Adeno-associated viral vector variants

Abstract

To provide: targeting peptides and vectors containing a sequence that encodes the targeting peptides that deliver agents to specific substructures in the brain; and methods for delivering agents to distinct brain structures of subjects.SOLUTION: A modified adeno-associated virus (AAV) capsid protein is provided which comprises a targeting peptide that targets a viral vector comprising the modified AAV capsid protein to a distinct organ or brain structure, where the targeting peptide is 3-10 amino acids in length.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Patent Application No. 62 / 939,315, filed November 22, 2019, and U.S. Provisional Patent Application No. 63 / 084,709, filed September 29, 2020, the entire contents of both applications being incorporated herein by reference.

[0002] Sequence Listing Reference This application contains a Sequence Listing that has been submitted in ASCII format via EFS-Web and is incorporated by reference herein in its entirety. The ASCII copy, created on November 19, 2020, is named CHOPP0038WO_ST25.txt and is 63.8 kilobytes in size.

[0003] 1. Field The present invention relates generally to the fields of medicine, virology, and neurology. More specifically, the present invention relates to targeting peptides that target the delivery of viral vectors to different structures in the brain. [Background technology]

[0004] 2. Description of Related Art Various strategies have been developed to generate AAV vector variants, including rational design and directed evolution. Rational design approaches use knowledge of the AAV capsid to create targeted changes to the capsid, such as tyrosine mutations on the capsid surface to increase transduction efficiency, thereby altering transduction efficiency or specificity. Directed evolution approaches do not require knowledge of the capsid structure and are performed through random mutagenesis, capsid shuffling, or random peptide insertion. These strategies generally use in vitro systems or mice, which are ideal for cell-based or mouse studies but do not translate to clinical applications. In fact, AAV variants do not specifically or efficiently target different brain structures. Therefore, there is a need for AAV variants that can target different primate brain structures. Summary of the Invention

[0005] overview Provided herein are viral vectors each comprising a modified capsid, the modified capsid comprising at least one amino acid sequence that allows the viral vector to be targeted to different brain structures.

[0006] In one embodiment, a modified adeno-associated virus (AAV) capsid protein is provided, the modified AAV capsid protein comprising a targeting peptide, wherein the targeting peptide targets a viral vector comprising the modified AAV capsid protein to a different organ or a different brain structure, and the targeting peptide is 3 to 10 amino acids in length. In some aspects, the modified AAV capsid protein is a modified AAV1 capsid protein, a modified AAV2 capsid protein, or a modified AAV9 capsid protein.

[0007] In some aspects, the modified AAV capsid protein is derived from the AAV1 capsid protein (see SEQ ID NO: 138), and the targeting peptide is inserted after residue 590 of the AAV1 capsid protein. In some aspects, the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length. In some aspects, the linker sequences are SSA on the N-terminal side of the targeting peptide and AS on the C-terminal side of the targeting peptide. In some aspects, the modified AAV1 capsid protein has a sequence at least 95% identical to SEQ ID NO: 141.

[0008] In some aspects, the modified AAV capsid protein is derived from the AAV2 capsid protein (see SEQ ID NO: 139), and the targeting peptide is inserted after residue 587 of the AAV2 capsid protein. In some aspects, the targeting peptide is flanked by linker sequences, and the linker sequences on either side of the targeting peptide are 2 or 3 amino acids in length. In some aspects, the linker sequences are AAA on the N-terminal side of the targeting peptide and AA on the C-terminal side of the targeting peptide. In some aspects, the modified AAV2 capsid protein has a sequence at least 95% identical to SEQ ID NO: 142.

[0009] In some aspects, the modified AAV capsid protein is derived from the AAV9 capsid protein (see SEQ ID NO: 140), and the targeting peptide is inserted after residue 588 of the AAV9 capsid protein. In some aspects, the targeting peptide is flanked by linker sequences, and the linker sequences on either side of the targeting peptide are 2 or 3 amino acids in length. In some aspects, the linker sequences are AAA on the N-terminal side of the targeting peptide and AS on the C-terminal side of the targeting peptide. In some aspects, the modified AAV9 capsid protein has a sequence at least 95% identical to SEQ ID NO: 143.

[0010] In some aspects, the targeting peptide comprises a sequence up to 10 amino acids in length, wherein the sequence has an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-137 and 144. In some aspects, the targeting peptide is 7 amino acids in length.

[0011] In some aspects, the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.In certain aspects, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from those listed in Table 1 to target the corresponding brain structure.In certain aspects, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from those listed in Table 2 to target the corresponding brain structure.In certain aspects, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from those listed in Table 3 to target the corresponding brain structure.

[0012] In some aspects, the different organs are brain, kidney, heart, liver, gonad, spleen, or liver.In certain aspects, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from those listed in Table 4 to target the corresponding organ.In certain aspects, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from those listed in Table 5 to target the corresponding organ.In certain aspects, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from those listed in Table 6 to target the corresponding organ.

[0013] In one embodiment, provided herein is a nucleic acid comprising a sequence encoding the modified capsid protein of any one of the present embodiments.

[0014] In one embodiment, a recombinant adeno-associated virus (rAAV) comprising any one of the modified capsid proteins of this embodiment is provided herein. In some aspects, a combination of rAAVs is provided. For example, the combination of rAAV with modified AAV1 capsid protein and the targeting peptide of SEQ ID NO: 21, rAAV with modified AAV2 capsid protein and the targeting peptide of SEQ ID NO: 53, rAAV with modified AAV2 capsid protein and the targeting peptide of SEQ ID NO: 80, and rAAV with modified AAV9 capsid protein and the targeting peptide of SEQ ID NO: 113 is provided.

[0015] In one embodiment, provided herein is a viral vector comprising the nucleic acid encoding any one of the modified capsid proteins of this embodiment.In some aspects, the viral vector further comprises the nucleic acid sequence encoding the nucleic acid of interest.In some aspects, the nucleic acid of interest is a therapeutic substance.In some aspects, the therapeutic substance is an enzyme or an RNAi molecule.

[0016] In one embodiment, provided herein is a cell comprising any one of the viral vectors of this embodiment. In some aspects, the cell is a mammalian cell, such as a human cell. In some aspects, the cell is in vitro or in vivo.

[0017] In one embodiment, provided herein is a pharmaceutical composition comprising a viral vector of this embodiment and a pharmaceutically acceptable carrier.

[0018] In one embodiment, provided herein is a method for delivering an active substance to different brain structures of a subject, comprising administering the virus of this embodiment to the subject.In some aspects, the different brain structures are brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.In certain aspects, rAAV with modified AAV1 capsid protein is used, and targeting peptide is selected from those listed in Table 1 to target corresponding brain structures.In certain aspects, rAAV with modified AAV2 capsid protein is used, and targeting peptide is selected from those listed in Table 2 to target corresponding brain structures.In certain aspects, rAAV with modified AAV9 capsid protein is used, and targeting peptide is selected from those listed in Table 3 to target corresponding brain structures.In various aspects, any combination of rAAV is used. For example, combinations of rAAV with a modified AAV1 capsid protein and a targeting peptide of SEQ ID NO: 21, rAAV with a modified AAV2 capsid protein and a targeting peptide of SEQ ID NO: 53, rAAV with a modified AAV2 capsid protein and a targeting peptide of SEQ ID NO: 80, and rAAV with a modified AAV9 capsid protein and a targeting peptide of SEQ ID NO: 113 are used.

[0019] In one embodiment, provided herein is a method for delivering an agent to different organs of a subject, comprising administering the virus of this embodiment to the subject.In some aspects, the organ is the brain, kidney, heart, liver, gonad, spleen, or liver.In certain aspects, an rAAV having a modified AAV1 capsid protein is used, and the targeting peptide is selected from those listed in Table 4 to target the corresponding organ.In certain aspects, an rAAV having a modified AAV2 capsid protein is used, and the targeting peptide is selected from those listed in Table 5 to target the corresponding organ.In certain aspects, an rAAV having a modified AAV9 capsid protein is used, and the targeting peptide is selected from those listed in Table 6 to target the corresponding organ.In various aspects, any combination of rAAVs is used.

[0020] In some aspects, the agent is siRNA, shRNA, miRNA, non-coding RNA, lncRNA, therapeutic protein, or CRISPR system.In some aspects, the administration is to the central nervous system.In some aspects, the administration is to the cisterna magna, intraventricular space, ependyma, ventricle, subarachnoid space, and / or intrathecal space.In some aspects, the ventricle is the rostral lateral ventricle, and / or the caudal lateral ventricle, and / or the right lateral ventricle, and / or the left lateral ventricle, and / or the right rostral lateral ventricle, and / or the left rostral lateral ventricle, and / or the right caudal lateral ventricle, and / or the left caudal lateral ventricle.

[0021] In some aspects, multiple viral particles are administered. In some aspects, the virus is administered at a concentration of about 1 x 10 vector genomes per kilogram. 6 ~Approx. 1×10 18 In some aspects, the virus is administered at a dose of about 1 x 10 vg / kg of patient. 7 ~1×10 17 , about 1×10 8 ~1×10 16, about 1×10 9 ~1×10 15 , about 1×10 10 ~1×10 14 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 11 , about 1×10 11 ~1×10 12 , about 1×10 12 ~×10 13 , or approximately 1 × 10 13 ~1×10 14 In some aspects, the subject is a human.

[0022] In one embodiment, provided herein is a method for treating disease in mammals, comprising administering the virus of this embodiment to mammals.In some aspects, the disease is neurodegenerative disease.In some aspects, the neurodegenerative disease is Huntington's disease, ALS, hereditary spastic hemiplegia, primary lateral sclerosis, spinal muscular atrophy, Kennedy's disease, Alzheimer's disease, polyglutamine repeat disease or Parkinson's disease.In some aspects, the mammal is human.

[0023] As used herein, "essentially free" of a specific component means that the specific component is not intentionally formulated into the composition and / or is present only as a contaminant or in trace amounts. The total amount of the specific component resulting from unintentional incorporation of the composition is therefore significantly less than 0.05%, preferably less than 0.01%. Most preferred is a composition in which the amount of such specific component cannot be detected by standard analytical methods.

[0024] As used herein, "a" or "an" can mean one or more. As used herein in the claims, the words "a" or "an," when used with the word "comprising," can mean one or more than one.

[0025] Although the use of the term "or" in the claims is used to mean "and / or," unless expressly indicated to refer to alternatives only or mutually exclusive alternatives, the present disclosure supports the definition of "and / or" referring to alternatives only. As used herein, "another" can mean at least a second or more.

[0026] Throughout this application, the term "about" is used to indicate that a value includes the inherent error variation of the device, the method being employed to determine the value, the variation that exists among study subjects, or a value that is within 10% of the stated value.

[0027] [The present invention 1001] A modified adeno-associated virus (AAV) capsid protein comprising a targeting peptide, the targeting peptide being 3 to 10 amino acids in length, that targets a viral vector comprising the modified AAV capsid protein to a different organ or a different brain structure. [The present invention 1002] The modified AAV capsid protein of the present invention 1001, wherein the modified AAV capsid protein is a modified AAV9 capsid protein having a sequence at least 95% identical to SEQ ID NO: 143, the targeting peptide is SEQ ID NO: 110, and the different brain structures are the brainstem, caudate nucleus, cerebellum, cochlea (ear), cortex, cerebral cortex, deep cerebellar nuclei, ependyma, globus pallidus, hippocampus, meninges, motor cortex, optic nerve, prefrontal cortex, putamen, spinal cord, substantia nigra, subthalamic nucleus, temporal cortex, thalamus, or visual cortex. [The present invention 1003] The modified AAV capsid protein of the present invention 1001, which is a modified AAV1 capsid protein, a modified AAV2 capsid protein, or a modified AAV9 capsid protein. [The present invention 1004] 1001. A modified AAV capsid protein of the present invention, wherein the modified AAV capsid protein is derived from the AAV1 capsid protein (see SEQ ID NO: 138) and the targeting peptide is inserted after residue 590 of the AAV1 capsid protein. [The present invention 1005] 1004. The modified AAV capsid protein of the present invention, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length. [The present invention 1006] 1005. The modified AAV capsid protein of the present invention, wherein the linker sequence is SSA at the N-terminus of the targeting peptide and AS at the C-terminus of the targeting peptide. [The present invention 1007] 1006. The modified AAV capsid protein of the present invention, wherein said modified AAV1 capsid protein has a sequence that is at least 95% identical to SEQ ID NO: 141. [The present invention 1008] 1001. A modified AAV capsid protein of the present invention, wherein the modified AAV capsid protein is derived from an AAV2 capsid protein (see SEQ ID NO: 139) and the targeting peptide is inserted after residue 587 of the AAV2 capsid protein. [The present invention 1009] 1008. The modified AAV capsid protein of the present invention, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length. [The present invention 1010] 1009. The modified AAV capsid protein of the present invention, wherein the linker sequence is AAA at the N-terminus of the targeting peptide and AA at the C-terminus of the targeting peptide. [The present invention 1011] 10. The modified AAV capsid protein of the present invention, wherein said modified AAV2 capsid protein has a sequence that is at least 95% identical to SEQ ID NO: 142. [The present invention 1012] 1001. A modified AAV capsid protein of the present invention, wherein the modified AAV capsid protein is derived from an AAV9 capsid protein (see SEQ ID NO: 140) and the targeting peptide is inserted after residue 588 of the AAV9 capsid protein. [The present invention 1013] 1012. The modified AAV capsid protein of the present invention, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length. [The present invention 1014] 1013. The modified AAV capsid protein of the present invention, wherein the linker sequence is AAA on the N-terminal side of the targeting peptide and AS on the C-terminal side of the targeting peptide. [The present invention 1015] 1014. The modified AAV capsid protein of the present invention, wherein said modified AAV9 capsid protein has a sequence that is at least 95% identical to SEQ ID NO: 143. [The present invention 1016] The modified AAV capsid protein of the present invention 1001, wherein the target peptide comprises a sequence of up to 10 amino acids in length, and the sequence has an amino acid sequence selected from the group consisting of SEQ ID NO: 1 to 137 or 144. [The present invention 1017] 1016. The modified AAV capsid protein of the present invention, wherein the targeting peptide is 7 amino acids in length. [The present invention 1018] The modified AAV capsid protein of any of claims 1001 to 1017, wherein the different brain structure is the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus. [The present invention 1019] The modified AAV capsid protein of the present invention, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1 to 9. [The present invention 1020] The modified AAV capsid protein of the present invention, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 5, 7, 10-16, 25, 26, 32, and 144. [The present invention 1021] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 4, 9, and 17 to 21. [The present invention 1022] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 5, 12, and 21 to 26. [The present invention 1023] The modified AAV capsid protein of the present invention, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2-4, 7, 9, 21, 22, 27, and 28. [The present invention 1024] The modified AAV capsid protein of the present invention, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 5, 12, 14, 16, 21, 22, and 29 to 31. [The present invention 1025] The modified AAV capsid protein of the present invention, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1-4, 7, and 32-34. [The present invention 1026] The modified AAV capsid protein of the present invention, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 5, 7, 9, 12, 21, and 35 to 37. [The present invention 1027] The modified AAV capsid protein of the present invention, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2, 3, 7, 14-16, 21, 31, and 38. [The present invention 1028] The modified AAV capsid protein of the present invention, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 4, 12, 13, 21, 30, and 39 to 42. [The present invention 1029] The modified AAV capsid protein of the present invention, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2 to 4, 7, 9, 21, 32, 33, and 43. [The present invention 1030] The modified AAV capsid protein of the present invention, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2, 3, 9, 44, and 45. [The present invention 1031] The modified AAV capsid protein of the present invention, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2 to 4, 12, 16, 30, 46, and 47. [The present invention 1032] The modified AAV capsid protein of the present invention, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 2, 8, 12, 21, 28, and 48 to 51. [The present invention 1033] The modified AAV capsid protein of the present invention, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52 to 60. [The present invention 1034] The modified AAV capsid protein of the present invention, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 59 and 61 to 69. [This invention 1035] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 56, 58, 60, and 70-75. [The present invention 1036] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 58, 60, 62, 63, 66, and 76-79. [This invention 1037] The modified AAV capsid protein of the present invention, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 60, 62, 63, 66, 74-77, and 80. [The present invention 1038] The modified AAV capsid protein of the present invention, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 60, 75, and 81 to 87. [This invention 1039] The modified AAV capsid protein of the present invention, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 55, 58, 60, 63, 76, 79, 88, and 89. [The present invention 1040] The modified AAV capsid protein of the present invention, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 58, 60, 66, 73, 76, 80, and 90-93. [The present invention 1041] The modified AAV capsid protein of the present invention, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 54, 57, 58, 60, 75, 79, 87, 88, and 94. [The present invention 1042] The modified AAV capsid protein of the present invention, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 55, 59, 60, 61, and 95 to 100. [This invention 1043] The modified AAV capsid protein of the present invention, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 58-61, 63, 77, 88, 95, and 101. [This invention 1044] The modified AAV capsid protein of the present invention, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52, 53, 57, 58, 75, 76, 87, 102, and 103. [This invention 1045] The modified AAV capsid protein of the present invention, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 57, 58, 60, 75, 79, 87, 88, 102, 104, and 105. [The present invention 1046] The modified AAV capsid protein of the present invention, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52, 55, 56, 74, 85, 88, and 106 to 109. [This invention 1047] The modified AAV capsid protein of the present invention, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110 to 117. [This invention 1048] The modified AAV capsid protein of the present invention, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 113, 115, 116, and 118 to 121. [This invention 1049] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 119, and 122 to 125. [The present invention 1050] The modified AAV capsid protein of the present invention, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 114, 116, and 125 to 127. [This invention 1051] The modified AAV capsid protein of the present invention, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 118-120, and 128. [This invention 1052] The modified AAV capsid protein of the present invention, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110 to 112, 114, 119, 120, and 129. [This invention 1053] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 116, 123, 125, 129, and 130. [This invention 1054] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 114, 118, 119, 122, and 131. [This invention 1055] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 114, 115, 117, 129, and 132. [The present invention 1056] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 112, 113, 116, 123, 127, 133, and 134. [This invention 1057] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 113, 119, 120, 122, 123, 128, and 134. [This invention 1058] The modified AAV capsid protein of the present invention, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110 to 114, 117, and 129. [This invention 1059] The modified AAV capsid protein of the present invention 1018, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 119, 120, 122, 132, and 135. [The present invention 1060] The modified AAV capsid protein of the present invention, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 112 to 114, 125, 133, 136, and 137. [This invention 1061] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 1, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, hippocampus, or thalamus. [This invention 1062] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 2, and the different brain structures are the brainstem, ependyma, hippocampus, optic nerve, spinal cord, substantia nigra, subthalamic nucleus, or thalamus. [This invention 1063] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 3, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, or subthalamic nucleus. [This invention 1064] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 4, and the different brain structures are the brainstem, cerebellar cortex, ependyma, hippocampus, putamen, spinal cord, or subthalamic nucleus. [This invention 1065] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 5, and the different brain structures are the brainstem, cerebral cortex, globus pallidus, or meninges. [The present invention 1066] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 6, and the different brain structure is the brainstem. [This invention 1067] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 7, and the different brain structures are the brainstem, caudate nucleus, ependyma, hippocampus, meninges, optic nerve, or spinal cord. [The present invention 1068] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 8, and the different brain structure is the brainstem or the thalamus. [This invention 1069] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 9, and the different brain structures are the brainstem, cerebellar cortex, ependyma, meninges, spinal cord, or substantia nigra. [The present invention 1070] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 10 or 11, and the different brain structure is the caudate nucleus. [This invention 1071] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 12, and the different brain structures are the caudate nucleus, cerebral cortex, globus pallidus, meninges, putamen, subthalamic nucleus, or thalamus. [This invention 1072] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 13, and the different brain structure is the caudate nucleus or putamen. [This invention 1073] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 14, and the different brain structures are the caudate nucleus, the globus pallidus, or the optic nerve. [This invention 1074] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 15, and the different brain structure is the caudate nucleus or the optic nerve. [This invention 1075] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 16, and the different brain structures are the caudate nucleus, the globus pallidus, the optic nerve, or the subthalamic nucleus. [This invention 1076] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 17 to 20, and the different brain structure is the cerebellar cortex. [This invention 1077] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 21, and the different brain structures are the cerebellar cortex, cerebral cortex, ependyma, globus pallidus, meninges, optic nerve, putamen, spinal cord, or thalamus. [This invention 1078] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 22, and the different brain structure is the cerebral cortex, the ependyma, or the globus pallidus. [This invention 1079] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 23 to 26, and the different brain structure is the cerebral cortex. [The present invention 1080] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 27, and the different brain structure is the ependyma. [This invention 1081] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 28, and the different brain structure is the ependyma or thalamus. [This invention 1082] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 29, and the different brain structure is the globus pallidus. [This invention 1083] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 30, and the different brain structures are the globus pallidus, putamen, or subthalamic nucleus. [This invention 1084] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 31, and the different brain structure is the globus pallidus or the optic nerve. [This invention 1085] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 32 or 33, and the different brain structure is the hippocampus or the spinal cord. [The present invention 1086] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 34, and the different brain structure is the hippocampus. [This invention 1087] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 35 to 37, and the different brain structure is the meninges. [This invention 1088] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 38, and the different brain structure is the optic nerve. [This invention 1089] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 39 to 42, and the different brain structure is the putamen. [The present invention 1090] The modified AAV capsid protein of invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 43, and the different brain structure is the spinal cord. [This invention 1091] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 44 or 45, and the different brain structure is the substantia nigra. [This invention 1092] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 46 or 47, and the different brain structure is the subthalamic nucleus. [This invention 1093] The modified AAV capsid protein of the present invention 1001 or 1007, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 48 to 51, and the different brain structure is the thalamus. [This invention 1094] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 52, and the different brain structure is the brainstem, the substantia nigra, or the thalamus. [This invention 1095] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 53, and the different brain structures are the brainstem, cerebral cortex, ependyma, hippocampus, meninges, optic nerve, spinal cord, or substantia nigra. [This invention 1096] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 54, and the different brain structure is the brainstem or the optic nerve. [This invention 1097] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 55, and the different brain structures are the brainstem, hippocampus, putamen, or thalamus. [This invention 1098] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 56, and the different brain structure is the brainstem, cerebellar cortex, or thalamus. [This invention 1099] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 57, and the different brain structures are the brainstem, optic nerve, substantia nigra, or subthalamic nucleus. [The present invention 1100] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 58, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, hippocampus, meninges, optic nerve, spinal cord, substantia nigra, or subthalamic nucleus. [The present invention 1101] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 59, and the different brain structures are the brainstem, the caudate nucleus, the putamen, or the spinal cord. [The present invention 1102] The modified AAV capsid protein of invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 60, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, or subthalamic nucleus. [The present invention 1103] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 61, and the different brain structure is the caudate nucleus, putamen, or spinal cord. [The present invention 1104] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 62, and the different brain structure is the caudate nucleus, the cerebral cortex, or the ependyma. [This invention 1105] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 63, and the different brain structures are the caudate nucleus, the cerebral cortex, the ependyma, the hippocampus, or the spinal cord. [The present invention 1106] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 64, 65, and 67 to 69, and the different brain structure is the caudate nucleus. [This invention 1107] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 66, and the different brain structures are the caudate nucleus, the cerebral cortex, the ependyma, or the meninges. [This invention 1108] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 70 to 72, and the different brain structure is the cerebellar cortex. [This invention 1109] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 73, and the different brain structure is the cerebellar cortex or the meninges. [The present invention 1110] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 74, and the different brain structure is the cerebellar cortex, the ependyma, or the thalamus. [The present invention 1111] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 75, and the different brain structures are the cerebellar cortex, the ependyma, the globus pallidus, the optic nerve, the substantia nigra, or the subthalamic nucleus. [The present invention 1112] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 76, and the different brain structures are the cerebral cortex, ependyma, hippocampus, meninges, or substantia nigra. [The present invention 1113] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 77, and the different brain structure is the cerebral cortex, the ependyma, or the spinal cord. [This invention 1114] The modified AAV capsid protein of invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 78, and the different brain structure is the cerebral cortex. [This invention 1115] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 79, and the different brain structures are the cerebral cortex, hippocampus, optic nerve, or subthalamic nucleus. [The present invention 1116] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 80, and the different brain structures are the ependyma, hippocampus, or meninges. [This invention 1117] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 81 to 84 and 86, and the different brain structure is the globus pallidus. [This invention 1118] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 85, and the different brain structure is the globus pallidus or the thalamus. [This invention 1119] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 87, and the different brain structures are the globus pallidus, the optic nerve, the substantia nigra, or the subthalamic nucleus. [The present invention 1120] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 88, and the different brain structures are the hippocampus, optic nerve, spinal cord, subthalamic nucleus, or thalamus. [This invention 1121] The modified AAV capsid protein of invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 89, and the different brain structure is the hippocampus. [This invention 1122] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 90 to 93, and the different brain structure is the meninges. [This invention 1123] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 94, and the different brain structure is the optic nerve. [This invention 1124] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 95, and the different brain structure is the putamen or the spinal cord. [Invention 1125] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 96 to 100, and the different brain structure is the putamen. [The present invention 1126] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 101, and the different brain structure is the spinal cord. [This invention 1127] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 102, and the different brain structure is the substantia nigra or the subthalamic nucleus. [This invention 1128] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 103, and the different brain structure is the substantia nigra. [This invention 1129] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 104 or 105, and the different brain structure is the subthalamic nucleus. [The present invention 1130] The modified AAV capsid protein of the present invention 1001 or 1011, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 106 to 109, and the different brain structure is the thalamus. [This invention 1131] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 110, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus. [This invention 1132] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 111, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, substantia nigra, or subthalamic nucleus. [This invention 1133] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 112, and the different brain structures are the brainstem, globus pallidus, putamen, substantia nigra, or thalamus. [This invention 1134] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 113, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, hippocampus, meninges, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus. [This invention 1135] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 114, and the different brain structures are the brainstem, cerebral cortex, globus pallidus, meninges, optic nerve, substantia nigra, or thalamus. [This invention 1136] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 115, and the different brain structure is the brainstem or the caudate nucleus. [This invention 1137] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 116, and the different brain structures are the brainstem, caudate nucleus, cerebral cortex, hippocampus, optic nerve, or putamen. [This invention 1138] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 117, and the different brain structure is the brainstem, the optic nerve, or the substantia nigra. [This invention 1139] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 118, and the different brain structure is the caudate nucleus, the ependyma, or the meninges. [The present invention 1140] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 119, and the different brain structures are the caudate nucleus, cerebellar cortex, ependyma, globus pallidus, meninges, spinal cord, or subthalamic nucleus. [This invention 1141] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 120, and the different brain structures are the caudate nucleus, the ependyma, the globus pallidus, the meninges, the spinal cord, or the subthalamic nucleus. [This invention 1142] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 121, and the different brain structure is the caudate nucleus. [This invention 1143] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 122, and the different brain structure is the cerebellar cortex, the meninges, the spinal cord, or the subthalamic nucleus. [This invention 1144] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 123, and the different brain structures are the cerebellar cortex, hippocampus, putamen, or spinal cord. [Invention 1145] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 124, and the different brain structure is the cerebellar cortex. [Invention 1146] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 125, and the different brain structures are the cerebellar cortex, the cerebral cortex, the hippocampus, or the thalamus. [This invention 1147] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 126, and the different brain structure is the cerebral cortex. [This invention 1148] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 127, and the different brain structure is the cerebral cortex or putamen. [This invention 1149] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 128, and the different brain structure is the ependyma or the spinal cord. [This invention 1150] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 129, and the different brain structures are the globus pallidus, hippocampus, optic nerve, or substantia nigra. [This invention 1151] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 130, and the different brain structure is the hippocampus. [This invention 1152] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 131, and the different brain structure is the meninges. [This invention 1153] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 132, and the different brain structure is the optic nerve or the subthalamic nucleus. [This invention 1154] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 133, and the different brain structure is the putamen or thalamus. [This invention 1155] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 134, and the different brain structure is the putamen or the spinal cord. [Invention 1156] The modified AAV capsid protein of invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 135, and the different brain structure is the subthalamic nucleus. [This invention 1157] The modified AAV capsid protein of the present invention 1001 or 1015, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 136 or 137, and the different brain structure is the thalamus. [This invention 1158] A nucleic acid comprising a sequence encoding any one of the modified capsid proteins of the present inventions 1001 to 1157. [This invention 1159] A recombinant adeno-associated virus (rAAV) virus comprising the modified capsid protein of any one of claims 1001 to 1157. [The present invention 1160] A viral vector comprising a nucleic acid encoding the modified capsid protein of any one of the present inventions 1001 to 1157. [This invention 1161] The viral vector of the present invention 1160 further comprising a nucleic acid sequence encoding a nucleic acid of interest. [This invention 1162] 1161. The viral vector of claim 1161, wherein said nucleic acid of interest is a therapeutic agent. [This invention 1163] 1162. The viral vector of claim 1162, wherein said therapeutic substance is an enzyme or an RNAi molecule. [This invention 1164] A cell comprising the viral vector of any one of the present inventions 1160 to 1163. [Invention 1165] The cell of the present invention 1164, which is a mammalian cell. [Invention 1166] The cell of the present invention 1164, which is a human cell. [This invention 1167] The cells of the present invention 1164, which are in vitro. [Invention 1168] The cells of the present invention 1164, which are in vivo. [This invention 1169] A pharmaceutical composition comprising a viral vector of the present invention and a pharmaceutically acceptable carrier. [This invention 1170] A method for delivering an agent to different brain structures in a subject, comprising administering to said subject a virus of the present invention. [This invention 1171] 1170. The method of claim 1170, wherein the method is for delivering an agent to the brainstem of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1-9, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52-60, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-117. [This invention 1172] 1170. The method of claim 1170, wherein the method is for delivering an agent to the caudate nucleus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 7, and 10-16, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 59 and 61-69, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 113, 115, 116, and 118-121. [This invention 1173] 1170. The method of claim 1170, wherein the method is for delivering an agent to the cerebellar cortex of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 4, 9, and 17-21, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 56, 58, 60, and 70-75, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 119, and 122-125. [This invention 1174] 1170. A method for delivering an agent to the cerebral cortex of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 5, 12, and 21-26, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58, 60, 62, 63, 66, and 76-79, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 114, 116, and 125-127. [This invention 1175] 1170. The method of claim 1170, wherein the method is for delivering an agent to the ependyma of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 7, 9, 21, 22, 27, and 28, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 60, 62, 63, 66, 74-77, and 80, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 118-120, and 128. [Invention 1176] 1170. A method for delivering an agent to the globus pallidus of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 5, 12, 14, 16, 21, 22, and 29-31, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 60, 75, and 81-87, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-112, 114, 119, 120, and 129. [This invention 1177] 1170. A method for delivering an agent to the hippocampus of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1-4, 7, and 32-34, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 55, 58, 60, 63, 76, 79, 80, 88, and 89; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 116, 123, 125, 129, and 130. [This invention 1178] 1170. A method for delivering an agent to the meninges of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 5, 7, 9, 12, 21, and 35-37, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58, 60, 66, 73, 76, 80, and 90-93; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 114, 118, 119, 122, and 131. [This invention 1179] 1170. The method of claim 1170, wherein the method is for delivering an agent to the optic nerve of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2, 3, 7, 14-16, 21, 31, and 38, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 54, 57, 58, 60, 75, 79, 87, 88, and 94, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 114, 115, 117, 129, and 132. [This invention 1180] 1170. A method for delivering an agent to the putamen of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 4, 12, 13, 21, 30, and 39-42, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 55, 59, 60, 61, and 95-100, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 112, 113, 116, 123, 127, 133, and 134. [This invention 1181] 1170. The method of claim 1170, wherein the method is for delivering an agent to the spinal cord of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 7, 9, 21, 32, 33, and 43, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58-61, 63, 77, 88, 95, and 101, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 113, 119, 120, 122, 123, 128, and 134. [This invention 1182] 1170. A method for delivering an agent to the substantia nigra of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2, 3, 9, 44, and 45, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52, 53, 57, 58, 75, 76, 87, 102, and 103; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-114, 117, and 129. [This invention 1183] 1170. A method for delivering an agent to the subthalamic nucleus of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 12, 16, 30, 46, and 47; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 57, 58, 60, 75, 79, 87, 88, 102, 104, and 105; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 119, 120, 122, 132, and 135. [This invention 1184] 1170. A method for delivering an agent to the thalamus of a subject, comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 2, 8, 12, 21, 28, and 48-51, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52, 55, 56, 74, 85, 88, and 106-109, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 112-114, 125, 133, 136, and 137. [This invention 1185] 118. The method of any of claims 1170 to 1184, wherein said agent is an siRNA, shRNA, miRNA, non-coding RNA, lncRNA, therapeutic protein, or CRISPR system. [Invention 1186] The method of any one of claims 1170 to 1184, wherein said administration is to the central nervous system. [This invention 1187] 1186. The method of claim 1186, wherein said administration is to the cisterna magna, intraventricular space, ependyma, ventricle, subarachnoid space, cochlea, and / or intrathecal space. [This invention 1188] The method of the present invention 1187, wherein the ventricles are the rostral lateral ventricle, and / or the caudal lateral ventricle, and / or the right lateral ventricle, and / or the left lateral ventricle, and / or the right rostral lateral ventricle, and / or the left rostral lateral ventricle, and / or the right caudal lateral ventricle, and / or the left caudal lateral ventricle. [This invention 1189] The method of any of claims 1170 to 1188, wherein a plurality of viral particles are administered. [This invention 1190] The virus has a vector genome content of about 1 x 10 per kilogram 6 ~Approx. 1×10 18 The method of the present invention, wherein the compound is administered at a dose of 1189 mg / kg. [This invention 1191] The virus is present in an amount of approximately 1 x 10 vg per kg of patient. 7 ~1×10 17 , about 1×10 8 ~1×10 16 , about 1×10 9 ~1×10 15 , about 1×10 10 ~1×10 14 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 11 , about 1×10 11 ~1×10 12 , about 1×10 12 ~×10 13 , or approximately 1 × 10 13 ~1×10 14 The method of the present invention, wherein the compound is administered in a single dose. [This invention 1192] The method of any one of claims 1170 to 1191, wherein the subject is a human. [This invention 1193] A method of treating a disease in a mammal comprising administering to said mammal a virus of the invention 1159. [This invention 1194] 1193. The method of claim 1193, wherein the disease is a neurodegenerative disease. [This invention 1195] 1194. The method of claim 1194, wherein said neurodegenerative disease is Huntington's disease, ALS, hereditary spastic hemiplegia, primary lateral sclerosis, spinal muscular atrophy, Kennedy's disease, Alzheimer's disease, polyglutamine repeat disease, or Parkinson's disease. [Invention 1196] 1193. The method of claim 1193, wherein the mammal is a human. Other objects, features, and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. [Brief explanation of the drawings]

