm ono aceseone, . an oona,ener comrses .e muuncona nanoarce o: c,am, wereneeener c:om,rses ::,,e-eu,a-a,a-,:a,-s,a-r, ore-:s., .e muuncona nanoarce o cam, wereneeener comrses :. :, :, or on.e or me morue ou ensceronraou nasnooarr excame oe a,n ce oanvea oe c wams an- ese,r waseer.enener comrses .e muuncona nanoarce o cam, werenener comrses
Figure imgf000066_0001
rou. on.e or more orazone n,a sneomacrarcaez oon aen, oonsee om cams-, werenener comrses acea,ea, oroeerrous, or carazone, cs-aconc ame, oroeser, rozaenesosome. a comnaonereo, or oer ac-aerousa are s.uee mruouu.ncona nanoarce o an one o cams-, werenener comrses a
- -- s.uee mruouu onrc anoonae nranroouar cceaev oa aen un oneer o re cuacmns co-n, wonesr.enener excues . oneen mruaceuuncaro onra ex nraancoeauracr ree oea asne a oenne. o cams-, wereneners ceavae .woe or m muoreunncraocneau naarno ora erxcraece ou anar r oeneea ose c aaemnsse-..,, w weerreenneners ceavaewo or more eren cemcarous eac ceavae a eren reeasee aenner. comrses se.ecee mroum aunn ecseorn,a narnaozoanre,c seem ccam , werenener comrses aeasworous oroeser, acea,ea, oeer,suare,a aznone a,eoseem, wcarazone, cs-aconc ame, amnoe acsseownee,n orw ao aenaswere erenee es aeaswo e ne a-mvneo am acnos aocns, oorn a.easour amno acson ore are. covea menu aunaccoenao naneoarrcoe oc anro ounes o o cams-, werene nucec ac ans aen su erou. e am esrou a oeer or a.acne mue nuucncecon aca nanoanarscoe oe carmo , wceroreun oee aomeer or ae ren su erou reacon o a ne or aene moe one.e., rso esusrom a ammonumrou an a o moe once nrouuce oc ace aman.ee.., one .roe mcuuncona nanoarce o cam , werene a ne or aene moe one en. rous o e am essro--n orro--en, orro--n, orro-- .roe mcurouunc oonae n amanoarcees or coa-m-n-, w-earmenno oer a anaem orn ao.ene moe one ca.aee o mu seecunvceonan nannoaor ac cee o su arnac oe a cnamens a-n,o wrerene nucec ac ans are --- ace muanuncaoanmae nra anoacareceo o aen ex one o cams-urer comrsn aaren nucec ans are caae o seecvenneroor a o cee surace, a wnereenn.earen nucec acs c.aaee m oueunnec reonuaa naonno, aanr wcee ore ann oen neu oce cca amcss- s, we,rene nucec acan mcro, or oereraeuc oonuceoe. zme, rozme,s c.aaee m ou seuenccvoena nannonaroc ae oro aenn o onre a o cams-, werene nucec acan e muuncona nanoarce o an one o carorae anor aca anre caaaameer cams-urer comrsn aaren nucec ans o a seaecceveoen exneroor a oroeen or a, c wareroenraee.aren nucec acar.ene m nuuceucn acconaa nnanosa cracaee o o c saemecv,e werennneo nucec acan anore seeceromerou conssn aroen, wereneroensransmemraneroens,on cann oes,u mmeomr-mraanreerrs,nerns, ce surace receors, cmercroens. ansorroen, enzmes, anoes, anar.ene m nuuceucn acconaa