JPWO2020106916A5

JPWO2020106916A5 -

Info

Publication number: JPWO2020106916A5
Application number: JP2021529002A
Authority: JP
Publication date: 2022-11-30

Claims

From 5' to 3',
5' inverted terminal repeat (ITR);
promoter;
a transgene sequence;
one or more polyadenylation signals; and a 3'ITR
Nucleic acids, including

an intronic sequence, wherein said intronic sequence comprises a sequence of any one of SEQ ID NOs: 13-16 , or a sequence that is at least 95% identical thereto;
a Kozak sequence immediately 5' of the transgene sequence, wherein said Kozak sequence comprises the sequence of SEQ ID NO: 17 or 18, or a sequence that is at least 95% identical thereto;
said promoter comprises a sequence selected from, or at least 95% identical to, any one of SEQ ID NOs: 6-12;
said transgene sequence encodes a frataxin (FXN) protein comprising the sequence of SEQ ID NO: 65, or a sequence that is at least 95% identical thereto;
said one or more polyadenylation signals comprise any one or more sequences of SEQ ID NOs: 21-24, or sequences that are at least 95% identical thereto; and/or
any combination of these
2. The nucleic acid of claim 1, comprising:

From 5' to 3',
5' inverted terminal repeat (ITR);
a promoter, wherein said promoter comprises a sequence selected from, or at least 95% identical to, any one of SEQ ID NOs: 6-12 ;
an intronic sequence, wherein said intronic sequence comprises a sequence of any one of SEQ ID NOs: 13-16, or a sequence that is at least 95% identical thereto ;
a Kozak sequence immediately 5' of the transgene sequence, wherein said Kozak sequence comprises the sequence of SEQ ID NO: 17 or 18, or a sequence that is at least 95% identical thereto;
a transgene sequence, wherein said transgene sequence encodes a frataxin (FXN) protein comprising the sequence of SEQ ID NO: 65, or a sequence that is at least 95% identical thereto;
one or more polyadenylation signals, wherein said one or more polyadenylation signals comprise the sequence of any one of SEQ ID NOs: 21-24, or a sequence that is at least 95% identical thereto; and
3' ITR
3. The nucleic acid of claim 1 or 2, comprising:

The nucleic acid of any one of claims 1-3, wherein the transgene sequence is CpG optimized.

5. The nucleic acid of any one of claims 1-4, wherein the nucleic acid comprises an enhancer, optionally wherein the enhancer comprises the sequence of SEQ ID NO: 4 or 5, or a sequence that is at least 95% identical thereto. nucleic acid.

6. The nucleic acid of any one of claims 1-5, wherein the nucleic acid comprises a sequence of any one of SEQ ID NOs: 28-64, or a sequence that is at least 85% identical thereto.

The nucleic acid of any one of claims 1-5, wherein the nucleic acid comprises a sequence of any one of SEQ ID NOs: 28-64, or a sequence that is at least 90% identical thereto.

6. The nucleic acid of any one of claims 1-5, wherein the nucleic acid comprises a sequence of any one of SEQ ID NOs: 28-64, or a sequence that is at least 95% identical thereto.

The nucleic acid of any one of claims 6-8, wherein the nucleic acid comprises or consists of the sequence of SEQ ID NO:51.

The nucleic acid of any one of claims 6-8, wherein the nucleic acid comprises or consists of the sequence of SEQ ID NO:49.

The nucleic acid of any one of claims 6-8, wherein the nucleic acid comprises or consists of the sequence of SEQ ID NO:32.

A plasmid comprising a nucleic acid according to any one of claims 1-11 .

A cell comprising a nucleic acid according to any one of claims 1 to 11 or a plasmid according to claim 12 .

A method of producing a recombinant AAV vector comprising contacting an AAV-producing cell with the nucleic acid of any one of claims 1-11 or the plasmid of claim 12 .

A recombinant AAV vector produced by the method of claim 14 .

Said recombinant AAV vector is single-stranded AAV (ssAAV), or self-complementary AAV (scAAV), optionally said recombinant AAV vector is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7 , AAV8, AAV9, AAV10, AAV11, AAV12, AAVrh8, AAVrh10 , AAVrh32.33, AAVrh74, avian AAV, or bovine AAV.

17. The recombinant AAV vector of claim 15 or 16 , wherein said AAV vector comprises a capsid protein from AAV9 .

The nucleic acid of any one of claims 1 to 11 , the plasmid of claim 12 , the cell of claim 13 , or the recombinant AAV vector of any one of claims 15 to 17 . A composition comprising:

A recombinant AAV vector, for use as a pharmaceutical, comprising a therapeutically effective amount of a nucleic acid according to any one of claims 1 to 11 or a plasmid according to claim 12 .

wherein said recombinant AAV vector is administered by direct injection into the subject's central nervous system (CNS), and optionally said administration causes expression of frataxin (FXN) protein in said subject's CNS and/or cardiac tissue; A recombinant AAV vector for use according to claim 19, which results in

21. A recombinant AAV vector for use according to claim 19 or 20, wherein said subject has Friedreich's ataxia, optionally said subject is a human subject.