[0028] The following drawings form part of the present specification and are provided to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein. [Figure 1] Schematic diagram of the AAV peptide display library. [Figure 2] Schematic of the in vivo screening strategy. [Figure 3] Graphical representation of input library diversity. Diversity of input viral libraries measured from aliquots of AAV1, AAV2, and AAV9 viral vectors before round 1 ICV injection. [Figure 4-1] Graphical representation of round-over-round barcode enrichment. Total number of unique barcodes recovered after round 1 and round 2 enrichment in Rhesus macaques per collected tissue. Round 2 values ​​for DNA and RNA are shown. [Figure 4-2] See description of Figure 4-1. [Figure 4-3] See description of Figure 4-1. [Figure 5-1] Illustrated round-over-round enrichment of barcodes in AAV1 serotypes in the cerebellar cortex. [Figure 5-2] Illustration of round-over-round enrichment of barcodes in AAV2 serotypes in the cerebellar cortex. [Figure 5-3] Illustrated round-over-round enrichment of barcodes in AAV9 serotypes in the cerebellar cortex. [Figure 6-1] Illustrated enrichment of AAV9 1999. Heatmap depiction of barcode enrichment from AAV9, with cells color-coded by the percentage of barcode detected in the indicated tissue. Barcodes recovered from DNA are shown on the left, and barcodes recovered from RNA are shown on the right. [Figure 6-2] See description of Figure 6-1. [Figure 7A] Heatmap depiction of opool barcode enrichment from AAV1. [Figure 7B] Heatmap depiction of o-pool barcode enrichment from AAV2. [Figure 7C] Heatmap depiction of o-pool barcode enrichment from AAV9. [Figure 8] AAV9 1999 in vivo rhesus monkey validation. The eGFP expression construct was packaged into AAV9 1999 driven by a CAG promoter. 1.5E13 vg of AAV9 1999 was delivered to a 5-year-old female rhesus macaque via ICV injection. Representative images of H&E-stained cerebellum depicting the transduction pattern of AAV9 1999 are shown. [Figure 9] Figure 9A-D. AAV9 1999 in vivo mouse validation. The eGFP expression construct was packaged into AAV9 1999 driven by the CAG promoter. AAV9 1999 containing the eGFP construct and AAV9 capsid were delivered to C57BL / 6 p0 mouse pups by ICV injection at 1E10 vg. Representative images of eGFP fluorescent signals are shown for the whole brain (Figure 9A), whole brain sagittal section (Figure 9B), S1 cortex section (Figure 9C, left), hippocampus section (Figure 9C, center), cerebellum sagittal section (Figure 9C, right), and lumbar spinal cord coronal section (Figure 9D). [Figure 10A] Fluorescence image of the AAV mixture in vivo in the rhesus monkey lateral ventricle. [Figure 10B] Fluorescence image of the in vivo rhesus monkey fourth ventricle with the AAV mixture. [Figure 10C] Fluorescence images of in vivo rhesus monkey meninges with AAV mixture. [Figure 11A]Fluorescence image of the cochlear turn after cochlear administration of AAV9 1999 capsid containing an eGFP construct into mice. [Figure 11B] Fluorescence image of inner hair cells after cochlear administration of AAV9 1999 capsid containing an eGFP construct into mice. [Figure 11C] Fluorescence images of the organ of Corti and distal modiolus after cochlear administration of AAV9 1999 capsid containing an eGFP construct into mice. DETAILED DESCRIPTION OF THE INVENTION

[0029] Detailed Description Provided herein are viral vectors, each of which comprises modified capsid, and each modified capsid comprises at least one amino acid sequence that allows viral vector to target different brain structures.In certain embodiments, the brain structure is brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus or thalamus.The targeting peptide for each brain structure is provided in Table 1-3.

[0030] In certain embodiments, the viral vector is an adeno-associated viral vector (AAV). In certain embodiments, the AAV is AAV1, AAV2, or AAV9. An exemplary wild-type reference AAV1 capsid protein sequence is provided in SEQ ID NO: 138. An exemplary wild-type reference AAV2 capsid protein sequence is provided in SEQ ID NO: 139. An exemplary wild-type reference AAV9 capsid protein sequence is provided in SEQ ID NO: 140. In certain aspects, the targeting peptide is inserted at position 590 of the AAV1 capsid, position 587 of the AAV2 capsid, or position 588 of the AAV9 capsid. An exemplary modified AAV1 capsid protein sequence is provided in SEQ ID NO: 141, which shows a targeting peptide insertion after position 590 as SSAX7AS, where the leading SSA and trailing AS are linker sequences, and X7 represents the targeting peptide. An exemplary modified AAV2 capsid protein sequence is provided in SEQ ID NO: 142, which shows a targeting peptide insertion after position 587 as AAAX7AA, where the leading AAA and trailing AA are linker sequences, and X7 represents the targeting peptide. An exemplary modified AAV9 capsid protein sequence is provided in SEQ ID NO: 143, which shows a targeting peptide insertion after position 588 as AAAX7AS, where the leading AAA and trailing AS are linker sequences, and X7 represents the targeting peptide.

[0031] Table 1. AAV1 targeting peptides for each brain structure TIFF2025148448000002.tif52154TIFF2025148448000003.tif227154TIFF2025 148448000004.tif227154TIFF2025148448000005.tif227154TIFF20251484480 00006.tif227154TIFF2025148448000007.tif227154TIFF2025148448000008.t if227154TIFF2025148448000009.tif227154TIFF2025148448000010.tif129154

[0032] Table 2. AAV2 targeting peptides for each brain structure TIFF2025148448000011.tif83154TIFF2025148448000012.tif222154TIFF2025 148448000013.tif232154TIFF2025148448000014.tif227154TIFF20251484480 00015.tif227154TIFF2025148448000016.tif227154TIFF2025148448000017.t if227154TIFF2025148448000018.tif227154TIFF2025148448000019.tif170154

[0033] Table 3. AAV9 targeting peptides for each brain structure TIFF2025148448000020.tif42154TIFF2025148448000021.tif227154TIFF2025 148448000022.tif227154TIFF2025148448000023.tif227154TIFF20251484480 00024.tif227154TIFF2025148448000025.tif227154TIFF2025148448000026.t if227154TIFF2025148448000027.tif227154TIFF2025148448000028.tif57154

[0034] Table 4. AAV1 targeting peptides for various organs TIFF2025148448000029.tif42154

[0035] Table 5. AAV2 targeting peptides for various organs TIFF2025148448000030.tif217154

[0036] Table 6. AAV9 targeting peptides for various organs TIFF2025148448000031.tif201154

[0037] I. Adeno-associated virus (AAV) vectors Adeno-associated viruses (AAVs) are small, nonpathogenic viruses in the Parvoviridae family. To date, numerous serologically distinct AAVs have been identified, with more than 12 identified in humans or primates. AAVs differ from other members of this family in their dependence on a helper virus for replication.

[0038] The AAV genome exists extrachromosomally without integrating into the host cell genome, has a broad host range, can transduce both dividing and non-dividing cells in vitro and in vivo, and can maintain high levels of transduced gene expression. AAV viral particles are thermostable, resistant to solvents, detergents, pH, and temperature changes, and can be column-purified and / or concentrated using CsCl gradients or other means. The AAV genome contains positive- or negative-sense single-stranded deoxyribonucleic acid (ssDNA). The approximately 4.7 kb AAV genome consists of a single segment of single-stranded DNA with either positive or negative polarity. Both ends of the genome are short inverted terminal repeats (ITRs) that can fold into hairpin structures and serve as origins of viral DNA replication.

[0039] An AAV "genome" refers to a recombinant nucleic acid sequence that is ultimately packaged or encapsidated to form an AAV particle. AAV particles often contain an AAV genome packaged by AAV capsid proteins. When a recombinant plasmid is used to construct or produce a recombinant vector, the AAV vector genome does not include portions of the "plasmid" that do not correspond to the vector genome sequence of the recombinant plasmid. This non-vector genome portion of the recombinant plasmid is referred to as the "plasmid backbone," which is important for plasmid cloning and amplification, processes necessary for plasmid growth and production, but is not itself packaged or encapsidated into a viral particle. Thus, an AAV vector "genome" refers to the nucleic acid that is packaged or encapsidated by the AAV capsid proteins.

[0040] AAV virions (particles) are non-enveloped icosahedral particles approximately 25 nm in diameter that contain the AAV capsid. AAV particles have icosahedral symmetry, consisting of three related capsid proteins, VP1, VP2, and VP3, which interact with each other to form the capsid. Most native AAV genomes contain two open reading frames (ORFs), sometimes referred to as the left-hand and right-hand ORFs. The right-hand ORF often encodes the capsid proteins VP1, VP2, and VP3. These proteins are often found in a 1:1:10 ratio, respectively, although the ratio can vary and all are derived from the right-hand ORF. The VP1, VP2, and VP3 capsid proteins differ from each other through alternative splicing and the use of unusual start codons. Deletion analysis has shown that removal or modification of VP1, which is translated from alternatively spliced ​​messages, results in reduced yields of infectious particles. Mutations within the VP3 coding region result in single-stranded progeny DNA or a failure to produce infectious particles. In certain embodiments, the genome of an AAV particle encodes one, two, or all three of the VP1, VP2, and VP3 polypeptides.

[0041] The left ORF often encodes the nonstructural Rep proteins Rep40, Rep52, Rep68, and Rep78, which are involved in regulating replication and transcription as well as producing single-stranded progeny genomes. Two of the Rep proteins have been associated with the preferential inclusion of the AAV genome in a region of the q arm of human chromosome 19. Rep68 / 78 have been shown to have NTP-binding activity in addition to DNA and RNA helicase activity. Some Rep proteins have a nuclear localization signal along with several potential phosphorylation sites. In certain embodiments, the genome of an AAV (e.g., rAAV) encodes some or all of the Rep proteins. In certain embodiments, the genome of an AAV (e.g., rAAV) does not encode a Rep protein. In certain embodiments, one or more of the Rep proteins can be delivered in trans and are therefore not included in AAV particles containing nucleic acids encoding polypeptides.

[0042] The termini of the AAV genome contain short inverted terminal repeats (ITRs) that have the potential to fold into T-shaped hairpin structures that serve as origins of viral DNA replication. Thus, the AAV genome contains one or more (e.g., a pair) ITR sequences flanking the single-stranded viral DNA genome. ITR sequences are often approximately 145 bases long each. Within the ITR region, two elements thought to be central to ITR function have been described: a GAGC repeat motif and a terminal resolution site (trs). The repeat motif has been shown to bind Rep when the ITR is in either a linear or hairpin conformation. This binding is thought to position Rep68 / 78 for site- and strand-specific cleavage at the trs. In addition to their role in replication, these two elements appear to be central to viral inclusion. The chromosome 19 integration locus contains a Rep binding site flanked by trs. These elements have been shown to be functional and required for locus-specific inclusion.