nnanosa cracaee o o c saemecv, werene nucec acan anore caroraes seecerom enno a carorae, werene cocax. erou conssn ocoroens, suar resues, anar.ene m nuuceucn acconaa nnanosa cracaee o o c saemec, werene nucec acan anore moe , zmes, rozmes, m,v sen, mncro,,, moe , ereo. s , an comnaonss c.aaee m ouuncona nanoarce o an one o cams-, werene nucec acan avneveo semecenvae snecnc coe a c seuracuer ann ae snesc orcoeveomena saee.., sae aumor ceaasumor-assocae anens orumor-se ac cec ann ae snesc, ancosreasee saee.., --- msu caaunce oon sae necanovearcne on ano a o cnee ou crams-, werenearen nucec acan avneveomena secc ce surace ann ae snesc orcoe av ceeomne an saec sacees.ea.s,e s saae aumor ceaasumor-assocae anens orumor-secc anens. ee.., .e muuncona nanoarce o an one o cams-, werene non-omerc amaoen.es areerve , werens .e muuncona
Figure imgf000069_0001
-, omerc am es areerve
Figure imgf000069_0002
or
Figure imgf000069_0003
, werensaoen. . o an one o cams-, werene non-omerc am e
Figure imgf000069_0004
, werensaoen. a.urae o meanaoens aonor am auraorou os, or o cams-, werene comrses ans are coor reaen-cmenasnosn,a wrearmeenwore m aeroaunonse ano osromane.c mo.re one2 m+,u2u+n,cao2n+,ae n3a+n,oea2r+,ce4+ o, cr4a+,m4+,, w2e+r,en+,e4 m+,ea3+,on2 c−o,m anrses3+ o.ne or
e3 uuncona nanoarce o cam, werene meaonsn2+,-2,a-- . e m + 2+, .+, ore e2+. muuncona nanoarce o cam, werene meaonsn2+. or. moe muuncona nanoarce o cam, werene orancan comrses one enzmre oazo-me,e-mema-z,o,e,-r-mazeoe,m-meazoe-,--meerazo-e, o-rm-mazoaez,omcaraoznoer,e. me.mea mzoue,u-nmceonam nanoazoare,ce-m oe ca-m-,m waezreone,me orazaonec, oranenz commarzsoese.- me.mea mzoue, ournc-monea nanmoaarzocee. o cam, werene orancan comrses- me.mea mzoue.uncona nanoarce o cam, werene orancans- or.ancean musu-nmceonam nanaozoaer.ce o cam, werene meaonsn2+, ane me.mea mzoueusn aceoona ncanmoaarzocaee or caammewor, w-eren-e.orme n2+ an- com.rsese o mneu or munocreo cnaao nnacnoarsc aen o a a onne o onre m oor cea nomn-c-ao,n wcereenereoss.ome co.eseroe- mauseunc.ona nanoarce o cam, werene one or more caoncss a com.rsese m-u-un'c,o'-nam neanoaamrcnoee oa cnaem-car,a wmoerecnoees cerooeseroro-casoere-o. --- no.n-caonec mueuenrconas c noamnorasresc ae o ana ones o neu craam osr zw-e,r wonerce anes onoeo orc maore. no.n-caonec mueuenrconas c noamnorasresce, o- anoe oone-s on- cacmesro---,o wseoreenanoea omnen oer more. u.racane monuuuanecona n,a anoar n-c oce ao oes a aenreo o r o cnoen o eu caa oem s,o a-, me.-urer comrsn a comnaonereo,ncororae ocoero conuae, or an . e muuncona nanoarce o cam, werene comrses a non-
Figure imgf000071_0001
. , omerc am e seece
Figure imgf000071_0002
Figure imgf000071_0003
