Claims (19)
1. Čestica rekombinantnog adeno-povezanog virusa (rAAV) za upotrebu u postupku isporuke rAAV-a u središnji živčani sustav (SŽS) sisavca, naznačena time što se čestica rAAV primjenjuje u striatum sisavca isporukom poboljšanom konvekcijom (CED), pri čemu čestica rAAV sadrži sadrži vektor rAAV koji kodira heterolognu nukleinsku kiselinu koja je eksprimirana barem u okcipitalnom korteksu i/ili sloju IV moždane kore i strijatuma sisavca, pri čemu čestica rAAV sadrži AAV serotip 2 (AAV2) kapsid, te pri čemu se čestica rAAV primjenjuje na najmanje jedno mjesto u kaudatnoj jezgri i dva mjesta u putamenu u svakoj hemisferi strijatuma.1. A recombinant adeno-associated virus (rAAV) particle for use in a method of delivering rAAV to the central nervous system (CNS) of a mammal, characterized in that the rAAV particle is administered to the mammalian striatum by convection-enhanced delivery (CED), wherein the rAAV particle comprises comprising an rAAV vector encoding a heterologous nucleic acid that is expressed at least in the occipital cortex and/or layer IV of the cerebral cortex and striatum of a mammal, wherein the rAAV particle comprises an AAV serotype 2 (AAV2) capsid, and wherein the rAAV particle is administered to at least one site in the caudate nucleus and two sites in the putamen in each hemisphere of the striatum.
2. Čestica rekombinantnog adeno-povezanog virusa (rAAV) za upotrebu u postupku liječenja poremećaja SŽS-a kod sisavca, naznačena time što se učinkovita količina čestice rAAV primjenjuje u striatum sisavca isporukom poboljšanom konvekcijom (CED), pri čemu čestica rAAV sadrži vektor rAAV koji kodira heterolognu nukleinsku kiselinu koja je eksprimirana u barem okcipitalnom korteksu i/ili sloju IV moždane kore i strijatuma sisavca, pri čemu čestica rAAV sadrži AAV2 kapsid, te pri čemu se čestica rAAV primjenjuje na najmanje jedno mjesto u kaudatnoj jezgri i dva mjesta u putamenu u svakoj hemisferi strijatuma.2. A recombinant adeno-associated virus (rAAV) particle for use in a method of treating a CNS disorder in a mammal, characterized in that an effective amount of the rAAV particle is administered to the striatum of the mammal by convection-enhanced delivery (CED), wherein the rAAV particle comprises an rAAV vector comprising encodes a heterologous nucleic acid that is expressed in at least the occipital cortex and/or layer IV of the cerebral cortex and striatum of a mammal, wherein the rAAV particle comprises an AAV2 capsid, and wherein the rAAV particle is administered to at least one site in the caudate nucleus and two sites in the putamen in each hemisphere of the striatum.
3. Čestica rAAV za upotrebu prema patentnom zahtjevu 2, naznačena time što je poremećaj SŽS-a Huntingtonova bolest, pri čemu po izboru heterologna nukleinska kiselina kodira terapeutski polipeptid ili terapeutsku nukleinsku kiselinu koja inhibira ekspresiju huntingtina (HTT) ili inhibira nakupljanje HTT u stanicama SŽS-a sisavca.3. The rAAV particle for use according to patent claim 2, characterized in that the CNS disorder is Huntington's disease, wherein optionally the heterologous nucleic acid encodes a therapeutic polypeptide or a therapeutic nucleic acid that inhibits the expression of huntingtin (HTT) or inhibits the accumulation of HTT in CNS cells - a mammal.
4. Čestica rAAV za upotrebu prema patentnom zahtjevu 3, naznačena time što heterologna nukleinska kiselina kodira terapeutsku miRNA koja cilja na huntingtin, i pri čemu po izboru huntingtin sadrži mutaciju povezanu s Huntingtonovom bolešću.4. The rAAV particle for use according to claim 3, characterized in that the heterologous nucleic acid encodes a therapeutic miRNA targeting huntingtin, and wherein the huntingtin optionally contains a mutation associated with Huntington's disease.
5. Čestica rAAV za upotrebu prema patentnom zahtjevu 2, naznačena time što je poremećaj SŽS-a Parkinsonova bolest, te pri čemu po izboru heterologna nukleinska kiselina kodira terapeutski polipeptid, pri čemu je po izboru terapeutski polipeptid faktor rasta koji potječe od glija (GDNF), faktor rasta koji potječe iz mozga (BDNF), tirozin hidroksalaza (TH), GTP-ciklohidrolaza (GTPCH), i/ili aminokiselinska dekarboksilaza (AADC).5. The rAAV particle for use according to patent claim 2, characterized in that the CNS disorder is Parkinson's disease, and wherein optionally the heterologous nucleic acid encodes a therapeutic polypeptide, wherein optionally the therapeutic polypeptide is glial-derived growth factor (GDNF). , brain-derived growth factor (BDNF), tyrosine hydroxylase (TH), GTP-cyclohydrolase (GTPCH), and/or amino acid decarboxylase (AADC).
6. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-5, naznačena time što je sisavac čovjek.6. The rAAV particle for use according to any one of claims 1-5, characterized in that the mammal is human.
7. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-5, naznačena time što se čestica rAAV primjenjuje na putamen i kaudatnu jezgru strijatuma, pri čemu je omjer čestica rAAV primijenjenih na putamen prema česticama rAAV primijenjenim u kaudatnu jezgru najmanje oko 2:1.7. The rAAV particle for use according to any one of claims 1-5, characterized in that the rAAV particle is administered to the putamen and the caudate nucleus of the striatum, wherein the ratio of rAAV particles administered to the putamen to rAAV particles administered to the caudate nucleus is at least about 2: 1.
8. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-7, naznačena time što:
(a) heterologna nukleinska kiselina je eksprimirana u prefrontalnim asocijacijskim kortikalnim područjima, premotornom korteksu, primarnim somatosenzornim kortikalnim područjima, senzornom motornom korteksu, parijetalnom korteksu, okcipitalnom korteksu i/ili primarnom motornom korteksu;
(b) čestica rAAV prolazi kroz retrogradni ili anterogradni transport u moždanoj kori; i/ili
(c) heterologna nukleinska kiselina je dalje eksprimirana u talamusu, subtalamičkoj jezgri, globus pallidus, crnoj tvari i/ili hipokampusu.8. An rAAV particle for use according to any one of claims 1-7, characterized in that:
(a) the heterologous nucleic acid is expressed in prefrontal association cortical areas, premotor cortex, primary somatosensory cortical areas, sensory motor cortex, parietal cortex, occipital cortex and/or primary motor cortex;
(b) the rAAV particle undergoes retrograde or anterograde transport in the cerebral cortex; and/or
(c) the heterologous nucleic acid is further expressed in the thalamus, subthalamic nucleus, globus pallidus, substantia nigra and/or hippocampus.
9. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-8, naznačena time što vektor rAAV sadrži heterolognu nukleinsku kiselinu okruženu jednom ili više sekvenci AAV invertiranog terminalnog ponavljanja (ITR), pri čemu po izboru:
(a) heterologna nukleinska kiselina je okružena s dva AAV ITR-a; i/ili
(b) AAV ITR-ovi su AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAVrh8, AAVrh8R, AAV9, AAV10, AAVrh10, AAV11, AAV12, AAV2R471A, AAV DJ, kozji AAV, goveđi AAV, ili mišji AAV serotip ITR-a.9. The rAAV particle for use according to any one of claims 1-8, characterized in that the rAAV vector contains a heterologous nucleic acid flanked by one or more AAV inverted terminal repeat (ITR) sequences, wherein optionally:
(a) the heterologous nucleic acid is flanked by two AAV ITRs; and/or
(b) AAV ITRs are AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAVrh8, AAVrh8R, AAV9, AAV10, AAVrh10, AAV11, AAV12, AAV2R471A, AAV DJ, goat AAV, bovine AAV, or murine AAV serotype of ITR.
10. Čestica rAAV za upotrebu prema patentnom zahtjevu 9, naznačena time što:
(a) jedna ili više ITR sekvenci i kapsid čestice rAAV virusa su iz istog AAV serotipa; ili
(b) jedna ili više ITR sekvenci i kapsid čestice rAAV virusa su iz različitih AAV serotipova.10. The rAAV particle for use according to claim 9, characterized in that:
(a) one or more ITR sequences and the capsid particle of the rAAV virus are from the same AAV serotype; or
(b) one or more ITR sequences and capsid particles of the rAAV virus are from different AAV serotypes.
11. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-10, naznačena time što je heterologna nukleinska kiselina operativno vezana na promotor, pri čemu po izboru:
(a) promotor eksprimira heterolognu nukleinsku kiselinu u stanici SŽS-a, pri čemu po izboru promotor eksprimira heterolognu nukleinsku kiselinu u stanici mozga;
(b) promotor eksprimira heterolognu nukleinsku kiselinu u neuronu i/ili glijalnoj stanici, pri čemu po izboru neuron je srednji bodljasti neuron kaudatne jezgre, srednji bodljasti neuron putamena, neuron sloja IV korteksa i/ili neuron sloja V korteksa;
(c) promotor je CBA promotor, minimalni CBA promotor, CMV promotor ili GUSB promotor; i/ili
(d) promotor se može inducirati.11. The rAAV particle for use according to any one of claims 1-10, characterized in that the heterologous nucleic acid is operably linked to the promoter, wherein optionally:
(a) the promoter expresses the heterologous nucleic acid in the SŽS cell, whereby the promoter optionally expresses the heterologous nucleic acid in the brain cell;
(b) the promoter expresses a heterologous nucleic acid in a neuron and/or a glial cell, wherein the neuron is optionally a medium spiny neuron of the caudate nucleus, a medium spiny neuron of the putamen, a neuron of layer IV cortex and/or a neuron of layer V cortex;
(c) the promoter is a CBA promoter, minimal CBA promoter, CMV promoter or GUSB promoter; and/or
(d) the promoter is inducible.
12. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-11, naznačena time što vektor rAAV sadrži jedan ili više pojačivača, par splice donor/splice akceptor, mjesto pričvršćivanja matriksa, i signal poliadenilacije.12. The rAAV particle for use according to any one of claims 1-11, characterized in that the rAAV vector contains one or more enhancers, a splice donor/splice acceptor pair, a matrix attachment site, and a polyadenylation signal.
13. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-12, naznačena time što je vektor rAAV samo-komplementaran vektor rAAV, pri čemu po izboru vektor sadrži prvu sekvencu nukleinske kiseline koja kodira heterolognu nukleinsku kiselinu i drugu sekvencu nukleinske kiseline koja kodira komplement heterologne nukleinske kiseline, pri čemu prva sekvenca nukleinske kiseline može formirati intralančane bazne parove s drugom sekvencom nukleinske kiseline duž većine ili cijele svoje duljine; i pri čemu su po izboru prva sekvenca nukleinske kiseline i druga sekvenca nukleinske kiseline povezane mutiranim AAV ITR, pri čemu mutirani AAV ITR sadrži deleciju D regije i sadrži mutaciju sekvence terminalne rezolucije.13. The rAAV particle for use according to any one of claims 1-12, characterized in that the rAAV vector is a self-complementary rAAV vector, wherein optionally the vector contains a first nucleic acid sequence that encodes a heterologous nucleic acid and a second nucleic acid sequence that encodes the complement of a heterologous nucleic acid, wherein the first nucleic acid sequence can form intrastrand base pairs with the second nucleic acid sequence along most or all of its length; and wherein optionally the first nucleic acid sequence and the second nucleic acid sequence are linked by a mutated AAV ITR, wherein the mutated AAV ITR comprises a D region deletion and comprises a terminal resolution sequence mutation.
14. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-2 i 6-13, naznačena time što heterologna nukleinska kiselina kodira:
(a) terapeutski polipeptid; ili
(b) terapeutsku nukleinsku kiselinu, pri čemu po izboru terapeutska nukleinska kiselina je siRNA, shRNA, RNAi, miRNA, protusmislena RNA, ribozim ili DNAzim.14. The rAAV particle for use according to any one of claims 1-2 and 6-13, characterized in that the heterologous nucleic acid encodes:
(a) a therapeutic polypeptide; or
(b) a therapeutic nucleic acid, wherein optionally the therapeutic nucleic acid is siRNA, shRNA, RNAi, miRNA, antisense RNA, ribozyme or DNAzyme.
15. Čestica rAAV za upotrebu prema patentnom zahtjevu 14, naznačena time što:
(a) terapeutski polipeptid je enzim, neurotrofni faktor, polipeptid koji ima nedostatak ili je mutiran kod pojedinca s poremećajem povezanim sa SŽS-om, antioksidans, anti-apoptotički faktor, anti-angiogeni faktor, te protu-upalni faktor, alfa-sinuklein, kisela beta-glukozidaza (GBA), beta-galaktozidaza-1 (GLB1), iduronat 2-sulfataza (IDS), galaktozilceramidaza (GALC), manozidaza, alfa-D-manozidaza (MAN2B1), beta-manozidaza (MANBA), pseudoarilsulfataza A (ARSA), N N-acetilglukozamin-1-fosfotransferaza (GNPTAB), kisela sfingomijelinaza (ASM), Niemann-Pick C protein (NPC1), kisela alfa-1,4-glukozidaza (GAA), podjedinica heksozaminidaze beta, HEXB, N-sulfoglukozamin sulfohidrolaza (MPS3A), N-alfa-acetilglukozaminidaza (NAGLU), heparin acetil-CoA, alfa-glukozaminidaza N-acetiltransferaza (MPS3C), N-acetilglukozamin-6-sulfataza (GNS), alfa-N-acetilgalaktozaminidaza (NAGA), beta-glukuronidaza (GUSB), podjedinica heksosaminidaze alfa (HEXA), huntingtin (HTT), lizosomska kisela lipaza (LIPA), aspartilglukozaminidaza, Alfa-galaktozidaza A, Palmitoil protein tioesteraza, tripeptidil peptidaza, lizosomalni transmembranski protein, cisteinski transporter, kisela ceramidaza, kisela alfa-L-fukozidaza, katepsin A, alfa-L-iduronidaza, arilsulfataza B, arilsulfataza A, N-acetilgalaktozamin-6-sulfat, kisela beta-galaktozidaza, ili alfa-neuramidaza; ili
(b) terapeutski polipeptid ili terapeutska nukleinska kiselina se koristi za liječenje poremećaja SŽS-a, pri čemu po izboru:
(i) poremećaj SŽS-a je Huntingtonova bolest, epilepsija, Parkinsonova bolest, Alzheimerova bolest, moždani udar, kortikobazalna degeneracija (CBD), kortikobazalna ganglijska degeneracija (CBGD), frontotemporalna demencija (FTD), multipla sistemska atrofija (MSA), progresivna supranuklearna paraliza (PSP) ili rak mozga; ili
(ii) poremećaj je lizosomska bolest nakupljanja odabrana iz skupine koju čine aspartilglukozaminurija, Fabryjeva bolest, infantilna Battenova bolest (CNL1), klasična kasna infantilna Battenova bolest (CNL2), juvenilna Battenova bolest (CNL3), Battenova bolest oblik CNL4, Battenova bolest oblik CNL5, Battenova bolest oblik CNL6, Battenova bolest oblik CNL7, Battenova bolest oblik CNL8, cistinoza, Farber, fukozidoza, galaktozidozijalidoza, Gaucherova bolest tip 1, Gaucherova bolest tip 2, Gaucherova bolest tip 3, GM1 gangliozidoza, Hunterova bolest, Krabbeova bolest, α manozidoza, β manozidoza, Maroteaux-Lamy, bolest metakromatske leukodistrofije, Morquio A, Morquio B, mukolipidoza tipa II/III, Niemann-Pickova bolest tip A, Niemann-Pickova bolest tip B, Niemann-Pickova bolest tip C, Pompeova bolest, Sandhoffova bolest, Sanfillipo A sindrom, Sanfillipo B sindrom, Sanfillipo C sindrom, Sanfillipo D sindrom, Schindlerova bolest, Schindler-Kanzaki, sijalidoza, Sly sindrom, Tay-Sachsova bolest, i Wolmanova bolest.15. The rAAV particle for use according to claim 14, characterized in that:
(a) the therapeutic polypeptide is an enzyme, a neurotrophic factor, a polypeptide that is deficient or mutated in an individual with a disorder associated with SŽS, an antioxidant, an anti-apoptotic factor, an anti-angiogenic factor, and an anti-inflammatory factor, alpha-synuclein, acid beta-glucosidase (GBA), beta-galactosidase-1 (GLB1), iduronate 2-sulfatase (IDS), galactosylceramidase (GALC), mannosidase, alpha-D-mannosidase (MAN2B1), beta-mannosidase (MANBA), pseudoarylsulfatase A (ARSA), N N-acetylglucosamine-1-phosphotransferase (GNPTAB), acid sphingomyelinase (ASM), Niemann-Pick C protein (NPC1), acid alpha-1,4-glucosidase (GAA), hexosaminidase beta subunit, HEXB, N -sulfoglucosamine sulfohydrolase (MPS3A), N-alpha-acetylglucosaminidase (NAGLU), heparin acetyl-CoA, alpha-glucosaminidase N-acetyltransferase (MPS3C), N-acetylglucosamine-6-sulfatase (GNS), alpha-N-acetylgalactosaminidase (NAGA) , beta-glucuronidase (GUSB), hexosaminidase alpha subunit (HEXA), huntingtin (HTT), lysosomal acid lipase (LIPA) , aspartylglucosaminidase, Alpha-galactosidase A, Palmitoyl protein thioesterase, tripeptidyl peptidase, lysosomal transmembrane protein, cysteine transporter, acid ceramidase, acid alpha-L-fucosidase, cathepsin A, alpha-L-iduronidase, arylsulfatase B, arylsulfatase A, N-acetylgalactosamine -6-sulfate, acid beta-galactosidase, or alpha-neuramidase; or
(b) the therapeutic polypeptide or therapeutic nucleic acid is used for the treatment of disorders of the SŽS, whereby optionally:
(i) a CNS disorder is Huntington's disease, epilepsy, Parkinson's disease, Alzheimer's disease, stroke, corticobasal degeneration (CBD), corticobasal ganglion degeneration (CBGD), frontotemporal dementia (FTD), multiple system atrophy (MSA), progressive supranuclear paralysis (PSP) or brain cancer; or
(ii) the disorder is a lysosomal storage disease selected from the group consisting of aspartylglucosaminuria, Fabry disease, infantile Batten disease (CNL1), classic late infantile Batten disease (CNL2), juvenile Batten disease (CNL3), Batten disease form CNL4, Batten disease form CNL5 , Batten disease form CNL6, Batten disease form CNL7, Batten disease form CNL8, cystinosis, Farber, fucosidosis, galactosidosialidosis, Gaucher disease type 1, Gaucher disease type 2, Gaucher disease type 3, GM1 gangliosidosis, Hunter disease, Krabbe disease, α mannosidosis , β mannosidosis, Maroteaux-Lamy, metachromatic leukodystrophy disease, Morquio A, Morquio B, mucolipidosis type II/III, Niemann-Pick disease type A, Niemann-Pick disease type B, Niemann-Pick disease type C, Pompe disease, Sandhoff disease , Sanfillipo A syndrome, Sanfillipo B syndrome, Sanfillipo C syndrome, Sanfillipo D syndrome, Schindler's disease, Schindler-Kanzaki, sialidosis, Sly syndrome, Tay-Sachs bo lest, and Wolman's disease.
16. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-15, naznačena time što je čestica rAAV u pripravku, pri čemu je po izboru pripravak farmaceutski pripravak koji sadrži farmaceutski prihvatljivu pomoćnu tvar.16. The rAAV particle for use according to any of claims 1-15, characterized in that the rAAV particle is in a preparation, wherein the preparation is optionally a pharmaceutical preparation containing a pharmaceutically acceptable excipient.
17. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-16, naznačena time što:
(a) čestica rAAV je proizvedena trostrukom transfekcijom nukleinske kiseline koja kodira vektor rAAV, nukleinske kiseline koja kodira AAV rep i cap, te nukleinske kiseline koja kodira pomoćne funkcije AAV virusa u stanici domaćina, pri čemu transfekcija nukleinskih kiselina na stanice domaćina stvara stanicu domaćina sposobnu za proizvodnju čestica rAAV; ili
(b) čestica rAAV je proizvedena od strane proizvodne stanične linije koja sadrži jednu ili više nukleinskih kiselina koje kodiraju vektor rAAV, nukleinsku kiselinu koja kodira AAV rep i cap p, i nukleinsku kiselinu koja kodira pomoćne funkcije AAV virusa.17. An rAAV particle for use according to any one of claims 1-16, characterized in that:
(a) the rAAV particle is produced by triple transfection of the nucleic acid encoding the rAAV vector, the nucleic acid encoding the AAV tail and cap, and the nucleic acid encoding the auxiliary functions of the AAV virus in the host cell, whereby the transfection of the nucleic acids into the host cells creates a host cell capable of for the production of rAAV particles; or
(b) the rAAV particle is produced by a production cell line containing one or more nucleic acids encoding the rAAV vector, nucleic acid encoding the AAV tail and cap p, and nucleic acid encoding the helper functions of the AAV virus.
18. Čestica rAAV za upotrebu prema bilo kojem od patentnih zahtjeva 1-17, naznačena time što se čestica rAAV isporučuje korištenjem CED sustava za isporuku.18. The rAAV particle for use according to any one of claims 1-17, characterized in that the rAAV particle is delivered using a CED delivery system.
19. Čestica rAAV za upotrebu prema patentnom zahtjevu 18, naznačena time što CED sustav sadrži:
(a) kanilu, pri čemu po izboru kanila je kanila otporna na refluks ili stepenasta kanila; i/ili
(b) pumpa, pri čemu je po izboru pumpa:
(i) ručna pumpa;
(ii) osmotska pumpa; ili
(iii) infuzijska pumpa.19. The rAAV particle for use according to claim 18, characterized in that the CED system contains:
(a) a cannula, where the choice of cannula is a reflux-resistant cannula or a stepped cannula; and/or
(b) a pump, where the pump is optionally:
(i) hand pump;
(ii) osmotic pump; or
(iii) infusion pump.