Hauv tsab xov xwm dhau los, peb tau tham txogAbsorption thiab Distribution of oligonucleotides (ONs).Hauv tsab xov xwm no, peb yuav txuas ntxiv kev sib tham los ntawm kev tsom mus rau Metabolism thiab Excretion ntawm ONs.
Tsis zoo li cov tshuaj molecule me me, oligonucleotide (ONs) cov khoom muaj qhov sib txawv me ntsis hauv cov metabolism ntawm cov tsiaj sib txawv, nrog qhov tshwm sim tsis tshua muaj tshwm sim ntawm cov metabolites tshwj xeeb tsuas yog pom hauv tib neeg lossis ntawm qhov sib npaug ntau dua hauv tib neeg. ONs feem ntau metabolized los ntawm endo- thiab exonucleases, enzymes uas qhia tsis muaj hom, poj niam txiv neej, lossis hnub nyoog- cuam tshuam txog qhov sib txawv ntawm cov tsiaj. Txawm li cas los xij, qhov no tsis txo qis qhov tseem ceeb ntawm kev kawm cov metabolism hauv ONs cov khoom. Cov metabolites tseem tshuav tom qab qhov kev tshem tawm ntawm ob peb nucleotides tseem tuaj yeem muaj kev ua haujlwm, thiab tsuas yog kuaj pom cov qauv ONs yuav ua rau qis dua qhov kev ua haujlwm ntawm molecule. Tsis tas li ntawd, cov metabolites me me tuaj yeem sib xyaw nrog kev tawm ntawm - lub hom phiaj sib lawv liag, kom tsis txhob xav txog cov tshuaj lom. Yog li, kev kawm txog cov metabolism ntawm ONs yog qhov tseem ceeb.
1.In Vitro Metabolism Stability
Nyob rau hauv vitro metabolism feem ntau yog siv thaum lub sij hawm tshawb nrhiav tshuaj los ntsuam xyuas qhov ruaj khov ntawm cov neeg sib tw molecules. Thaum hom metabolites thoob plaws hom feem ntau zoo sib xws, cov metabolism hauv yuav txawv. Qhov zoo tshaj plaws hauv vitro kev ntsuam xyuas yog qhov tseem ceeb rau kev kwv yees qhov kev ruaj ntseg hauv vivo ntawm cov khoom. Feem ntau, plasma, lub hom phiaj cov ntaub so ntswg, lossis homogenates nyiam dua. Rau kev tswj hwm hauv zos, cov matrices hauv zos raug pom zoo.
2.In Vitro Metabolite Identification
Raws li tau hais ua ntej, kev txheeb xyuas cov metabolite hauv vitro yog qhov tseem ceeb rau cov molecules me me, tab sis rau ONs, qhov tseem ceeb ntawm txoj kev tshawb no raug txwv vim qhov zoo sib xws hauv cov metabolism ntawm cov tsiaj. Qee lub tuam txhab tsis xa cov ntaub ntawv ntawm kev txheeb xyuas cov metabolite nyob rau theem IND, uas tsis cuam tshuam rau qhov pib ntawm kev sim tshuaj. Lwm tus, txhawm rau txo qis kev pheej hmoo rau kev sau npe, tsis txhob ua hauv vitro kuaj thiab tsuas yog muab cov ntaub ntawv hauv vivo metabolism. Qee lub tuam txhab muab cov ntaub ntawv -raws li cov ntaub ntawv pov thawj, piav qhia meej tias qhov kev pheej hmoo ntawm kev tsis ua qhov kev tshawb fawb no tsis tseem ceeb.
3.In Vivo Metabolism
Xav txog cov txheej txheem metabolic uas xav tau ntawm ONs cov khoom, OSWG thiab ICH cov lus qhia qhia muab cov ntaub ntawv no ua ntej Phase III kev sim tshuaj.