[0043] The term "recombinant" as a modifier of a vector, such as a recombinant viral vector, e.g., a recombinant lentivirus or recombinant parvovirus (e.g., AAV) vector, and as a modifier of a sequence, such as a recombinant nucleic acid sequence or a recombinant polypeptide, means that the composition has been manipulated (i.e., engineered) in a manner that does not normally occur in nature. Particular examples of recombinant vectors, such as AAV, retrovirus, or lentivirus vectors, are those in which a nucleic acid sequence not normally present in the wild-type viral genome has been inserted into the viral genome. An example of a recombinant nucleic acid sequence is one in which the nucleic acid (e.g., gene) encodes an inhibitory RNA cloned into the vector with or without the 5', 3', and / or intron regions normally associated with that gene in the viral genome. While the term "recombinant" is not always used herein with respect to vectors, such as viral vectors, and sequences, such as polynucleotides, "recombinant" forms, including nucleic acid sequences, polynucleotides, transgenes, and the like, are expressly encompassed despite such omissions.

[0044] A recombinant viral "vector" is derived from a wild-type viral genome by using molecular methods to remove a portion of the wild-type genome from the virus and replace it with a non-native nucleic acid, such as a nucleic acid sequence. Typically, in the case of AAV, for example, one or both inverted terminal repeat (ITR) sequences of the AAV genome are retained in the recombinant AAV vector. A "recombinant" viral vector (e.g., rAAV) is distinguished from a viral (e.g., AAV) genome in that a portion of the viral genome is replaced with a non-native sequence, such as a nucleic acid encoding a transactivator, a nucleic acid encoding an inhibitory RNA, or a nucleic acid encoding a therapeutic protein, compared to the viral genome nucleic acid. Therefore, the incorporation of such a non-native nucleic acid sequence defines the viral vector as a "recombinant" vector, and in the case of AAV, it can be called a "rAAV vector."

[0045] In certain embodiments, an AAV (e.g., rAAV) comprises two ITRs. In certain embodiments, an AAV (e.g., rAAV) comprises a pair of ITRs. In certain embodiments, an AAV (e.g., rAAV) comprises a pair of ITRs adjacent to a nucleic acid sequence encoding a polypeptide having at least a function or activity (i.e., at the 5' end and the 3' end of the nucleic acid sequence, respectively).

[0046] An AAV vector (e.g., a rAAV vector) can be packaged and is referred to herein as an "AAV particle" for infecting (transducing) cells ex vivo, in vitro, or in vivo. When a recombinant AAV vector is enclosed or packaged in an AAV particle, the particle can also be referred to as an "rAAV particle." In certain embodiments, the AAV particle is a rAAV particle. The rAAV particle often comprises an rAAV vector or a portion thereof. The rAAV particle can be one or more rAAV particles (e.g., multiple AAV particles). The rAAV particle typically comprises proteins (e.g., capsid proteins) that enclose or package the rAAV vector genome. It should be noted that reference to a rAAV vector can also be used to refer to a rAAV particle.

[0047] Any suitable AAV particle (e.g., rAAV particle) can be used in the methods or uses herein. The rAAV particle and / or the genome contained therein can be derived from any suitable serotype or strain of AAV. The rAAV particle and / or the genome contained therein can be derived from two or more serotypes or strains of AAV. Thus, the rAAV can contain proteins and / or nucleic acids or portions thereof of any serotype or strain of AAV, and the AAV particle is suitable for infecting and / or transducing mammalian cells. Non-limiting examples of AAV serotypes include AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, AAV-rh74, AAV-rh10, and AAV-2i8.

[0048] In certain embodiments, the plurality of rAAV particles comprises particles of the same strain or serotype (or subgroup or variant), or particles derived from the same strain or serotype (or subgroup or variant). In certain embodiments, the plurality of rAAV particles comprises a mixture of two or more different (e.g., different serotypes and / or different strains) rAAV particles.

[0049] As used herein, the term "serotype" refers to an AAV having a capsid that is serologically distinct from other AAV serotypes. Serological distinctiveness is determined based on the lack of cross-reactivity between antibodies to one AAV compared to another AAV. Such differences in cross-reactivity are usually due to differences in capsid protein sequences / antigenic determinants (e.g., differences in the VP1, VP2, and / or VP3 sequences of AAV serotypes). Although AAV variants, including capsid variants, may not be serologically distinct from a reference AAV serotype or other AAV serotypes, they differ by at least one nucleotide or amino acid residue compared to the reference AAV serotype or other AAV serotypes.

[0050] In certain embodiments, an rAAV vector based on a first serotype genome corresponds to one or more serotypes of the capsid proteins that package the vector. For example, the serotype of one or more AAV nucleic acids (e.g., ITRs) that make up the AAV vector genome corresponds to the serotype of the capsid that makes up the rAAV particle.

[0051] In certain embodiments, the rAAV vector genome can be based on an AAV (e.g., AAV2) serotype genome, in which one or more serotypes of the AAV capsid proteins that package the vector are derived from. For example, the rAAV vector genome can include nucleic acid (e.g., ITR) derived from AAV2, while at least one or more of the three capsid proteins are derived from a different serotype, such as AAV1, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, Rh10, Rh74, or AAV-2i8 serotype or a variant thereof.

[0052] In certain embodiments, an rAAV particle or its vector genome related to a reference serotype has a polynucleotide, polypeptide, or subsequence thereof that comprises or consists of a sequence that is at least 60% or more (e.g., 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc.) identical to a polynucleotide, polypeptide, or subsequence of an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, Rh10, Rh74, or AAV-2i8 particle. In certain embodiments, an rAAV particle or its vector genome related to a reference serotype has a capsid or ITR sequence that comprises or consists of a sequence that is at least 60% or more (e.g., 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc.) identical to the capsid or ITR sequence of an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, Rh10, Rh74, or AAV-2i8 serotype.

[0053] In certain embodiments, the methods herein include the use, administration, or delivery of rAAV1, rAAV2, rAAV3, rAAV4, rAAV5, rAAV6, rAAV7, rAAV8, rAAV9, rAAV10, rAAV11, rAAV12, rRh10, rRh74, or rAAV-2i8 particles.

[0054] In certain embodiments, the methods herein include the use, administration, or delivery of rAAV2 particles. In certain embodiments, the rAAV2 particles include AAV2 capsids. In certain embodiments, the rAAV2 particles include one or more capsid proteins (e.g., VP1, VP2, and / or VP3) that are at least 60%, 65%, 70%, 75% or more identical to the corresponding capsid proteins of native or wild-type AAV2 particles, for example, 80%, 85%, 85%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical. In certain embodiments, the rAAV2 particles comprise VP1, VP2, and VP3 capsid proteins that are at least 75% or more identical to the corresponding capsid proteins of native or wild-type AAV2 particles, e.g., 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical. In certain embodiments, the rAAV2 particles are variants of native or wild-type AAV2 particles. In some aspects, one or more capsid proteins of the AAV2 variant have 1, 2, 3, 4, 5, 5-10, 10-15, 15-20, or more amino acid substitutions compared to the capsid proteins of a native or wild-type AAV2 particle.

[0055] In certain embodiments, the rAAV9 particles comprise an AAV9 capsid. In certain embodiments, the rAAV9 particles comprise one or more capsid proteins (e.g., VP1, VP2, and / or VP3) that are at least 60%, 65%, 70%, 75%, or more identical, e.g., 80%, 85%, 85%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical, to the corresponding capsid protein of a native or wild-type AAV9 particle. In certain embodiments, the rAAV9 particles comprise VP1, VP2, and VP3 capsid proteins that are at least 75% or more identical to the corresponding capsid proteins of native or wild-type AAV9 particles, e.g., 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical. In certain embodiments, the rAAV9 particles are variants of native or wild-type AAV9 particles. In some aspects, one or more capsid proteins of the AAV9 variant have 1, 2, 3, 4, 5, 5-10, 10-15, 15-20, or more amino acid substitutions compared to the capsid proteins of a native or wild-type AAV9 particle.

[0056] In certain embodiments, rAAV particles are designed so that they contain one or more desired ITR functions (e.g., the ability to form a hairpin that allows DNA replication, the ability to integrate AAV into the host cell genome, etc.). and / or AAV-2i8, so long as the ITRs retain the ability to infect the host cell, including one or two ITRs (e.g., a pair of ITRs) that are at least 75% or more identical, e.g., 80%, 85%, 85%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical, to the corresponding ITRs of native or wild-type AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, AAV-rh74, AAV-rh10, or AAV-2i8, so long as the ITRs retain the ability to infect the host cell, including the inclusion of DNA, and / or packaging, if desired.

[0057] In certain embodiments, the rAAV2 particles comprise one or two ITRs (e.g., a pair of ITRs) that are at least 75% or more identical, e.g., 80%, 85%, 85%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical, to the corresponding ITRs of native or wild-type AAV2 particles, so long as they retain one or more desired ITR functions (e.g., the ability to form a hairpin to enable DNA replication, inclusion of the AAV DNA into the host cell genome, and / or packaging, if desired).

[0058] In certain embodiments, the rAAV9 particles comprise one or two ITRs (e.g., a pair of ITRs) that are at least 75% or more identical, e.g., 80%, 85%, 85%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, etc., up to 100% identical, to the corresponding ITRs of native or wild-type AAV2 particles, so long as they retain one or more desired ITR functions (e.g., the ability to form a hairpin to allow DNA replication, inclusion of the AAV DNA into the host cell genome, and / or packaging, if desired).

[0059] rAAV particles can comprise ITRs with any suitable number of "GAGC" repeats. In certain embodiments, the ITRs of AAV2 particles comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 or more "GAGC" repeats. In certain embodiments, the rAAV2 particles comprise ITRs with three "GAGC" repeats. In certain embodiments, the rAAV2 particles comprise ITRs with fewer than four "GAGC" repeats. In certain embodiments, the rAAV2 particles comprise ITRs with more than four "GAGC" repeats. In certain embodiments, the ITRs of rAAV2 particles comprise Rep binding sites in which the fourth nucleotide in the first two "GAGC" repeats is C rather than T.

[0060] Examples of suitable lengths of DNA that can be incorporated into rAAV vectors for packaging / encapsidation into rAAV particles can be about 5 kilobases (kb) or less. In certain embodiments, the length of the DNA is less than about 5 kb, less than about 4.5 kb, less than about 4 kb, less than about 3.5 kb, less than about 3 kb, or less than about 2.5 kb.

[0061] rAAV vectors containing nucleic acid sequences directing the expression of RNAi or polypeptides can be produced using appropriate recombinant techniques known in the art (see, for example, Sambrook et al., 1989). Recombinant AAV vectors are typically packaged into transducible AAV particles and propagated using an AAV viral packaging system. Transducible AAV particles have the ability to bind to and enter mammalian cells and subsequently deliver their nucleic acid cargo (e.g., a heterologous gene) to the nucleus of the cells. Thus, intact transducible rAAV particles are configured to transduce mammalian cells. rAAV particles configured to transduce mammalian cells are often not replicative and require additional protein machinery for self-replication. Therefore, rAAV particles configured to transduce mammalian cells are engineered to bind to and enter mammalian cells and deliver nucleic acids to the cells, with the delivered nucleic acid often being located between a pair of AAV ITRs in the rAAV genome.

[0062] Suitable host cells for producing transducible AAV particles include, but are not limited to, microorganisms, yeast cells, insect cells, and mammalian cells that can be used as recipients of heterologous rAAV vectors or have been used as recipients of heterologous rAAV vectors. The stable human cell line HEK293 (e.g., readily available from the American Type Culture Collection under accession number ATCC CRL1573) can be used. In certain embodiments, modified human embryonic kidney cell lines (e.g., HEK293) transformed with adenovirus type 5 DNA fragments to express the adenovirus E1a and E1b genes are used to produce recombinant AAV particles. The modified HEK293 cell line is easily transfected, making it a particularly convenient platform for producing rAAV particles. Methods for producing high-titer AAV particles capable of transducing mammalian cells are known in the art. For example, AAV particles can be produced as described in Wright, 2008 and Wright, 2009.

[0063] In certain embodiments, AAV helper functions are introduced into host cells by transfecting an AAV helper construct into the host cell before or simultaneously with transfection of the AAV expression vector. Thus, in some cases, AAV helper constructs are used to at least transiently express the AAV rep and / or cap genes to complement missing AAV functions necessary for productive AAV transduction. AAV helper constructs often lack AAV ITRs and are unable to replicate or package themselves. These constructs can take the form of a plasmid, phage, transposon, cosmid, virus, or virion. Several AAV helper constructs have been described, including the commonly used plasmids pAAV / Ad and pIM29+45, which encode both Rep and Cap expression products. Several other vectors encoding Rep and / or Cap expression products are also known.

[0064] An "expression vector" is a specific vector that contains a gene or nucleic acid sequence together with the necessary regulatory regions required for expression in a host cell. Expression vectors contain at least an origin of replication for propagation within a cell and may optionally contain additional elements, such as heterologous nucleic acid sequences, expression control elements (e.g., promoters, enhancers), introns, ITRs, and polyadenylation signals.

[0065] II. Therapeutic Substances In some embodiments, viral gene transfer methods can be used to introduce nucleic acids into mammalian cells or target tissues. Such methods can be used to administer nucleic acids encoding inhibitory RNAs, non-coding RNAs, and / or therapeutic proteins to cells in culture or in a host organism.

[0066] A. Inhibitory RNA "RNA interference (RNAi)" is the process of sequence-specific post-transcriptional gene silencing initiated by siRNA. During RNAi, siRNA induces the degradation of target mRNA, resulting in sequence-specific inhibition of gene expression.

[0067] " Inhibitory RNA ", "RNAi ", "small interfering RNA " or "short interfering RNA " or " siRNA " molecule, "short hairpin RNA " or " shRNA " molecule, or "miRNA " is an RNA duplex of nucleotides that targets a nucleic acid sequence of interest. As used herein, the term " siRNA " is a general term that encompasses a subset of shRNA and miRNA. " RNA duplex " refers to the structure formed by the complementary pairing between two regions of an RNA molecule. The nucleotide sequence of the duplex part of siRNA is complementary to the nucleotide sequence of the target gene, so that siRNA " targets " that gene. In certain embodiments, siRNA targets the sequence encoding Huntington. In some embodiments, the length of the duplex of siRNA is less than 30 base pairs. In some embodiments, the duplex can be 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, or 10 base pairs in length. In some embodiments, the length of the duplex is 19 to 25 base pairs. In certain embodiments, the length of the duplex is 19 or 21 base pairs. The RNA duplex portion of the siRNA can be part of a hairpin structure. The hairpin structure contains, in addition to the duplex portion, a loop portion located between the two sequences forming the duplex. The length of the loop can vary. In some embodiments, the loop is 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides long. In certain embodiments, the loop is 18 nucleotides long. The hairpin structure can also contain a 3' overhang and / or a 5' overhang. In some embodiments, the overhang is 0, 1, 2, 3, 4, or 5 nucleotides long.

[0068] shRNAs are composed of a stem-loop structure designed to contain a 5' flanking region, an siRNA region segment, a loop region, a 3' siRNA region, and a 3' flanking region. Most RNAi expression strategies have used short hairpin RNAs (shRNAs) driven by strong PolIII promoters. Many shRNAs have demonstrated effective knockdown of target sequences both in vitro and in vivo, but some shRNAs that demonstrate effective knockdown of target genes have also been found to be toxic in vivo.

[0069] miRNA is a small cellular RNA (approximately 22nt) that is processed from precursor stem-loop transcript.Known miRNA stem-loop can be modified to contain RNAi sequence specific to gene of interest.miRNA molecule may be preferable to shRNA molecule because miRNA is endogenously expressed.Therefore, miRNA molecule is less likely to induce dsRNA-responsive interferon pathway, is processed more efficiently than shRNA, and has been shown to silence 80% more effectively than shRNA.

[0070] An alternative approach that has recently been discovered is the use of artificial miRNAs (pri-miRNA scaffolds that shuttle siRNA sequences) as RNAi vectors. Artificial miRNAs are more naturally similar to endogenous RNAi substrates and are more amenable to Pol-II transcription (e.g., allowing tissue-specific expression of RNAi) and polycistronic strategies (e.g., allowing delivery of multiple siRNA sequences). See U.S. Patent No. 10,093,927, incorporated herein by reference.

[0071] The transcription unit of an "shRNA" consists of a sense sequence and an antisense sequence connected by a loop of unpaired nucleotides. shRNAs are exported from the nucleus by exportin-5 and, once in the cytoplasm, are processed by Dicer to generate functional siRNAs. The stem-loop of an "miRNA" consists of a sense sequence and an antisense sequence connected by a loop of unpaired nucleotides and is typically expressed as part of a larger primary transcript (pri-miRNA), which is excised by the Drosha-DGCR8 complex to generate an intermediate known as a pre-miRNA, which is then exported from the nucleus by exportin-5 and, once in the cytoplasm, is processed by Dicer to generate functional siRNAs. As used interchangeably herein, "artificial miRNA" or "artificial miRNA shuttle vector" refers to a primary miRNA transcript in which the double-stranded stem-loop region (at least approximately 9-20 nucleotides) excised by Drosha and Dicer processing is replaced with an siRNA sequence for the target gene while preserving the structural elements within the stem-loop necessary for effective Drosha processing. The term "artificial" derives from the fact that the flanking sequences (approximately 35 nucleotides upstream and approximately 40 nucleotides downstream) are derived from restriction enzyme sites within the multiple cloning site of the siRNA. As used herein, the term "miRNA" encompasses both naturally occurring miRNA sequences and artificially engineered miRNA shuttle vectors.

[0072] siRNA can be encoded by a nucleic acid sequence, and this nucleic acid sequence can also include a promoter.This nucleic acid sequence can also include a polyadenylation signal.In some embodiments, the polyadenylation signal is a synthetic minimal polyadenylation signal or a sequence of six T's.

[0073] In the design of RNAi, there are several factors that need to be considered, such as the nature of siRNA, the durability of silencing effect, and the choice of delivery system.To produce RNAi effect, siRNA introduced into organisms typically contains exonic sequences.Furthermore, because the RNAi process depends on homology, the sequence must be carefully selected to maximize gene specificity while minimizing the possibility of cross-interference between homologous but non-gene-specific sequences.Preferably, siRNA exhibits greater than 80%, 85%, 90%, 95%, or 98% identity, or even 100% identity, between the sequence of siRNA and the gene to be inhibited.Sequences that are less than about 80% identical to the target gene are significantly less effective.Therefore, the higher the homology between siRNA and the gene to be inhibited, the less likely the expression of unrelated genes will be affected.

[0074] In addition, the size of the siRNA is also an important consideration. In some embodiments, the present invention relates to siRNA molecules that contain at least about 19 to 25 nucleotides and are capable of modulating gene expression. In the context of the present invention, the siRNA is preferably less than 500, 200, 100, 50, or 25 nucleotides in length. More preferably, the siRNA is about 19 to about 25 nucleotides in length.

[0075] siRNA target generally refers to the polynucleotide that comprises the region that codes for a polypeptide, or the polynucleotide that comprises the region that regulates the replication, transcription or translation or other processes that are important for the expression of a polypeptide, or the polynucleotide that comprises both the region that codes for a polypeptide and the region that is functionally linked to regulate its expression.Any gene that is expressed in cells can be targeted.Preferably, target gene is involved or related to the progression of cellular activity that is important for disease or the progression of cellular activity that is particularly interesting for research.

[0076] B. Non-coding RNA As evidenced by cDNA cloning projects and genome tiling arrays, over 90% of the human genome is transcribed but does not encode proteins. These transcripts are referred to as non-protein-coding RNAs (ncRNAs). Various ncRNA transcripts, such as ribosomal RNAs, transfer RNAs, competitive endogenous RNAs (ceRNAs), small nuclear RNAs (snRNAs), and small nucleolar RNAs (snoRNAs), are essential for cellular function. Similarly, numerous short ncRNAs, such as microRNAs (miRNAs), endogenous short interfering RNAs (siRNAs), PIWI-interacting RNAs (piRNAs), and small nucleolar RNAs (snoRNAs), are also known to play important regulatory roles in eukaryotic cells. Recent studies have demonstrated a group of long ncRNA (lncRNA) transcripts that exhibit cell-type-specific expression and localize to specific subcellular compartments. lncRNAs are also known to play important roles during cellular development and differentiation, supporting the notion that they have been selected for during evolution.

[0077] lncRNAs appear to have many different functions. In many cases, they appear to play a role in regulating protein activity or localization or function as the organizational framework for intracellular structures. In other cases, lncRNAs may be processed to produce multiple small RNAs or may regulate how other RNAs are processed. The latest version of data generated by the public research consortium GenCode (version number 27) catalogs just under 16,000 lncRNAs in the human genome, generating nearly 28,000 transcripts; when other databases are included, more than 40,000 lncRNAs are known.

[0078] Interestingly, lncRNAs can affect the expression of specific target proteins at specific genomic loci, modulate the activity of protein-binding partners, recruit chromatin-modifying complexes to their sites of action, and undergo post-transcriptional processing to generate numerous 5'-capped small RNAs. Epigenetic pathways can also regulate the differential expression of lncRNAs.

[0079] Increasing evidence suggests that aberrantly expressed lncRNAs play important roles in normal physiological processes and multiple disease states. lncRNAs are misregulated in a variety of diseases, including ischemia, cardiac disease, Alzheimer's disease, psoriasis, and spinocerebellar ataxia type 8. This misregulation has also been shown in various types of cancer, such as breast cancer, colon cancer, prostate cancer, hepatocellular carcinoma, and leukemia. Some lncRNAs, such as gadd74 and lncRNA-RoR5, regulate cell cycle regulators such as cyclins, cyclin-dependent kinases (CDKs), CDK inhibitors, and p53, thus providing additional flexibility and robustness to cell cycle progression. Furthermore, some lncRNAs are associated with mitotic processes, such as centromeric satellite RNAs, which are essential for kinetochore formation and therefore important for chromosome segregation during mitosis in humans and flies. Another nuclear lncRNA, MA-lincl, regulates M-phase exit by functioning in cis to repress the expression of the neighboring gene Pura, a regulator of cell proliferation.

[0080] lncRNAs are a group generally defined as transcripts of more than 200 nucleotides (e.g., about 200 to about 1200 nt, about 2500 nt, or more) that lack an extended open reading frame (ORF). The term "non-coding RNA" (ncRNA) includes lncRNAs and short transcripts of less than about 200 nt, e.g., about 30 to 200 nt.