, werensaoen an aoae oonuceoe. o.onuceoeseween aouo o nuc eeoer oes cname or, wereneoae en, aou an nuceoesnen, aou n, aou an nuceoesn an nuceoesnen, aou an nuc an nuceoesnen, aou nen, aou an nuceoesn eoesnen, aou an nuceoes an nuceoesnen. en, aou an nuceoesnen, or aou
--- o.onuceeo muesunocona nanoarce o an o cams-, wereneoae oonuceouesu renaccoen2. . e m 0 a w nanoea nrocn-eo o amner ocn aem o came v-a a, woerenen cec coeae reacon. msr rea.ce we mue nounn-cooname nracno amarce oe c vaam ao, wneer cenc ceemosare re oacoonnu.ceoes com.rsese o mneu or munocreo ona a naenroaaerucce nu oc aen one o cam-, wereneeraeuc aen romrou co c ac, arooc sma moecueru seece an ancoauanns,s ann an oa aen an-cancer aen, an anoc, an anvra, an anarasc aen, cannenor, an an-nscam amena,o arn, a an meseaeoc aruen, a,n an anon c comannenaooneneareoor,. anon ca. e muuncona nanoarce o cam, wereneeraeuc aen comrses csmaon,e ccn,oxorucn,aunorucn, vncrsne,acaxe, neocarznosan, caceamcn,amoxen,a eroruacnn,, o mxaeoaren, saraan,coan,uroecan, annamcn,oceaxe, a-, or an comnaonexreaoe,. vnasn, vncrsn,ooecan,rensone,rensoone, ac. comers mnu onuenc oro mnoare na onomaracen oer ceranm, wereneeraeuc aens a nucec nsenseonuceoes s, rozmes, aas ,cro m , arn s ,asms, sneu mers, m, crcuarze m,or or an comnaonereo. e ,ce-wcn onuceoes s, co. e muuncona nanoarce o an one o cams-, omnr,soecso aearon,o csoce aseenro,, w a sceros a, anuo eroecroorne, a raoaee w nuecreeonee, n aa rnaooarsocoee, comnaonereo. ensea an a mea ceaor, or an
- --ar.eeoe a m suecuncc ceona rec neanoor.arce o an one o cams-, werene nanoarces e. como csomono csomonrosrnre aaneas or onree ovenen m ausuenacse,o ansorer, or smomn a suec, - or a armaceuca acceae sa ereo an aeaa nanoarce o an one o cams vece anor excen. s one armaceuca acceae e.reo an m eeecov oe amreoaunn o asee mase orsorer, comrsn amnserno a suecn nee comoson o cam , wereneununcona nanoarce o an one o cams- ore exraceuar reease aenrese ers ceavae one or morenraceuar or anosc aen. nne suec,us reeasn eeraeuc aen or mo.reane a moeuo o, n coa mmore, wane areon no morean aou , no morean aou , no o eners ceaven an exraceuuar e,n nvor moonrmeena.n aou , or no morean aou no.es e meo o cam or , weren noessan aou , noessan aou , osaen anoeurs ce,av neoenss anannr aacoeuuar e,n nvoroensmsenan. aou , or noessan aouum.or cese. meo o cam , werene reease mecansmss an enzme exresse en.osomee a meen,o an oo arn cav oeo caaem asen.-, werene reease aens asosome aen,
PCT/US2024/034387 2023-06-16 2024-06-17 Nucleic acid nanocapsules for drug delivery Pending WO2024259439A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202363521369P 2023-06-16 2023-06-16
US63/521,369 2023-06-16