Tshwj tsis yog qhov sib txawv ntawm poj niam txiv neej nyob rau hauv qhov raug ntawm cov tshuaj tiv thaiv yog pom nyob rau hauv lwm yamPK/TK kev kawm, nyob rau hauv vivo metabolism feem ntau yog kuaj nyob rau hauv ib tug poj niam txiv neej. Plasma los yog zis kuaj feem ntau yog sau rau cov metabolism. Plasma qhia tau zoo sib xws nrog cov ntaub so ntswg, thaum tso zis ntxiv rau lub raum metabolism. Cov ntaub so ntswg kuj yog ib qho kev xaiv, xav txog cov khoom nruab nrog cev uas ONs feem ntau xa los yog cov khoom nruab nrog cev qhia PD lub hom phiaj lossis muaj peev xwm ua tau ntev - lub sijhawm khaws cia. Txawm li cas los xij, kev tshuaj xyuas cov metabolites hauv cov ntaub so ntswg tsis yog kev cai tswj hwm thiab nyob ntawm lub tuam txhab cov hom phiaj kev tshawb fawb. LC-MS feem ntau yog siv rau hauv vivo metabolism hauv kev tshawb nrhiav, uas tuaj yeem nrog cov kev tshawb fawb toxicology.
4.Metabolite Kev Ntsuas Kev Nyab Xeeb
FDA cov lus qhia txog cov tshuaj metabolites xav tau kev txhawj xeeb yog tias cov metabolite raug nyob ruaj khov- xeev siab tshaj 10% ntawm cov qauv tshuaj. Qhov no siv rau ONs cov khoom thiab.
Rau cov khoom siv LNP tshiab, peptides, lossis cov khoom sib txuas tsis siv rau hauv cov tshuaj pom zoo, lawv yuav tsum tau soj ntsuam kev nyab xeeb.
5. Kev tso tawm
OSWG thiab ICH M3 pom zoo muab cov ntaub ntawv tso tawm ua ntej Phase III kev sim tshuaj. Txoj kev excretion rau ONs zoo sib xws. Feem ntau ONs feem ntau yog metabolized hauv cov ntaub so ntswg, tsuas yog ib feem me me tawm raws li cov qauv tshuaj.
Kev tshem tawm tuaj yeem kuaj xyuas hauv PK cov kev tshawb fawb cais lossis koom nrog kev tshawb fawb toxicology.Tib - noj tshuajnyob rau hauv ib hom yog feem ntau txaus. Cov nas tsuag feem ntau nyiam. Ob qho tshuaj tiv thaiv thiab cov tshuaj metabolites yuav tsum tau kuaj. Ib qho - koob tshuaj ntau npaum li cas ntawm cov tshuaj uas tau npaj tseg yog feem ntau txaus, tab sis yog tias kev txheeb xyuas cov ntaub so ntswg, yuav xav tau koob tshuaj ntau dua. Feem ntau, tsuas yog cov zis tso zis yuav tsum tau kuaj. Txawm li cas los xij, yog tias cov zis tsis yog thawj txoj hauv kev tawm mus, yuav tsum tau sau lwm cov qauv xws li cov kua tsib. Yog hais tias mob plab hnyuv los yog pulmonary toxicity tshwm sim, kev kuaj ntxiv ntawm cov quav thiab ua pa yog pom zoo.
Txawm hais tias radiolabeling tsis yog yuav tsum tau los ntawm cov koom haum tswj hwm, feem ntau cov lag luam ONs siv txoj kev no los ntsuas cov khoom sib npaug. Txawm li cas los xij, OSWG ntseeg hais tias ONs ' metabolism thiab excretion twb muaj cov ntaub ntawv tshawb fawb txaus, thiab qhov zoo sib xws ntawm cov khoom zoo sib xws tsis lees paub tus nqi ntxiv los ntawm radiolabeling. Tsis tas li ntawd, yog siv radiolabeling, nws tsis tuaj yeem ua nrog rau kev tshawb fawb toxicology. Txawm li cas los xij, rau cov xwm txheej uas lwm txoj hauv kev tsis muaj kev nkag siab zoo, lossis rau hom tshiab ntawm ONs cov khoom, lossis rau cov ntaub so ntswg nrog lub sijhawm khaws cia ntev uas cuam tshuam cov kev tshawb fawb soj ntsuam raws li kev tso tawm radiolabeled, radiolabeling hauv cov tsiaj muaj qee qhov tseem ceeb. Raws li tau hais ua ntej, yog siv radiolabeling, txoj hauj lwm daim ntawv lo yuav tsum zoo dua nyob rau hauv cov cheeb tsam ruaj khov rau nucleases.
Mis Mis Secretion nyob ntawm seb cov khoom raug kis thiab lub sijhawm, qhov ncauj bioavailability, thiab seb cov neeg lub hom phiaj puas suav nrog cov poj niam lactating. Raws li ICH S5(R3), txoj kev tshawb no feem ntau tsis tas yuav tsum tau ua. Cov txiaj ntsig tau los ntawm cov kev tshawb fawb txog kev tso kua mis niam uas tau ua los ntawm cov khoom lag luam ONs tau pom tias muaj cov ntsiab lus tsawg heev lossis tsis tuaj yeem kuaj pom.
Tsis tas li ntawd, zoo ib yam li cov kev cai rau kev nqus, kev faib tawm, thiab cov metabolism, yog tias cov conjugates lossis formulations muaj cov khoom tshiab sib nrug los ntawm ONs, lawv qhov kev kuaj pom yuav tsum tau ntxiv rau cov kev xeem excretion. Thaum kawg, rau cov khoom siv kho mob mRNA, vim lawv qhov tsis ruaj khov, tsis tas yuav muaj kev tshawb fawb tawm.
6.In Vitro Drug Interactions
DDI feem ntau cuam tshuam nrog metabolic enzymes thiab thauj. Cov tshuaj tuaj yeem ua "tus neeg raug tsim txom" lossis "tus neeg ua txhaum cai" cuam tshuam rau PK ntawm lwm yam tshuaj. Los ntawm tus neeg raug tsim txom txoj kev xav, ONs yuav tsum yog cov substrates rau Phase I lossis II metabolic enzymes lossis thauj. Txawm li cas los xij, ONs feem ntau cuam tshuam cov metabolism ntawm nucleases thiab tsis cuam tshuam rau CYPs metabolic enzymes lossis cov thauj khoom. Ntxiv mus, ONs feem ntau tsis cuam tshuam rau me me plasma protein khi. Yog li ntawd, ONs cov khoom feem ntau tsis tas yuav tsum muaj cov kev tshawb fawb cuam tshuam raws li cov substrates rau metabolic enzymes lossis cov thauj khoom. Txawm li cas los xij, rau conjugated ONs lossis LNP formulations nrog cov khoom tshiab, kev tshawb fawb enzyme substrate yuav raug txiav txim siab. Rau ONs feem ntau tawm ntawm lub raum txoj hauv kev, kev ntsuam xyuas seb lawv puas yog cov substrates rau lub raum thauj khoom tuaj yeem ntsuas. Rau GalNAc-conjugated ONs cov khoom, yuav tsum tau saib xyuas seb lawv puas yog substrates rau hepatic efflux transporters.
Los ntawm tus neeg ua txhaum txoj kev xav, ONs tiag tiag muaj peev xwm ua tsis tau tshwj xeeb khi rau qee qhov loj molecules, txawm hais tias muaj kev sib raug zoo tsis muaj zog, lawv tseem tuaj yeem tuav cov chaw nquag ntawm cov enzymes lossis cov thauj khoom, cuam tshuam rau PK ntawm lwm cov tshuaj. Tsis yog ONs 'cov qauv nkaus xwb tab sis kuj yog cov khoom sib txuas lossis cov qauv tsim ua rau muaj kev pheej hmoo. Yog tias cov khoom tsim los siv cov cuab yeej siv tshuab, DDI cov ntaub ntawv keeb kwm yav dhau tuaj yeem raug xa mus. Piv txwv li, yog tias cov khoom lag luam ua ntej pom zoo DDI, cov khoom tom ntej tsim los ntawm lub platform kuj xav tau cov kev tshawb fawb cuam tshuam. Yog tias qhov tsis zoo, lwm yam khoom siv thiab cov ntaub ntawv tuaj yeem muab tau yam tsis tas yuav muaj kev tshawb fawb ntxiv DDI. Tsis tas li ntawd, DDI feem ntau ntsuam xyuas tag nrho - cov khoom siv ntsuas ntev, nrog cov metabolites tsis suav nrog qhov kev kuaj xyuas. Txij li tsuas yog ib feem me me ntawm ONs khiav tawm lysosomes thiab nkag mus rau cytoplasm tom qab cellular uptake, cuam tshuam rau cov metabolism hauv cov enzymes, hepatocytes zoo dua simulate tag nrho cov txheej txheem no, thaum qhov tsis zoo yog feem ntau nrog hepatic microsomes. Saib ntawm cov neeg thauj khoom, kev lag luam ONs tuaj yeem cuam tshuam qee cov neeg thauj khoom, xws li casimersen inhibiting MATE1 thiab eteplirsen inhibiting OCT1 thiab OATP1B1. Yog li, nws raug nquahu kom ua cov kev tshawb fawb txog kev cuam tshuam ntawm efflux thiab influx transporters los ntawm ONs.
Dhau li ncaj qha ua rau DDI los ntawm kev khi, ONs kuj tseem tuaj yeem cuam tshuam rau kev qhia ntawm CYPs lossis cov neeg thauj khoom los ntawm lawv cov teebmeem PD. Yog li ntawd, kev tshawb fawb cov ntaub ntawv zoo yog xav tau los ntsuam xyuas qhov cuam tshuam ntawm cov khoom tsim los ntawm PD cov teebmeem ntawm kev nthuav tawm ntawm cov enzymes metabolic thiab thauj. FDA tseem txhawj xeeb txog ONs tsis ncaj qha cuam tshuam rau CYPs kev ua haujlwm. Piv txwv li, cytokines tuaj yeem cuam tshuam rau CYPs qhia, thiab ONs tuaj yeem tswj hwm cytokine concentrations. Hauv cov xwm txheej zoo li no, cov qauv kev ntsuas hauv vitro CYPs tsis siv tau, thiab kev soj ntsuam siv cov neeg mob ntshav ntshav tom qab kev tswj hwm rau hepatocytes tuaj yeem txiav txim siab.
FDA (2020) thiab EMA (2012) cov lus qhia qhia ua kom tiav DDI cov kev tshawb fawb rau cov tshuaj me me ua ntej Phase III kev sim tshuaj, uas kuj siv rau ONs cov khoom.
7. Cov lus xaus
Cov tshuaj Oligonucleotide yog ib pawg tshiab ntawm kev kho mob, thiab tsis muaj cov lus qhia tshwj xeeb rau ADME cov kev tshawb fawb rau cov khoom lag luam no. Cov koom haum muaj xws li OSWG tau ua lub luag haujlwm tseem ceeb hauv kev muab cov lus qhia ntxaws ntxaws rau ADME cov kev tshawb fawb. Txawm hais tias feem ntau cov qauv tsim qauv rau cov tshuaj oligonucleotide zoo ib yam li cov tshuaj me me thiab biologics, lawv muaj lawv tus kheej tshwj xeeb. Piv txwv li, kev pheej hmoo ntawm kev sib txawv ntawm poj niam txiv neej hauv kev nqus yog qhov tsawg, tso cai rau ib leeg - hom PK kev tshawb fawb. Kev sau cov qauv tshuaj lom neeg yuav tsum xav txog ib nrab -lub neej hauv cov ntaub so ntswg. Kev tshawb fawb ntawm cov ntaub so ntswg rau cov tshuaj oligonucleotide yog qhov tseem ceeb thiab yuav tsum tau ua ntej qhov tseem ceeb tshaj li cov tshuaj me me thiab biologics. Tsis tas li ntawd, vim qhov sib xws ntawm cov enzymes metabolic thoob plaws txhua hom, qhov tseem ceeb ntawm cov tshuaj oligonucleotide cov tshuaj metabolism hauv kev tshawb fawb tsis yog siab npaum li cov molecules me. Cov tshuaj Oligonucleotide feem ntau tsis yog cov substrates rau cov kab mob siab xws li CYPs, tab sis lawv yeej muaj feem cuam tshuam rau kev ua haujlwm ntawm CYPs lossis cov neeg thauj khoom, yuav tsum tau saib xyuas DDI txaus ntshai. Txawm li cas los xij, tsis yog txhua qhov kev tshawb fawb yuav tsum ua kom tiav ua ntej IND; Muaj ntau qhov kev sim tuaj yeem muab tau thaum kuaj mob. Cov khoom lag luam Oligonucleotide kuj tseem cuam tshuam txog kev pheej hmoo los ntawm cov khoom sib txuas, cov khoom xa khoom hauv tsheb, thiab lwm yam, uas yuav tsum tau ua tib zoo xav hauv kev tshawb fawb tsim vim tias lawv tuaj yeem cuam tshuam rau kev kuaj pom.