[0081] Thus, in some embodiments, delivery of ncRNA, for example to a specific brain structure of interest, corrects abnormal RNA expression levels or modulates levels of disease-causing lncRNA. Thus, in some embodiments, the present invention provides rAAVs in which the viral genome has been engineered to encode a therapeutic non-coding RNA (ncRNA). In some embodiments, the ncRNA is a long non-coding RNA (lncRNA) about 200 nucleotides (nt) or longer in length. In some embodiments, the therapeutic agent is an ncRNA about 25 nt or about 30 nt to about 200 nt in length. In some embodiments, the lncRNA is about 200 nt to about 1,200 nt in length. In some embodiments, the lncRNA is about 200 nt to about 1,100, about 1,000, about 900, about 800, about 700, about 600, about 500, about 400, or about 300 nt in length.

[0082] C. CRISPR Systems Gene editing is a technique that allows for targeted gene modification within living cells. In recent years, the performance of on-demand gene editing using the bacterial CRISPR immune system has revolutionized the way scientists approach genome editing. The Cas9 protein of the CRISPR system, an RNA-guided DNA endonuclease, can be relatively easily engineered to target new sites by altering its guide RNA sequence. This discovery made sequence-specific gene editing functionally enabled.

[0083] In general, "CRISPR system" refers collectively to transcripts and other elements involved in directing the expression or activity of CRISPR-associated ("Cas") genes, such as sequences encoding Cas genes, tracr (trans-activating CRISPR) sequences (e.g., tracrRNA or active partial tracrRNA), tracr-mate sequences (including "direct repeats" and, in the case of endogenous CRISPR systems, tracrRNA-processed partial direct repeats), guide sequences (also referred to as "spacers" in the case of endogenous CRISPR systems), and / or other sequences and transcripts from a CRISPR locus.

[0084] A CRISPR / Cas nuclease or CRISPR / Cas nuclease system can include a non-coding RNA molecule (guide) RNA that binds to DNA in a sequence-specific manner and a Cas protein (e.g., Cas9) with nuclease functionality (e.g., two nuclease domains). One or more elements of a CRISPR system can be derived from a Type I, Type II, or Type III CRISPR system, and can be derived from a particular organism that contains an endogenous CRISPR system, such as Streptococcus pyogenes.

[0085] As discussed herein, the CRISPR system can induce a double-stranded break (DSB) at the target site followed by a split. In another embodiment, a Cas9 variant considered a "nickase" is used to nick one strand at the target site. For example, to improve specificity, a pair of nickases can be used, each directed by a pair of different gRNAs that target a sequence such that a 5' overhang is introduced when the nicks are introduced simultaneously. In another embodiment, catalytically inactive Cas9 is fused to a heterologous effector domain, such as a transcriptional repressor (e.g., KRAB) or a transcriptional activator, to affect gene expression. Alternatively, the CRISPR system using catalytically inactive Cas9 further comprises a transcriptional repressor or transcriptional activator fused to a ribosome-binding protein.

[0086] In some aspects, a Cas nuclease and a gRNA (comprising a fusion of a target sequence-specific crRNA and an invariant tracrRNA) are introduced into a cell. Generally, a target site at the 5' end of the gRNA targets the Cas nuclease to the target site, e.g., a gene, using complementary base pairing. The target site can be selected based on its location immediately 5' to a protospacer adjacent motif (PAM) sequence, e.g., typically NGG or NAG. In this regard, the gRNA targets the desired sequence by modifying the first 20, 19, 18, 17, 16, 15, 14, 14, 12, 11, or 10 nucleotides of the guide RNA to correspond to the target DNA sequence. Generally, CRISPR systems are characterized by elements that promote the formation of a CRISPR complex at the site of the target sequence. Generally, "target sequence" generally refers to the sequence that guide sequence is designed to have complementarity with, and the hybridization between target sequence and guide sequence promotes the formation of CRISPR complex.Perfect complementarity is not necessarily required, as long as there is sufficient complementarity to cause hybridization and promote the formation of CRISPR complex.

[0087] The target sequence can comprise any polynucleotide, such as a DNA polynucleotide or an RNA polynucleotide. The target sequence can be located in the nucleus or cytoplasm of a cell, such as in a cellular organelle. Generally, a sequence or template that can be used for recombination into a target locus that contains a target sequence is referred to as an "editing template" or an "editing polynucleotide" or an "editing sequence." In some aspects, an exogenous template polynucleotide can be referred to as an editing template. In some aspects, the recombination is homologous recombination.

[0088] Typically, in endogenous CRISPR systems, formation of a CRISPR complex (including a guide sequence hybridized to a target sequence to form a complex with one or more Cas proteins) results in cleavage of one or both strands within or near the target sequence (e.g., within 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 50, or more base pairs of the target sequence). A tracr sequence, which may comprise or consist of all or a portion of a wild-type tracr sequence (e.g., about 20, 26, 32, 45, 48, 54, 63, 67, 85, or more nucleotides of the wild-type tracr sequence, or more than about 20, 26, 32, 45, 48, 54, 63, 67, 85, or more nucleotides of the wild-type tracr sequence), may also form part of a CRISPR complex, for example, by hybridization along at least a portion of the tracr sequence to all or a portion of a tracr mate sequence operably linked to the guide sequence. The tracr sequence has sufficient complementarity to the tracr mate sequence to hybridize and participate in the formation of a CRISPR complex, e.g., at least 50%, 60%, 70%, 80%, 90%, 95%, or 99% sequence complementarity over the entire length of the tracr mate sequence when optimally aligned.

[0089] One or more vectors driving the expression of one or more elements of the CRISPR system can be introduced into cells, so that the expression of these elements of the CRISPR system directs the formation of CRISPR complexes at one or more target sites. Components can also be delivered to cells as proteins and / or RNA. For example, a Cas enzyme, a guide sequence linked to a tracr-mate sequence, and a tracr sequence can each be operably linked to separate regulatory elements on separate vectors. The Cas enzyme can be a target gene that is controlled by regulatory alternative splicing events, as disclosed herein as a chimeric target gene minigene or as a target gene for a chimeric minigene transactivator. The gRNA can be controlled by a constitutive promoter.

[0090] Alternatively, two or more elements expressed from the same or different regulatory elements may be combined in one vector, and one or more additional vectors may provide any components of the CRISPR system that are not included in the first vector. The vector may contain one or more insertion sites, such as restriction endonuclease recognition sequences (also referred to as "cloning sites"). In some embodiments, the one or more insertion sites are located upstream and / or downstream of one or more sequence elements of one or more vectors. The use of multiple different guide sequences allows a single expression construct to target CRISPR activity to multiple corresponding target sequences in a cell.

[0091] The vector may include regulatory elements operably linked to an enzyme coding sequence encoding a CRISPR enzyme, such as a Cas protein, non-limiting examples of which include Cas1, Cas1B, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, and Cas9. (also known as Csn1 and Csx12), Cas10, Csy1, Csy2, Csy3, Csel, Cse2, Csc1, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmr1, Cmr3, Cmr4, Cmr5, Cmr6, Csb1, Csb2, Csb3, Csx17, Csx14, Csx10, Csx16, CsaX, Csx3, Csx1, Csx15, Csfl, Csf2, Csf3, Csf4, homologs thereof, or modified forms thereof. These enzymes are known. For example, the amino acid sequence of the S. pyogenes Cas9 protein can be found in the SwissProt database under accession number Q99ZW2.

[0092] The CRISPR enzyme can be Cas9 (e.g., from Streptococcus pyogenes or S. pneumoniae). CRISPR enzymes can direct cleavage of one or both strands at the location of a target sequence, e.g., within the target sequence and / or within the complementary strand of the target sequence. The vector can encode a mutated CRISPR enzyme such that, compared to the corresponding wild-type enzyme, the mutant CRISPR enzyme lacks the ability to cleave one or both strands of a target polynucleotide containing the target sequence. For example, an aspartic acid to alanine substitution (D10A) in the RuvC I catalytic domain of Cas9 from Streptococcus pyogenes converts Cas9 from a nuclease that cleaves both strands to a nickase (one that cleaves one strand). In some embodiments, Cas9 nickase can be used in combination with a guide sequence, e.g., two guide sequences that target the sense and antisense strands of a DNA target, respectively. This combination allows both strands to be nicked and used to induce NHEJ or HDR.

[0093] In some embodiments, the enzyme coding sequence encoding the CRISPR enzyme is codon-optimized for expression in a specific cell, such as a eukaryotic cell. The eukaryotic cell may be or be derived from a specific organism, such as a mammal, including but not limited to, a human, mouse, rat, rabbit, dog, or non-human primate. Generally, codon optimization refers to the process of modifying a nucleic acid sequence to enhance expression in a target host cell by replacing at least one codon of the native sequence with a codon that is more frequently or most frequently used in the host cell's genes while maintaining the native amino acid sequence. Different species exhibit specific biases toward certain codons for specific amino acids. Codon bias (differences in codon usage between organisms) is often correlated with the translation efficiency of messenger RNA (mRNA), and is believed to depend, among other things, on the characteristics of the codon being translated and the availability of specific transfer RNA (tRNA) molecules. The dominance of a selected tRNA in a cell generally reflects the codon most frequently used in peptide synthesis. Thus, genes can be tailored for optimal gene expression in a given organism based on codon optimization.

[0094] Generally, a guide sequence is any polynucleotide sequence that has sufficient complementarity with a target polynucleotide sequence to hybridize with the target sequence and direct the sequence-specific binding of a CRISPR complex to the target sequence. In some embodiments, the degree of complementarity between a guide sequence and its corresponding target sequence is about 50%, 60%, 75%, 80%, 85%, 90%, 95%, 97.5%, 99% or more, or greater than about 50%, 60%, 75%, 80%, 85%, 90%, 95%, 97.5%, 99% or more, when optimally aligned using a suitable alignment algorithm.

[0095] Optimal alignment can be determined using any suitable algorithm for aligning sequences, non-limiting examples of such algorithms include the Smith-Waterman algorithm, the Needleman-Wunsch algorithm, algorithms based on the Burrows-Wheeler transform (e.g., Burrows Wheeler Aligner), Clustal W, Clustal X, BLAT, Novoalign (Novocraft Technologies), ELAND (Illumina, San Diego, CA), SOAP (available at soap.genomics.org.cn), and Maq (available at maq.sourceforge.net).

[0096] CRISPR enzymes may be part of a fusion protein containing one or more heterologous protein domains. CRISPR enzyme fusion proteins may contain any additional protein sequences and, optionally, a linker sequence between any two domains. Examples of protein domains that can be fused to CRISPR enzymes include, but are not limited to, epitope tags, reporter gene sequences, and protein domains with one or more of the following activities: methylase activity, demethylase activity, transcription activation activity, transcription repression activity, transcription termination factor activity, histone modification activity, RNA cleavage activity, and nucleic acid binding activity. Non-limiting examples of epitope tags include histidine (His) tags, V5 tags, FLAG tags, influenza hemagglutinin (HA) tags, Myc tags, VSV-G tags, and thioredoxin (Trx) tags. Examples of reporter genes include, but are not limited to, glutathione-5-transferase (GST), horseradish peroxidase (HRP), chloramphenicol acetyltransferase (CAT), β-galactosidase, β-glucuronidase, luciferase, green fluorescent protein (GFP), HcRed, DsRed, cyan fluorescent protein (CFP), yellow fluorescent protein (YFP), and autofluorescent proteins including blue fluorescent protein (BFP). CRISPR enzymes can be fused to gene sequences encoding proteins or fragments of such proteins that bind to DNA molecules or other cellular molecules, such as, but not limited to, maltose binding protein (MBP), S-tag, Lex A DNA binding domain (DBD) fusions, GAL4A DNA binding domain fusions, and herpes simplex virus (HSV) BP16 protein fusions. Further domains that can form part of fusion proteins comprising CRISPR enzymes are described in US 20110059502, which is incorporated herein by reference.

[0097] D. Therapeutic Proteins Some embodiments relate to the expression of recombinant proteins and polypeptides. In some aspects, proteins or polypeptides may be modified to increase serum stability. Thus, when the present application refers to the function or activity of a "modified protein" or "modified polypeptide," it will be understood by those skilled in the art that this encompasses proteins or polypeptides that have additional advantages over, for example, the unmodified protein or polypeptide. It is specifically contemplated that embodiments relating to "modified proteins" can also be implemented with respect to "modified polypeptides," and vice versa.

[0098] Recombinant proteins may possess amino acid deletions and / or substitutions. Thus, proteins with deletions, proteins with substitutions, and proteins with deletions and substitutions are modified proteins. In some embodiments, these proteins may further comprise inserted or added amino acids, such as fusion proteins or proteins with linkers. A "modified deletion protein" lacks one or more residues of the native protein but may retain the specificity and / or activity of the native protein. A "modified deletion protein" may also have reduced immunogenicity or antigenicity. An example of a modified deletion protein is one in which amino acid residues are deleted from at least one antigenic region, i.e., from a region of the protein determined to be antigenic in a particular organism, e.g., the organism to which the modified protein is administered.

[0099] Substitution or replacement variants typically contain the exchange of one amino acid for another at one or more sites within the protein and can be designed to modulate one or more properties of the polypeptide, particularly its effector functions and / or bioavailability. Substitutions may or may not be conservative, i.e., one amino acid is replaced with one of similar shape and charge. Conservative substitutions are well known in the art and include, for example, alanine to serine, arginine to lysine, asparagine to glutamine or histidine, aspartic acid to glutamic acid, cysteine ​​to serine, glutamine to asparagine, glutamic acid to aspartic acid, glycine to proline, histidine to asparagine or glutamine, isoleucine to leucine or valine, leucine to valine or isoleucine, lysine to arginine, methionine to leucine or isoleucine, phenylalanine to tyrosine, leucine or methionine, serine to threonine, threonine to serine, tryptophan to tyrosine, tyrosine to tryptophan or phenylalanine, and valine to isoleucine or leucine.

[0100] In addition to deletions or substitutions, modified proteins may also contain residue insertions. This typically involves the addition of at least one residue in the polypeptide. This may include the insertion of a targeting peptide or targeting polypeptide, or simply the insertion of a single residue. Terminal additions, called fusion proteins, are discussed below.

[0101] The term "biologically functional equivalent" is well understood in the art and is further described herein. Thus, sequences in which about 70% to about 80%, or about 81% to about 90%, or even about 91% to about 99% of the amino acids are identical or functionally equivalent to those of a reference polypeptide are encompassed, so long as the biological activity of the protein is maintained. A recombinant protein may, in certain respects, be a biologically functional equivalent to the corresponding native protein.

[0102] It will also be understood that amino acid and nucleic acid sequences can include additional residues, such as additional N- or C-terminal amino acids, or 5' or 3' sequences, and still be essentially as set forth in one of the sequences disclosed herein, so long as the sequence meets the above criteria, including, where protein expression is concerned, maintenance of biological protein activity. The addition of terminal sequences is particularly applicable to nucleic acid sequences, which may include, for example, various non-coding sequences adjacent to either the 5' or 3' portion of the coding region, or various internal sequences, i.e., introns, that are known to be present within genes.

[0103] As used herein, a protein or peptide generally refers to, but is not limited to, a protein of more than about 200 amino acids, up to the full-length sequence, translated from a gene; a polypeptide of more than about 100 amino acids; and / or a peptide of about 3 to about 100 amino acids. For convenience, the terms "protein," "polypeptide," and "peptide" are used interchangeably herein.

[0104] As used herein, "amino acid residue" refers to any naturally occurring amino acid, any amino acid derivative, or any amino acid mimic known in the art. In certain embodiments, the residues of a protein or peptide are contiguous, without any non-amino acid sequences interrupting the sequence of amino acid residues. In other embodiments, the sequence may contain one or more non-amino acid moieties. In certain embodiments, the sequence of residues of a protein or peptide may be interrupted by one or more non-amino acid moieties.

[0105] Thus, the term "protein or peptide" encompasses amino acid sequences that include at least one of the 20 common amino acids found in naturally occurring proteins, or at least one modified or unusual amino acid.

[0106] Certain aspects of the present invention relate to fusion proteins. These molecules can have a therapeutic protein linked at the N- or C-terminus to a heterologous domain. For example, fusions can also use leader sequences from other species to allow recombinant expression of the protein in a heterologous host. Other useful fusions include the addition of a protein affinity tag, preferably cleavable, such as a serum albumin affinity tag or six histidine residues, or an immunologically active domain, such as an antibody epitope, to facilitate protein purification. Non-limiting examples of affinity tags include polyhistidine, chitin-binding protein (CBP), maltose-binding protein (MBP), and glutathione-S-transferase (GST).

[0107] Methods for making fusion proteins are well known to those skilled in the art. Such proteins can be produced, for example, by de novo synthesis of an entire fusion protein, or by the attachment of a DNA sequence encoding a heterologous domain followed by expression of the intact fusion protein.

[0108] The production of fusion proteins that restore the functional activity of the parent protein can be facilitated by ligating the genes with a bridging DNA segment encoding a peptide linker joined between the tandemly linked polypeptides, the linker being of sufficient length to allow proper folding of the resulting fusion protein.

[0109] III. Method of Administration In some aspects, viral vectors can be administered directly to patients (in vivo), or can be used to treat cells in vitro or ex vivo and then administered to patients. The term "vector" refers to a small carrier nucleic acid molecule, a plasmid, a virus (e.g., AAV vector, retroviral vector, lentiviral vector), or other vehicle that can be manipulated by inserting or incorporating a nucleic acid. Vectors such as viral vectors can be used to introduce / transfer nucleic acids into cells so that the nucleic acid sequence within the nucleic acid is transcribed by the cell and, if it encodes a protein, subsequently translated.

[0110] Any suitable cell or mammal can be administered or treated by the methods or uses described herein. Typically, a mammal in need of the methods described herein is suspected of having or expressing an abnormal or aberrant protein associated with a disease state. Alternatively, the recipient mammal may have a condition suitable for gene replacement therapy. As used herein, "gene replacement therapy" refers to the administration of exogenous genetic material encoding a therapeutic agent to a recipient and the subsequent in situ expression of the administered genetic material. Thus, the phrase "condition suitable for gene replacement therapy" encompasses conditions such as genetic diseases (i.e., disease states resulting from one or more gene defects), acquired pathologies (i.e., pathological conditions not resulting from congenital defects), cancer, and preventative processes (i.e., prevention of disease or undesirable medical conditions). Thus, as used herein, the term "therapeutic agent" refers to any agent or material that has a beneficial effect on a recipient mammal. Thus, "therapeutic agent" encompasses both therapeutic and preventative molecules having nucleic acid or protein components.

[0111] Non-limiting examples of mammals include humans, non-human primates (such as apes, gibbons, chimpanzees, orangutans, monkeys, macaques, etc.), livestock (e.g., dogs and cats), farm animals (e.g., horses, cows, goats, sheep, pigs), and laboratory animals (e.g., mice, rats, rabbits, guinea pigs). In certain embodiments, the mammal is a human. In certain embodiments, the mammal is a non-rodent mammal (e.g., a human, pig, goat, sheep, horse, dog, etc.). In certain embodiments, the non-rodent mammal is a human. The mammal can be of any age or at any stage of development (e.g., an adult, teenager, child, infant, or mammal in utero). The mammal can be male or female. In certain embodiments, the mammal can be an animal disease model, e.g., an animal model having or expressing an aberrant or abnormal protein associated with a disease state, or an animal model in which insufficient expression of a protein causes a disease.

[0112] Mammals (subjects) treated with the methods or compositions described herein include adults (18 years of age or older) and children (under 18 years of age). Adults include the elderly. A typical adult is 50 years of age or older. Children range in age from 1-2 years or 2-4 years, 4-6 years, 6-18 years, 8-10 years, 10-12 years, 12-15 years, and 15-18 years. Children also include infants. Infants are typically in the 1-12 month age range.

[0113] In certain embodiments, the method comprises administering a plurality of viral particles to a mammal as described herein to reduce, reduce, prevent, inhibit, or delay the severity, frequency, progression, or time to onset of one or more symptoms of a disease state, such as a neurodegenerative disease. In certain embodiments, the method comprises administering a plurality of viral particles to a mammal to treat adverse symptoms of a disease state, such as a neurodegenerative disease. In certain embodiments, the method comprises administering a plurality of viral particles to a mammal to stabilize, delay, or prevent the worsening or progression or reversal of a disease state, such as a neurodegenerative disease, and adverse symptoms.

[0114] In certain embodiments, the method includes administering a plurality of viral particles to the central nervous system or a portion thereof, as described herein, to reduce, decrease, prevent, inhibit, or delay by at least about 5 to about 10 days, about 10 to about 25 days, about 25 to about 50 days, or about 50 to about 100 days the severity, frequency, progression, or time to onset of one or more symptoms of a disease state, such as a neurodegenerative disease.

[0115] In certain embodiments, the symptoms or adverse effects include early, intermediate or late symptoms, behavioral, personality or language symptoms, swallowing, movement, seizures, tremors or restlessness, ataxia, and / or cognitive symptoms such as memory and ability to organize.

[0116] IV. Pharmaceutical Compositions As used herein, the terms "pharmaceutically acceptable" and "physiologically acceptable" refer to a biologically acceptable composition, formulation, liquid or solid, or mixture thereof, suitable for one or more routes of administration, in vivo delivery, or in vivo contact. A "pharmaceutically acceptable" or "physiologically acceptable" composition is a material that is not biologically or otherwise undesirable; e.g., the material can be administered to a subject without causing substantial undesirable biological effects. Such compositions, "pharmaceutically acceptable" and "physiologically acceptable" formulations and compositions can be sterile. Such pharmaceutical formulations and compositions can be used, for example, when administering viral particles to a subject.

[0117] Such formulations and compositions include solvents (aqueous or non-aqueous), solutions (aqueous or non-aqueous), emulsions (e.g., oil-in-water or water-in-oil), suspensions, syrups, elixirs, dispersion and suspending media, coatings, isotonicity adjusting and absorption enhancing or absorption delaying agents that are compatible with pharmaceutical administration or in vivo contact or delivery. Aqueous and non-aqueous solvents, solutions and suspensions may include suspending agents and thickening agents. Supplementary active compounds (e.g., preservatives, antibacterial, antiviral and antifungal agents) can also be incorporated into the formulations and compositions.

[0118] Pharmaceutical compositions typically contain a pharmaceutically acceptable excipient. Such excipients include any pharmaceutical agent that does not itself induce the production of antibodies harmful to the individual receiving the composition and that may be administered without undue toxicity. Pharmaceutically acceptable excipients include, but are not limited to, sorbitol, Tween 80, and liquids such as water, saline, glycerol, and ethanol. Pharmaceutically acceptable salts may also be included, such as mineral acid salts such as hydrochlorides, hydrobromides, phosphates, and sulfates, and salts of organic acids such as acetates, propionates, malonates, and benzoates. Additionally, auxiliary substances, such as surfactants, wetting or emulsifying agents, pH buffering substances, and the like, may also be present in such vehicles.

[0119] Pharmaceutical compositions can be formulated to be compatible with particular routes of administration or delivery described herein or known to those skilled in the art, and thus include carriers, diluents, or excipients suitable for administration or delivery by various routes.

[0120] Pharmaceutical forms suitable for injection of virus particles include sterile aqueous solutions or dispersions suitable for the extemporaneous preparation of sterile injectable or infusible solutions or dispersions, optionally encapsulated in liposomes. In all cases, the final dosage form should be sterile, fluid, and stable under the conditions of manufacture, use, and storage. Liquid carriers or vehicles can be solvents or liquid dispersion media containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), vegetable oils, non-toxic glyceryl esters, and suitable mixtures thereof. Proper fluidity can be maintained, for example, by the formation of liposomes, by maintaining the required particle size in the case of dispersions, or by the use of surfactants. Isotonicity agents, such as sugars, buffers, or salts (e.g., sodium chloride), can be included. Prolonged absorption of injectable compositions can be achieved by using agents delaying absorption, such as aluminum monostearate and gelatin, in the composition.

[0121] The solution or suspension of viral particles may optionally contain one or more of the following components: sterile diluents, such as water for injection, saline solutions, such as phosphate-buffered saline (PBS), artificial CSF, surfactants, fixed oils, polyols (such as glycerol, propylene glycol, and liquid polyethylene glycol), glycerin, or other synthetic solvents, antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, ascorbic acid, and the like; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid; buffers, such as acetates, citrates, or phosphates, and agents for adjusting tonicity, such as sodium chloride or dextrose.

[0122] Pharmaceutical formulations, compositions and delivery systems suitable for the compositions, methods and uses of the present invention are known in the art (see, e.g., Remington: The Science and Practice of Pharmacy (2003) 20 th ed., Mack Publishing Co., Easton, PA; Remington's Pharmaceutical Sciences (1990) 18 th ed., Mack Publishing Co., Easton, PA; The Merck Index (1996) 12 th ed., Merck Publishing Group, Whitehouse, NJ; Pharmaceutical Principles of Solid Dosage Forms (1993), Technonic Publishing Co., Inc., Lancaster, Pa.; Ansel and Stoklosa, Pharmaceutical Calculations (2001) 11 th ed., Lippincott Williams & Wilkins, Baltimore, MD; and Poznansky et al., Drug Delivery Systems (1980), R.L. Juliano, ed., Oxford, NY, pp. 253-315).

[0123] Viral particles and their compositions can be formulated into dosage unit form to facilitate administration and ensure uniformity of dosage.As used herein, dosage unit form refers to a physically discrete unit suitable as a unitary dosage for an individual to be treated, each unit containing a predetermined amount of active compound calculated to produce a desired therapeutic effect together with the necessary pharmaceutical carrier.The dosage unit form depends on the number of viral particles that are considered necessary to produce a desired effect.The required amount can be formulated in a single administration or in multiple dosage units.The dosage can be adjusted to an appropriate viral particle concentration, optionally combined with an anti-inflammatory agent, and packaged for use.

[0124] In one embodiment, the pharmaceutical composition will contain sufficient genetic material to provide a therapeutically effective amount, i.e., an amount sufficient to reduce or ameliorate the symptoms or deleterious effects of the disease state in question, or an amount sufficient to provide the desired benefit.

[0125] As used herein, the term "unit dosage form" refers to a physically discrete unit suitable as a unitary dosage for a subject to be treated, each unit containing a predetermined amount calculated to produce a desired effect (e.g., a prophylactic or therapeutic effect) when administered one or more times, optionally together with a pharmaceutical carrier (excipient, diluent, vehicle, or filler). Unit dosage forms may be contained, for example, in ampoules and vials that may contain liquid compositions or compositions in a freeze-dried or lyophilized state, to which, for example, a sterile liquid carrier can be added prior to in vivo administration or delivery. Individual unit dosage forms may be included in multi-dose kits or containers. Thus, for example, viral particles and pharmaceutical compositions thereof can be packaged in single or multiple unit dosage forms for ease of administration and uniformity of dosage.

[0126] Formulations containing viral particles typically contain an effective amount, which can be easily determined by one skilled in the art. The viral particles can typically range from about 1% to about 95% (w / w) of the composition, or even higher if appropriate. The amount administered depends on factors such as the age, weight, and health of the mammalian or human subject being treated. Those skilled in the art can establish effective dosages through routine testing to establish dose-response curves.

[0127] V. Definition The terms "polynucleotide," "nucleic acid," and "transgene" are used interchangeably herein to refer to all forms of nucleic acid, oligonucleotide, including deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) and polymers thereof. Polynucleotides include genomic DNA, cDNA, and antisense DNA, as well as spliced ​​and unspliced ​​mRNA, rRNA, tRNA, and inhibitory DNA or RNA (RNAi, e.g., small or short hairpin (sh) RNA, microRNA (miRNA), small or short interfering (si) RNA, trans-splicing RNA, or antisense RNA). Polynucleotides can include natural, synthetic, and intentionally modified or altered polynucleotides (e.g., variant nucleic acids). Polynucleotides can be single-stranded, double-stranded, or triplexed, linear, or circular, and can be of any suitable length. When discussing polynucleotides, the sequence or structure of a particular polynucleotide may be described herein according to the convention of describing the sequence in the 5' to 3' direction.

[0128] A nucleic acid that encodes a polypeptide often includes an open reading frame that encodes the polypeptide. Unless otherwise indicated, a particular nucleic acid sequence also includes degenerate codon substitutions.

[0129] The nucleic acid can include one or more expression control or expression regulatory elements operably linked to the open reading frame, which one or more regulatory elements are configured to direct the transcription and translation of the polypeptide encoded by the open reading frame in mammalian cells. Non-limiting examples of expression control / regulatory elements include transcription initiation sequences (e.g., promoters, enhancers, TATA boxes, etc.), translation initiation sequences, mRNA stability sequences, polyA sequences, secretion sequences, etc. Expression control / regulatory sequences can be obtained from the genome of any suitable organism.

[0130] A "promoter" refers to a nucleotide sequence, usually located upstream (5') of a coding sequence, that directs and / or controls the expression of the coding sequence by providing recognition sites for RNA polymerase and other factors required for proper transcription. Pol II promoters contain a minimal promoter, a short DNA sequence consisting of a TATA box and, optionally, other sequences that serve to specify the transcription start site, to which regulatory elements are added to control expression. Type 1 Pol III promoters contain three cis-acting sequence elements downstream of the transcription start site: a) a 5' sequence element (A block); b) an intermediate sequence element (I block); and c) a 3' sequence element (C block). Type 2 Pol III promoters contain two essential cis-acting sequence elements downstream of the transcription start site: a) an A box (5' sequence element); and b) a B box (3' sequence element). Type 3 Pol III promoters contain several cis-acting promoter elements upstream of the transcription start site, including a conventional TATA box, a proximal sequence element (PSE), and a distal sequence element (DSE).

[0131] An "enhancer" is a DNA sequence that can stimulate transcriptional activity and can be a promoter-specific or heterologous element that enhances the level or tissue specificity of expression. It can operate in either orientation (5'→3' or 3'→5') and has the ability to function whether placed upstream or downstream of a promoter.

[0132] A promoter and / or enhancer may be derived in its entirety from a native gene, may be composed of different elements derived from different elements found in nature, or may even be composed of synthetic DNA segments. A promoter or enhancer may contain DNA sequences that are involved in the binding of protein factors that regulate / control the effectiveness of transcription initiation in response to stimuli, physiological or developmental conditions.

[0133] Non-limiting examples of promoters include the SV40 early promoter, mouse mammary tumor virus LTR promoter, adenovirus major late promoter (Ad MLP), herpes simplex virus (HSV) promoter, cytomegalovirus (CMV) promoter, such as the CMV immediate-early promoter region (CMVIE), Rous sarcoma virus (RSV) promoter, pol II promoter, pol III promoter, synthetic promoters, hybrid promoters, and the like. Additionally, sequences derived from non-viral genes, such as the mouse metallothionein gene, are also considered useful herein. Exemplary constitutive promoters include promoters of the following genes encoding certain constitutive or "housekeeping" functions, as well as other constitutive promoters known to those skilled in the art: hypoxanthine phosphoribosyltransferase (HPRT), dihydrofolate reductase (DHFR), adenosine deaminase, phosphoglycerol kinase (PGK), pyruvate kinase, phosphoglycerol mutase, actin promoter, and U6. Additionally, many viral promoters function constitutively in eukaryotic cells. These include, among others, the early and late promoters of SV40, the long terminal repeats (LTRs) of Moloney leukemia virus and other retroviruses, and the thymidine kinase promoter of herpes simplex virus.In addition, sequences derived from intronic miRNA promoters, such as miR107, miR206, miR208b, miR548f-2, miR569, miR590, miR566, and miR128 promoters, may also find use herein (see, for example, Monteys et al., 2010).Therefore, any of the above-mentioned constitutive promoters can be used to control the transcription of heterologous gene inserts.

[0134] "Transgene" is used herein for convenience to refer to a nucleic acid sequence / polynucleotide that is or has been introduced into a cell or organism. Transgenes include any nucleic acid, such as an inhibitory RNA or a gene encoding a polypeptide or protein, which is generally heterologous to the native AAV genome sequence.

[0135] The term "transduction" refers to the introduction of a nucleic acid sequence into a cell or host organism by a vector (e.g., a viral particle). Thus, the introduction of a transgene into a cell by a viral particle can be referred to as "transduction" of the cell. The transgene may or may not be integrated into the genomic nucleic acid of the transduced cell. If the introduced transgene is integrated into the nucleic acid (genomic DNA) of the recipient cell or recipient organism, it can be stably maintained in the cell or organism and further transmitted or inherited by the recipient cell's or recipient organism's progeny cells or progeny organisms. Finally, the introduced transgene may be present extrachromosomally or only transiently in the recipient cell or recipient host organism. Therefore, a "transduced cell" is a cell into which a transgene has been introduced by transduction. Thus, a "transduced" cell is a cell into which a transgene has been introduced or its progeny. The transduced cell can be propagated to transcribe the transgene and express the encoded inhibitory RNA or protein. For gene therapy uses and methods, the transduced cell can be present in a mammal.

[0136] A transgene under the control of an inducible promoter is expressed only in the presence of an inducer, or is expressed more strongly in the presence of an inducer (e.g., transcription under the control of a metallothionein promoter is significantly increased in the presence of certain metal ions). Inducible promoters contain a responsive element (RE) that stimulates transcription when the respective inducer binds. Examples include REs for serum factors, steroid hormones, retinoic acid, and cyclic AMP. A promoter containing a specific RE can be selected to obtain an inducible response, and in some cases, the RE itself can be linked to a different promoter to confer inducibility to the recombinant gene. Thus, by selecting the appropriate promoter (constitutive or inducible, strong or weak), it is possible to control both the presence and expression level of a polypeptide in genetically engineered cells. When a gene encoding a polypeptide is under the control of an inducible promoter, delivery of the polypeptide in situ can be triggered by exposing the genetically engineered cells in situ to conditions that allow transcription of the polypeptide, e.g., by intraperitoneal injection of a specific inducer of the inducible promoter that controls transcription of the agent. For example, in situ expression by genetically modified cells of polypeptides encoded by genes under the control of a metallothionein promoter is enhanced by contacting the genetically modified cells in situ with a solution containing appropriate (i.e., inducible) metal ions.

[0137] A nucleic acid / transgene is "operably linked" when it is placed into a functional relationship with another nucleic acid sequence. The nucleic acid / transgene encoding an RNAi or polypeptide or the nucleic acid directing the expression of the polypeptide may contain an inducible promoter or a tissue-specific promoter to control transcription of the encoded polypeptide. A nucleic acid operably linked to an expression control element may also be referred to as an expression cassette.

[0138] In certain embodiments, the methods and uses described herein use CNS-specific or inducible promoters, enhancers, etc. Non-limiting examples of CNS-specific promoters include those isolated from the genes of myelin basic protein (MBP), glial fibrillary acidic protein (GFAP), and neuron-specific enolase (NSE). Non-limiting examples of inducible promoters include DNA responsive elements to ecdysone, tetracycline, hypoxia, and IFN.

[0139] In certain embodiments, the expression control element comprises a CMV enhancer. In certain embodiments, the expression control element comprises a β-actin promoter. In certain embodiments, the expression control element comprises a chicken β-actin promoter. In certain embodiments, the expression control element comprises a CMV enhancer and a chicken β-actin promoter.

[0140] As used herein, the term " modified " or " variant " and its grammatical variants refer to nucleic acid, polypeptide or its subsequence that deviates from reference sequence.Therefore, the expression level, activity or function of modified and variant sequences may be substantially the same, greater or less than that of reference sequence, but at least partially retain the activity or function of reference sequence.A specific type of variant is mutant protein, which refers to the protein encoded by a gene that has mutations such as missense mutation or nonsense mutation.

[0141] A "nucleic acid" variant or "polynucleotide" variant refers to a modified sequence in which genes are altered compared to the wild-type. A sequence can be genetically altered without changing the encoded protein sequence. Alternatively, a sequence can be genetically altered to encode a variant protein. A nucleic acid variant or polynucleotide variant can also refer to a combination sequence in which codons have been altered to encode a protein that still retains at least partial sequence identity to a reference sequence, such as a wild-type protein sequence, and also to encode a variant protein. For example, some codons in such a nucleic acid variant are altered in a way that does not change the amino acids of the protein encoded thereby, and some codons in the nucleic acid variant are altered, resulting in a change in the amino acids of the protein it encodes.

[0142] The terms "protein" and "polypeptide" are used interchangeably herein. The "polypeptide" encoded by the "nucleic acid" or "polynucleotide" or "transgene" disclosed herein includes partial or full-length native sequences, as well as naturally occurring wild-type and functional polymorphic proteins, their functional subsequences (fragments), and sequence variants thereof, so long as the polypeptide retains some function or activity. Thus, in the methods and uses of the present invention, such polypeptides encoded by nucleic acid sequences need not be identical to endogenous proteins that are defective, or whose activity, function, or expression is insufficient, deficient, or absent in the mammal being treated.

[0143] Non-limiting examples of modifications include substitution of one or more nucleotides or amino acids (e.g., about 1 to about 3, about 3 to about 5, about 5 to about 10, about 10 to about 15, about 15 to about 20, about 20 to about 25, about 25 to about 30, about 30 to about 40, about 40 to about 50, about 50 to about 100, about 100 to about 150, about 150 to about 200, about 200 to about 250, about 250 to about 500, about 500 to about 750, about 750 to about 1000, or more nucleotides or residues).

[0144] Examples of amino acid modifications are conservative amino acid substitutions or deletions. In certain embodiments, the modified or variant sequence retains at least some of the function or activity of the unmodified sequence (e.g., the wild-type sequence).

[0145] Another example of an amino acid modification is a targeting peptide introduced into the capsid protein of the viral particle. Peptides have been identified that target recombinant viral vectors to the central nervous system, for example, to different brain regions.

[0146] Such modified recombinant viruses may preferentially bind to one type of tissue (e.g., CNS tissue) over another type of tissue (e.g., liver tissue). In certain embodiments, recombinant viruses bearing modified capsid proteins can "target" cerebrovascular epithelial tissue by binding at a higher level than comparable unmodified capsid proteins. For example, recombinant viruses with modified capsid proteins may bind to cerebrovascular epithelial tissue at a level 50% to 100% higher than unmodified recombinant viruses.

[0147] A "nucleic acid fragment" is a portion of a given nucleic acid molecule. In most organisms, deoxyribonucleic acid (DNA) is the genetic material, while ribonucleic acid (RNA) is responsible for transferring the information contained in DNA into proteins. Fragments and variants of the disclosed nucleotide sequences and the proteins or partial-length proteins encoded thereby are also encompassed by the present invention. By "fragment" or "portion" is meant a full-length or less-than-full-length nucleotide sequence encoding a polypeptide or protein or an amino acid sequence of a polypeptide or protein. In certain embodiments, the fragment or portion is biologically functional (i.e., retains 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of wild-type activity or function).

[0148] A "variant" of a molecule is a sequence that is substantially similar to that of the native molecule. In the case of nucleotide sequences, variants include sequences that, due to the degeneracy of the genetic code, encode the same amino acid sequence as the native protein. Such natural allelic variants can be identified using molecular biology techniques, such as polymerase chain reaction (PCR) and hybridization techniques. Variant nucleotide sequences also include synthetically derived nucleotide sequences, such as those generated using site-directed mutagenesis, that encode native proteins as well as those that encode polypeptides with amino acid substitutions. Generally, nucleotide sequence variants of the invention will have at least 40%, 50%, 60% or 70%, for example 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78% or 79%, typically at least 80%, for example 81% to 84%, at least 85%, for example 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97% or 98% sequence identity to the native (endogenous) nucleotide sequence. In certain embodiments, the variant is biologically functional (i.e., retains 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 99%, or 100% of the wild-type activity or function).

[0149] "Conservative substitutions" of a particular nucleic acid sequence refer to nucleic acid sequences that encode identical or essentially identical amino acid sequences. Because the genetic code is degenerate, any given polypeptide can be encoded by a large number of functionally identical nucleic acids. For example, the codons CGT, CGC, CGA, CGG, AGA, and AGG all encode the amino acid arginine. Thus, wherever arginine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded protein. Such nucleic acid variations are "silent variations," which are a species of "conservatively modified variations." All nucleic acid sequences described herein that encode polypeptides represent all possible silent variations, unless otherwise noted. Those skilled in the art will recognize that each codon in a nucleic acid (except ATG, which is usually the only methionine codon) can be altered using standard techniques to result in a functionally identical molecule. Thus, each "silent variation" of a nucleic acid that encodes a polypeptide is implicit in each described sequence.

[0150] The term "substantial identity" of a polynucleotide sequence means that the polynucleotide comprises a sequence having at least 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, or 79%, or at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, or 89%, or at least 90%, 91%, 92%, 93%, or 94%, or even at least 95%, 96%, 97%, 98%, or 99% sequence identity when compared to a reference sequence using one of the alignment programs described with standard parameters. Those skilled in the art will understand that these values ​​can be adjusted appropriately to determine the corresponding identity of proteins encoded by two nucleotide sequences, taking into account codon degeneracy, amino acid similarity, reading frame position, etc. For these purposes, substantial amino acid sequence identity typically means at least 70%, at least 80%, 90%, or even at least 95% sequence identity.

[0151] The term "substantial identity" with respect to a polypeptide indicates that the polypeptide comprises a sequence having at least 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, or 79%, or 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, or 89%, or at least 90%, 91%, 92%, 93%, or 94%, or even 95%, 96%, 97%, 98%, or 99%, sequence identity to a reference sequence over a specified comparison window. An indication that two polypeptide sequences are identical is that one polypeptide immunologically reacts with an antibody raised against the other polypeptide. Thus, for example, one polypeptide is identical to another polypeptide if the only difference between the two polypeptides is a conservative substitution.

[0152] The terms "treat" and "treatment" refer to both therapeutic treatment and prophylactic or preventative measures, the purpose of which is to prevent, inhibit, reduce, or lessen an undesirable physiological change or physiological disorder, e.g., the onset, progression, or worsening of a disorder. For purposes of this invention, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, diminishment of the extent of the disease, stabilization (i.e., no worsening or progression) of symptoms or adverse effects of the disease, delay or slowing of disease progression, improvement or palliation of the disease state, and remission (whether partial or complete), whether detectable or undetectable. "Treatment" can also mean prolonging survival as compared to expected survival if not receiving treatment. Those in need of treatment include those already with the condition or disorder, as well as those with a predisposition (e.g., as determined by genetic assay).

[0153] VI. Kits The present invention provides kits with packaging and one or more components therein. The kits typically include a label or insert that describes the components contained therein or includes instructions for in vitro, in vivo, or ex vivo use of the components contained therein. The kits can include a collection of such components, such as nucleic acids, recombinant vectors, and / or viral particles.

[0154] A kit refers to a physical structure that contains one or more components of the kit. The packaging can maintain the sterility of the components and can be made of materials commonly used for such purposes (e.g., paper, cardboard, glass, plastic, foil, ampoules, vials, tubes, etc.).

[0155] The label or package insert may include the identity of one or more components contained therein, dosage, and the clinical pharmacology of the active ingredient, e.g., mechanism of action, pharmacokinetics, and pharmacodynamics. The label or package insert may include information identifying the manufacturer, lot number, location and date of manufacture, and expiration date. The label or package insert may include information identifying the manufacturer information, lot number, location and date of manufacture. The label or package insert may include information regarding the disease for which the kit components may be used. The label or package insert may include instructions for a clinician or subject for using one or more of the kit components in a method, use, or treatment protocol or therapeutic regimen. The instructions may include dosage, frequency or duration of administration, and instructions for practicing any of the methods, uses, treatment protocols, or prophylactic or therapeutic regimens described herein.

[0156] The label or package insert can include information about a benefit the component may provide, such as a prophylactic or therapeutic benefit. The label or package insert can include information about potentially adverse side effects, complications, or reactions, such as warnings to the subject or clinician about situations in which using a particular composition may not be appropriate. Because adverse side effects or complications may also occur if the subject has previously taken, will take, or is currently taking one or more other medications that may be incompatible with the composition, or if the subject has previously taken, will take, or is currently taking another treatment protocol or therapeutic regimen that may be incompatible with the composition, the instructions can also include information about such incompatibilities.

[0157] Labels or package inserts include "printed matter," e.g., paper or cardboard, either separate or attached to a component, kit, or packaging (e.g., a box), or attached to the ampoule, tube, or vial that the kit component is in. Labels or package inserts can further include computer-readable media, e.g., a printed label with a barcode, a disk, an optical disk, e.g., a CD- or DVD-ROM / RAM, a DVD, an MP3, or electronic storage media, e.g., RAM and ROM, or hybrids thereof, e.g., magnetic / optical storage media, flash memory, hybrid, and memory-type cards. [Example]

[0158] VII. Working Examples The following examples are provided to demonstrate preferred embodiments of the invention. As will be appreciated by those skilled in the art, the techniques disclosed in the examples below represent techniques discovered by the inventors to function well in the practice of the invention, and can therefore be considered to constitute preferred modes for its implementation. However, those skilled in the art will, in light of this disclosure, appreciate that many changes can be made to the specific embodiments disclosed and still obtain like or similar results without departing from the spirit and scope of the invention.

[0159] Example 1 - Identification of AAV variants that target the brain parenchyma We developed a highly barcoded AAV library using AAV1, AAV2, and AAV9 capsids as starting platforms. AAV1, AAV2, and AAV9 peptide display libraries were generated by inserting random sequences into positions 590 of the AAV1 capsid, 587 of the AAV2 capsid, and 588 of the AAV9 capsid, respectively (Figure 1). The libraries consisted of 1 × 10 7 The diversity of the clones was 100% (Figure 3).

[0160] To test the utility of the library, a pilot study was conducted using bench-grade (low titer, low purity) capsid-modified AAV2. The AAV2 library was inoculated into two C57BL / 6 mice at 8 × 10 vector genomes per animal. 10 The virus was intravenously injected at 1000 x 1000 cells / animal. 72 hours later, the cerebral cortex, cerebellum, and spinal cord were dissected. Of note, the heart, skeletal muscle, and diaphragm were collected separately to identify muscle tropism. Viral genomic DNA was isolated, and the recovered random oligonucleotide sequences were amplified by PCR. The PCR products from the brain were pooled to generate a second-round library, which was then inoculated into two mice with 4 x 10 vector genomes per animal. 10 After the second injection, vector genomes were collected as before and subjected to NexGen sequencing along with the starting library and first-round tissues. To test whether sequences showing enrichment in brain tissues could actually deliver AAV2 to the brain, individual hits were cloned into an AAV2 capsid packaging plasmid to generate AAV2 expressing eGFP. Bench-grade vectors were generated, and 3 x 10 vector genomes of AAV2-based capsid-modified viruses were packaged. 10 were injected into mice. After 4 weeks, even these low-titer variants showed eGFP fluorescence in the brain.

[0161] These highly barcoded AAV libraries were used to identify AAV variants capable of targeting different primate brain structures in non-human primates. AAV1, AAV2, and AAV9 libraries were delivered to a single non-human primate via intracerebroventricular injection (Figure 2). 72 hours after injection, brain regions were microdissected for viral DNA isolation, and AAV DNA was amplified by PCR. The products were pooled and used to package a second-round library, which was then injected into additional NHPs. Brain regions were then microdissected 12 days after injection. After two rounds of panning, vector genomes were recovered and subjected to next-generation sequencing. Specifically, genomic DNA extracted from tissues from rounds 1 and 2 was PCR-amplified to generate Illumina amplicon sequencing libraries at the vector barcode locations. The resulting libraries were pooled and run on a single lane of an Illumina HiSeq 4000 using 100 bp single-end read chemistry. To illustrate the utility of this approach, several target regions were tested as examples: the ependyma, meninges, and cerebellum. In general, the sequences targeting AAVx to the ependyma, meninges, and cerebellum were different and varied among various serotypes.

[0162] Round-over-round enrichment graphs (Figure 4) and heat maps (Figures 5 and 6) were generated for the following tissues: brainstem, caudate nucleus, cerebellar cortex (Figure 5), cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, and thalamus. These illustrate the enrichment of the indicated barcodes at baseline (round 0) and after rounds 1 and 2 of in vivo passage in rhesus macaques. To generate these, fastq result files for each tissue and round combination were processed using custom Python scripts designed to extract and quantify the unique barcode configurations observed at the DNA level. Custom R scripts were used to calculate the percentage of barcodes present in each sample and convert DNA barcodes to amino acid barcodes. Table 1 corresponds to samples treated with the AAV1-derived library; Table 2 represents tissues treated with the AAV2-derived library; and Table 3 corresponds to samples treated with the AAV9-derived library. The top hits from these three libraries were selected and assembled into a validation library containing barcodes derived from 50 (AAV1), 58 (AAV2), and 30 (AAV9). This validation library was delivered to additional rhesus macaques via ICV injection. Tissues were again collected and processed to facilitate recovery of barcode abundance by deep sequencing. Barcode abundance was assessed in the collected tissues and the input viral library. Enrichment values ​​for each barcode were calculated relative to their abundance in the input viral library. The resulting relative enrichment values ​​are a robust indicator of vector performance across the various tissues evaluated and facilitate the identification of broad and specific AAV vector variants (Figure 7A-C).

[0163] To verify the identified cell-type specificity, AAV9-1999 (KGGGFHG; SEQ ID NO: 110) was selected for in vivo validation. The eGFP expression construct was packaged into AAV9-1999 driven by the CAG promoter. Five-year-old female rhesus macaques were administered 1.5E13 vg of AAV9-1999 via ICV injection into the left lateral ventricle. Brains were collected 30 days after injection for histological analysis. Cerebellar slices were stained with H&E to depict the transduction pattern of AAV9-1999 (Figure 8). The cochlea was also collected from this animal and, surprisingly, showed robust transduction of hair cells. Furthermore, AAV9-1999 and AAV9 capsids containing the eGFP construct were delivered to C57BL / 6 p0 mouse pups via ICV injection at 1E10 vg per hemisphere. After 21 days, mice were perfused. Whole-mount brains (Figure 9A), 40 μm whole-brain sagittal sections (Figure 9B), 40 μm S1 cortex sections (Figure 9C, left), 40 μm hippocampal sections (Figure 9C, center), 40 μm cerebellar sagittal sections (Figure 9C, right), and 40 μm lumbar spinal cord coronal sections (Figure 9D) were imaged for eGFP fluorescence signal. AAV9-1999 injected into Bl / 6 neonatal mouse pups demonstrated more ubiquitous expression than dose-matched injections of AAV9.

[0164] One adult rhesus macaque was injected with a mixture of four modified AAVs: AAV9 with the RGDLQWV (SEQ ID NO: 113) targeting peptide sequence and an mTAGBFP2 tag; AAV1 with the ERDRTRG (SEQ ID NO: 21) targeting peptide sequence as mTFP1; AAV2 with the GRGAPGG (SEQ ID NO: 80) targeting peptide sequence and an mNG tag; and AAV2 with the DDPSARR (SEQ ID NO: 53) targeting peptide sequence and an mRuby3 tag. The viruses were directly mixed in equal amounts to achieve the respective final total doses, as follows: AAV9.RGDL mTagBFP2 6.13E12 All vg AAV1.ERDR mTFP1 1.23E13 All vg AAV2.GRGA mNG 8.8E12 all vg AAV2.DDPS mRuby3 1.32E13 All vg

[0165] Brains were collected 30 days after injection for fluorescence imaging. Lateral ventricle sections (Figure 10A), fourth ventricle sections (Figure 10B), and meningeal sections (Figure 10C) were imaged for mTagBFP2, mTFP2, mNG, and mRuby3 fluorescent signals.

[0166] Additional experiments were performed by injecting AAV9-1999 into the cochlea of ​​rhesus monkeys. Based on the results of cochlear transduction, AAV9-1999 was administered to the lateral ventricle of the animals. A single animal received AAV9-1999 3E11 vg injected directly into its fenestra cochlea via a fenestration procedure (Figure 11A-C).

[0167] All of the methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of the present invention have been described in terms of preferred embodiments, it will be apparent to those skilled in the art that various modifications can be made to the methods and the steps or sequence of steps of the methods described herein without departing from the concept, spirit, and scope of the invention. More specifically, it will be apparent that certain agents that are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope, and concept of the invention as defined by the appended claims.

[0168] References The following references, to the extent that they provide exemplary procedural or other details supplementary to those set forth herein, are specifically incorporated herein by reference. TIFF2025148448000032.tif189150TIFF2025148448000033.tif32150

[0169] Array information SEQUENCE LISTING <110> The Children's Hospital of Philadelphia <120> ADENO-ASSOCIATED VIRAL VECTOR VARIANTS <150> US 63 / 084,709 <151> 2020-09-29 <150> US 62 / 939,315 <151> 2019-11-22 <160> 144 <170> PatentIn version 3.5 [[ID=​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​​ <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 3 Arg Pro Gly Arg Glu Ser Ala 1 5 <210> 4 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 4 Asn Glu Ser Leu Lys Lys Lys 1 5 <210> 5 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 5 Asp Lys Thr Arg Ala Gly Ser 1 5 <210> 6 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 6 Thr Ala Lys Ser Lys Gln Ala 1 5 <210> 7 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 7 Pro Val Lys Lys Lys Asp Ala 1 5 <210> 8 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 8 Gly Arg Glu Thr Leu Lys Gly 1 5 <210> 9 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 9 Pro Ile Pro Ala Gly Lys Lys 1 5 <210> 10 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 10 Asn Val Val Arg Ala Gly Thr 1 5 <210> 11 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 11 Lys Ala Thr Ala Asn Thr Arg 1 5 <210> 12 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 12 Arg Asp Ala Thr Arg Ser Ser 1 5 <210> 13 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 13 Val Pro Thr Lys Ser Pro Lys 1 5 <210> 14 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 14 Ala Gly Val Ala Arg Ser Lys 1 5 <210> 15 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 15 Arg Ser Arg Ser Glu Val Leu 1 5 <210> 16 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 16 Glu Val Lys Gly Lys Gly Lys 1 5 <210> 17 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 17 Ala Lys Leu Asn Lys Ser Ser 1 5 <210> 18 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 18 Asn Ser Val His Asn Thr Ala 1 5 <210> 19 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 19 Asn Val Val Arg Gly Gly Ala 1 5 <210> 20 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 20 Asn Arg Leu Val Ala Gly Gly 1 5 <210> 21 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 21 Glu Arg Asp Arg Thr Arg Gly 1 5 <210> 22 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 22 Val Gln Gly Ser Lys Met Lys 1 5 <210> 23 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 23 Asn Ser Val Arg Pro Leu Thr 1 5 <210> 24 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 24 Asn Lys Ile His Ala Asn Pro 1 5 <210> 25 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 25 Thr Ala Pro Lys Ser Leu Lys 1 5 <210> 26 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 26 Arg Asp Ser Thr Arg Gln Leu 1 5 <210> 27 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 27 Asn Ser Val Lys Ser Val Leu 1 5 <210> 28 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 28 Asn Val Thr Ile Lys Ser Lys 1 5 <210> 29 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 29 Val Ser Leu Lys Glu Arg Val 1 5 <210> 30 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 30 Asp Glu Thr Ser Arg Leu Val 1 5 <210> 31 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 31 Asp Arg Leu Lys Gly Ile Val 1 5 <210> 32 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 32 Ser Gly Val Leu Val Gln Arg 1 5 <210> 33 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 33 Ser Gly Thr Phe Val Lys Ala 1 5 <210> 34 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 34 Asn Ser Ile Ala Arg Pro Val 1 5 <210> 35 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 35 Asn Arg Ala Arg Ala Gly Glu 1 5 <210> 36 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 36 Ala Arg His Ala Leu Gly Gly 1 5 <210> 37 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 37 His Ser Ser Arg Pro Val Ala 1 5 <210> 38 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 38 Lys Thr Gly Thr Ala Arg Leu 1 5 <210> 39 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 39 Val Lys Ala Leu Gly Arg Pro 1 5 <210> 40 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 40 Asn Asp Val Arg Ala Lys Gly 1 5 <210> 41 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 41 Gln Gly Val Leu Val Val Arg 1 5 <210> 42 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 42 Lys Gln Tyr Ala Gly Ser Gln 1 5 <210> 43 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 43 Thr Asn Arg Met Ala Leu Ser 1 5 <210> 44 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 44 Gly Ile Thr Leu Gly Arg Leu 1 5 <210> 45 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 45 Ala Gly Ile Met Val Arg Val 1 5 <210> 46 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 46 Lys Ala Ala Gly Arg Thr Val 1 5 <210> 47 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 47 His Val Ile Arg Leu Pro Ser 1 5 <210> 48 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 48 Ser Val Ala Ser Ala Lys Lys 1 5 <210> 49 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 49 Thr Ala Thr Pro Arg Lys Gly 1 5 <210> 50 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 50 Thr Lys Thr Gly Leu Lys Leu 1 5 <210> 51 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 51 Lys Gly Leu Arg Thr Pro Thr 1 5 <210> 52 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 52 Leu Thr Ser Arg Thr Ser Pro 1 5 <210> 53 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 53 Asp Asp Pro Ser Ala Arg Arg 1 5 <210> 54 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 54 Gly Glu Gln Asp Leu Arg Arg 1 5 <210> 55 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 55 Val Ser Thr Ala Leu Pro Arg 1 5 <210> 56 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 56 Arg Asp Asp Val Pro Leu Arg 1 5 <210> 57 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 57 Thr Arg Val Gly Thr Ala Gly 1 5 <210> 58 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 58 Ser Ser Ser Lys Thr Gly Ser 1 5 <210> 59 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 59 Ser Leu Ser Thr Gly Pro Lys 1 5 <210> 60 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 60 Val Gln Gly Arg Gln Gly Gly 1 5 <210> 61 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 61 Arg Gly Ala Ser Gly Ala Val 1 5 <210> 62 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 62 Asn Ala Arg Ala Gln Gly Val 1 5 <210> 63 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 63 Thr Ser Asn Arg Gly Gln Val 1 5 <210> 64 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 64 Ala Val Arg Gly Gly Met Ala 1 5 <210> 65 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 65 Arg Gly Leu Asp Lys Gly Thr 1 5 <210> 66 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 66 Lys Gly Val Asp Leu Lys Pro 1 5 <210> 67 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 67 Thr Ala Val Arg Glu Glu Arg 1 5 <210> 68 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 68 Gly Asn Ala Gly Ile Thr Lys 1 5 <210> 69 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 69 Ser Ala Arg Ala Gly Ala Pro 1 5 <210> 70 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 70 Ser Gly Glu Phe Val Gly Arg 1 5 <210> 71 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 71 Ser Gly Arg Lys Leu Glu Val 1 5 <210> 72 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 72 Ser Ala Arg Ser Gly Ser Val 1 5 <210> 73 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 73 Glu Ser Thr Gly Arg Glu Arg 1 5 <210> 74 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 74 Val Thr Gln Ser Lys Gly Ala 1 5 <210> 75 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 75 Arg Gly Ser Gly Ser Ala Val 1 5 <210> 76 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 76 Asn Ala Arg Pro Val Ser Ala 1 5 <210> 77 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 77 Thr Ala Arg Gly Gly Gly Gly 1 5 <210> 78 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 78 Gly Arg Ser Ala Ser Gly Ser 1 5 <210> 79 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 79 Lys Ala Gln Gly Val Gly Gly 1 5 <210> 80 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 80 Gly Arg Gly Ala Pro Gly Gly 1 5 <210> 81 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 81 Val Pro Gly Arg Thr Ala Gly 1 5 <210> 82 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 82 Ala Arg Gly Ser Gly Val Asn 1 5 <210> 83 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 83 Ser Val Arg Val Gly Gly Gln 1 5 <210> 84 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 84 Arg Ala Val Pro Ala Gly Gly 1 5 <210> 85 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 85 Val Met Ser Ser Gly Lys Pro 1 5 <210> 86 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 86 Ser Thr Pro Ala Pro Lys Ser 1 5 <210> 87 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 87 Arg Gly Gly Ala Gln Val Val 1 5 <210> 88 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 88 Lys Ala Ser Gly Ala Gly Gly 1 5 <210> 89 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 89 Thr Gly Thr Ala Gly Leu Lys 1 5 <210> 90 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 90 Arg Ser Asn Ala Pro Gln Thr 1 5 <210> 91 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 91 Glu Ala Gln Ser His Pro Arg 1 5 <210> 92 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 92 Thr Lys Ser Leu Ser Ser Gly 1 5 <210> 93 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 93 Ala Ala Gly Ala Lys Val Met 1 5 <210> 94 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 94 Arg Gly Ser Thr Gln Val Gly 1 5 <210> 95 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 95 Gly Arg Ser Thr Gly Met Thr 1 5 <210> 96 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 96 Arg Ala Thr Ser Gln Ser Thr 1 5 <210> 97 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 97 Val Gly Arg Ser Val Gly Ala 1 5 <210> 98 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 98 Gly Glu Gly Gly Gly Gly Arg 1 5 <210> 99 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 99 Thr Ala Ala Gly Gly Gln Arg 1 5 <210> 100 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 100 Gly Arg Gly Gly Ala Ala Leu 1 5 <210> 101 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 101 Val Ala Pro Ile Ser Lys Ser 1 5 <210> 102 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 102 Ala Pro Pro Val Lys Leu Ser 1 5 <210> 103 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 103 His Asp Gly Gly Ala Ser Arg 1 5 <210> 104 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 104 Arg Ser Gly Gly Ala Ala Val 1 5 <210> 105 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 105 Gly Ser Arg Ala Gly Val Gly 1 5 <210> 106 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 106 Lys Leu Ser Ile Ser Gly Asn 1 5 <210> 107 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 107 Gly Ala Val Gly Gly Val Lys 1 5 <210> 108 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 108 Lys Asn Glu Ser Gly Lys Val 1 5 <210> 109 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 109 Ala Gly Gln Leu Ala Gly Arg 1 5 <210> 110 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 110 Lys Gly Gly Gly Phe His Gly 1 5 <210> 111 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 111 Arg Ala Lys Pro Gly Met Glu 1 5 <210> 112 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 112 Gly Arg Asp Val Thr Arg Ser 1 5 <210> 113 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 113 Arg Gly Asp Leu Gln Trp Val 1 5 <210> 114 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 114 Gly Gly Asp Arg Thr Arg Gly 1 5 <210> 115 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 115 Arg Gly Asp Leu Ala Ser Val 1 5 <210> 116 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 116 Arg Asp Thr Thr Arg Asn Leu 1 5 <210> 117 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 117 Lys Gly Gly Gly Val His Gly 1 5 <210> 118 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 118 Arg Gly Asp Met Tyr Arg Val 1 5 <210> 119 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 119 Arg Gly Asp Arg Pro Val Ser 1 5 <210> 120 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 120 Arg Ser Asp Val Gly Ser Leu 1 5 <210> 121 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 121 Ala Gly Val Lys Pro Gly Arg 1 5 <210> 122 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 122 Arg Gly Asp Trp Pro Arg Gly 1 5 <210> 123 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 123 Gly Gly Arg Pro Gly Ser Trp 1 5 <210> 124 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 124 Arg Gly Asp Tyr Pro Arg Ser 1 5 <210> 125 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 125 Arg Gly Asp Leu Arg Phe Ile 1 5 <210> 126 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 126 Arg Gly Gly Gly Val Tyr Gly 1 5 <210> 127 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 127 Gly Ala Asp Arg Thr Arg Gly 1 5 <210> 128 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 128 Arg Gly Asp Phe Met Gly Leu 1 5 <210> 129 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 129 Arg Arg Asp Glu Thr Arg Thr 1 5 <210> 130 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 130 Ala Glu Ser Pro Trp Glu Arg 1 5 <210> 131 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 131 Ala Trp Asp Gly Thr Arg Val 1 5 <210> 132 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 132 Ala Arg Gly Asp Gly Trp Arg 1 5 <210> 133 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 133 Gly Arg Asp Tyr Thr Arg Leu 1 5 <210> 134 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 134 Arg Arg Gly Asp Ala Trp Ser 1 5 <210> 135 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 135 Met Asp Leu Thr Lys Ala Val 1 5 <210> 136 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 136 Met Met Gly Arg Pro Gly Arg 1 5 <210> 137 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 137 Thr Gly Arg Pro Gly Val Trp 1 5 <210> 138 <211> 736 <212> PRT <213> adeno-associated virus 1 <400> 138 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Asn Leu Ser 1 5 10 15 Glu Gly Ile Arg Glu Trp Trp Asp Leu Lys Pro Gly Ala Pro Lys Pro 20 25 30 Lys Ala Asn Gln Gln Lys Gln Asp Asp Gly Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Phe Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Ala Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Gln Gln Leu Lys Ala Gly Asp Asn Pro Tyr Leu Arg Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Gln Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Val Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Gly Ala Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu Gln Ser Pro Gln Glu Pro Asp Ser Ser Ser Gly Ile Gly 145 150 155 160 Lys Thr Gly Gln Gln Pro Ala Lys Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Ser Glu Ser Val Pro Asp Pro Gln Pro Leu Gly Glu Pro Pro 180 185 190 Ala Thr Pro Ala Ala Val Gly Pro Thr Thr Met Ala Ser Gly Gly Gly 195 200 205 Ala Pro Met Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Asn Ala 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Thr Trp Leu Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Ser Ala Ser Thr Gly Ala Ser Asn Asp Asn His 260 265 270 Tyr Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg Phe 275 280 285 His Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn Asn 290 295 300 Trp Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile Gln 305 310 315 320 Val Lys Glu Val Thr Thr Asn Asp Gly Val Thr Thr Ile Ala Asn Asn 325 330 335 Leu Thr Ser Thr Val Gln Val Phe Ser Asp Ser Glu Tyr Gln Leu Pro 340 345 350 Tyr Val Leu Gly Ser Ala His Gln Gly Cys Leu Pro Pro Phe Pro Ala 355 360 365 Asp Val Phe Met Ile Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asn Gly 370 375 380 Ser Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe Pro 385 390 395 400 Ser Gln Met Leu Arg Thr Gly Asn Asn Phe Thr Phe Ser Tyr Thr Phe 405 410 415 Glu Glu Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu Asp 420 425 430 Arg Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Asn Arg 435 440 445 Thr Gln Asn Gln Ser Gly Ser Ala Gln Asn Lys Asp Leu Leu Phe Ser 450 455 460 Arg Gly Ser Pro Ala Gly Met Ser Val Gln Pro Lys Asn Trp Leu Pro 465 470 475 480 Gly Pro Cys Tyr Arg Gln Gln Arg Val Ser Lys Thr Lys Thr Asp Asn 485 490 495 Asn Asn Ser Asn Phe Thr Trp Thr Gly Ala Ser Lys Tyr Asn Leu Asn 500 505 510 Gly Arg Glu Ser Ile Ile Asn Pro Gly Thr Ala Met Ala Ser His Lys 515 520 525 Asp Asp Glu Asp Lys Phe Phe Pro Met Ser Gly Val Met Ile Phe Gly 530 535 540 Lys Glu Ser Ala Gly Ala Ser Asn Thr Ala Leu Asp Asn Val Met Ile 545 550 555 560 Thr Asp Glu Glu Glu Ile Lys Ala Thr Asn Pro Val Ala Thr Glu Arg 565 570 575 Phe Gly Thr Val Ala Val Asn Phe Gln Ser Ser Ser Thr Asp Pro Ala 580 585 590 Thr Gly Asp Val His Ala Met Gly Ala Leu Pro Gly Met Val Trp Gln 595 600 605 Asp Arg Asp Val Tyr Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His 610 615 620 Thr Asp Gly His Phe His Pro Ser Pro Leu Met Gly Gly Phe Gly Leu 625 630 635 640 Lys Asn Pro Pro Pro Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala 645 650 655 Asn Pro Pro Ala Glu Phe Ser Ala Thr Lys Phe Ala Ser Phe Ile Thr 660 665 670 Gln Tyr Ser Thr Gly Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln 675 680 685 Lys Glu Asn Ser Lys Arg Trp Asn Pro Glu Val Gln Tyr Thr Ser Asn 690 695 700 Tyr Ala Lys Ser Ala Asn Val Asp Phe Thr Val Asp Asn Asn Gly Leu 705 710 715 720 Tyr Thr Glu Pro Arg Pro Ile Gly Thr Arg Tyr Leu Thr Arg Pro Leu 725 730 735 <210> 139 <211> 735 <212> PRT <213> adeno-associated virus 2 <400> 139 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Thr Leu Ser 1 5 10 15 Glu Gly Ile Arg Gln Trp Trp Lys Leu Lys Pro Gly Pro Pro Pro Pro 20 25 30 Lys Pro Ala Glu Arg His Lys Asp Asp Ser Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Phe Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Glu Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Arg Gln Leu Asp Ser Gly Asp Asn Pro Tyr Leu Lys Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Lys Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Val Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Pro Val Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu His Ser Pro Val Glu Pro Asp Ser Ser Ser Gly Thr Gly 145 150 155 160 Lys Ala Gly Gln Gln Pro Ala Arg Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Ala Asp Ser Val Pro Asp Pro Gln Pro Leu Gly Gln Pro Pro 180 185 190 Ala Ala Pro Ser Gly Leu Gly Thr Asn Thr Met Ala Thr Gly Ser Gly 195 200 205 Ala Pro Met Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Asn Ser 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Thr Trp Met Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Ser Gln Ser Gly Ala Ser Asn Asp Asn His Tyr 260 265 270 Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg Phe His 275 280 285 Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn Asn Trp 290 295 300 Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile Gln Val 305 310 315 320 Lys Glu Val Thr Gln Asn Asp Gly Thr Thr Thr Ile Ala Asn Asn Leu 325 330 335 Thr Ser Thr Val Gln Val Phe Thr Asp Ser Glu Tyr Gln Leu Pro Tyr 340 345 350 Val Leu Gly Ser Ala His Gln Gly Cys Leu Pro Pro Phe Pro Ala Asp 355 360 365 Val Phe Met Val Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asn Gly Ser 370 375 380 Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe Pro Ser 385 390 395 400 Gln Met Leu Arg Thr Gly Asn Asn Phe Thr Phe Ser Tyr Thr Phe Glu 405 410 415 Asp Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu Asp Arg 420 425 430 Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Ser Arg Thr 435 440 445 Asn Thr Pro Ser Gly Thr Thr Thr Gln Ser Arg Leu Gln Phe Ser Gln 450 455 460 Ala Gly Ala Ser Asp Ile Arg Asp Gln Ser Arg Asn Trp Leu Pro Gly 465 470 475 480 Pro Cys Tyr Arg Gln Gln Arg Val Ser Lys Thr Ser Ala Asp Asn Asn 485 490 495 Asn Ser Glu Tyr Ser Trp Thr Gly Ala Thr Lys Tyr His Leu Asn Gly 500 505 510 Arg Asp Ser Leu Val Asn Pro Gly Pro Ala Met Ala Ser His Lys Asp 515 520 525 Asp Glu Glu Lys Phe Phe Pro Gln Ser Gly Val Leu Ile Phe Gly Lys 530 535 540 Gln Gly Ser Glu Lys Thr Asn Val Asp Ile Glu Lys Val Met Ile Thr 545 550 555 560 Asp Glu Glu Glu Ile Arg Thr Thr Asn Pro Val Ala Thr Glu Gln Tyr 565 570 575 Gly Ser Val Ser Thr Asn Leu Gln Arg Gly Asn Arg Gln Ala Ala Thr 580 585 590 Ala Asp Val Asn Thr Gln Gly Val Leu Pro Gly Met Val Trp Gln Asp 595 600 605 Arg Asp Val Tyr Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His Thr 610 615 620 Asp Gly His Phe His Pro Ser Pro Leu Met Gly Gly Phe Gly Leu Lys 625 630 635 640 His Pro Pro Pro Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala Asn 645 650 655 Pro Ser Thr Thr Phe Ser Ala Ala Lys Phe Ala Ser Phe Ile Thr Gln 660 665 670 Tyr Ser Thr Gly Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln Lys 675 680 685 Glu Asn Ser Lys Arg Trp Asn Pro Glu Ile Gln Tyr Thr Ser Asn Tyr 690 695 700 Asn Lys Ser Val Asn Val Asp Phe Thr Val Asp Thr Asn Gly Val Tyr 705 710 715 720 Ser Glu Pro Arg Pro Ile Gly Thr Arg Tyr Leu Thr Arg Asn Leu 725 730 735 <210> 140 <211> 736 <212> PRT <213> adeno-associated virus 9 <400> 140 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Asn Leu Ser 1 5 10 15 Glu Gly Ile Arg Glu Trp Trp Ala Leu Lys Pro Gly Ala Pro Gln Pro 20 25 30 Lys Ala Asn Gln Gln His Gln Asp Asn Ala Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Gly Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Ala Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Gln Gln Leu Lys Ala Gly Asp Asn Pro Tyr Leu Lys Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Lys Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Leu Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Ala Ala Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu Gln Ser Pro Gln Glu Pro Asp Ser Ser Ala Gly Ile Gly 145 150 155 160 Lys Ser Gly Ala Gln Pro Ala Lys Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Thr Glu Ser Val Pro Asp Pro Gln Pro Ile Gly Glu Pro Pro 180 185 190 Ala Ala Pro Ser Gly Val Gly Ser Leu Thr Met Ala Ser Gly Gly Gly 195 200 205 Ala Pro Val Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Ser Ser 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Gln Trp Leu Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Asn Ser Thr Ser Gly Gly Ser Ser Asn Asp Asn 260 265 270 Ala Tyr Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg 275 280 285 Phe His Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn 290 295 300 Asn Trp Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile 305 310 315 320 Gln Val Lys Glu Val Thr Asp Asn Asn Gly Val Lys Thr Ile Ala Asn 325 330 335 Asn Leu Thr Ser Thr Val Gln Val Phe Thr Asp Ser Asp Tyr Gln Leu 340 345 350 Pro Tyr Val Leu Gly Ser Ala His Glu Gly Cys Leu Pro Pro Phe Pro 355 360 365 Ala Asp Val Phe Met Ile Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asp 370 375 380 Gly Ser Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe 385 390 395 400 Pro Ser Gln Met Leu Arg Thr Gly Asn Asn Phe Gln Phe Ser Tyr Glu 405 410 415 Phe Glu Asn Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu 420 425 430 Asp Arg Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Ser 435 440 445 Lys Thr Ile Asn Gly Ser Gly Gln Asn Gln Gln Thr Leu Lys Phe Ser 450 455 460 Val Ala Gly Pro Ser Asn Met Ala Val Gln Gly Arg Asn Tyr Ile Pro 465 470 475 480 Gly Pro Ser Tyr Arg Gln Gln Arg Val Ser Thr Thr Val Thr Gln Asn 485 490 495 Asn Asn Ser Glu Phe Ala Trp Pro Gly Ala Ser Ser Trp Ala Leu Asn 500 505 510 Gly Arg Asn Ser Leu Met Asn Pro Gly Pro Ala Met Ala Ser His Lys 515 520 525 Glu Gly Glu Asp Arg Phe Phe Pro Leu Ser Gly Ser Leu Ile Phe Gly 530 535 540 Lys Gln Gly Thr Gly Arg Asp Asn Val Asp Ala Asp Lys Val Met Ile 545 550 555 560 Thr Asn Glu Glu Glu Ile Lys Thr Thr Asn Pro Val Ala Thr Glu Ser 565 570 575 Tyr Gly Gln Val Ala Thr Asn His Gln Ser Ala Gln Ala Gln Ala Gln 580 585 590 Thr Gly Trp Val Gln Asn Gln Gly Ile Leu Pro Gly Met Val Trp Gln 595 600 605 Asp Arg Asp Val Tyr Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His 610 615 620 Thr Asp Gly Asn Phe His Pro Ser Pro Leu Met Gly Gly Phe Gly Met 625 630 635 640 Lys His Pro Pro Pro Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala 645 650 655 Asp Pro Pro Thr Ala Phe Asn Lys Asp Lys Leu Asn Ser Phe Ile Thr 660 665 670 Gln Tyr Ser Thr Gly Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln 675 680 685 Lys Glu Asn Ser Lys Arg Trp Asn Pro Glu Ile Gln Tyr Thr Ser Asn 690 695 700 Tyr Tyr Lys Ser Asn Asn Val Glu Phe Ala Val Asn Thr Glu Gly Val 705 710 715 720 Tyr Ser Glu Pro Arg Pro Ile Gly Thr Arg Tyr Leu Thr Arg Asn Leu 725 730 735 <210> 141 <211> 748 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <220> <221> MISC_FEATURE <222> (594)..(600) <223> X is any amino acid <400> 141 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Asn Leu Ser 1 5 10 15 Glu Gly Ile Arg Glu Trp Trp Asp Leu Lys Pro Gly Ala Pro Lys Pro 20 25 30 Lys Ala Asn Gln Gln Lys Gln Asp Asp Gly Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Phe Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Ala Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Gln Gln Leu Lys Ala Gly Asp Asn Pro Tyr Leu Arg Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Gln Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Val Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Gly Ala Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu Gln Ser Pro Gln Glu Pro Asp Ser Ser Ser Gly Ile Gly 145 150 155 160 Lys Thr Gly Gln Gln Pro Ala Lys Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Ser Glu Ser Val Pro Asp Pro Gln Pro Leu Gly Glu Pro Pro 180 185 190 Ala Thr Pro Ala Ala Val Gly Pro Thr Thr Met Ala Ser Gly Gly Gly 195 200 205 Ala Pro Met Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Asn Ala 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Thr Trp Leu Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Ser Ala Ser Thr Gly Ala Ser Asn Asp Asn His 260 265 270 Tyr Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg Phe 275 280 285 His Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn Asn 290 295 300 Trp Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile Gln 305 310 315 320 Val Lys Glu Val Thr Thr Asn Asp Gly Val Thr Thr Ile Ala Asn Asn 325 330 335 Leu Thr Ser Thr Val Gln Val Phe Ser Asp Ser Glu Tyr Gln Leu Pro 340 345 350 Tyr Val Leu Gly Ser Ala His Gln Gly Cys Leu Pro Pro Phe Pro Ala 355 360 365 Asp Val Phe Met Ile Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asn Gly 370 375 380 Ser Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe Pro 385 390 395 400 Ser Gln Met Leu Arg Thr Gly Asn Asn Phe Thr Phe Ser Tyr Thr Phe 405 410 415 Glu Glu Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu Asp 420 425 430 Arg Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Asn Arg 435 440 445 Thr Gln Asn Gln Ser Gly Ser Ala Gln Asn Lys Asp Leu Leu Phe Ser 450 455 460 Arg Gly Ser Pro Ala Gly Met Ser Val Gln Pro Lys Asn Trp Leu Pro 465 470 475 480 Gly Pro Cys Tyr Arg Gln Gln Arg Val Ser Lys Thr Lys Thr Asp Asn 485 490 495 Asn Asn Ser Asn Phe Thr Trp Thr Gly Ala Ser Lys Tyr Asn Leu Asn 500 505 510 Gly Arg Glu Ser Ile Ile Asn Pro Gly Thr Ala Met Ala Ser His Lys 515 520 525 Asp Asp Glu Asp Lys Phe Phe Pro Met Ser Gly Val Met Ile Phe Gly 530 535 540 Lys Glu Ser Ala Gly Ala Ser Asn Thr Ala Leu Asp Asn Val Met Ile 545 550 555 560 Thr Asp Glu Glu Glu Ile Lys Ala Thr Asn Pro Val Ala Thr Glu Arg 565 570 575 Phe Gly Thr Val Ala Val Asn Phe Gln Ser Ser Ser Thr Asp Ser Ser 580 585 590 Ala Xaa Xaa Xaa Xaa Xaa Xaa Xaa Ala Ser Pro Ala Thr Gly Asp Val 595 600 605 His Ala Met Gly Ala Leu Pro Gly Met Val Trp Gln Asp Arg Asp Val 610 615 620 Tyr Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His Thr Asp Gly His 625 630 635 640 Phe His Pro Ser Pro Leu Met Gly Gly Phe Gly Leu Lys Asn Pro Pro 645 650 655 Pro Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala Asn Pro Pro Ala 660 665 670 Glu Phe Ser Ala Thr Lys Phe Ala Ser Phe Ile Thr Gln Tyr Ser Thr 675 680 685 Gly Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln Lys Glu Asn Ser 690 695 700 Lys Arg Trp Asn Pro Glu Val Gln Tyr Thr Ser Asn Tyr Ala Lys Ser 705 710 715 720 Ala Asn Val Asp Phe Thr Val Asp Asn Asn Gly Leu Tyr Thr Glu Pro 725 730 735 Arg Pro Ile Gly Thr Arg Tyr Leu Thr Arg Pro Leu 740 745 <210> 142 <211> 747 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <220> <221> MISC_FEATURE <222> (591)..(597) <223> X is any amino acid <400> 142 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Thr Leu Ser 1 5 10 15 Glu Gly Ile Arg Gln Trp Trp Lys Leu Lys Pro Gly Pro Pro Pro Pro 20 25 30 Lys Pro Ala Glu Arg His Lys Asp Asp Ser Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Phe Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Glu Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Arg Gln Leu Asp Ser Gly Asp Asn Pro Tyr Leu Lys Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Lys Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Val Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Pro Val Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu His Ser Pro Val Glu Pro Asp Ser Ser Ser Gly Thr Gly 145 150 155 160 Lys Ala Gly Gln Gln Pro Ala Arg Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Ala Asp Ser Val Pro Asp Pro Gln Pro Leu Gly Gln Pro Pro 180 185 190 Ala Ala Pro Ser Gly Leu Gly Thr Asn Thr Met Ala Thr Gly Ser Gly 195 200 205 Ala Pro Met Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Asn Ser 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Thr Trp Met Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Ser Gln Ser Gly Ala Ser Asn Asp Asn His Tyr 260 265 270 Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg Phe His 275 280 285 Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn Asn Trp 290 295 300 Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile Gln Val 305 310 315 320 Lys Glu Val Thr Gln Asn Asp Gly Thr Thr Thr Ile Ala Asn Asn Leu 325 330 335 Thr Ser Thr Val Gln Val Phe Thr Asp Ser Glu Tyr Gln Leu Pro Tyr 340 345 350 Val Leu Gly Ser Ala His Gln Gly Cys Leu Pro Pro Phe Pro Ala Asp 355 360 365 Val Phe Met Val Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asn Gly Ser 370 375 380 Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe Pro Ser 385 390 395 400 Gln Met Leu Arg Thr Gly Asn Asn Phe Thr Phe Ser Tyr Thr Phe Glu 405 410 415 Asp Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu Asp Arg 420 425 430 Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Ser Arg Thr 435 440 445 Asn Thr Pro Ser Gly Thr Thr Thr Gln Ser Arg Leu Gln Phe Ser Gln 450 455 460 Ala Gly Ala Ser Asp Ile Arg Asp Gln Ser Arg Asn Trp Leu Pro Gly 465 470 475 480 Pro Cys Tyr Arg Gln Gln Arg Val Ser Lys Thr Ser Ala Asp Asn Asn 485 490 495 Asn Ser Glu Tyr Ser Trp Thr Gly Ala Thr Lys Tyr His Leu Asn Gly 500 505 510 Arg Asp Ser Leu Val Asn Pro Gly Pro Ala Met Ala Ser His Lys Asp 515 520 525 Asp Glu Glu Lys Phe Phe Pro Gln Ser Gly Val Leu Ile Phe Gly Lys 530 535 540 Gln Gly Ser Glu Lys Thr Asn Val Asp Ile Glu Lys Val Met Ile Thr 545 550 555 560 Asp Glu Glu Glu Ile Arg Thr Thr Asn Pro Val Ala Thr Glu Gln Tyr 565 570 575 Gly Ser Val Ser Thr Asn Leu Gln Arg Gly Asn Ala Ala Ala Xaa Xaa 580 585 590 Xaa Xaa Xaa Xaa Xaa Ala Ala Arg Gln Ala Ala Thr Ala Asp Val Asn 595 600 605 Thr Gln Gly Val Leu Pro Gly Met Val Trp Gln Asp Arg Asp Val Tyr 610 615 620 Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His Thr Asp Gly His Phe 625 630 635 640 His Pro Ser Pro Leu Met Gly Gly Phe Gly Leu Lys His Pro Pro Pro 645 650 655 Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala Asn Pro Ser Thr Thr 660 665 670 Phe Ser Ala Ala Lys Phe Ala Ser Phe Ile Thr Gln Tyr Ser Thr Gly 675 680 685 Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln Lys Glu Asn Ser Lys 690 695 700 Arg Trp Asn Pro Glu Ile Gln Tyr Thr Ser Asn Tyr Asn Lys Ser Val 705 710 715 720 Asn Val Asp Phe Thr Val Asp Thr Asn Gly Val Tyr Ser Glu Pro Arg 725 730 735 Pro Ile Gly Thr Arg Tyr Leu Thr Arg Asn Leu 740 745 <210> 143 <211> 748 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <220> <221> MISC_FEATURE <222> (592)..(598) <223> X is any amino acid <400> 143 Met Ala Ala Asp Gly Tyr Leu Pro Asp Trp Leu Glu Asp Asn Leu Ser 1 5 10 15 Glu Gly Ile Arg Glu Trp Trp Ala Leu Lys Pro Gly Ala Pro Gln Pro 20 25 30 Lys Ala Asn Gln Gln His Gln Asp Asn Ala Arg Gly Leu Val Leu Pro 35 40 45 Gly Tyr Lys Tyr Leu Gly Pro Gly Asn Gly Leu Asp Lys Gly Glu Pro 50 55 60 Val Asn Ala Ala Asp Ala Ala Ala Leu Glu His Asp Lys Ala Tyr Asp 65 70 75 80 Gln Gln Leu Lys Ala Gly Asp Asn Pro Tyr Leu Lys Tyr Asn His Ala 85 90 95 Asp Ala Glu Phe Gln Glu Arg Leu Lys Glu Asp Thr Ser Phe Gly Gly 100 105 110 Asn Leu Gly Arg Ala Val Phe Gln Ala Lys Lys Arg Leu Leu Glu Pro 115 120 125 Leu Gly Leu Val Glu Glu Ala Ala Lys Thr Ala Pro Gly Lys Lys Arg 130 135 140 Pro Val Glu Gln Ser Pro Gln Glu Pro Asp Ser Ser Ala Gly Ile Gly 145 150 155 160 Lys Ser Gly Ala Gln Pro Ala Lys Lys Arg Leu Asn Phe Gly Gln Thr 165 170 175 Gly Asp Thr Glu Ser Val Pro Asp Pro Gln Pro Ile Gly Glu Pro Pro 180 185 190 Ala Ala Pro Ser Gly Val Gly Ser Leu Thr Met Ala Ser Gly Gly Gly 195 200 205 Ala Pro Val Ala Asp Asn Asn Glu Gly Ala Asp Gly Val Gly Ser Ser 210 215 220 Ser Gly Asn Trp His Cys Asp Ser Gln Trp Leu Gly Asp Arg Val Ile 225 230 235 240 Thr Thr Ser Thr Arg Thr Trp Ala Leu Pro Thr Tyr Asn Asn His Leu 245 250 255 Tyr Lys Gln Ile Ser Asn Ser Thr Ser Gly Gly Ser Ser Asn Asp Asn 260 265 270 Ala Tyr Phe Gly Tyr Ser Thr Pro Trp Gly Tyr Phe Asp Phe Asn Arg 275 280 285 Phe His Cys His Phe Ser Pro Arg Asp Trp Gln Arg Leu Ile Asn Asn 290 295 300 Asn Trp Gly Phe Arg Pro Lys Arg Leu Asn Phe Lys Leu Phe Asn Ile 305 310 315 320 Gln Val Lys Glu Val Thr Asp Asn Asn Gly Val Lys Thr Ile Ala Asn 325 330 335 Asn Leu Thr Ser Thr Val Gln Val Phe Thr Asp Ser Asp Tyr Gln Leu 340 345 350 Pro Tyr Val Leu Gly Ser Ala His Glu Gly Cys Leu Pro Pro Phe Pro 355 360 365 Ala Asp Val Phe Met Ile Pro Gln Tyr Gly Tyr Leu Thr Leu Asn Asp 370 375 380 Gly Ser Gln Ala Val Gly Arg Ser Ser Phe Tyr Cys Leu Glu Tyr Phe 385 390 395 400 Pro Ser Gln Met Leu Arg Thr Gly Asn Asn Phe Gln Phe Ser Tyr Glu 405 410 415 Phe Glu Asn Val Pro Phe His Ser Ser Tyr Ala His Ser Gln Ser Leu 420 425 430 Asp Arg Leu Met Asn Pro Leu Ile Asp Gln Tyr Leu Tyr Tyr Leu Ser 435 440 445 Lys Thr Ile Asn Gly Ser Gly Gln Asn Gln Gln Thr Leu Lys Phe Ser 450 455 460 Val Ala Gly Pro Ser Asn Met Ala Val Gln Gly Arg Asn Tyr Ile Pro 465 470 475 480 Gly Pro Ser Tyr Arg Gln Gln Arg Val Ser Thr Thr Val Thr Gln Asn 485 490 495 Asn Asn Ser Glu Phe Ala Trp Pro Gly Ala Ser Ser Trp Ala Leu Asn 500 505 510 Gly Arg Asn Ser Leu Met Asn Pro Gly Pro Ala Met Ala Ser His Lys 515 520 525 Glu Gly Glu Asp Arg Phe Phe Pro Leu Ser Gly Ser Leu Ile Phe Gly 530 535 540 Lys Gln Gly Thr Gly Arg Asp Asn Val Asp Ala Asp Lys Val Met Ile 545 550 555 560 Thr Asn Glu Glu Glu Ile Lys Thr Thr Asn Pro Val Ala Thr Glu Ser 565 570 575 Tyr Gly Gln Val Ala Thr Asn His Gln Ser Ala Gln Ala Ala Ala Xaa 580 585 590 Xaa Xaa Xaa Xaa Xaa Xaa Ala Ser Ala Gln Ala Gln Thr Gly Trp Val 595 600 605 Gln Asn Gln Gly Ile Leu Pro Gly Met Val Trp Gln Asp Arg Asp Val 610 615 620 Tyr Leu Gln Gly Pro Ile Trp Ala Lys Ile Pro His Thr Asp Gly Asn 625 630 635 640 Phe His Pro Ser Pro Leu Met Gly Gly Phe Gly Met Lys His Pro Pro 645 650 655 Pro Gln Ile Leu Ile Lys Asn Thr Pro Val Pro Ala Asp Pro Pro Thr 660 665 670 Ala Phe Asn Lys Asp Lys Leu Asn Ser Phe Ile Thr Gln Tyr Ser Thr 675 680 685 Gly Gln Val Ser Val Glu Ile Glu Trp Glu Leu Gln Lys Glu Asn Ser 690 695 700 Lys Arg Trp Asn Pro Glu Ile Gln Tyr Thr Ser Asn Tyr Tyr Lys Ser 705 710 715 720 Asn Asn Val Glu Phe Ala Val Asn Thr Glu Gly Val Tyr Ser Glu Pro 725 730 735 Arg Pro Ile Gly Thr Arg Tyr Leu Thr Arg Asn Leu 740 745 <210> 144 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> Synthetic polypeptide <400> 144 Arg Pro Gly Arg Glu Ala Ser 1 5

Claims

1. A modified adeno-associated virus (AAV) capsid protein comprising a targeting peptide, the targeting peptide being 3 to 10 amino acids in length, which targets a viral vector comprising the modified AAV capsid protein to a different organ or a different brain structure.

2. 2. The modified AAV capsid protein of claim 1, wherein the modified AAV capsid protein is a modified AAV9 capsid protein having a sequence at least 95% identical to SEQ ID NO: 143, the targeting peptide is SEQ ID NO: 110, and the different brain structures are the brainstem, caudate nucleus, cerebellum, cochlea (ear), cortex, cerebral cortex, deep cerebellar nuclei, ependyma, globus pallidus, hippocampus, meninges, motor cortex, optic nerve, prefrontal cortex, putamen, spinal cord, substantia nigra, subthalamic nucleus, temporal cortex, thalamus, or visual cortex.

3. The modified AAV capsid protein of claim 1, which is a modified AAV1 capsid protein, a modified AAV2 capsid protein, or a modified AAV9 capsid protein.

4. The modified AAV capsid protein of claim 1, wherein the modified AAV capsid protein is derived from the AAV1 capsid protein (see SEQ ID NO: 138) and the targeting peptide is inserted after residue 590 of the AAV1 capsid protein.

5. The modified AAV capsid protein of claim 4, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length.

6. The modified AAV capsid protein of claim 5, wherein the linker sequence is SSA on the N-terminal side of the targeting peptide and AS on the C-terminal side of the targeting peptide.

7. The modified AAV capsid protein of claim 6, wherein the modified AAV1 capsid protein has a sequence that is at least 95% identical to SEQ ID NO:

141.

8. The modified AAV capsid protein of claim 1, wherein the modified AAV capsid protein is derived from the AAV2 capsid protein (see SEQ ID NO: 139) and the targeting peptide is inserted after residue 587 of the AAV2 capsid protein.

9. 9. The modified AAV capsid protein of claim 8, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length.

10. 10. The modified AAV capsid protein of claim 9, wherein the linker sequence is AAA at the N-terminus of the targeting peptide and AA at the C-terminus of the targeting peptide.

11. The modified AAV2 capsid protein of claim 10, wherein the modified AAV2 capsid protein has a sequence that is at least 95% identical to SEQ ID NO:

142.

12. The modified AAV capsid protein of claim 1, wherein the modified AAV capsid protein is derived from the AAV9 capsid protein (see SEQ ID NO: 140) and the targeting peptide is inserted after residue 588 of the AAV9 capsid protein.

13. 13. The modified AAV capsid protein of claim 12, wherein the targeting peptide is flanked by linker sequences, and the linker sequences on both sides of the targeting peptide are 2 or 3 amino acids in length.

14. 14. The modified AAV capsid protein of claim 13, wherein the linker sequence is AAA at the N-terminus of the targeting peptide and AS at the C-terminus of the targeting peptide.

15. The modified AAV capsid protein of claim 14, wherein the modified AAV9 capsid protein has a sequence that is at least 95% identical to SEQ ID NO:

143.

16. 2. The modified AAV capsid protein of claim 1, wherein the target peptide comprises a sequence of up to 10 amino acids in length, and the sequence has an amino acid sequence selected from the group consisting of SEQ ID NOs: 1-137 or 144.

17. 17. The modified AAV capsid protein of claim 16, wherein the targeting peptide is 7 amino acids in length.

18. 18. The modified AAV capsid protein of any one of claims 1 to 17, wherein the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.

19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1-9.

20. The modified AAV capsid protein of claim 18, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 5, 7, 10-16, 25, 26, 32, and 144.

21. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 4, 9, and 17-21.

22. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 3, 5, 12, and 21-26.

23. The modified AAV capsid protein of claim 18, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2-4, 7, 9, 21, 22, 27, and 28.

24. The modified AAV capsid protein of claim 18, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 5, 12, 14, 16, 21, 22, and 29-31.

25. The modified AAV capsid protein of claim 18, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1-4, 7, and 32-34.

26. The modified AAV capsid protein of claim 18, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 5, 7, 9, 12, 21, and 35-37.

27. The modified AAV capsid protein of claim 18, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2, 3, 7, 14-16, 21, 31, and 38.

28. The modified AAV capsid protein of claim 18, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 3, 4, 12, 13, 21, 30, and 39-42.

29. The modified AAV capsid protein of claim 18, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2-4, 7, 9, 21, 32, 33, and 43.

30. The modified AAV capsid protein of claim 18, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2, 3, 9, 44, and 45.

31. The modified AAV capsid protein of claim 18, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 2-4, 12, 16, 30, 46, and 47.

32. The modified AAV capsid protein of claim 18, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV1 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 1, 2, 8, 12, 21, 28, and 48-51.

33. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52-60.

34. The modified AAV capsid protein of claim 18, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 59 and 61-69.

35. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 56, 58, 60, and 70-75.

36. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 58, 60, 62, 63, 66, and 76-79.

37. The modified AAV capsid protein of claim 18, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 60, 62, 63, 66, 74-77, and 80.

38. The modified AAV capsid protein of claim 18, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 60, 75, and 81-87.

39. The modified AAV capsid protein of claim 18, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 55, 58, 60, 63, 76, 79, 88, and 89.

40. The modified AAV capsid protein of claim 18, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 58, 60, 66, 73, 76, 80, and 90-93.

41. The modified AAV capsid protein of claim 18, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 54, 57, 58, 60, 75, 79, 87, 88, and 94.

42. The modified AAV capsid protein of claim 18, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 55, 59, 60, 61, and 95-100.

43. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 53, 58-61, 63, 77, 88, 95, and 101.

44. The modified AAV capsid protein of claim 18, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52, 53, 57, 58, 75, 76, 87, 102, and 103.

45. The modified AAV capsid protein of claim 18, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 57, 58, 60, 75, 79, 87, 88, 102, 104, and 105.

46. The modified AAV capsid protein of claim 18, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV2 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 52, 55, 56, 74, 85, 88, and 106-109.

47. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the brainstem, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110-117.

48. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the caudate nucleus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 113, 115, 116, and 118-121.

49. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebellar cortex, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 119, and 122-125.

50. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the cerebral cortex, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 114, 116, and 125-127.

51. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the ependyma, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 118-120, and 128.

52. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the globus pallidus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110-112, 114, 119, 120, and 129.

53. The modified AAV capsid protein of claim 18, wherein the different brain structure is the hippocampus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 116, 123, 125, 129, and 130.

54. The modified AAV capsid protein of claim 18, wherein the different brain structure is the meninges, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 114, 118, 119, 122, and 131.

55. The modified AAV capsid protein of claim 18, wherein the different brain structure is the optic nerve, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 114, 115, 117, 129, and 132.

56. The modified AAV capsid protein of claim 18, wherein the different brain structure is the putamen, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 112, 113, 116, 123, 127, 133, and 134.

57. The modified AAV capsid protein of claim 18, wherein the different brain structure is the spinal cord, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 113, 119, 120, 122, 123, 128, and 134.

58. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the substantia nigra, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110-114, 117, and 129.

59. The modified AAV capsid protein of claim 18, wherein the different brain structure is the subthalamic nucleus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 111, 113, 119, 120, 122, 132, and 135.

60. 19. The modified AAV capsid protein of claim 18, wherein the different brain structure is the thalamus, the modified AAV capsid protein is a modified AAV9 capsid protein, and the targeting peptide is selected from SEQ ID NOs: 110, 112-114, 125, 133, 136, and 137.

61. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 1, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, hippocampus, or thalamus.

62. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 2, and the different brain structures are the brainstem, ependyma, hippocampus, optic nerve, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.

63. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 3, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, or subthalamic nucleus.

64. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 4, and the different brain structures are the brainstem, cerebellar cortex, ependyma, hippocampus, putamen, spinal cord, or subthalamic nucleus.

65. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 5, and the different brain structures are the brainstem, cerebral cortex, globus pallidus, or meninges.

66. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 6, and the different brain structure is the brainstem.

67. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 7, and the different brain structures are the brainstem, caudate nucleus, ependyma, hippocampus, meninges, optic nerve, or spinal cord.

68. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 8, and the different brain structure is the brainstem or the thalamus.

69. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 9, and the different brain structures are the brainstem, cerebellar cortex, ependyma, meninges, spinal cord, or substantia nigra.

70. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 10 or 11, and the different brain structure is the caudate nucleus.

71. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 12, and the different brain structures are the caudate nucleus, cerebral cortex, globus pallidus, meninges, putamen, subthalamic nucleus, or thalamus.

72. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 13, and the different brain structure is the caudate nucleus or putamen.

73. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 14, and the different brain structures are the caudate nucleus, the globus pallidus, or the optic nerve.

74. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 15, and the different brain structure is the caudate nucleus or the optic nerve.

75. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 16, and the different brain structures are the caudate nucleus, the globus pallidus, the optic nerve, or the subthalamic nucleus.

76. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 17 to 20, and the different brain structure is the cerebellar cortex.

77. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 21, and the different brain structures are the cerebellar cortex, cerebral cortex, ependyma, globus pallidus, meninges, optic nerve, putamen, spinal cord, or thalamus.

78. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 22, and the different brain structures are the cerebral cortex, the ependyma, or the globus pallidus.

79. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 23 to 26, and the different brain structure is the cerebral cortex.

80. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 27, and the different brain structure is the ependyma.

81. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 28, and the different brain structure is the ependyma or the thalamus.

82. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 29, and the different brain structure is the globus pallidus.

83. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 30, and the different brain structures are the globus pallidus, putamen, or subthalamic nucleus.

84. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 31, and the different brain structure is the globus pallidus or the optic nerve.

85. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 32 or 33, and the different brain structure is the hippocampus or the spinal cord.

86. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 34, and the different brain structure is the hippocampus.

87. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 35 to 37, and the different brain structure is the meninges.

88. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 38, and the different brain structure is the optic nerve.

89. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 39 to 42, and the different brain structure is the putamen.

90. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 43, and the different brain structure is the spinal cord.

91. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 44 or 45, and the different brain structure is the substantia nigra.

92. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is SEQ ID NO: 46 or 47, and the different brain structure is the subthalamic nucleus.

93. The modified AAV capsid protein of claim 1 or 7, wherein the modified AAV capsid protein is a modified AAV1 capsid protein, the targeting peptide is any one of SEQ ID NOs: 48 to 51, and the different brain structure is the thalamus.

94. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 52, and the different brain structures are the brainstem, the substantia nigra, or the thalamus.

95. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 53, and the different brain structures are the brainstem, cerebral cortex, ependyma, hippocampus, meninges, optic nerve, spinal cord, or substantia nigra.

96. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 54, and the different brain structures are the brainstem or the optic nerve.

97. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 55, and the different brain structures are the brainstem, hippocampus, putamen, or thalamus.

98. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 56, and the different brain structures are the brainstem, cerebellar cortex, or thalamus.

99. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 57, and the different brain structures are the brainstem, optic nerve, substantia nigra, or subthalamic nucleus.

100. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 58, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, hippocampus, meninges, optic nerve, spinal cord, substantia nigra, or subthalamic nucleus.

101. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 59, and the different brain structures are the brainstem, the caudate nucleus, the putamen, or the spinal cord.

102. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 60, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, or subthalamic nucleus.

103. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 61, and the different brain structures are the caudate nucleus, putamen, or spinal cord.

104. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 62, and the different brain structures are the caudate nucleus, the cerebral cortex, or the ependyma.

105. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 63, and the different brain structures are the caudate nucleus, cerebral cortex, ependyma, hippocampus, or spinal cord.

106. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 64, 65, and 67-69, and the different brain structure is the caudate nucleus.

107. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 66, and the different brain structures are the caudate nucleus, the cerebral cortex, the ependyma, or the meninges.

108. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 70 to 72, and the different brain structure is the cerebellar cortex.

109. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 73, and the different brain structure is the cerebellar cortex or the meninges.

110. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 74, and the different brain structures are the cerebellar cortex, the ependyma, or the thalamus.

111. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 75, and the different brain structures are the cerebellar cortex, ependyma, globus pallidus, optic nerve, substantia nigra, or subthalamic nucleus.

112. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 76, and the different brain structures are the cerebral cortex, ependyma, hippocampus, meninges, or substantia nigra.

113. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 77, and the different brain structures are the cerebral cortex, the ependyma, or the spinal cord.

114. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 78, and the different brain structure is the cerebral cortex.

115. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 79, and the different brain structures are the cerebral cortex, hippocampus, optic nerve, or subthalamic nucleus.

116. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 80, and the different brain structures are the ependyma, hippocampus, or meninges.

117. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 81 to 84 and 86, and the different brain structure is the globus pallidus.

118. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 85, and the different brain structure is the globus pallidus or the thalamus.

119. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 87, and the different brain structures are the globus pallidus, optic nerve, substantia nigra, or subthalamic nucleus.

120. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 88, and the different brain structures are the hippocampus, optic nerve, spinal cord, subthalamic nucleus, or thalamus.

121. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 89, and the different brain structure is the hippocampus.

122. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 90 to 93, and the different brain structure is the meninges.

123. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 94, and the different brain structure is the optic nerve.

124. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 95, and the different brain structure is the putamen or the spinal cord.

125. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 96-100, and the different brain structure is the putamen.

126. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 101, and the different brain structure is the spinal cord.

127. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 102, and the different brain structure is the substantia nigra or the subthalamic nucleus.

128. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 103, and the different brain structure is the substantia nigra.

129. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is SEQ ID NO: 104 or 105, and the different brain structure is the subthalamic nucleus.

130. The modified AAV capsid protein of claim 1 or 11, wherein the modified AAV capsid protein is a modified AAV2 capsid protein, the targeting peptide is any one of SEQ ID NOs: 106 to 109, and the different brain structure is the thalamus.

131. 16. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 110, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.

132. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 111, and the different brain structures are the brainstem, cerebellar cortex, cerebral cortex, ependyma, globus pallidus, hippocampus, meninges, optic nerve, substantia nigra, or subthalamic nucleus.

133. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 112, and the different brain structures are the brainstem, globus pallidus, putamen, substantia nigra, or thalamus.

134. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 113, and the different brain structures are the brainstem, caudate nucleus, cerebellar cortex, cerebral cortex, ependyma, hippocampus, meninges, putamen, spinal cord, substantia nigra, subthalamic nucleus, or thalamus.

135. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 114, and the different brain structures are the brainstem, cerebral cortex, globus pallidus, meninges, optic nerve, substantia nigra, or thalamus.

136. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 115, and the different brain structure is the brainstem or the caudate nucleus.

137. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 116, and the different brain structures are the brainstem, caudate nucleus, cerebral cortex, hippocampus, optic nerve, or putamen.

138. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 117, and the different brain structures are the brainstem, the optic nerve, or the substantia nigra.

139. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 118, and the different brain structures are the caudate nucleus, the ependyma, or the meninges.

140. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 119, and the different brain structures are the caudate nucleus, cerebellar cortex, ependyma, globus pallidus, meninges, spinal cord, or subthalamic nucleus.

141. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 120, and the different brain structures are the caudate nucleus, the ependyma, the globus pallidus, the meninges, the spinal cord, or the subthalamic nucleus.

142. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 121, and the different brain structure is the caudate nucleus.

143. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 122, and the different brain structures are the cerebellar cortex, meninges, spinal cord, or subthalamic nucleus.

144. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 123, and the different brain structures are the cerebellar cortex, hippocampus, putamen, or spinal cord.

145. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 124, and the different brain structure is the cerebellar cortex.

146. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 125, and the different brain structures are the cerebellar cortex, the cerebral cortex, the hippocampus, or the thalamus.

147. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 126, and the different brain structure is the cerebral cortex.

148. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 127, and the different brain structure is the cerebral cortex or putamen.

149. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 128, and the different brain structure is the ependyma or the spinal cord.

150. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 129, and the different brain structures are the globus pallidus, hippocampus, optic nerve, or substantia nigra.

151. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 130, and the different brain structure is the hippocampus.

152. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 131, and the different brain structure is the meninges.

153. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 132, and the different brain structure is the optic nerve or the subthalamic nucleus.

154. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 133, and the different brain structure is the putamen or thalamus.

155. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 134, and the different brain structure is the putamen or the spinal cord.

156. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 135, and the different brain structure is the subthalamic nucleus.

157. The modified AAV capsid protein of claim 1 or 15, wherein the modified AAV capsid protein is a modified AAV9 capsid protein, the targeting peptide is SEQ ID NO: 136 or 137, and the different brain structure is the thalamus.

158. 158. A nucleic acid comprising a sequence encoding the modified capsid protein of any one of claims 1 to 157.

159. 158. A recombinant adeno-associated virus (rAAV) virus comprising the modified capsid protein of any one of claims 1-157.

160. 158. A viral vector comprising a nucleic acid encoding the modified capsid protein of any one of claims 1-157.

161. 161. The viral vector of claim 160, further comprising a nucleic acid sequence encoding a nucleic acid of interest.

162. 162. The viral vector of claim 161, wherein said nucleic acid of interest is a therapeutic agent.

163. 163. The viral vector of claim 162, wherein the therapeutic agent is an enzyme or an RNAi molecule.

164. A cell comprising the viral vector of any one of claims 160-163.

165. The cell of claim 164, which is a mammalian cell.

166. The cell of claim 164, which is a human cell.

167. The cell of claim 164, which is in vitro.

168. The cell of claim 164, which is in vivo.

169. 160. A pharmaceutical composition comprising the viral vector of claim 159 and a pharmaceutically acceptable carrier.

170. 160. A method for delivering an agent to different brain structures in a subject, comprising administering to the subject the virus of claim 159.

171. The method of claim 170, wherein the method is for delivering an agent to the brainstem of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1-9, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52-60, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-117.

172. The method of claim 170, wherein the method is for delivering an agent to the caudate nucleus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 7, and 10-16, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 59 and 61-69, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 113, 115, 116, and 118-121.

173. The method of claim 170, wherein the method is for delivering an agent to the cerebellar cortex of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 4, 9, and 17-21, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 56, 58, 60, and 70-75, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 119, and 122-125.

174. The method of claim 170, wherein the method is for delivering an agent to the cerebral cortex of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 3, 5, 12, and 21-26, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58, 60, 62, 63, 66, and 76-79, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 114, 116, and 125-127.

175. The method of claim 170, wherein the method is for delivering an agent to the ependyma of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 7, 9, 21, 22, 27, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 60, 62, 63, 66, 74-77, and 80; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 118-120, and 128.

176. The method of claim 170, wherein the method is for delivering an agent to the globus pallidus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 5, 12, 14, 16, 21, 22, and 29-31, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 60, 75, and 81-87, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-112, 114, 119, 120, and 129.

177. 171. The method of claim 170, wherein the method is for delivering an agent to the hippocampus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1-4, 7, and 32-34, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 55, 58, 60, 63, 76, 79, 80, 88, and 89; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 116, 123, 125, 129, and 130.

178. The method of claim 170, wherein the method is for delivering an agent to the meninges of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 5, 7, 9, 12, 21, and 35-37, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58, 60, 66, 73, 76, 80, and 90-93; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 114, 118, 119, 122, and 131.

179. The method of claim 170, wherein the method is for delivering an agent to the optic nerve of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2, 3, 7, 14-16, 21, 31, and 38, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 54, 57, 58, 60, 75, 79, 87, 88, and 94, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 114, 115, 117, 129, and 132.

180. The method of claim 170, wherein the method is for delivering an agent to the putamen of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 3, 4, 12, 13, 21, 30, and 39-42, an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 55, 59, 60, 61, and 95-100, or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 112, 113, 116, 123, 127, 133, and 134.

181. 171. The method of claim 170, wherein the method is for delivering an agent to the spinal cord of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 7, 9, 21, 32, 33, and 43; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 53, 58-61, 63, 77, 88, 95, and 101; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 113, 119, 120, 122, 123, 128, and 134.

182. The method of claim 170, wherein the method is for delivering an agent to the substantia nigra of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2, 3, 9, 44, and 45, and 28; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52, 53, 57, 58, 75, 76, 87, 102, and 103; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110-114, 117, and 129.

183. The method of claim 170, wherein the method is for delivering an agent to the subthalamic nucleus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 2-4, 12, 16, 30, 46, and 47; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 57, 58, 60, 75, 79, 87, 88, 102, 104, and 105; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 111, 113, 119, 120, 122, 132, and 135.

184. The method of claim 170, wherein the method is for delivering an agent to the thalamus of a subject, the method comprising administering an AAV1 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 1, 2, 8, 12, 21, 28, and 48-51; an AAV2 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 52, 55, 56, 74, 85, 88, and 106-109; or an AAV9 virus comprising a modified capsid protein having a targeting peptide selected from SEQ ID NOs: 110, 112-114, 125, 133, 136, and 137.

185. The method of any one of claims 170-184, wherein said agent is an siRNA, shRNA, miRNA, non-coding RNA, lncRNA, therapeutic protein, or a CRISPR system.

186. 185. The method of any one of claims 170-184, wherein said administering is to the central nervous system.

187. 187. The method of claim 186, wherein said administration is to the cisterna magna, intraventricular space, ependyma, ventricle, subarachnoid space, cochlea, and / or intrathecal space.

188. 188. The method of claim 187, wherein said ventricles are the rostral lateral ventricle, and / or the caudal lateral ventricle, and / or the right lateral ventricle, and / or the left lateral ventricle, and / or the right rostral lateral ventricle, and / or the left rostral lateral ventricle, and / or the right caudal lateral ventricle, and / or the left caudal lateral ventricle.

189. The method of any one of claims 170-188, wherein a plurality of viral particles is administered.

190. The virus has a vector genome content of about 1 x 10 per kilogram 6 ~Approx. 1×10 18 190. The method of claim 189, wherein the dose is administered in a dose of (vg / kg).

191. The virus is present in an amount of approximately 1 x 10 vg per kg of patient. 7 ~1×10 17 , about 1×10 8 ~1×10 16 , about 1×10 9 ~1×10 15 , about 1×10 10 ~1×10 14 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 13 , about 1×10 10 ~1×10 11 , about 1×10 11 ~1×10 12 , about 1×10 12 ~×10 13 , or approximately 1 × 10 13 ~1×10 14 190. The method of claim 189, wherein the dose is administered in individual doses.

192. The method of any one of claims 170 to 191, wherein the subject is a human.

193. 160. A method of treating a disease in a mammal comprising administering to the mammal the virus of claim 159.

194. 194. The method of claim 193, wherein the disease is a neurodegenerative disease.

195. 195. The method of claim 194, wherein the neurodegenerative disease is Huntington's disease, ALS, hereditary spastic hemiplegia, primary lateral sclerosis, spinal muscular atrophy, Kennedy's disease, Alzheimer's disease, polyglutamine repeat disease, or Parkinson's disease.

196. 194. The method of claim 193, wherein the mammal is a human.

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