Publications (1)

Publication Number Publication Date
WO2024259439A1 true WO2024259439A1 (en) 2024-12-19

Family

ID=93852729

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2024/034387 Pending WO2024259439A1 (en) 2023-06-16 2024-06-17 Nucleic acid nanocapsules for drug delivery

Country Status (1)

Country Link
WO (1) WO2024259439A1 (en)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20160271268A1 (en) * 2013-11-08 2016-09-22 Dana-Farber Cancer Institute, Inc. Nucleic acid nanostructures for in vivo agent delivery
US20210038528A1 (en) * 2017-06-23 2021-02-11 University Of Connecticut Nucleic Acid Nanocapsules for Drug Delivery and Targeted Gene Knockdown
US20210236651A1 (en) * 2017-07-13 2021-08-05 Northwestern University General and Direct Method for Preparing Oligonucleotide-Functionalized Metal-Organic Framework Nanoparticles

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20160271268A1 (en) * 2013-11-08 2016-09-22 Dana-Farber Cancer Institute, Inc. Nucleic acid nanostructures for in vivo agent delivery
US20210038528A1 (en) * 2017-06-23 2021-02-11 University Of Connecticut Nucleic Acid Nanocapsules for Drug Delivery and Targeted Gene Knockdown
US20210236651A1 (en) * 2017-07-13 2021-08-05 Northwestern University General and Direct Method for Preparing Oligonucleotide-Functionalized Metal-Organic Framework Nanoparticles

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
GAVITT TYLER D., HARTMANN ALYSSA K., SAWANT SHRADDHA S., MARA ARLIND B., SZCZEPANEK STEVEN M., ROUGE JESSICA L.: "A GATA3 Targeting Nucleic Acid Nanocapsule for In Vivo Gene Regulation in Asthma", ACS NANO, vol. 15, no. 7, 27 July 2021 (2021-07-27), US , pages 11192 - 11201, XP093255227, ISSN: 1936-0851, DOI: 10.1021/acsnano.0c07781 *
HERBERT FABIAN C., ABEYRATHNA SAMEERA S., ABEYRATHNA NISANSALA S., WIJESUNDARA YALINI H., BROHLIN OLIVIA R., CARRARO FRANCESCO, AM: "Stabilization of supramolecular membrane protein–lipid bilayer assemblies through immobilization in a crystalline exoskeleton", NATURE COMMUNICATIONS, vol. 12, no. 1, UK, pages 1 - 13, XP093255225, ISSN: 2041-1723, DOI: 10.1038/s41467-021-22285-y *

Similar Documents

Publication Publication Date Title
Ganote et al. The nature of D-serine-induced nephrotoxicity
Rossato et al. Mitochondrial cumulative damage induced by mitoxantrone: late onset cardiac energetic impairment
WO2024259439A1 (en) Nucleic acid nanocapsules for drug delivery
GRAHAM JR et al. Cytochemical demonstration of peroxidase activity with 3-amino-9-ethylcarbazole
Tang et al. Fluorescence microscopic localization of actin in pollen tubes: comparison of actin antibody and phalloidin staining
Menaker et al. Standardization of 2, 6-dichlorophenolindophenol
Laforenza et al. Thiamine outflow from the enterocyte: a study using basolateral membrane vesicles from rat small intestine.
Hörstadius Vegetalization of sea urchin larvae by chloramphenicol
Dirksen et al. Microtubule sliding in cilia of the rabbit trachea and oviduct
Hinrichs Modification of development on the basis of differential susceptibility to radiation I. Fundulus heteroclitus and ultraviolet radiation
Goodrum et al. Lipid droplets in Schwann cells during tellurium neuropathy are derived from newly synthesized lipid
Nachman-Clewner et al. Injury-induced remodelling and regeneration of the ribbon presynaptic terminal in vitro
Buck et al. A new method for the standardization of the dye used for the determination of cevitamic acid (vitamin C)
McCORD et al. Experimental chromium dermatitis
Van Erp et al. Eye irritancy screening for classification of chemicals
Fiedler et al. Toward the reconstruction of the Guang gender system
Chung Production and perception of English vowel length depending on the following consonant voicing by Korean learners of English
de Souza et al. Effects of SQ109 on Trichomonas vaginalis
Noel et al. Morphogenesis during division and griseofulvin-induced changes of the microtubular cytoskeleton in the parasitic protist, Trichomonas vaginalis
Kristjánsdóttir et al. Anti-fascist discourses, practices and confrontations in 1930s Iceland
Lachenaud Liber 5 Caput 22
WO2025059533A1 (en) Crispr enzymes and systems
Ishii et al. Processing of emotional utterances: Is vocal tone really more significant than verbal content in Japanese?
WO2024206196A2 (en) Rag and reminder rings: surfacing memories for humans and llms
Jones Ethnohistory of Chiquitos: Una etnohistoria de Chiquitos: más allá del horizonte jesuítico. By Cecilia Martínez,. Cochabamba: Instituto de Misionología–ILAMIS and Itinerarios Editorial, 2018. $42.00 paper.

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 24824363

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE