Compositions and methods for the treatment of disorders related to cdkl5 deficiency

Abstract

The present disclosure relates to compositions and methods for altering, e.g., enhancing, expression of the CDKL5 protein via delivery of adeno-associated virus (AAV) capsid variants. The compositions and methods of the present disclosure are useful for treating subjects diagnosed with or suspected of having CDD or another CDKL5-related disorder.

Description

(19) State Intellectual Property Office (12) Invention Patent Application (10) Application Publication Number (43) Application Publication Date (21) Application Number 202480027232.8 (22) Application Date 2024.05.01 (30) Priority Data 63 / 499,933 2023.05.03 US 63 / 593,830 2023.10.27 US 63 / 564,457 2024.03.12 US (85) PCT International Application Entering National Phase Date 2025.10.21 (86) PCT International Application Application Data PCT / US2024 / 027313 2024.05.01 (87) PCT International Application Publication Data WO2024 / 229163 EN 2024.11.07 (71) Applicant: Voyager Therapeutics, Inc. Address: Massachusetts, USA (72) Inventors: M.E. Nonnenmacher, T.C. Moyer, J. Lee, D.R. Lax (74) Patent Agency: Longtian Intellectual Property Agency Co., Ltd., 72003 Patent Attorney: Fu Wenchuan (51) Int.Cl. C07K 14 / 005 (2006.01) C12N 15 / 86 (2006.01) (54) Invention Title: Compositions and Methods for Treating Diseases Related to CDKL5 Deficiency (57) Abstract: This disclosure relates to compositions and methods for altering, for example enhancing, the expression of CDKL5 protein by delivery via an adeno-associated virus (AAV) capsid variant. The compositions and methods of this disclosure are suitable for treating subjects diagnosed with or suspected of having CDD or another CDKL5-related disease. Claims 9 pages, Description 285 pages, Sequence Listing (electronic publication), Drawings 12 pages, CN 121079310 A 2025.12.05 CN 1 21 07 93 10 A 1. An adeno-associated virus (AAV) particle comprising: a) an AAV capsid variant comprising an amino acid sequence having the following formula: [N1]-[N2]-[N3], wherein: (i) optionally, [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G; (ii) [N2] comprises an amino acid sequence of SPH; and (iii) [N3] comprises X4, X5, and X6, wherein at least one of X4, X5, or X6 is a basic amino acid; and b) a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence. 2. The AAV particle of claim 1, wherein the amino acid sequence [N1]-[N2]-[N3] is located in the hypervariable ring IV of the AAV capsid variant. 3. The AAV particle of claim 1 or claim 2, wherein the AAV capsid variant is an AAV9 capsid variant.4. The AAV particle of any one of claims 1 to 3, wherein [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G. 5. The AAV particle of any one of claims 1 to 4, wherein [N2]-[N3] comprises the amino acid sequence of SPHSKA (SEQ ID NO: 941). 6. An adeno-associated virus (AAV) particle comprising a viral genome containing a cyclin-dependent kinase-like 5 (CDKL5) coding sequence and an AAV9 capsid variant comprising the amino acid sequence of SPHSKA (SEQ ID NO: 941). 7. The AAV particle of claim 6, wherein the amino acid sequence of SPHSKA (SEQ ID NO: 941) is located in the hypervariable loop IV of the AAV9 capsid variant. 8. The AAV particle of claim 6 or claim 7, wherein the amino acid sequence of SPHSKA (SEQ ID NO: 941) is immediately following the amino acid position corresponding to position 455 of SEQ ID NO: 4 or SEQ ID NO: 36. 9. The AAV particle of any one of claims 6 to 8, wherein the AAV9 capsid variant further comprises one, two, or all of the following: N at the amino acid position corresponding to position 452 of SEQ ID NO: 4, E at the amino acid position corresponding to position 451 of SEQ ID NO: 4, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO: 4. 10. The AAV particle of any one of claims 6 to 9, wherein the AAV9 capsid variant comprises the amino acid sequence KTENVSGSPHSKAQNQQT (SEQ ID NO: 3272). 11. The AAV particle of any one of claims 6 to 10, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence having at least 90% identity with positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising an amino acid sequence having at least 90% identity with positions 203-742 of SEQ ID NO: 4. 12. The AAV particle of any one of claims 6 to 11, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 95% identity with SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence having at least 95% identity with positions 138-742 of SEQ ID NO: 4; and / or (iii)VP3 protein, comprising an amino acid sequence having at least 95% identity with the amino acid sequence at positions 203-742 of SEQ ID NO: 4. (Claim 1 / 9 page 2 CN 121079310 A) 13. The AAV particle of any one of claims 6 to 12, wherein the AAV9 capsid variant comprises: (i) a VP1 protein, comprising an amino acid sequence having at least 99% identity with the amino acid sequence at positions 138-742 of SEQ ID NO: 4; (ii) a VP2 protein, comprising an amino acid sequence having at least 99% identity with the amino acid sequence at positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein, comprising an amino acid sequence having at least 99% identity with the amino acid sequence at positions 203-742 of SEQ ID NO: 4. 14. The AAV particle of any one of claims 6 to 13, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 4. 15. The AAV particle of any one of claims 6 to 13, wherein the AAV9 capsid variant comprises: (i) the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 4; (ii) an E at the amino acid position corresponding to position 451 of SEQ ID NO: 4 and a V at the amino acid position corresponding to position 453 of SEQ ID NO: 4; and (iii) no other modifications relative to wild-type AAV9. 16. The AAV particle of any one of claims 6 to 8, wherein the AAV9 capsid variant further comprises one, two, or all of the following: E ​​at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO: 36. 17. The AAV particle of any one of claims 6 to 8 and 16, wherein the AAV9 capsid variant comprises the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589). 18. The AAV particle of any one of claims 6 to 8, 16, and 17, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 36; (ii)19. The AAV particle of any one of claims 6 to 8 and 16 to 18, wherein the AAV9 capsid variant comprises: (i) a VP1 protein containing an amino acid sequence having at least 95% identity with SEQ ID NO: 36; (ii) a VP2 protein containing an amino acid sequence having at least 95% identity with SEQ ID NO: 36; and / or (iii) a VP3 protein containing an amino acid sequence having at least 95% identity with SEQ ID NO: 36. 20. The AAV particle according to any one of claims 6 to 8 and 16 to 19, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 99% identity with SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence having at least 99% identity with positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence having at least 99% identity with positions 203-742 of SEQ ID NO: 36. 21. The AAV particle according to any one of claims 6 to 8 and 16 to 20, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 36. 22. The AAV particle of any one of claims 6 to 8 and 16 to 20, wherein the AAV9 capsid variant comprises: (i) the amino acid sequence SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 36; (ii) E at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and V at the amino acid position corresponding to position 453 of SEQ ID NO: 36; and (iii) no other modifications relative to wild-type AAV9.23. The AAV particle of any one of claims 1 to 4, wherein [N1]-[N2]-[N3] are immediately following the position corresponding to amino acid position 452 of SEQ ID NO: 982; and wherein the AAV capsid variant comprises an amino acid sequence that is at least 90% identical to the amino acid sequence of SEQ ID NO: 982, for example, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence at positions 203-742 of SEQ ID NO: 982. 24. The AAV particle of claim 23, wherein [N1] comprises GHD. 25. The AAV particle of claim 23 or claim 24, wherein [N1] comprises amino acid G at position 453 of SEQ ID NO: 138 or SEQ ID NO: 982, amino acid H at position 454 of SEQ ID NO: 138 or SEQ ID NO: 982, and amino acid D at position 455. 26. The AAV particle of any one of claims 23 to 25, wherein [N3] comprises KSG. 27. The AAV particle of any one of claims 23 to 26, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity with positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity with positions 203-742 of SEQ ID NO: 982. 28. The AAV particle of any one of claims 23 to 27, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with positions 203-742 of SEQ ID NO: 982.The amino acid sequence at positions 203-742 of SEQ ID NO: 982 has at least 95% identity. 29. The AAV particle of any one of claims 23 to 28, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with SEQ ID NO: 982. 30. The AAV particle of any one of claims 23 to 29, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 982. 31. The AAV particle of any one of claims 1 to 30, wherein the viral genome encodes a wild-type CDKL5 protein or a fragment thereof. 32. The AAV particle of any one of claims 1 to 31, wherein the viral genome encodes a human CDKL5 protein. 33. The AAV particle of claim 31 or claim 32, wherein the CDKL5 protein comprises the amino acid sequence of SEQ ID NO: 6413. 34. The AAV particle of any one of claims 1 to 33, wherein the CDKL5 coding sequence comprises a nucleotide sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to SEQ ID NO: 6414. 35. The AAV particle of claim 34, wherein the CDKL5 coding sequence comprises a nucleotide sequence that is at least 95% identical to SEQ ID NO: 6414. 36. The AAV particle of claim 35, wherein the CDKL5 coding sequence comprises a nucleotide sequence that is at least 99% identical to SEQ ID NO: 6414. 37. The AAV particle of claim 36, wherein the CDKL5 coding sequence comprises a nucleotide sequence that is at least 99% identical to SEQ ID NO: 6414.38. The AAV particle of claim 37, wherein the CDKL5 coding sequence comprises the nucleotide sequence of SEQ ID NO: 6414. 39. The AAV particle of any one of claims 1 to 38, wherein the viral genome contains a promoter effectively linked to the CDKL5 coding sequence. 40. The AAV particle of claim 39, wherein the promoter is a human elongation factor 1α-subunit (EF1α), a cytomegalovirus (CMV) immediate early enhancer and / or promoter, a chicken β-actin (CBA) promoter, a CAG promoter, a CAG derivative promoter, a β-glucuronidase (GUSB) promoter, a ubiquitin C (UBC) promoter, a neuron-specific enolase (NSE) promoter, a platelet-derived growth factor (PDGF) promoter, a platelet-derived growth factor B chain (PDGF-β) promoter, an intercellular adhesion molecule 2 (ICAM-2) promoter, a synaptic protein (Syn) promoter, a synaptic protein 1 (Syn1) promoter, a methyl-CpG-binding protein 2 (MeCP2) promoter, a Ca2+ / calmodulin-dependent protein kinase II (CaMKII) promoter, or a metabolite glutamate receptor 2. (mGluR2) promoter, neurofilament light chain (NFL) or heavy chain (NFH) promoter, β-globin small gene nβ2 promoter, proenkephalinogenin (PPE) promoter, enkephalin (Enk) and excitatory amino acid transporter 2 (EAAT2) promoter, glial fibrillary acidic protein (GFAP) promoter, myelin basic protein (MBP) promoter, cardiovascular promoters (e.g., αMHC, cTnT, and CMV-MLC2k), liver promoters (e.g., hAAT, TBG), skeletal muscle promoters (e.g., desmin, MCK, C512), or fragments thereof, such as truncated or functional variants. 41. The AAV particle according to any one of claims 1 to 40, wherein the viral genome further comprises a reversed end-of-claim A-terminal repeat (ITR) sequence. 42. The AAV particle of claim 41, wherein the viral genome comprises an ITR sequence located at 5' relative to the CDKL5 coding sequence. 43. The AAV particle of claim 41 or claim 42, wherein the viral genome comprises an ITR sequence located at 3' relative to the CDKL5 coding sequence. 44. The AAV particle of any one of claims 41 to 43, wherein the viral genome comprises an ITR sequence located at 5' relative to the CDKL5 coding sequence and an ITR sequence located at 3' relative to the CDKL5 coding sequence. 45. An adeno-associated virus (AAV) particle comprising a cyclin-dependent kinase-like 5'-12 ...A viral genome encoding a cyclin-dependent kinase-like 5 (CDKL5) sequence, and an AAV capsid variant comprising: (i) a VP1 protein containing the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein containing the amino acid sequence of positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein containing the amino acid sequence of positions 203-742 of SEQ ID NO: 4. 46. An adeno-associated virus (AAV) particle comprising a viral genome encoding a cyclin-dependent kinase-like 5 (CDKL5) sequence, and an AAV capsid variant comprising: (i) a VP1 protein containing the amino acid sequence of SEQ ID NO: 36; (ii) a VP2 protein containing the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein containing the amino acid sequence of positions 203-742 of SEQ ID NO: 36. 47. A cell comprising AAV particles as described in any one of claims 1 to 46, optionally wherein the cell is a mammalian cell (e.g., HEK293 cell), an insect cell (e.g., Sf9 cell), or a bacterial cell. 48. A method for preparing AAV particles as described in any one of claims 1 to 46, the method comprising: (i) providing a cell comprising a viral genome containing a CDKL5 coding sequence and nucleic acid encoding the AAV capsid variant; and (ii) culturing the cell under conditions suitable for encapsulating the viral genome within the AAV capsid variant; thereby preparing the AAV particles. 49. The method of claim 48, wherein: (a) the viral genome comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical); and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 4; and (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 4 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 4. Positions 138-742 have at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, to...).(iii) An amino acid sequence having at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with SEQ ID NO: 4; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 4 or an amino acid sequence having at least 90% identity with positions 203-742 of SEQ ID NO: 4 (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity). 50. The method of claim 49, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 4, the amino acid sequence at positions 138-742 of SEQ ID NO: 4, and / or the amino acid sequence at positions 203-742 of SEQ ID NO: 4. 51. The method of claim 48, wherein: (a) the viral genome comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical); and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 36; and (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 36 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: The amino acid sequence at positions 138-742 of SEQ ID NO: 36 has at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity); or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 36 or an amino acid sequence having at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity). 52. The method of claim 51, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 36, SEQ ID NO:The amino acid sequence at position 138-742 of SEQ ID NO: 36, and / or the amino acid sequence at position 203-742 of SEQ ID NO: 36. 53. The method of claim 48, wherein: (a) the viral genome comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical); and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity) with SEQ ID NO: 982; and (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity) with SEQ ID NO: The amino acid sequence at positions 138-742 of SEQ ID NO: 982 has at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity); or (iii) the VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or having at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity) with the amino acid sequence at positions 203-742 of SEQ ID NO: 982. 54. The method of claim 53, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 982, the amino acid sequence at positions 138-742 of SEQ ID NO: 982, and / or the amino acid sequence at positions 203-742 of SEQ ID NO: 982. 55. The method of any one of claims 48 to 54, further comprising introducing a first nucleic acid molecule comprising the viral genome into the cell prior to step (i). 56. The method of any one of claims 48 to 55, wherein the cell comprises a second nucleic acid molecule encoding the AAV capsid variant, optionally further comprising introducing the second nucleic acid molecule into the cell prior to step (i). 57. The method of any one of claims 48 to 56, wherein the cell comprises mammalian cells (e.g.,58. A pharmaceutical composition comprising AAV particles as described in any one of claims 1 to 46 and a pharmaceutically acceptable excipient. 59. A pharmaceutical composition comprising AAV particles as described in any one of claims 5 to 22 and a pharmaceutically acceptable excipient. 60. A pharmaceutical composition comprising AAV particles as described in any one of claims 9 to 15 and 45 and a pharmaceutically acceptable excipient. 61. A pharmaceutical composition comprising AAV particles as described in any one of claims 16 to 22 and 46 and a pharmaceutically acceptable excipient. 62. A method of delivering AAV particles encoding the CDKL5 protein to a subject, comprising administering to the subject an effective amount of the pharmaceutical composition as described in any one of claims 58 to 61 or the AAV particles as described in any one of claims 1 to 46. 63. The method of claim 62, wherein the subject has, has been diagnosed with, or is at risk of having a CDKL5-related condition, optionally wherein the CDKL5-related condition is a CDKL5-related neurodegeneration or neuromuscular disease. 64. The method of claim 62 or claim 63, wherein the subject has, has been diagnosed with, or is at risk of having: CDD, developmental and epileptic encephalopathy 2, or atypical Rett syndrome. 65. A method of treating a subject who has or has been diagnosed with a CDKL5-related condition, comprising administering to the subject an effective amount of the pharmaceutical composition of any one of claims 58 to 61 or AAV particles of any one of claims 1 to 46. 66. A method of treating a subject who has or has been diagnosed with a CDKL5-related condition, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 60 or AAV particles of any one of claims 9 to 15 and 45. 67. A method of treating a subject who has or has been diagnosed with a CDKL5-related disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 61 or AAV particles as described in any one of claims 16 to 22 and 46. 68. A method of treating a subject who has or has been diagnosed with a CDKL5-related disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 59 or AAV particles as described in any one of claims 5 to 22. 69. The method of claim 35, wherein the CDKL5-related disorder is a CDKL5-related neurodegeneration or neuromuscular disorder. 70. The method of claim 69, wherein the CDKL5-related neurodegeneration or neuromuscular disorder is CDD, developmental and epileptic encephalopathy 2, or atypical Rett syndrome.71. A method of treating a subject suffering from or diagnosed with CDKL5 deficiency (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition as described in any one of claims 58 to 61 or AAV particles as described in any one of claims 1 to 46. 72. A method of treating a subject suffering from or diagnosed with CDKL5 deficiency (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition as described in claim 60 or AAV particles as described in any one of claims 9 to 15 and 45. 73. A method of treating a subject suffering from or diagnosed with CDKL5 deficiency (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition as described in claim 61 or AAV particles as described in any one of claims 16 to 22 and 46. 74. A method of treating a subject with or diagnosed with CDKL5 deficiency (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition of claim 59 or AAV particles of any one of claims 5 to 22. 75. The method of any one of claims 32 to 38, wherein the subject has one or more mutations in the CDKL5 gene. 76. The method of any one of claims 62 to 75, wherein the subject has a reduced CDKL5 activity level compared to a reference level for a subject without CDKL5-related disease. 77. The method of any one of claims 65 to 76, wherein the administration halts the progression of the subject's disease. 78. The method of any one of claims 65 to 77, wherein the administration improves at least one symptom of the disease and / or alters one or more biomarkers of the disease. 79. The method of claim 78, wherein the one or more biomarkers comprise markers of neurofilament light chain or CDKL5 activity, such as those measured by phosphorylation levels of substrate proteins, such as MECP2, or as measured by mass spectrometry. 80. The method of any one of claims 65 to 77, wherein the treatment improves at least one symptom, optionally, wherein the at least one symptom includes epilepsy (e.g., early-onset epilepsy), autism, cognitive deficits, motor skill limitations, sleep difficulties, visual impairment, low muscle tone, gastrointestinal reflux, behavioral symptoms (including unexplained laughter or crying, touch sensitivity, and sleep disturbances), facial appearance changes (including microcephaly, high and wide forehead, large and deep-set eyes, a smaller-than-normal distance between the nose and upper lip, upturned nose, full lips, and wide-set teeth), difficulty standing and walking, small and cold feet,Lack or reduced eye contact, frequent strabismus, cortical visual impairment or cortical blindness, bruxism, speech limitation or aphasia, difficulty eating, stereotypes, limited fine motor skills, gastroesophageal reflux, constipation, or a combination thereof. 81. The method of any one of claims 62 to 80, wherein the subject is a human. 82. The method of any one of claims 62 to 81, wherein the AAV particles are delivered to cells, tissues, or regions of the CNS, such as regions of the brain or spinal cord, such as the parenchyma, cortex, substantia nigra, caudate nucleus, cerebellum, striatum, corpus callosum, cerebellum, brainstem, caudate nucleus-putamen, thalamus, superior colliculus, spinal cord, or a combination thereof, and / or delivered to neurons, or a combination thereof. 83. The method of any one of claims 62 to 82, further comprising assessing, for example, measuring, the level of CDKL5 expression in the subject, such as the level of CDKL5 gene expression, CDKL5 mRNA expression, and / or CDKL5 protein expression. 84. The method of claim 83, wherein the CDKL5 protein expression level is measured by ELISA, Western blotting, or immunohistochemistry. 85. The method of claim 83 or claim 84, wherein the CDKL5 expression level is assessed before and after administration of the AAV particles, optionally, wherein the CDKL5 expression level before administration is compared with the CDKL5 expression level after administration. 86. The method of any one of claims 83 to 85, comprising assessing the CDKL5 expression level in cells or tissues of the central nervous system (e.g., the parenchyma). 87. The method of any one of claims 83 to 86, wherein the CDKL5 protein expression level of the subject increases after administration relative to the CDKL5 protein expression level of the subject before administration. 88. The method of any one of claims 62 to 87, further comprising assessing, for example, measuring, the CDKL5 activity level in the subject, for example, in the subject's cells or tissues. 89. The method of any one of claims 65 to 88, wherein the administration results in an increase in: (i) CDK L5 activity in the subject's cells, tissues (e.g., cells or tissues of the CNS, such as the cortex, striatum, thalamus, cerebellum, and / or brainstem) and / or body fluids (e.g., CSF and / or serum) relative to CDK L5 activity in the subject prior to the administration; (ii) the level of viral genome (VG) in each cell of the subject's CNS tissues (e.g., cortex, striatum, thalamus, cerebellum, brainstem, and / or spinal cord) relative to the level of viral genome (VG) in each cell of the subject's peripheral tissues; and / or (iii) CDK L5 activity compared to CDK L5 activity in the subject prior to the administration.mRNA expression, CDKL5 mRNA expression in the subject's cells or tissues (e.g., cells or tissues of the CNS, such as the cortex, thalamus, and / or brainstem). 90. The method of any one of claims 65 to 89, further comprising administering to the subject an additional agent suitable for treating or preventing CDKL5-related conditions; optionally, wherein the additional agent comprises one or more antiepileptic drugs (e.g., valproic acid, levetiracetam, clobazian, lamotrigine, ganaxolone, topiramate, or combinations thereof). 91. The method of any one of claims 65 to 90, further comprising administering to the subject an immunosuppressant. 92. The method of claim 91, wherein the immunosuppressant comprises a corticosteroid (e.g., prednisone, prednisolone, methylprednisolone, and / or dexamethasone), rapamycin, mycophenolate mofetil, tacrolimus, rituximab, and / or ikuzumab hydroxychloroquine. 93. The pharmaceutical composition of any one of claims 58 to 61 or the AAV particles of any one of claims 1 to 46, for the treatment of CDKL5-related disorders; optionally, wherein the CDKL5-related disorder is CDKL5 deficiency disorder (CDD), developmental and epileptic encephalopathy 2, or atypical Rett syndrome. 94. Use of the pharmaceutical composition of any one of claims 58 to 61 or the AAV particles of any one of claims 1 to 46 in the preparation of a medicament for the treatment of CDKL5-related disorders; optionally, wherein the CDKL5-related disorder is CDKL5 deficiency disorder (CDD), developmental and epileptic encephalopathy 2, or atypical Rett syndrome. Claims 9 / 9 Page 10 CN 121079310 A Composition and method for treating conditions related to CDKL5 deficiency

[0001] Related Applications

[0002] This application claims the benefit and priority of U.S. Provisional Application Serial No. 63 / 499,933, filed May 3, 2023; U.S. Provisional Application Serial No. 63 / 593,830, filed October 27, 2023; and U.S. Provisional Application Serial No. 63 / 564,457, filed March 12, 2024, the contents of each of which are incorporated herein by reference in their entirety.

[0003] Serial List

[0004] This application is filed with an electronic serial list. The serial list file named 14640_0086-00304_SL.xml was created on March 27, 2024, and is 5,493,990 bytes in size. Information in the electronic sequence listing is incorporated herein by reference in its entirety. Technical Field

[0005] This document describes and delivers polynucleotides, such as those encoding cyclin-dependent kinase-like 5 (CDKL5) proteins.Compositions and methods relating to adeno-associated virus (AAV) viral particles of polynucleotides (“CDKL5 protein”) and peptides (“CDKL5 peptides”) for the treatment of CDKL5 deficiency (CDD) and other CDKL5-related conditions, including developmental and epileptic encephalopathy 2 and atypical Rett syndrome (collectively, “CDKL5-related conditions”). In some embodiments, the compositions described herein may be used to treat subjects in need, such as human subjects diagnosed with CDKL5-related conditions or other conditions caused by a deficiency in the amount and / or function of CDKL5 protein.

[0006] Prior Art

[0007] Cyclin-dependent kinase-like 5 (CDKL5) is a serine / threonine protein kinase and is also known as serine / threonine kinase 9 (STK9). Other aliases for CDKL5 include EIEE2, ISSX, CFAP247, and DEE2. It is encoded by the gene CDKL5 (Ensembl gene ID ENSG00000008086) located on the X chromosome.

[0008] The CDKL5 protein is an enzyme and is believed to play an important role in brain development and the regulation of responses to oxidative stress. CDKL5 is believed to be expressed throughout the cell, including in the cell body and the nucleus and cytoplasm of the dendrites.

[0009] CDKL5 is responsible for the phosphorylation of many targets. CDKL5 can target and phosphorylate the gene MECP2, which has been characterized as essential for the function of neurons and other brain cells and the maintenance of neuronal synapses, and is believed to be the causative agent of Rett syndrome. CDKL5 can also target and phosphorylate CEP131, which is believed to play a role in cell proliferation. CDKL5 can also target and phosphorylate MAP1S, DLG5, EB2 and / or ARHGEF, which are microtubule-associated proteins and are believed to play a role in neuronal division, differentiation, migration and neurite growth. CDKL5 can also target and phosphorylate AKT and mTOR, which are believed to play a role in cell proliferation, migration, and development.

[0010] Without wishing to be bound by any particular theory, CDKL5 may be involved in the formation, growth, and migration of neurons. It may also play a role in cell division and / or chemical signaling at neuronal synapses.

[0011] Mutations in CDKL5 are known to cause disease in subjects, such as human subjects. CDKL5 mutations result in CDKL5 deficiency syndrome (CDD). CDD is a neurodevelopmental disorder. It is characterized by neurological symptoms, including epilepsy (e.g., early-onset epilepsy), autism, cognitive deficits, motor skill limitations, sleep difficulties, and / or visual impairments. It is also characterized by low muscle tone and gastrointestinal reflux.

[0012] CDD can present with a wide range of clinical manifestations. These include behavioral symptoms such as episodes of laughing or crying for no reason, hypersensitivity to touch, and sleep disturbances. Clinical manifestations also include facial appearance changes, including microcephaly, a high, broad forehead, large, deep-set eyes, a smaller-than-normal distance between the nose and upper lip, an upturned nose, full lips, and widely spaced teeth. Other clinical manifestations include difficulty standing and walking, small, cold feet, lack of or poor eye contact, frequent sideways glances, and cortical visual disturbances or cortical blindness. Other manifestations include bruxism, speech impairment or aphasia, difficulty eating, stereotypies, limited ability to make small, focused hand movements, gastroesophageal reflux, and constipation.

[0013] The incidence of CDD in the United States is 1 in 42,000 newborns, and 85% of cases occur in females. Typically, patients with CDD are completely dependent on caregivers for life.

[0014] CDD is usually caused by a de novo mutation in CDKL5. CDKL5 mutations in CDD patients result in a reduction in the amount of functional CDKL5 protein.

[0015] Current treatments for CDD patients focus on relieving symptoms, such as seizures. To date, treatments available for CDD patients are limited, and delivering therapeutic drugs to the central nervous system (CNS) remains a major challenge in developing new and effective therapies.

[0016] The current standard of care for CDD is first-line treatment with antiepileptic drugs. Antiepileptic drugs are known in this technology and include pentylpropionate and levetiracetam. Second-line treatment for refractory patients includes first-line therapy plus additional antiepileptic drugs such as clobazam and / or lamotrigine and ganaxolone. However, there are reports of short-duration tolerance of ganaxolone in patients with CDD. Third...Line therapy introduces additional antiepileptic drugs, such as topiramate, and / or alternative interventions, such as a ketogenic diet and / or vagus nerve stimulation. However, seizures in patients with CDD are often refractory to known treatments.

[0017] Therefore, there is still a need for modified therapies and treatments that target the etiology of CDD. Specifically, there is still a need for pharmaceutical compositions and methods for treating CDD that can be delivered to the CNS to treat CDD and improve CDKL5 deficiency in subjects (e.g., human subjects).

[0018] Previous attempts to provide AAV capsids with modified properties (e.g., modified tropism suitable for delivery to the brain or CNS) have had limited success. Therefore, there is still a need for effective treatments using AAV capsid variants capable of delivering the payload of interest (e.g., CDKL5) to target cells or tissues (e.g., CNS cells or tissues).

[0019] In some embodiments, this disclosure provides an adeno-associated virus (AAV) particle comprising: a) an AAV capsid variant comprising an amino acid sequence having the formula [N1]-[N2]-[N3], wherein: (i) optionally [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G; (ii) [N2] comprises an amino acid sequence of SPH; and (iii) [N3] comprises X4, X5, and X6, wherein at least one of X4, X5, or X6 is a basic amino acid; and b) a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence. In some embodiments, the amino acid sequence [N1]-[N2]-[N3] is located in the hypervariable loop IV of the AAV capsid variant. In some embodiments, the AAV capsid variant is an AAV9 capsid variant. In some embodiments, [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G. In some embodiments, [N2]-[N3] comprises the amino acid sequence of SPHSKA (SEQ ID NO: 941).

[0020] In some embodiments, this disclosure provides an adeno-associated virus (AAV) particle comprising a viral genome containing a cyclin-dependent kinase-like 5 (CDKL5) coding sequence and an AAV9 capsid variant comprising the amino acid sequence of SPHSKA (SEQ ID NO: 941). In some embodiments, the amino acid sequence of SPHSKA (SEQ ID NO: 941) is located in the hypervariable loop IV of the AAV9 capsid variant on page 2 / 285 of the specification, CN 121079310 A. In some embodiments, the amino acid sequence of SPHSKA (SEQ ID NO: 941) is immediately following SEQ ID NO: 4 or SEQ ID NO:The amino acid position corresponding to position 455 of SEQ ID NO: 36 is present after the position.

[0021] In some embodiments, the AAV9 capsid variant comprises one, two, or all of the following: N at the amino acid position corresponding to position 452 of SEQ ID NO: 4, E at the amino acid position corresponding to position 451 of SEQ ID NO: 4, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO: 4. In some embodiments, the AAV9 capsid variant comprises the amino acid sequence KTENVSGSPHSKAQNQQT (SEQ ID NO: 3272).

[0022] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein containing an amino acid sequence having at least 90% identity with SEQ ID NO: 4; (ii) a VP2 protein containing an amino acid sequence having at least 90% identity with positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein containing an amino acid sequence having at least 90% identity with positions 203-742 of SEQ ID NO: 4.

[0023] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein containing an amino acid sequence having at least 95% identity with SEQ ID NO: 4; (ii) a VP2 protein containing an amino acid sequence having at least 95% identity with positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein containing an amino acid sequence having at least 95% identity with positions 203-742 of SEQ ID NO: 4.

[0024] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 99% identity with SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence having at least 99% identity with positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising an amino acid sequence having at least 99% identity with positions 203-742 of SEQ ID NO: 4.

[0025] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 4.

[0026] In some embodiments, the AAV9 capsid variant comprises: (i) the amino acid sequence of SPHSKA (SEQ ID NO:941), wherein the amino acid sequence is present immediately following the amino acid position corresponding to position 455 of SEQ ID NO: 4; (ii) E at the amino acid position corresponding to position 451 of SEQ ID NO: 4 and V at the amino acid position corresponding to position 453 of SEQ ID NO: 4; and (iii) no other modifications relative to wild-type AAV9.

[0027] In some embodiments, the AAV9 capsid variant further comprises one, two, or all of the following: E ​​at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO: 36. In some embodiments, the AAV9 capsid variant comprises the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589).

[0028] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 90% identity with SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence having at least 90% identity with positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence having at least 90% identity with positions 203-742 of SEQ ID NO: 36.

[0029] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence having at least 95% identity with SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence having at least 95% identity with positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence having at least 95% identity with positions 203-742 of SEQ ID NO: 36.

[0030] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein containing an amino acid sequence having at least 99% identity with SEQ ID NO: 36; (ii) a VP2 protein containing an amino acid sequence having at least 99% identity with positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein containing an amino acid sequence having at least 99% identity with positions 203-742 of SEQ ID NO: 36.

[0031] In some embodiments, the AAV9 capsid variant comprises: (i) a VP1 protein containing an amino acid sequence having at least 99% identity with SEQ ID NO: 36; (ii) a VP2 protein containing an amino acid sequence having at least 99% identity with positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein containing an amino acid sequence having at least 99% identity with positions 203-742 of SEQ ID NO: 36.(ii) the amino acid sequence of SEQ ID NO: 36; (iii) the VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) the VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 36.

[0032] In some embodiments, the AAV9 capsid variant comprises: (i) the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 36; (ii) E at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and V at the amino acid position corresponding to position 453 of SEQ ID NO: 36; and (iii) no other modifications relative to wild-type AAV9.

[0033] In some embodiments, this disclosure provides an AAV particle comprising an AAV capsid variant comprising an amino acid sequence having the following formula: [N1]-[N2]-[N3], wherein [N1]-[N2]-[N3] is immediately following the position corresponding to amino acid position 452 of SEQ ID NO: 982; and wherein the AAV capsid variant comprises an amino acid sequence that is at least 90% identical (e.g., at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to the amino acid sequence of SEQ ID NO: 982, for example, to positions 203-742 of SEQ ID NO: 982.

[0034] In some embodiments, [N1] comprises GHD. In some embodiments, [N1] comprises amino acid G at position 453 of SEQ ID NO: 138 or SEQ ID NO: 982, amino acid H at position 454 of SEQ ID NO: 138 or SEQ ID NO: 982, and amino acid D at position 455 of SEQ ID NO: 138 or SEQ ID NO: 982. In some embodiments, [N3] comprises KSG.

[0035] In some embodiments, the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 90% identity with positions 138-742 of SEQ ID NO: 982.

[0036] In some embodiments, the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 95% identity with SEQ ID NO: 982.

[0037] In some embodiments, the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or an amino acid sequence having at least 99% identity with positions 203-742 of SEQ ID NO: 982.

[0038] In some embodiments, the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982.

[0039] In some embodiments, this disclosure provides a specification 4 / 285 pages 14 CN 121079310 A AAV particle comprising a viral genome encoding a wild-type CDKL5 protein. In some embodiments, the viral genome encodes a human CDKL5 protein. In some embodiments, the CDKL5 protein comprises the amino acid sequence of SEQ ID NO: 6413.

[0040] In some embodiments, the CDKL5 coding sequence comprises the amino acid sequence of SEQ ID NO:The CDKL5 coding sequence is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to SEQ ID NO: 6414. In some embodiments, the CDKL5 coding sequence contains at least 95% identical nucleotide sequences to SEQ ID NO: 6414. In some embodiments, the CDKL5 coding sequence contains at least 99% identical nucleotide sequences to SEQ ID NO: 6414. In some embodiments, the CDKL5 coding sequence comprises the nucleotide sequence of SEQ ID NO: 6414. In some embodiments, the CDKL5 coding sequence consists of the nucleotide sequence of SEQ ID NO: 6414.

[0041] In some embodiments, the viral genome contains a promoter effectively linked to the CDKL5 coding sequence. In some implementations, the promoters are human elongation factor 1α-subunit (EF1α) promoter, cytomegalovirus (CMV) immediate early enhancer and / or promoter, chicken β-actin (CBA) promoter, CAG promoter, CAG derivative promoter, β-glucuronidase (GUSB) promoter, ubiquitin C (UBC) promoter, neuron-specific enolase (NSE) promoter, platelet-derived growth factor (PDGF) promoter, platelet-derived growth factor B chain (PDGF-β) promoter, intercellular adhesion molecule 2 (ICAM-2) promoter, synaptic protein (Syn) promoter, synaptic protein 1 (Syn1) promoter, methyl-CpG binding protein 2 (MeCP2) promoter, Ca2+ / calmodulin-dependent protein kinase II (CaMKII) promoter, and metabolite glutamate receptor 2. (mGluR2) promoter, neurofilament light chain (NFL) or heavy chain (NFH) promoter, β-globin small gene nβ2 promoter, proenkephalinogenin (PPE) promoter, enkephalin (Enk) and excitatory amino acid transporter 2 (EAAT2) promoter, glial fibrillary acidic protein (GFAP) promoter, myelin basic protein (MBP) promoter, cardiovascular promoters (e.g., αMHC, cTnT, and CMV-MLC2k), liver promoters (e.g., hAAT, TBG), skeletal muscle promoters (e.g., desmin, MCK, C512) or fragments thereof (e.g., truncated) or functional variants thereof.

[0042] In some embodiments, the viral genome further comprises an inverted terminal repeat (ITR) sequence. In some embodiments, the viral genome comprises an ITR sequence located at the 5' of the CDKL5 coding sequence. In some embodiments, the viral genome comprises an ITR sequence located at the 3' of the CDKL5 coding sequence. In some embodiments, the viral genomeIt includes an ITR sequence located at the 5' position relative to the CDKL5 coding sequence and an ITR sequence located at the 3' position relative to the CDKL5 coding sequence.

[0043] In some embodiments, this disclosure provides an AAV particle comprising a viral genome containing a cyclin-dependent kinase-like 5 (CDKL5) coding sequence and comprising an AAV capsid variant comprising: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of SEQ ID NO: 4 at positions 138-742; and / or (iii) a VP3 protein comprising the amino acid sequence of SEQ ID NO: 4 at positions 203-742.

[0044] In some embodiments, this disclosure provides an AAV particle comprising a viral genome containing a cyclin-dependent kinase-like 5 (CDKL5) coding sequence and an AAV capsid variant comprising: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 36.

[0045] In some embodiments, this disclosure provides a cell comprising the AAV particle described herein. In some embodiments, the cell is a mammalian cell (e.g., HEK293 cell), an insect cell (e.g., Sf9 cell), or a bacterial cell.

[0046] In some embodiments, this disclosure provides a method for preparing the AAV particles described herein, the method comprising: (i) providing a cell containing a viral genome, the viral genome containing a CDKL5 coding sequence and a nucleic acid encoding an AAV capsid variant; and (ii) culturing the cell under conditions suitable for encapsulating the viral genome in the AAV capsid variant; thereby preparing the AAV particles. Specification 5 / 285 pages 15 CN 121079310 A

[0047] In some embodiments, the viral genome of the AAV particle contains the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%); and the AAV capsid variant of the AAV particle contains: (i) a VP1 protein that contains the amino acid sequence of SEQ ID NO: 4 or has at least 90% identity with SEQ ID NO: 4 (e.g., at least 90%).(i) an amino acid sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with SEQ ID NO: 4; or (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 4 or having at least 90% identity with positions 138-742 of SEQ ID NO: 4 (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with SEQ ID NO: 4); or (iii) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 4 or having at least 90% identity with positions 203-742 of SEQ ID NO: 4. The amino acid sequence at positions 203-742 of SEQ ID NO: 4 has at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity).

[0048] In some embodiments, the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 4, the amino acid sequence at positions 138-742 of SEQ ID NO: 4, and / or the amino acid sequence at positions 203-742 of SEQ ID NO: 4.

[0049] In some embodiments, the viral genome of the AAV particle contains the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical); and the AAV capsid variant of the AAV particle contains (i) a VP1 protein containing the amino acid sequence of SEQ ID NO: 36 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 36; and (ii) a VP2 protein containing the amino acid sequence at positions 138-742 of SEQ ID NO: 36 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 36. The amino acid sequence at positions 138-742 of SEQ ID NO: 36 has at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity); or (iii) the VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 36 or the amino acid sequence at positions 203-742 of SEQ ID NO: 36.Positions 203-742 of SEQ ID NO: 36 have an amino acid sequence with at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity).

[0050] In some embodiments, the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 36, the amino acid sequence of positions 138-742 of SEQ ID NO: 36, and / or the amino acid sequence of positions 203-742 of SEQ ID NO: 36.

[0051] In some embodiments, the viral genome of the AAV particle comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to it; and the AAV capsid variant of the AAV particle comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 982; and (ii) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982 or an amino acid sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: The amino acid sequence at positions 138-742 of SEQ ID NO: 982 has at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity); or (iii) the VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982 or having at least 90% identity (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity) with SEQ ID NO: 982.

[0052] In some embodiments, the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 982, the amino acid sequence at positions 138-742 of SEQ ID NO: 982, and / or the amino acid sequence at positions 203-742 of SEQ ID NO: 982. Specification 6 / 285 pages 16 CN 121079310 A

[0053] In some embodiments, the preparation method further includes, prior to step (i), the first...A nucleic acid molecule is introduced into a cell.

[0054] In some embodiments, the cell contains a second nucleic acid molecule encoding an AAV capsid variant. In some embodiments, the preparation method further includes introducing the second nucleic acid molecule into the cell prior to step (i).

[0055] In some embodiments, the cell contains mammalian cells (e.g., HEK293 cells), insect cells (e.g., Sf9 cells), or bacterial cells.

[0056] In some embodiments, this disclosure provides a pharmaceutical composition comprising AAV particles as described herein and pharmaceutically acceptable excipients.

[0057] In some embodiments, this disclosure provides a method of delivering AAV particles encoding the CDKL5 protein to a subject, comprising administering to the subject an effective amount of the pharmaceutical composition or AAV particles described herein.

[0058] In some embodiments, this disclosure provides a method of treating a subject who has or has been diagnosed with a CDKL5-related condition, comprising administering to the subject an effective amount of the pharmaceutical composition or AAV particles described herein.

[0059] In some embodiments, this disclosure provides a pharmaceutical composition or AAV particles of a pharmaceutical composition or AAV particles described herein for treating a subject who has or is diagnosed with a CDKL5-related condition.

[0060] In some embodiments, this disclosure provides the use of a pharmaceutical composition or AAV particles of a pharmaceutical composition or AAV particles described herein in the preparation of a medicament for treating a subject who has or is diagnosed with a CDKL5-related condition.

[0061] In some embodiments, a CDKL5-related condition is a CDKL5-related neurodegeneration or neuromuscular disease.

[0062] In some embodiments, a CDKL5-related neurodegeneration or neuromuscular disease is CDKL5 deficiency (CDD), developmental and epileptic encephalopathy 2, or atypical Rett syndrome.

[0063] In some embodiments, this disclosure provides a method of treating a subject who has or is diagnosed with CDD, comprising administering to the subject an effective amount of a pharmaceutical composition or AAV particles described herein.

[0064] In some embodiments, the subject has one or more mutations in the CDKL5 gene.

[0065] In some embodiments, the subject's CDKL5 activity level is reduced compared to a reference level in a subject without CDKL5-related disease.

[0066] In some embodiments, the treatment halts the progression of the subject's disease. In some embodiments, the treatment improves the disease. In some embodiments, the treatment causes an alteration in one or more biomarkers of the disease. In some embodiments, one or more biomarkers comprise markers of neurofilament light chains or CDKL5 activity, such as those measured by phosphorylation levels of substrate proteins, such as MECP2, or those measured by mass spectrometry.

[0067] In some embodiments, the treatment improves at least one symptom of the condition. In some embodiments, at least one symptom includes epilepsy (e.g., early-onset epilepsy), autism, cognitive deficits, limited motor skills, sleep difficulties, visual impairment, low muscle tone, gastrointestinal reflux, behavioral symptoms (including unexplained laughter or crying, touch sensitivity, and sleep disturbances), facial appearance changes (including microcephaly, high and wide forehead, large and deep-set eyes, a smaller-than-normal distance between the nose and upper lip, upturned nose, full lips, and wide-set teeth), difficulty standing and walking, small and cold feet, lack of or poor eye contact, frequent strabismus, cortical visual impairment or cortical blindness, bruxism, limited speech or aphasia, difficulty eating, stereotypes, limited fine motor skills, gastroesophageal reflux, constipation, or a combination thereof.

[0068] In some embodiments, the subject is a human.

[0069] In some embodiments, AAV particles are delivered to cells, tissues, or regions of the CNS, such as regions of the brain or spinal cord, such as parenchyma, cortex, substantia nigra, caudate nucleus, cerebellum, striatum, corpus callosum, cerebellum, brainstem, caudate nucleus-putamen, thalamus, epithalamus, spinal cord, or combinations thereof, and / or delivered to neurons or combinations thereof.

[0070] In some embodiments, the delivery or treatment method further includes assessment, such as measuring the level of CDKL5 expression in a subject, such as the subject's cells, tissues, or body fluids, including, for example, the level of CDKL5 gene expression, CDKL5 mRNA expression, and / or CDKL5 protein expression. In some embodiments, the CDKL5 protein expression level is measured by ELISA, Western blot, or immunohistochemistry.

[0071] In some embodiments, CDKL5 expression levels are assessed before and after administration of AAV particles, optionally wherein CDKL5 expression levels before treatment are compared with CDKL5 expression levels after administration.

[0072] In some embodiments, CDKL5 expression levels are assessed in cells or tissues of the central nervous system (e.g., parenchyma) of the subject.

[0073] In some embodiments, CDKL5 protein expression levels in the subject after administration are increased relative to CDKL5 protein expression levels in the subject before administration.

[0074] In some embodiments, administration of the pharmaceutical composition or AAV particles causes an increase in: (i) CDKL5 activity in the subject's cells, tissues (e.g., cells or tissues of the CNS, such as the cortex, striatum, thalamus, cerebellum, and / or brainstem) and / or body fluids (e.g., CSF and / or serum) relative to CDKL5 activity in the subject before administration; (ii) CDKL5 activity relative to CDKL5 activity in the subject before administration.The content of viral genome (VG) per cell in the subject's peripheral tissues, the content of VG per cell in the subject's CNS tissues (e.g., cortex, striatum, thalamus, cerebellum, brainstem, and / or spinal cord); and / or (iii) CDKL5 mRNA expression in the subject compared to prior administration, CDKL5 mRNA expression in the subject's cells or tissues (e.g., cells or tissues of the CNS, such as the cortex, thalamus, and / or brainstem).

[0075] In some embodiments, the treatment method further includes administering to the subject an additional agent suitable for treating or preventing CDKL5-related conditions. In some embodiments, the additional agent comprises one or more antiepileptic drugs (e.g., levetiracetam, phenobarbital, clonazepam, topiramate), adrenocorticotropic hormone, or combinations thereof. In some embodiments, the method further includes administering an immunosuppressant to the subject. In some implementations, the immunosuppressant comprises a corticosteroid (e.g., prednisone, prednisolone, methylprednisolone and / or dexamethasone), rapamycin, mycophenolate mofetil, tacrolimus, rituximab and / or eculizumab hydroxychloroquine).

[0076] The listed embodiments

[0077] 1. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence having the following formula: [N1]-[N2]-[N3], wherein: (i) optionally [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G; (ii) [N2] comprises an amino acid sequence of SPH; and (iii) [N3] comprises X4, X5, and X6, wherein at least one of X4, X5, or X6 is a basic amino acid, such as K or R.

[0078] 2. The AAV particle of embodiment 1, wherein X4, X5, or both of [N3] are K.

[0079] 3. The AAV particle of embodiment 1 or 2, wherein X4, X5, or X6 of [N3] is R.

[0080] 4. The AAV particle of any one of embodiments 1 to 3, wherein: (a) X4 of [N3] is: K, S, A, V, T, G, F, W, V, N, or R; (b)X5 of [N3] is: S, K, T, F, I, L, Y, H, M or R; and / or the specification 8 / 285 page 18 CN 121079310 A (c) X6 of [N3] is: G, A, R, M, I, N, T, Y, D, P, V, L, E, W, N, Q, K or S; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(c), for example, a conservative substitution.

[0081] 5. AAV particles as described in any one of embodiments 1 to 4, wherein [N3] comprises SK, KA, KS, AR, RM, VK, AS, SR, VK, KR, KK, KN, VR, RS, RK, KT, TS, KF, FG, KI, IG, KL, LG, TT, TY, KY, YG, KD, KP, TR, RG, VR, GA, SL, SS, FL, WK, SA, RA, LR, KW, RR, GK, TK, NK, AK, KV, KG, KH, KM, TG, SE, SV, SW, SN, HG, SQ, LW, MG, MA or SG.

[0082] 6. An AAV particle as described in any one of embodiments 1 to 5, wherein [N3] is or includes SKA, KSG, ARM, VKS, ASR, VKI, KKN, VRM, RKA, KTS, KFG, KIG, KLG, KTT, KTY, KYG, SKD, SKP, TRG, VRG, KRG, GAR, KSA, KSR, SKL, SRA, SKR, SLR, SRG, SSR, FLR, SKW, SKS, WKA, VRR, SKV, SKT, SKG, GKA, TKA, NKA, SKL, SKN, AKA, KTG, KSL, KSE, KSV, KSW, KSN, KHG, KSQ, KSK, KLW, WKG, KMG, KMA, or RSG.

[0083] 7. AAV particles as described in any one of embodiments 1 to 6, wherein [N2]-[N3] comprises SPHSK (SEQ ID NO: 4701), SPHKS (SEQ ID NO: 4704), SPHAR (SEQ ID NO: 4705), SPHVK (SEQ ID NO: 4706), SPHAS (SEQ ID NO: 4707), SPHKK (SEQ ID NO: 4708), SPHVR (SEQ ID NO: 4709), SPHRK (SEQ ID NO: 4710), SPHKT (SEQ ID NO: 4711), SPHKF (SEQ ID NO: 4712), SPHKI (SEQ ID NO: 4713), SPHKL (SEQ ID NO: 4714), SPHKY(SEQ ID NO: 4715), SPHTR (SEQ ID NO: 4716), SPHKR (SEQ ID NO: 4717), SPHGA (SEQ ID NO: 4718), SPHSR (SEQ ID NO: 4719), SPHSL (SEQ ID NO: 4720), SPHSS (SEQ ID NO: 4721), SPHFL (SEQ ID NO: 4722), SPHWK (SEQ ID NO: 4723), SPHGK (SEQ ID NO: 4724), SPHTK (SEQ ID NO: 4725), SPHNK (SEQ ID NO: 4726), SPHAK (SEQ ID NO: 4727), SPHKH (SEQ ID NO: 4728), SPHKM (SEQ ID NO: 4729), or SPHRS (SEQ ID NO: 4730).

[0084] 8. An AAV particle according to any one of embodiments 1 to 7, wherein [N2]-[N3] is or comprises: (i) SPHSKA (SEQ ID NO: 941), SPHKSG (SEQ ID NO: 946), SPHARM (SEQ ID NO: 947), SPHVKS (SEQ ID NO: 948), SPHASR (SEQ ID NO: 949), SPHVKI (SEQ ID NO: 950), SPHKKN (SEQ ID NO: 954), SPHVRM (SEQ ID NO: 955), SPHRKA (SEQ ID NO: 956), SPHKFG (SEQ ID NO: 957), SPHKIG (SEQ ID NO: 958), SPHKLG (SEQ ID NO: 959), SPHKTS (SEQ ID NO: 963), SPHKTT (SEQ ID NO: 964), SPHKT Y (SEQ ID NO: 965), SPHKYG (SEQ ID NO: 966), SPHSKD (SEQ ID NO: 967), SPHSKP (SEQ ID NO: 968), SPHTRG (SEQ ID NO: 972), SPHVRG (SEQ ID NO: 973), SPHKRG (SEQ ID NO: 974), SPHGAR (SEQ ID NO: 975), SPHKSA (SEQ ID NO: 977), SPHKSR (SEQ IDNO: 951), SPHSKL (SEQ ID NO: 960), SPHSRA (SEQ ID NO: 969), SPHSKR (SEQ ID NO: 978), SPHSLR (SEQ ID NO: 952), SPHSRG (SEQ ID NO: 961), SPHSSR (SEQ ID NO: 970), SPHFLR (SEQ ID NO: 979), SPHSKW (SEQ ID NO: 953), SPHSKS (SEQ ID NO: 962), SPHWKA (SEQ ID NO: 971), SPHVRR (SEQ ID NO: 980), SPHSKT (SEQ ID NO: 4731), SPHSKG (SEQ ID NO: 4732), SPHGKA (SEQ ID NO: 4733), SPHNKA (SEQ ID NO: 4734), SPHSKN (SEQ ID NO: 4735), SPHAKA (SEQ ID NO: 4736), SPHSKV (SEQ ID NO: 4737), SPHKTG (SEQ ID NO: 4738), SPHTKA (SEQ ID NO: 4739), SPHKSL (SEQ ID NO: 4740), SPHKSE (SEQ ID NO: 4741), SPHKSV (SEQ ID NO: 4742), SPHKSW (SEQ ID NO: 4743), instruction sheet 9 / 285 page 19 CN 121079310 A SPHKSN (SEQ ID NO: 4744), SPHKHG (SEQ ID NO: 4745), SPHKSQ (SEQ ID NO: 4746), SPHKSK (SEQ ID NO: 4747), SPHKLW (SEQ ID NO: 4748), SPHWKG (SEQ ID NO: 4749), SPHKMG (SEQ ID NO: 4750), SPHKMA (SEQ ID NO: (ii) any portion of the amino acid sequence in (i), such as any 2, 3, 4, or 5 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence containing one, two, or three, but no more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv)Relative to any of the amino acid sequences in (i), an amino acid sequence comprising one, two, or three, but not more than four, different amino acids.

[0085] 9. An AAV particle as described in any of embodiments 1 to 8, wherein the AAV capsid variant comprises an amino acid other than G (e.g., V, R, D, E, M, T, I, S, A, N, L, K, H, P, W, or C) at position 453, an amino acid other than S (e.g., V, L, N, D, H, R, P, G, T, I, A, E, Y, M, or Q) at position 454, and / or an amino acid other than G (e.g., C, L, D, E, Y, H, V, A, N, P, or S) at position 455.

[0086] 10. An AAV particle of any one of embodiments 1 to 8, wherein the AAV capsid variant comprises amino acid G at position 453, amino acid S at position 454, and amino acid G at position 455, as specified in SEQ ID NO: 138 or 981.

[0087] 11. An AAV particle of any one of embodiments 1 to 9, wherein the AAV capsid variant comprises amino acid G at position 453, amino acid H at position 454, and amino acid D at position 455, as specified in SEQ ID NO: 138 or 982.

[0088] 12. An AAV particle of any one of embodiments 1 to 11, wherein [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G.

[0089] 13. An AAV particle as described in any one of embodiments 1 to 12, wherein: (a) X1 of [N1] is: G, V, R, D, E, M, T, I, S, A, N, L, K, H, P, W or C; (b) X2 of [N1] is: S, V, L, N, D, H, R, P, G, T, I, A, E, Y, M or Q; and / or (c) X3 of [N1] is: G, C, L, D, E, Y, H, V, A, N, P or S; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(c), for example, a conservative substitution.

[0090] 14. An AAV particle as described in any one of embodiments 1 to 13, wherein [N1] comprises GS, SG, GH, HD, GQ, QD, VS, CS, GR, RG, QS, SH, MS, RN, TS, IS, GP, ES, SS, GN, AS, NS, LS, GG, KS, GT, PS, RS, GI, WS, DS, ID, GL, DA, DG, ME, EN, KN, KE, AI, NG, PG, TG, SV, IG, LG, AG, EG, SA, YD, HE, HG, RD, ND, PD, MG,QV, DD, HN, HP, GY, GM, GD or HS.

[0091] 15. AAV particles as described in any one of embodiments 1 to 14, wherein [N1] is or includes GSG, GHD, GQD, VSG, CSG, GRG, CSH, GQS, GSH, RVG, GSC, GLL, GDD, GHE, GNY, MSG, RNG, TSG, ISG, GPG, ESG, SSG, GNG, ASG, NSG, LSG, GGG, KSG, HSG, GTG, PSG, GSV, RSG, GIG, WSG, DSG, IDG, GLG, DAG, DGG, MEG, ENG, GSA, KNG, KEG, AIG, GYD, GHG, GRD, GND, GPD, GMG, GQV, GHN, GHP or GHS.

[0092] 16. AAV particles as described in any one of embodiments 1 to 15, wherein [N1]-[N2] comprises: (i) SGSPH (SEQ ID NO: 4752), HDSPH (SEQ ID NO: 4703), QDSPH (SEQ ID NO: 4753), RGSPH (SEQ ID NO: 4754), SHSPH (SEQ ID NO: 4755), QSSPH (SEQ ID NO: 4756), DDSPH (SEQ ID NO: 4757), HESPH (SEQ ID NO: 4758), NYSPH (SEQ ID NO: 4759), VGSPH (SEQ ID NO: 4760), SCSPH (SEQ ID NO: 4761), LLSPH (SEQ ID NO: 4762), NGSPH (SEQ ID NO: 4763), PGSPH (SEQ ID NO: 4764), GGSPH (SEQ ID NO: 4765), ... Instructions 10 / 285 Page 20 CN 121079310 A 4765), TGSPH (SEQ ID NO: 4766), SVSPH (SEQ ID NO: 4767), IGSPH (SEQ ID NO: 4768), DGSPH (SEQ ID NO: 4769), LGSPH (SEQ ID NO: 4770), AGSPH (SEQ ID NO: 4771), EGSPH (SEQ ID NO: 4772), SASPH (SEQ ID NO: 4773), YDSPH (SEQ ID NO: 4774), HGSPH (SEQ ID NO: 4775), RDSPH (SEQ ID NO:(ii) any part of the amino acid sequence in (i), such as any 2, 3 or 4 amino acids, such as an amino acid sequence of consecutive amino acids; (iii) a modified amino acid sequence containing one, two or three but no more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) a modified amino acid sequence containing one, two or three but no more than four different amino acids relative to any of the amino acid sequences in (i).

[0093] 17. AAV particles as described in any one of embodiments 1 to 16, wherein [N1]-[N2] are or comprise: (i) GSGSPH (SEQ ID NO: 4695), GHDSPH (SEQ ID NO: 4784), GQDSPH (SEQ ID NO: 4785), VSGSPH (SEQ ID NO: 4786), CSGSPH (SEQ ID NO: 4787), GRGSPH (SEQ ID NO: 4788), CSHSPH (SEQ ID NO: 4789), GQSSPH (SEQ ID NO: 4790), GSHSPH (SEQ ID NO: 4791), GDDSPH (SEQ ID NO: 4792), GHESPH (SEQ ID NO: 4793), GNYSPH (SEQ ID NO: 4794), RVGSPH (SEQ ID NO: 4795), GSCSPH (SEQ ID NO: 4796), GLLSPH (SEQ ID NO: 4797), MSGSPH (SEQ ID NO: 4798), RNGSPH (SEQ ID NO: 4799), TSGSPH (SEQ ID NO: 4800), ISGSPH (SEQ ID NO: 4801), GPGSPH (SEQ ID NO: 4802), ESGSPH (SEQ ID NO: 4803), SSGSPH (SEQ ID NO: 4801) NO: 4804), GNGSPH (SEQ ID NO: 4805), ASGSPH (SEQ ID NO: 4806), NSGSPH (SEQID NO: 4807), LSGSPH (SEQ ID NO: 4808), GGGSPH (SEQ ID NO: 4809), KSGSPH (SEQ ID NO: 4810), HSGSPH (SEQ ID NO: 4811), GTGSPH (SEQ ID NO: 4812), PSGSPH (SEQ ID NO: 4813), GSVSPH (SEQ ID NO: 4814), RSGSPH (SEQ ID NO: 4815), GIGSPH (SEQ ID NO: 4816), WSGSPH (SEQ ID NO: 4817), DSGSPH (SEQ ID NO: 4818), IDGSPH (SEQ ID NO: 4819), GLGSPH (SEQ ID NO: 4820), DAGSPH (SEQ ID NO: 4821), DGGSPH (SEQ ID NO: 4822), MEGSPH (SEQ ID NO: 4823), ENGSPH (SEQ ID NO: (ii) any portion of the amino acid sequence in (i), such as any 2, 3, 4, or 5 amino acids therein, such as a sequence of consecutive amino acids; (iii) GSASPH (SEQ ID NO: 4824), KNGSPH (SEQ ID NO: 4826), KEGSPH (SEQ ID NO: 4827), AIGSPH (SEQ ID NO: 4828), GYDSPH (SEQ ID NO: 4829), GHGSPH (SEQ ID NO: 4830), GRDSPH (SEQ ID NO: 4831), GNDSPH (SEQ ID NO: 4832), GPDSPH (SEQ ID NO: 4833), GMGSPH (SEQ ID NO: 4834), GQVSPH (SEQ ID NO: 4835), GHNSPH (SEQ ID NO: 4836), GHPSPH (SEQ ID NO: 4837), or GHSSPH (SEQ ID NO: 4838); Relative to any of the amino acid sequences in (i), there is an amino acid sequence containing one, two, or three, but no more than four modified amino acids; or (iv) relative to any of the amino acid sequences in (i), there is an amino acid sequence containing one, two, or three, but no more than four different amino acids. Specification 11 / 285 pages 21 CN 121079310 A

[0094] 18.An AAV particle as in any one of embodiments 1 to 17, wherein [N1]-[N2]-[N3] comprises: (i) SGSPHSK (SEQ ID NO: 4839), HDSPHKS (SEQ ID NO: 4840), SGSPHAR (SEQ ID NO: 4841), SGSPHVK (SEQ ID NO: 4842), QDSPHKS (SEQ ID NO: 4843), SGSPHKK (SEQ ID NO: 4844), SGSPHVR (SEQ ID NO: 4845), SGSPHAS (SEQ ID NO: 4846), SGSPHRK (SEQ ID NO: 4847), SGSPHKT (SEQ ID NO: 4848), SHSPHKS (SEQ ID NO: 4849), QSSPHRS (SEQ ID NO: 4850), RGSPHAS (SEQ ID NO: 4851), RGSPHSK (SEQ ID NO: 4852), SGSPHKF (SEQ ID NO: 4853), SGSPHKI (SEQ ID NO: 4854), SGSPHKL (SEQ ID NO: 4855), SGSPHKY (SEQ ID NO: 4856), SGSPHTR (SEQ ID NO: 4857), SHSPHKR (SEQ ID NO: 4858), SGSPHGA (SEQ ID NO: 4859), HDSPHKR (SEQ ID NO: 4860), DDSPHKS (SEQ ID NO: 4861), HESPHKS (SEQ ID NO: 4862), NYSPHKI (SEQ ID NO: 4863), SGSPHSR (SEQ ID NO: 4864), SGSPHSL (SEQ ID NO: 4865), SGSPHSS (SEQ ID NO: 486,6), VGSPHSK (SEQ ID NO: 4867), SCSPHRK (SEQ ID NO: 4868), SGSPHFL (SEQ ID NO: 4869), LLSPHWK (SEQ ID NO: 4870), NGSPHSK (SEQ ID NO: 4871), PGSPHSK (SEQ ID NO: 4872), GGSPHSK (SEQ ID NO: 4873), TGSPHSK (SEQ ID NO: 4874), SVSPHGK (SEQ ID NO:4875), SSGSPHTK (SEQ ID NO: 4876), IGSPHSK (SEQ ID NO: 4877), DGSPHSK (SEQ ID NO: 4878), SGSPHNK (SEQ ID NO: 4879), LGSPHSK (SEQ ID NO: 4880), AGSPHSK (SEQ ID NO: 4881), EGSPHSK (SEQ ID NO: 4882), SASPHSK (SEQ ID NO: 4883), SGSPHAK (SEQ ID NO: 4884), HDSPHKI (SEQ ID NO: 4885), YDSPHKS (SEQ ID NO: 4886), HDSPHKT (SEQ ID NO: 4887), RGSPHKR (SEQ ID NO: 4888), HGSPHSK (SEQ ID NO: 4889), RDSPHKS (SEQ ID NO: 4890), NDSPHKS (SEQ ID NO: 4891), QDSPHKI (SEQ ID NO: 4892), PDSPHKI (SEQ ID NO: 4893), PDSPHKS (SEQ ID NO: 4894), MGSPHSK (SEQ ID NO: 4895), HDSPHKH (SEQ ID NO: 4896), QVSPHKS (SEQ ID NO: 4897), HNSPHKS (SEQ ID NO: 4898), NGSPHKR (SEQ ID NO: 4899), HDSPHKY (SEQ ID NO: 4900), NDSPHKI (SEQ ID NO: 4901), HDSPHKL (SEQ ID NO: 4902), HPSPHWK (SEQ ID NO: 4903), HDSPHKM (SEQ ID NO: 4904) or HSSPHRS (SEQ ID NO: 4905); (ii) (i) any portion of the amino acid sequence, such as any 2, 3, 4, 5, or 6 amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) an amino acid sequence containing one, two, or three, but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) an amino acid sequence containing one, two, or three, but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0095] 19.Compound 1 contains 18 individual AAV fragments, including [N1]‐[N2]‐[N3]: (i) GSGSPHSKA (SEQ ID NO: 4697) and GHDSPHKSG (SEQ ID NO: 4698), GSGSPHARM (SEQ ID NO: 4906), GSGSPHVKS (SEQ ID NO: 4907), GQDSPHKSG (SEQ ID NO: 4908), GSGSPHASR (SEQ ID NO: 4909), GSGSPHVKI (SEQ ID NO: 4909) 4910) GSPHKKN (SEQ ID NO: 4911), GSGSPHVRM (SEQ ID NO: 4912), VSGSPHSKA (SEQ ID NO: 4913), CSGSPHSKA (SEQ ID NO: 4914), GSGSPHRKA (SEQ ID NO: 4915), CSGSPHKTS (SEQ CSHSPHKSG (SEQ ID NO: 4917), GQSSPHRSG (SEQ ID NO: 4918), GRGSPHASR (SEQ ID NO: 4919), GRGSPHSKA (SEQ ID NO: 4920), GSGSPHKFG (SEQ ID NO: 4920) 4921) GSGSPHKIG (SEQ ID NO: 4922) GSGSPHKLG (SEQ ID NO: 4923) GSGSPHKTS (SEQ ID NO: 4924) SPECIFICATION 12 / 285 SHEET 22 CN 121079310 A GSGSPHKTT (SEQ ID NO: 4925), GSGSPHKTY (SEQ ID NO: 4926), GSGSPHKYG (SEQ ID NO: 4927), GSGSPHSKD (SEQ ID NO: 4928), GSGSPHSKP (SEQ ID NO: 4929), GSGSPHTRG (SEQ ID NO: 4930), GSGSPHVRG (SEQ ID NO: 4931), GSHSPHKRG (SEQ ID NO: 4932), GSHSPHKSG (SEQ ID NO: 4933), VSGSPHASR (SEQ ID NO: 4934), VSGSPHGAR (SEQ ID NO: 4935), VSGSPHKFG (SEQ ID NO: 4935).4936)、GHDSPHKRG (SEQ ID NO: 4937)、GDDSPHKSG (SEQ ID NO: 4938)、GHESPHKSA (SEQ ID NO: 4939)、GHDSPHKSA (SEQ ID NO: 4940)、GNYSPHKIG (SEQ ID NO: 4941)、GHDSPHKSR (SEQ ID NO: 4942)、GSGSPHSKL (SEQ ID NO: 4943)、GSGSPHSRA (SEQ ID NO: 4944)、GSGSPHSKR (SEQ ID NO: 4945)、GSGSPHSLR (SEQ ID NO: 4946)、GSGSPHSRG (SEQ ID NO: 4947)、GSGSPHSSR (SEQ ID NO: (SEQ ID NO: 4954)、GSGSPHVRR (SEQ ID NO: 4955). (SEQ ID NO: 4960). ID NO: 4966)、NSGSPHSKA (SEQ ID NO: 4967)、LSGSPHSKA (SEQ ID NO: 4968)、GGGSPHSKA (SEQ ID NO: 4969)、KSGSPHSKA (SEQ ID NO: 4970)、GGGSPHSKS (SEQ ID NO: 4971)、GSGSPHSKG(SEQ ID NO: 4972)、 HSGSPHSKA (SEQ ID NO: 4973)、GTGSPHSKA (SEQ ID NO: 4974)、PSGSPHSKA (SEQ ID NO: 4975)、GSVSPHGKA (SEQ ID NO: 4976)、RSGSPHSKA (SEQ ID NO: 4977)、GSGSPHTKA (SEQ ID NO: 4978)、GIGSPHSKA (SEQ ID NO: 4979)、WSGSPHSKA (SEQ ID NO: 4980)、 DSGSPHSKA (SEQ ID NO: 4981)、IDGSPHSKA (SEQ ID NO: 4982)、GSGSPHNKA (SEQ ID NO: 4983)、GLGSPHSKS (SEQ ID NO: 4984)、DAGSPHSKA (SEQ ID NO: 4985)、DGGSPHSKA (SEQ ID NO: 4986)、MEGSPHSKA (SEQ ID NO: 4987)、ENGSPHSKA (SEQ ID NO: 4988)、 GSASPHSKA (SEQ ID NO: 4989)、GNGSPHSKS (SEQ ID NO: 4990)、KNGSPHSKA (SEQ ID NO: 4991)、KEGSPHSKA (SEQ ID NO: 4992)、AIGSPHSKA (SEQ ID NO: 4993)、GSGSPHSKN (SEQ ID NO: 4994)、GSGSPHAKA (SEQ ID NO: 4995)、GHDSPHKIG (SEQ ID NO: 4996)、 GYDSPHKSG (SEQ ID NO: 4997)、GHESPHKSG (SEQ ID NO: 4998)、GHDSPHKTG (SEQ ID NO: 4999)、GRGSPHKRG (SEQ ID NO: 5000)、GQDSPHKSG (SEQ ID NO: 4908)、GHDSPHKSL (SEQ ID NO: 5001)、GHGSPHSKA (SEQ ID NO: 5002)、GHDSPHKSE (SEQ ID NO: 5003)、 VSGSPHSKA (SEQ ID NO: 4913)、GRDSPHKSG (SEQ ID NO: 5004)、GNDSPHKSV (SEQ ID NO:5005), GQDSPHKIG (SEQ ID NO: 5006), GHDSPHKSV (SEQ ID NO: 5007), GPDSPHKIG (SEQ ID NO: 5008), GPDSPHKSG (SEQ ID NO: 5009), GHDSPHKSW (SEQ ID NO: 5010), GHDSPHKSN (SEQ ID NO: 5011), GMGSPHSKT (SEQ ID NO: 5012), GHDSPHKHG (SEQ ID NO: 5013), GQVSPHKSG (SEQ ID NO: 5014), GDDSPHKSV (SEQ ID NO: 5015), GHNSPHKSG (SEQ ID NO: 5016), GGNSPHKRG (SEQ ID NO: 5017), GHDSPHKYG (SEQ ID NO: 5018), GHDSPHKSQ (SEQ ID NO: 5019), GNDSPHKIG (SEQ ID NO: 5020), GHDSPHKSK (SEQ ID NO: 5021), GHDSPHKLW (SEQ ID NO: 5022), GHPSPHWKG (SEQ ID NO: 5023), GHDSPHKMG (SEQ ID NO: 5024), GHDSPHKMA (SEQ ID NO: 5025) or GHSSPHRSG (SEQ ID NO: 5026); Specification 13 / 285 pages 23 CN 121079310 A (ii) comprising any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7 or 8 amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) comprising one, two or three but not more than four modified amino acid sequences relative to any of the amino acid sequences in (i); or (iv) comprising one, two or three but not more than four different amino acid sequences relative to any of the amino acid sequences in (i).

[0096] 20. An AAV particle as described in any one of embodiments 1 to 19, wherein [N3] comprises SK, KA, KS, or SG.

[0097] 21. An AAV particle as described in any one of embodiments 1 to 20, wherein [N3] is or comprises SKA, KSG, or KYG.

[0098] 22. An AAV particle as described in any one of embodiments 1 to 21, wherein [N2]-[N3] comprises SPHSK (SEQ ID NO: 4701), SPHKS (SEQ ID NO: 4701), or SPHKS (SEQ ID NO: 4701).4704) or SPHKY (SEQ ID NO: 4715).

[0099] 23. An AAV particle as described in any one of embodiments 1 to 22, wherein [N2]-[N3] is or contains SPHSKA (SEQ ID NO: 941).

[0100] 24. An AAV particle as described in any one of embodiments 1 to 22, wherein [N2]-[N3] is or contains SPHKSG (SEQ ID NO: 946).

[0101] 25. An AAV particle as described in any one of embodiments 1 to 22, wherein [N2]-[N3] is or contains SPHKYG (SEQ ID NO: 966).

[0102] 26. An AAV particle as described in any one of embodiments 1 to 25, wherein [N1] contains GS, SG, GH, or HD.

[0103] 27. An AAV particle as described in any one of embodiments 1 to 26, wherein [N1] is or contains GSG.

[0104] 28. An AAV particle as described in any one of embodiments 1 to 26, wherein [N1] is or contains GHD.

[0105] 29. An AAV particle as described in any one of embodiments 1 to 23 or 26 to 27, wherein [N1]-[N2]-[N3] contains SGSPHSK (SEQ ID NO: 4839).

[0106] 30. An AAV particle as described in any one of embodiments 1 to 22, 24, 26 or 28, wherein [N1]-[N2]-[N3] contains HDSPHKS (SEQ ID NO: 4840).

[0107] 31. An AAV particle as described in any one of embodiments 1 to 22 or 25 to 27, wherein [N1]-[N2]-[N3] contains SGSPHKYG (SEQ ID NO: 5027).

[0108] 32. An AAV particle as described in any one of embodiments 1 to 8, 10, 12 to 23, 26 to 27 or 29, wherein [N1]-[N2]-[N3] is or contains GGSPHSKA (SEQ ID NO: 4697).

[0109] 33. An AAV particle as described in any one of embodiments 1 to 9, 11 to 22, 24, 26, 28 or 30, wherein [N1]-[N2]-[N3] is or contains GHDSPHKSG (SEQ ID NO: 4698).

[0110] 34. An AAV particle as described in any one of embodiments 1 to 8, 10, 12 to 22, 25 to 27 or 31, wherein [N1]-[N2]-[N3] is or contains GGSPHKYG (SEQ ID NO: 4927).

[0111] 35. AAV particles as described in any one of embodiments 1 to 34, wherein [N1]-[N2]-[N3] replace according to SEQ ID NO:The position of number 138 is 453-455.

[0112] 36. An AAV particle as described in any one of embodiments 1 to 35, wherein the AAV capsid variant comprises an amino acid other than Q at position 456 (e.g., W, K, R, G, L, V, S, P, H, K, I, M, A, E, or F), an amino acid other than N at position 457 (e.g., Y, C, K, T, H, R, D, V, S, P, G, W, E, F, A, I, M, Q, or L), an amino acid other than Q at position 458 (e.g., G, K, H, R, T, L, D, A, P, I, F, V, M, W, Y, S, E, N, or Y), and / or an amino acid other than Q at position 459 (e.g., H, L, R, W, K, A, P, E, M, I, S, G, N, Y, C, V, T, D, or V).

[0113] 37. An AAV particle as described in any one of embodiments 1 to 36, wherein the AAV capsid variant comprises an amino acid other than Q (e.g., W, K, R, G, L, V, S, P, H, K, I, M, A, E, or F) at position 462, as specified in SEQ ID NO: 981, 982, 36, 37, 39, 40, 42-46, 48, 49, 50, 52, 53, 56, or 57, according to page 14 / 285 of the specification, CN 121079310 A ID NO: 981, 982, 36, 37, 39, 40, 42-46, 48, 49, 50, 52, 53, 56, or 57; an amino acid other than N (e.g., Y, C, K, T, H, R, D, V, S, P, G, W, E, F, A, I, M, Q, or L) at position 463; and an amino acid other than Q (e.g., G, ...) at position 464. K, H, R, T, L, D, A, P, I, F, V, M, W, Y, S, E, N or Y) and / or amino acids other than Q at position 465 (e.g., H, L, R, W, K, A, P, E, M, I, S, G, N, Y, C, V, T, D or V).

[0114] 38. An AAV particle of any one of embodiments 1 to 37, wherein the AAV capsid variant comprises: (a) amino acid Q at position 456, amino acid N at position 457, amino acid Q at position 458, and / or amino acid Q at position 459 according to SEQ ID NO: 138; or (b) amino acid Q at position 462, amino acid N at position 463, amino acid Q at position 464, and / or amino acid Q at position 465 according to SEQ ID NO: 981, 982, 36, 37, 39, 40, 42-46, 48, 49, 50, 52, 53, 56, or 57.

[0115] 39. An AAV particle of any one of embodiments 1 to 38, wherein the AAV capsid variant further comprises [N4], wherein [N4] comprises X7 X8 X9.X10, wherein: (a) X7 is: Q, W, K, R, G, L, V, S, P, H, K, I, M, A, E, or F; (b) X8 is: N, Y, C, K, T, H, R, D, V, S, P, G, W, E, F, A, I, M, Q, or L; (c) X9 is: Q, G, K, H, R, T, L, D, A, P, I, F, V, M, W, Y, S, E, N, or Y; and (d) X10 is: Q, H, L, R, W, K, A, P, E, M, I, S, G, N, Y, C, V, T, D, or V; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(d), for example, a conservative substitution.

[0116] 40. The AAV particle of embodiment 39, wherein: (a) X7 of [N4] is Q or R; (b) X8 of [N4] is N or R; (c) X9 of [N4] is Q or R; and (d) X10 of [N4] is Q, L or R.

[0117] 41. AAV particles as described in embodiment 39 or 40, wherein [N4] is or comprises: (i) QNQQ (SEQ ID NO: 5028), WNQQ (SEQ ID NO: 5029), QYYV (SEQ ID NO: 5030), RRQQ (SEQ ID NO: 5031), GCGQ (SEQ ID NO: 5032), LRQQ (SEQ ID NO: 5033), RNQQ (SEQ ID NO: 5034), VNQQ (SEQ ID NO: 5035), FRLQ (SEQ ID NO: 5036), FNQQ (SEQ ID NO: 5037), LLQQ (SEQ ID NO: 5038), SNQQ (SEQ ID NO: 5039), RLQQ (SEQ ID NO: 5040), LNQQ (SEQ ID NO: 5041), QRKL (SEQ ID NO: 5042), LRRQ (SEQ ID NO: 5043), VNQQ (SEQ ID NO: 5034), FRLQ ... LRRQ (SEQ ID NO: 5036), FNQQ (SEQ ID NO: 5037), LLLQQ (SEQ ID NO: 5038), SNQQ (SEQ ID NO: 5039), RLQQ (SEQ ID NO: 5040), LNQQ (SEQ ID NO: ID NO: 5043), QRLR (SEQ ID NO: 5044), QRRL (SEQ ID NO: 5045), RRLQ (SEQ ID NO: 5046), RLRQ (SEQ ID NO: 5047), SKRQ (SEQ ID NO: 5048), QLYR (SEQ ID NO: 5049), QLTV (SEQ ID NO: 5050), QNKQ (SEQ ID NO: 5051), KNQQ (SEQ ID NO: 5052), QKQQ (SEQ ID NO: 5053), QTQQ(SEQ ID NO: 5054), QNHQ (SEQ ID NO: 5055), QHQQ (SEQ ID NO: 5056), QNQH (SEQ ID NO: 5057), QHRQ (SEQ ID NO: 5058), LTQQ (SEQ ID NO: 5059), QNQW (SEQ ID NO: 5060), QNTH (SEQ ID NO: 5061), RRRQ (SEQ ID NO: 5062), QYQQ (SEQ ID NO: 5063), QNDQ (SEQ ID NO: 5064), QNRH (SEQ ID NO: 5065), RDQQ (SEQ ID NO: 5066), PNLQ (SEQ ID NO: 5067), HVRQ (SEQ ID NO: 5068), PNQH (SEQ ID NO: 5069), HNQQ (SEQ ID NO: 5070), QSQQ (SEQ ID NO: 5071), QPAK (SEQ ID NO: 5072), Description 15 / 285 pages 25 CN 121079310 A QNLA (SEQ ID NO: 5073), QNQL (SEQ ID NO: 5074), QGQQ (SEQ ID NO: 5075), LNRQ (SEQ ID NO: 5076), QNPP (SEQ ID NO: 5077), QNLQ (SEQ ID NO: 5078), QDQE (SEQ ID NO: 5079), QDQQ (SEQ ID NO: 5080), HWQQ (SEQ ID NO: 5081), PNQQ (SEQ ID NO: 5082), PEQQ (SEQ ID NO: 5083), QRTM (SEQ ID NO: 5084), LHQH (SEQ ID NO: 5085), QHRI (SEQ ID NO: 5086), QYIH (SEQ ID NO: 5087), QKFE (SEQ ID NO: 5088), QFPS (SEQ ID NO: 5089), QNPL (SEQ ID NO: 5090), QAIK (SEQ ID NO: 5091), QNRQ (SEQ ID NO: 5092), QYQH (SEQ ID NO: 5093), QNPQ (SEQ ID NO: 5094), QHQL (SEQ ID NO:5095)、QSPP (SEQ ID NO: 5096)、QAKL (SEQ ID NO: 5097)、KSQQ (SEQ ID NO: 5098)、 QDRP (SEQ ID NO: 5099)、QNLG (SEQ ID NO: 5100)、QAFH (SEQ ID NO: 5101)、QNAQ (SEQ ID NO: 5102)、HNQL (SEQ ID NO: 5103)、QKLN (SEQ ID NO: 5104)、QNVQ (SEQ ID NO: 5105)、QAQQ (SEQ ID NO: 5106)、QTPP (SEQ ID NO: 5107)、QPPA (SEQ ID NO: 5108)、QERP (SEQ ID NO: 5109)、QDLQ (SEQ ID NO: 5110)、QAMH (SEQ ID NO: 5111)、 QHPS (SEQ ID NO: 5112)、PGLQ (SEQ ID NO: 5113)、QGIR (SEQ ID NO: 5114)、QAPA (SEQ ID NO: 5115)、QIPP (SEQ ID NO: 5116)、QTQL (SEQ ID NO: 5117)、QAPS (SEQ ID NO: 5118)、QNTY (SEQ ID NO: 5119)、QDKQ (SEQ ID NO: 5120)、QNHL (SEQ ID NO: 5121)、QIGM (SEQ ID NO: 5122)、LNKQ (SEQ ID NO: 5123)、PNQL (SEQ ID NO: 5124)、 QLQQ (SEQ ID NO: 5125)、QRMS (SEQ ID NO: 5126)、QGIL (SEQ ID NO: 5127)、QDRQ (SEQ ID NO: 5128)、RDWQ (SEQ ID NO: 5129)、QERS (SEQ ID NO: 5130)、QNYQ (SEQ ID NO: 5131)、QRTC (SEQ ID NO: 5132)、QIGH (SEQ ID NO: 5133)、QGAI (SEQ ID NO: 5134)、QVPP (SEQ ID NO: 5135)、QVQQ (SEQ ID NO: 5136)、LMRQ (SEQ ID NO: 5137)、 QYSV (SEQ ID NO:5138), QAIT (SEQ ID NO: 5139), QKTL (SEQ ID NO: 5140), QLHH (SEQ ID NO: 5141), QNII (SEQ ID NO: 5142), QGHH (SEQ ID NO: 5143), QSKV (SEQ ID NO: 5144), QLPS (SEQ ID NO: 5145), IGKQ (SEQ ID NO: 5146), QAIH (SEQ ID NO: 5147), QHGL (SEQ ID NO: 5148), QFMC (SEQ ID NO: 5149), QNQM (SEQ ID NO: 5150), QHLQ (SEQ ID NO: 5151), QPAR (SEQ ID NO: 5152), QSLQ (SEQ ID NO: 5153), QSQL (SEQ ID NO: 5154), HSQQ (SEQ ID NO: 5155)、QMPS (SEQ ID NO: 5156)、QGSL (SEQ ID NO: 5157)、QVPA (SEQ ID NO: 5158)、HYQQ (SEQ ID NO: 5159)、QVPS (SEQ ID NO: 5160)、RGEQ (SEQ ID NO: 5161)、PGQQ (SEQ ID NO: 5162)、LEQQ (SEQ ID NO: 5163)、QNQS (SEQ ID NO: 5164)、QKVI (SEQ ID NO: 5165)、QNND (SEQ ID NO: 5166)、QSVH (SEQ ID NO: 5167)、QPLG (SEQ ID NO: 5168)、HNQE (SEQ ID NO: 5169)、QIQQ (SEQ ID NO: 5170)、QVRN (SEQ ID NO: 5171), PSNQ (SEQ ID NO: 5172)、QVGH (SEQ ID NO: 5173)、QRDI (SEQ ID NO: 5174)、QMPN (SEQ ID NO: 5175)、RGLQ (SEQ ID NO: 5176)、PSLQ (SEQ ID NO: 5177)、QRDQ (SEQ ID NO: 5178)、QAKG (SEQ ID NO: 5179)、QSAH (SEQ ID NO: 5180), QSTM (SEQ ID NO:5181), QREM (SEQ ID NO: 5182), QYRA (SEQ ID NO: 5183), QRQQ (SEQ ID NO: 5184), QWQQ (SEQ ID NO: 5185), QRMN (SEQ ID NO: 5186), GDSQ (SEQ ID NO: 5187), QKIS (SEQ ID NO: 5188), PSMQ (SEQ ID NO: 5189), SPRQ (SEQ ID NO: 5190), MEQQ (SEQ ID NO: 5191), QYQN (SEQ ID NO: 5192), QIRQ (SEQ ID NO: 5193), QSVQ (SEQ ID NO: 5194), RSQQ (SEQ ID NO: 5195), QNKL (SEQ ID NO: 5196), QIQH (SEQ ID NO: 5197), PRQQ (SEQ ID NO: 5198), HTQQ (SEQ ID NO: Specification Page 16 / 285 26 CN 121079310 A 5199), QRQH (SEQ ID NO: 5200), RNQE (SEQ ID NO: 5201), QSKQ (SEQ ID NO: 5202), QNQP (SEQ ID NO: 5203), QSPQ (SEQ ID NO: 5204), QTRQ (SEQ ID NO: 5205), QNLH (SEQ ID NO: 5206), QNQE (SEQ ID NO: 5207), LNQP (SEQ ID NO: 5208), QNQD (SEQ ID NO: 5209), QNLL (SEQ ID NO: 5210), QLVI (SEQ ID NO: 5211), RTQE (SEQ ID NO: 5212), QTHQ (SEQ ID NO: 5213), QDQH (SEQ ID NO: 5214), QSQH (SEQ ID NO: 5215), VRQQ (SEQ ID NO: 5216), AWQQ (SEQ ID NO: 5217), QSVP (SEQ ID NO: 5218), QNIQ (SEQ ID NO: 5219), LDQQ (SEQ ID NO: 5220), PDQQ (SEQ ID NO: 5221), ESQQ (SEQ ID NO: 5222), QRQL (SEQ IDNO: 5223), QIIV (SEQ ID NO: 5224), QKQS (SEQ ID NO: 5225), QSHQ (SEQ ID NO: 5226), QFVV (SEQ ID NO: 5227), QSQP (SEQ ID NO: 5228), QNEQ (SEQ ID NO: 5229), INQQ (SEQ ID NO: 5230), RNRQ (SEQ ID NO: 5231), RDQK (SEQ ID NO: 5232), QWKR (SEQ ID NO: 5233), ENRQ (SEQ ID NO: 5234), QTQP (SEQ ID NO: 5235), QKQL (SEQ ID NO: 5236), RNQL (SEQ ID NO: 5230), 5237), ISIQ (SEQ ID NO: 5238), QTVC (SEQ ID NO: 5239), QQIM (SEQ ID NO: (ii) any part of the amino acid sequence in (i), such as any two or three amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) an amino acid sequence containing one, two or three but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) an amino acid sequence containing one, two or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0118] 42. An AAV particle as described in any one of embodiments 39 to 41, wherein [N1]-[N2]-[N3]-[N4] is or comprises: (i) an amino acid sequence of any one of SEQ ID NO: 1800-2241; (ii) an amino acid sequence comprising any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 amino acids thereon, such as a sequence of consecutive amino acids; (iii) a sequence comprising one, two, or three, but not more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) a sequence comprising one, two, or three, but not more than four different amino acids relative to any one of the amino acid sequences in (i).

[0119] 43.AAV particles of any one of embodiments 39 to 42, wherein [N1]-[N2]-[N3]-[N4] are or contain GSGSPHSKAQNQQ (SEQ ID NO: 1801).

[0120] 44. AAV particles of any one of embodiments 39 to 42, wherein [N1]-[N2]-[N3]-[N4] are or contain GHDSPHKSGQNQQ (SEQ ID NO: 1800).

[0121] 45. AAV particles of any one of embodiments 39 to 42, wherein [N1]-[N2]-[N3]-[N4] are or contain GSGSPHKYGQNQQT (SEQ ID NO: 910).

[0122] 46. An AAV particle as described in any one of embodiments 1 to 45, wherein the AAV capsid variant comprises an amino acid other than T (e.g., S, Y, M, A, C, I, R, L, D, F, V, Q, N, H, E, or G) at position 450, an amino acid other than I (e.g., M, P, E, N, D, S, A, T, G, Q, F, V, L, C, H, R, W, or L) at position 451, and / or an amino acid other than N (e.g., M, E, G, Y, W, T, I, Q, F, V, A, L, I, P, K, R, H, S, D, or S) at position 452.

[0123] 47. An AAV particle as described in any one of embodiments 1 to 46, wherein the AAV capsid variant comprises amino acid T at position 450, amino acid I at position 451, and / or amino acid N at position 452, according to any one of SEQ ID NO: 138, 981, or 982.

[0124] 48. An AAV particle as described in any one of embodiments 1 to 47, wherein the AAV capsid variant further comprises [NO], wherein [NO] comprises XA, XB, and XC, and wherein: (a) XA is: T, S, Y, M, A, C, I, R, L, D, F, V, Q, N, H, E, or G; (b) XB is: I, M, P, E, N, D, S, A, T, G, Q, F, V, L, C, H, R, W, or L; and (c) XC is: N, M, E, G, Y, W, T, I, Q, F, V, A, L, I, P, K, R, H, S, D, or S; and optionally, wherein the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(c), for example, a conservative substitution.

[0125] 49. AAV particles as described in embodiment 48, wherein [NO] is or includes TIN, SMN, TIM, YLS, GLS, MPE,MEG, MEY, AEW, CEW, ANN, IPE, ADM, IEY, ADY, IET, MEW, CEY, RIN, MEI, LEY, ADW, IEI, DIM, FEQ, MEF, CDQ, LPE, IEN, MES, AEI, VEY, IIN, TSN, IEV, MEM, AEV, MDA, VEW, AEQ, LEW, MEL, MET, MEA, IES, MEV, CEI, ATN, MDG, QEV, ADQ, NMN, IEM, ISN, TGN, QQQ, HDW, IEG, TII, TFP, TEK, EIN, TVN, TFN, SIN, TER, TSY, ELH, AIN, SVN, TDN, TFH, TVH, TEN, TSS, TID, TCN, NIN, TEH, AEM, AIK, TDK, TFK, SDQ, TEI, NTN, TET, SIK, TEL, TEA, TAN, TIY, TFS, TES, TTN, TED, TNN, EVH, TIS, TVR, TDR, TIK, NHI, TIP, ESD, TDL, TVP, TVI, AEH, NCL, TVK, NAD, TIT, NCV, TIR, NAL, VIN, TIQ, TEF, TRE, QGE, SEK, NVN, GGE, EFV, SDK, TEQ, EVQ, TEY, NCW, TDV, SDI, NSI, NSL, EVV, TEP, SEL, TWQ, TEV, AVN, GVL, TLN, TEG, TRD, NAI, AEN, AET, ETA, NNL or any dipeptide thereof.

[0126] 50. The AAV particle of embodiment 48 or 49, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or comprises: (i) an amino acid sequence of any one of SEQ ID NO: 2242-2886; (ii) any portion of the amino acid sequence of (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 amino acids thereof, such as an amino acid sequence of consecutive amino acids; (iii) a modified amino acid sequence comprising one, two or three but not more than four modified amino acid sequences relative to any one of the amino acid sequences of (i); or (iv) a modified amino acid sequence comprising one, two or three but not more than four different amino acids relative to any one of the amino acid sequences of (i).

[0127] 51. An AAV particle as described in any one of embodiments 48 to 50, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains TINGSGSPHSKAQNQQ (SEQ ID NO: 2242).

[0128] 52.53. An AAV particle as described in any one of embodiments 48 to 50, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains TINGHDSPHKSGQNQQ (SEQ ID NO: 2243).

[0129] 54. An AAV particle as described in any one of embodiments 48 to 52, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains TINGSGSPHKYGQNQQT (SEQ ID NO: 5246).

[0130] 55. An AAV particle as described in any one of embodiments 1 to 53, wherein [N1]-[N2]-[N3] is present in ring IV.

[0131] 56. An AAV particle as described in any one of embodiments 48 to 54, wherein [N0] and [N4] are present in ring IV.

[0132] 56. An AAV particle as described in any one of embodiments 48 to 55, wherein [NO] is present immediately after position 449 according to the numbering of SEQ ID NO: 138. Specification 18 / 285 pages 28 CN 121079310 A

[0133] 57. An AAV particle as described in any one of embodiments 48 to 56, wherein [NO] is present immediately after position 449 according to the numbering of any one of SEQ ID NO: 36-59, 981 or 982.

[0134] 58. An AAV particle as described in any one of embodiments 48 to 57, wherein [NO] replaces positions 450, 451 and 452 according to the numbering of SEQ ID NO: 138 (e.g., T450, I451 and N452).

[0135] 59. An AAV particle as described in any one of embodiments 48 to 58, wherein [NO] replaces positions 450-452 (e.g., T450, I451, and N452) numbered according to any one of SEQ ID NO: 36-59, 981, or 982.

[0136] 60. An AAV particle as described in any one of embodiments 48 to 59, wherein [NO] corresponds to positions 450-452 of any one of SEQ ID NO: 36-59, 138, 981, or 982.

[0137] 61. An AAV particle as described in any one of embodiments 48 to 60, wherein [NO] is present immediately after position 449, and wherein [NO] replaces positions 450-452 (e.g., T450, I451, and N452) numbered according to SEQ ID NO: 138.

[0138] 62. An AAV particle as described in any of embodiments 48 to 61, wherein [NO] is present immediately after position 449, and wherein [NO] replaces positions 450-452 (e.g., T450, I451, and N452), these positions according to SEQ ID NO:36-59, 981, or 982.

[0139] 63. An AAV particle as described in any of embodiments 1 to 62, wherein [N1] is present immediately after position 452 of the amino acid sequence number according to SEQ ID NO: 138.

[0140] 64. An AAV particle as described in any of embodiments 1 to 63, wherein [N1] is present immediately after position 452 of the number according to SEQ ID NO: 981 or 982.

[0141] 65. An AAV particle as described in any of embodiments 1 to 61, wherein [N1] replaces positions 453-455 of the number according to SEQ ID NO: 138 (e.g., G453, S454, and G455).

[0142] 66. An AAV particle as described in any of embodiments 1 to 64, wherein [N1] replaces position 453 of the number according to SEQ ID NO: 138 (e.g., G453).

[0143] 67. An AAV particle as described in any one of embodiments 1 to 65, wherein [N1] replaces positions 453-455 (e.g., G453, S454, and G455) numbered according to SEQ ID NO: 981.

[0144] 68. An AAV particle as described in any one of embodiments 1 to 65 or 67, wherein [N1] replaces positions 453-455 numbered according to SEQ ID NO: 982.

[0145] 69. An AAV particle as described in any one of embodiments 1 to 65, 67, or 68, wherein [N1] is present immediately after position 452, and wherein [N1] replaces positions 453-455 (e.g., G453, S454, and G455) numbered according to SEQ ID NO: 138.

[0146] 70. An AAV particle as described in any one of embodiments 1 to 64 or 66, wherein [N1] is present immediately after position 452, and wherein [N1] replaces position 453 (e.g., G453), these positions being numbered according to SEQ ID NO: 138.

[0147] 71. An AAV particle as described in any one of embodiments 1 to 64, 66 or 70, wherein [N1] is present immediately after position 452, and wherein [N1] replaces positions 453-455, these positions being numbered according to SEQ ID NO: 4, 36, 981 or 982.

[0148] 72. An AAV particle as described in any one of embodiments 1 to 71, wherein [N1] corresponds to positions 453-455 numbered according to any one of SEQ ID NO: 4, 36-59, 981 or 982.

[0149] 73. An AAV particle as described in any one of embodiments 1 to 72, wherein the AAV capsid variant comprises according to SEQ ID NO:The amino acid other than S at position 454 of number 138, 981, or 982 and / or the amino acid other than G at position 455.

[0150] 74. An AAV particle as described in any of embodiments 1 to 73, wherein the AAV capsid variant comprises amino acid H at position 454 and amino acid D at position 455 according to SEQ ID NO: 138 or 982, page 19 / 285, CN 121079310.

[0151] 75. An AAV particle as described in any of embodiments 1 to 74, wherein the AAV capsid variant comprises a substitution (e.g., S454H) at position 454 according to SEQ ID NO: 138 and / or a substitution (e.g., G455D) at position 455.

[0152] 76. An AAV particle of any one of embodiments 1 to 75, wherein the AAV capsid variant comprises amino acid H at position 454 and amino acid D at position 455, and further comprises the amino acid sequence SPHSKA (SEQ ID NO: 941) immediately following position 455, these positions being numbered according to SEQ ID NO: 138.

[0153] 77. An AAV particle of any one of embodiments 1 to 76, wherein the AAV capsid variant comprises amino acid H at position 454 and amino acid D at position 455, numbered according to SEQ ID NO: 982.

[0154] 78. An AAV particle of any one of embodiments 1 to 77, wherein the AAV capsid variant comprises amino acid H at position 454 and amino acid D at position 455, and further comprises the amino acid sequence SPHSKA (SEQ ID NO: 941) immediately following position 455, these positions being numbered according to SEQ ID NO: 982.

[0155] 79. An AAV particle as described in any one of embodiments 1 to 72, wherein the AAV capsid variant comprises amino acid S at position 454 and amino acid G at position 455 according to SEQ ID NO: 138.

[0156] 80. An AAV particle as described in any one of embodiments 1 to 72 or 79, wherein the AAV capsid variant comprises amino acid S at position 454 and amino acid G at position 455, and further comprises the amino acid sequence SPHSKA (SEQ ID NO: 941) immediately following position 455, these positions being numbered according to SEQ ID NO: 138.

[0157] 81. An AAV particle as described in any one of embodiments 1 to 72, 79 or 80, wherein the AAV capsid variant comprises amino acid S at position 454 and amino acid G at position 455 according to SEQ ID NO: 981.

[0158] 82.AAV particles of any one of embodiments 1 to 72 or 79 to 81, wherein the AAV capsid variant comprises amino acid S at position 454 and amino acid G at position 455, and further comprises the amino acid sequence SPHSKA (SEQ ID NO: 941) immediately following position 455, these positions being numbered according to SEQ ID NO: 981.

[0159] 83. AAV particles of any one of embodiments 1 to 82, wherein [N2] is present immediately following position 455 numbered according to SEQ ID NO: 138.

[0160] 84. AAV particles of any one of embodiments 1 to 83, wherein [N2] corresponds to positions 456-458 (e.g., S456, P457, H458) of SEQ ID NO: 981 or 982.

[0161] 85. An AAV particle as described in any one of embodiments 1 to 83, wherein [N2] corresponds to position 456-458 of any one of SEQ ID NO: 4 or 36-59 (e.g., S456, P457, H458).

[0162] 86. An AAV particle as described in any one of embodiments 1 to 85, wherein [N2]-[N3] are present immediately after position 455 according to SEQ ID NO: 138.

[0163] 87. An AAV particle as described in any one of embodiments 1 to 86, wherein [N2] is present immediately after position 455 according to SEQ ID NO: 4, 36, 981 or 982.

[0164] 88. An AAV particle as described in any one of embodiments 1 to 87, wherein [N2]-[N3] are present immediately after position 455 according to SEQ ID NO: 4, 36, 981 or 982.

[0165] 89. An AAV particle as described in any one of embodiments 1 to 88, wherein [N2]-[N3] correspond to positions 456-461 of SEQ ID NO: 981 (e.g., S456, P457, H458, S459, K460, A461).

[0166] 90. An AAV particle as described in any one of embodiments 1 to 88, wherein [N2]-[N3] correspond to positions 456-461 of SEQ ID NO: 982 (e.g., S456, P457, H458, K459, S460, G461).

[0167] 91. An AAV particle as described in any one of embodiments 1-90, wherein [N2] is present immediately after [N1].

[0168] 92. An AAV particle as described in any one of embodiments 1 to 64, 66, 70 or 71, wherein [N3] is present immediately after [N2]. (Specification page 20 / 285, 30 CN 121079310 A)Positions 454 and 455 (e.g., S454 and G455) exist and replace positions 454 and 455, which are numbered according to SEQ ID NO: 138.

[0169] 93. AAV particles as in any of embodiments 1 to 64, 66, 70, 71 or 92, wherein [N3] exists immediately after [N1]-[N2] and replaces positions 454 and 455 (e.g., S454 and G455), which are numbered according to SEQ ID NO: 138.

[0170] 94. AAV particles as in any of embodiments 39 to 93, wherein [N4] exists immediately after position 455, which is numbered according to SEQ ID NO: 138.

[0171] 95. An AAV particle as described in any one of embodiments 39 to 94, wherein [N4] replaces positions 456-459 (e.g., Q456, N457, Q458, and Q459) according to SEQ ID NO: 138.

[0172] 96. An AAV particle as described in any one of embodiments 39 to 95, wherein [N4] corresponds to positions 462-465 (e.g., Q462, N463, Q464, and Q465) according to SEQ ID NO: 4, 36, 981, or 982.

[0173] 97. An AAV particle as described in any one of embodiments 39 to 96, wherein [N2]-[N3]-[N4] replaces positions 456-459 (e.g., Q456, N457, Q458, and Q459) according to SEQ ID NO: 138.

[0174] 98. An AAV particle as described in any of embodiments 39 to 97, wherein [N2]-[N3]-[N4] is present immediately after position 455, and wherein [N2]-[N3]-[N4] replaces positions 456-459 (e.g., Q456, N457, Q458, and Q459), these positions being numbered according to SEQ ID NO: 138.

[0175] 99. An AAV particle as described in any of embodiments 39 to 98, wherein [N2]-[N3]-[N4] corresponds to positions 456-465 of SEQ ID NO: 981 (e.g., S456, P457, H458, S459, K460, A461, Q462, N463, Q464, and Q465).

[0176] 100. AAV particles as described in any one of embodiments 39 to 98, wherein [N2]-[N3]-[N4] correspond to positions 456-465 of SEQ ID NO: 982 (e.g., S456, P457, H458, K459, S460, G461, Q462, N463, Q464, Q465).

[0177] 101.AAV particles as in any of embodiments 39 to 98, wherein [N2]-[N3]-[N4] correspond to positions 456-465 of SEQ ID NO: 4 or any of 36-59.

[0178] 102. AAV particles as in any of embodiments 39 to 101, wherein [N1]-[N2]-[N3]-[N4] replace positions 453-459 (e.g., G453, S454, G455, Q456, N457, Q458 and Q459) numbered according to SEQ ID NO: 138.

[0179] 103. An AAV particle as described in any of embodiments 39 to 102, wherein [N1]-[N2]-[N3]-[N4] is present immediately after position 452, and wherein [N1]-[N2]-[N3]-[N4] replaces positions 453-459 (e.g., G453, S454, G455, Q456, N457, Q458 and Q459), which are numbered according to SEQ ID NO: 138.

[0180] 104. AAV particles as described in any one of embodiments 39 to 99, 102 or 103, wherein [N1]-[N2]-[N3]-[N4] correspond to positions 453-465 of SEQ ID NO: 981 (e.g., G453, S454, G455, S456, P457, H458, S459, K460, A461, Q462, N463, Q464, Q465).

[0181] 105. AAV particles as described in any one of embodiments 39 to 98, 100, 102 or 103, wherein [N1]-[N2]-[N3]-[N4] correspond to positions 453-465 of SEQ ID NO: 982 (e.g., G453, H454, D455, S456, P457, H458, K459, S460, G461, Q462, N463, Q464, Q465).

[0182] 106. AAV particles as described in any one of embodiments 39 to 98, 102 or 103, wherein [N1]-[N2]-[N3]-[N4] correspond to positions 453-465 of any one of SEQ ID NO: 4 or 36-59.

[0183] 107. AAV particles as described in any one of embodiments 1 to 99 or 102 to 104, wherein [N1]-[N2]-[N3] correspond to positions 453-461 of SEQ ID NO: 981 (e.g., G453, S454, G455, S456, P457, H458, S459, K460, A461). Specification 21 / 285 pages 31 CN 121079310 A

[0184] 108.AAV particles of any one of embodiments 1 to 98, 100, 102, 103 or 105, wherein [N1]-[N2]-[N3] correspond to positions 453-461 of SEQ ID NO: 982 (e.g., G453, H454, D455, S456, P457, H458, K459, S460, G461).

[0185] 109. AAV particles of any one of embodiments 39 to 98, 102, 103 or 106, wherein [N1]-[N2]-[N3] correspond to positions 453-461 of any one of SEQ ID NO: 36-59.

[0186] 110. AAV particles as described in any of embodiments 48 to 109, wherein [N0]-[N1]-[N2]-[N3]-[N4] replace positions 450-459 numbered according to SEQ ID NO: 138 (e.g., T450, I451, N452, G453, S454, G455, Q456, N457, Q458 and Q459).

[0187] 111. An AAV particle as described in any of embodiments 48 to 110, wherein [N0]-[N1]-[N2]-[N3]-[N4] is present immediately after position 449, and wherein [N0]-[N1]-[N2]-[N3]-[N4] replaces positions 450-459 (e.g., T450, I451, N452, G453, S454, G455, Q456, N457, Q458 and Q459), these positions being numbered according to SEQ ID NO: 138.

[0188] 112. AAV particles as described in any one of embodiments 48 to 99, 102 to 104 or 106, wherein [N0]-[N1]-[N2]-[N3]-[N4] correspond to positions 450-465 of SEQ ID NO: 981 (e.g., T450, I451, N452, G453, S454, G455, S456, P457, H458, S459, K460, A461, Q462, N463, Q464, Q465).

[0189] 113. AAV particles as described in any one of embodiments 48 to 98, 100, 102, 103, 105 or 108, wherein [N0]-[N1]-[N2]-[N3]-[N4] correspond to positions 450-465 of SEQ ID NO: 982 (e.g., T450, I451, N452, G453, H454, D455, S456, P457, H458, K459, S460, G461, Q462, N463, Q464, Q465).

[0190] 114.AAV particles of any one of embodiments 48 to 98, 102, 103, 106 or 109, wherein [N0]-[N1]-[N2]-[N3]-[N4] correspond to positions 450-465 of any one of SEQ ID NO: 36-59.

[0191] 115. AAV particles of any one of embodiments 39 to 114, wherein [N4] replaces positions 462-465 (e.g., Q462, N463, Q464 and Q465) numbered according to SEQ ID NO: 4, 36, 981 or 982.

[0192] 116. An AAV particle as described in any one of embodiments 39 to 115, wherein [N2]-[N3]-[N4] replaces positions 462-465 (e.g., Q462, N463, Q464, and Q465) numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0193] 117. An AAV particle as described in any one of embodiments 39 to 116, wherein [N2]-[N3]-[N4] is present immediately after position 455, and wherein [N2]-[N3]-[N4] replaces positions 462-465 (e.g., Q462, N463, Q464, and Q465) numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0194] 118. An AAV particle as described in any one of embodiments 1 to 117, wherein [N3] is present immediately following [N2].

[0195] 119. An AAV particle as described in any one of embodiments 1 to 118, wherein the AAV capsid variant comprises [N2]-[N3] from the N-terminus to the C-terminus.

[0196] 120. An AAV particle as described in any one of embodiments 1 to 119, wherein the AAV capsid variant comprises [N1]-[N2]-[N3] from the N-terminus to the C-terminus.

[0197] 121. An AAV particle as described in any one of embodiments 48 to 120, wherein the AAV capsid variant comprises [NO]-[N1]-[N2]-[N3] from the N-terminus to the C-terminus.

[0198] 122. An AAV particle as described in any one of embodiments 39 to 121, wherein the AAV capsid variant comprises [N1]-[N2]-[N3]-[N4] from the N-terminus to the C-terminus.

[0199] 123. An AAV particle as described in any one of embodiments 48 to 122, wherein the AAV capsid variant comprises [N0]-[N1]-[N2]-[N3]-[N4] from the N-terminus to the C-terminus.

[0200] 124. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises [N0]-[N1]-[N2]-[N3]-[N4] according to SEQ ID NO: 22 / 285 pages 32 CN 121079310 A ID NO:

[0201] 125. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises an amino acid other than T (e.g., N, I, C, H, R, L, D, Y, A, M, Q, I, E, K, P, G, or S) at position 460 according to SEQ ID NO: 138.

[0202] 126. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises an amino acid other than T (e.g., N, I, C, H, R, L, D, Y, A, M, Q, I, E, K, P, G, or S) at position 466 according to any of SEQ ID NO: 36-59, 981, or 982.

[0203] 127. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises amino acid N, I, C, H, R, L, D, Y, A, M, Q, I, E, K, P, G, or S at position 466 according to any of the numbers in SEQ ID NO: 36-59, 981, or 982.

[0204] 128. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises amino acid other than K (e.g., E, N, or T) at position 449 according to any of the numbers in SEQ ID NO: 36-59, 138, 981, or 982.

[0205] 129. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises amino acid E, N, or T at position 449 according to any of the numbers in SEQ ID NO: 36-59, 138, 981, or 982.

[0206] 130. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises [A][B] (SEQ ID NO: 4694), wherein: (i) [A] comprises the amino acid sequence GSGSPH (SEQ ID NO: 4695); and (ii) [B] comprises X1 X2 X3 X4 X5 X6 X7, wherein: (a) X1 is: S, C, F, or V; (b) X2 is: K, L, R, I, E, Y, V, or S; (c) X3 is: A, R, L, G, I, Y, S, F, or W; (d) X4 is: W, Q, R, G, L, V, S, or F; (e) X5 is: N, Y, R, C, K, or L; (f) X6 is: Q, G, K, R, T, L, or Y; and(g) X7 is: Q, L, R or V; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(g), for example, a conservative substitution.

[0207] 131. The AAV particle of embodiment 130, wherein

[0208] (a) X1 is S; (b) X2 is K or L; (c) X3 is: A, R or L; (d) X4 is: Q or R; (e) X5 is: N or R; (f) X6 is: Q or R; and (g) X7 is: Q, L or R.

[0209] 132. AAV particles as described in embodiment 130 or 131, wherein [B] comprises: (i) SLLWNQQ (SEQ ID NO: 5247), SKAQYYV (SEQ ID NO: 5248), SKLRRQQ (SEQ ID NO: 5249), SIWQNQQ (SEQ ID NO: 5250), SKAGCGQ (SEQ ID NO: 5251), SRAQNQQ (SEQ ID NO: 5252), SKRLRQQ (SEQ ID NO: 5253), SLRRNQQ (SEQ ID NO: 5254), SRGRNQQ (SEQ ID NO: 5255), SEIVNQQ (SEQ ID NO: 5256), SSRRNQQ (SEQ ID NO: 5257), CLLQNQQ (SEQ ID NO: 5258), SLRRNQQ (SEQ ID NO: 5259 ... 5258), SKAFRLQ (SEQ ID NO: 5259), CLAQNQQ (SEQ ID NO: 5260), FLRQNQQ (SEQ ID NO: 5261), SLRFNQQ (SEQ ID NO: 5262), SYLRNQQ (SEQ ID NO: 5263), CSLQNQQ (SEQ ID NO: 5264), VLWQNQQ (SEQ ID NO: 5265), SKWLLQQ (SEQ ID NO: 5266), SLWSNQQ (SEQ ID NO: 5267), SKRRLQQ (SEQ ID NO: 5268), SVYLNQQ (SEQ ID NO: 5269), SLWLNQQ (SEQ ID NO: 5270), SKAQRKL (SEQ ID NO: 5271), SKALRRQ (SEQ ID NO: 5272), SKAQRLR (SEQ ID NO: 5273), SKAQNQQ (SEQ ID NO:(ii) any portion of the amino acid sequence in (i), such as any 2, 3, 4, 5, or 6 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence containing one, two, or three, but no more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) a sequence containing one, two, or three, but no more than four modified amino acid sequences. An amino acid sequence containing one, two, or three, but no more than four different amino acids, relative to any one of the amino acid sequences in (i).

[0210] 133. AAV particles as described in any one of embodiments 130 to 132, wherein [A][B] comprises: (i) GSGSPHSLLWNQQ (SEQ ID NO: 5285), GSGSPHSKAQYYV (SEQ ID NO: 2060), GSGSPHSKLRRQQ (SEQ ID NO: 2061), GSGSPHSIWQNQQ (SEQ ID NO: 5286), GSGSPHSKAGCGQ (SEQ ID NO: 2062), GSGSPHSRAQNQQ (SEQ ID NO: 2063), GSGSPHSKRLRQQ (SEQ ID NO: 2064), GSGSPHSLRRNQQ (SEQ ID NO: 2065), GSGSPHSRGRNQQ (SEQ ID NO: 2066), GSGSPHSEIVNQQ (SEQ ID NO: 5287), GSGSPHSSRRNQQ (SEQ ID NO: 5288), GSGSPHSSRRNQQ (SEQ ID NO: 5288), GSGSPHSSRRNQQ (SEQ ID NO: 5288), GSGSPHSLR ... 2067), GGSSPHCLLQNQQ (SEQ ID NO: 5288), GSGSPHSKAFRLQ (SEQ ID NO: 2068), GGSSPHCLAQNQQ (SEQ ID NO: 5289), GGSSPHFLRQNQQ (SEQ ID NO: 2070), GSGSPHSLRFNQQ (SEQ ID NO: 2071),GSGSPHSYLRNQQ (SEQ ID NO: 5290), GGSSPHCSLQNQQ (SEQ ID NO: 5291), GGSSPHVLWQNQQ (SEQ ID NO: 5292), GSGSPHSKWLLQQ (SEQ ID NO: 2072), GGSPHSLWSNQQ (SEQ ID NO: 5293), GGSSPHSKRRLQQ (SEQ ID NO: 2073), GSGSPHSVYLNQQ (SEQ ID NO: 5294), GSGSPHSLWLNQQ (SEQ ID NO: 5295), GSGSPHSKAQRKL (SEQ ID NO: 2074), GSGSPHSKALRRQ (SEQ ID NO: 2075), GGSPHSKALRRQ (SEQ ID NO: 2075), GSGSPHSKAQRLR (SEQ ID NO: 2076), GGSSPHSKAQNQQ (SEQ ID NO: 1801), GGSSPHSKAQRRL (SEQ ID NO: (ii) Any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 amino acids, such as a sequence of consecutive amino acids; (iii) Relative to any of the amino acid sequences in (i), comprising one, two, or three but not more than four modified amino acid sequences; or (iv) relative to any of the amino acid sequences in (i), comprising one, two, or three but not more than four different amino acid sequences.

[0211] 134. An AAV particle as described in any of embodiments 130 to 133, wherein the AAV capsid variant further comprises according to SEQ ID NO:

[0212] 135. An AAV particle as described in any one of embodiments 130 to 134, wherein the AAV capsid variant further comprises S at position 450 (other than T, e.g., S, Y, or G), amino acid at position 451 (other than I, e.g., M or L), and / or amino acid at position 452 (other than N, e.g., S).

[0213] 136. An AAV particle as described in any one of embodiments 130 to 134, wherein the AAV capsid variant further comprises Y at position 450, L at position 451, and S at position 452, according to SEQ ID NO: 138.

[0214] 137. An AAV particle as described in any one of embodiments 130 to 134, wherein the AAV capsid variant further comprises G at position 450, L at position 451, and S at position 452, as specified in SEQ ID NO: 138.

[0215] 138. An AAV particle as described in any one of embodiments 130 to 137, wherein [A] is present in ring IV.

[0216] 139. An AAV particle as described in any one of embodiments 130 to 138, wherein [A] is present immediately after position 452, as specified in SEQ ID NO: 138.

[0217] 140. An AAV particle as described in any one of embodiments 130 to 139, wherein [A] replaces positions 453-455, as specified in SEQ ID NO: 138 (e.g., G453, S454, G455).

[0218] 141. An AAV particle as described in any of embodiments 130 to 140, wherein [A] is present immediately after position 452, and wherein [A] replaces positions 453-455 (e.g., G453, S454, G455), which are numbered according to SEQ ID NO: 138.

[0219] 142. An AAV particle as described in any of embodiments 130 to 141, wherein [B] is present immediately after [A].

[0220] 143. An AAV particle as described in any of embodiments 130 to 142, wherein [B] replaces positions 456-459 (e.g., Q456, N457, Q458, Q459) numbered according to SEQ ID NO: 138.

[0221] 144. AAV particles as described in any one of embodiments 130 to 143, wherein [A][B] replaces positions 453-459 (e.g., G453, S454, G455, Q456, N457, Q458, Q459) according to SEQ ID NO: 138.

[0222] 145.AAV particle as described in any of embodiments 130 to 144, wherein [A][B] is present immediately after position 452, and wherein [A][B] replaces positions 453-459 (e.g., G453, S454, G455, Q456, N457, Q458, Q459), which are numbered according to SEQ ID NO: 138.

[0223] 146. AAV particle as described in any of embodiments 130 to 145, wherein the AAV capsid variant comprises [A][B] from the N-terminus to the C-terminus.

[0224] 147. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises [A][B] (SEQ ID NO: 4699), wherein: (i) [A] comprises X1 X2 X3 X4 X5 X6, wherein (a) X1 is T, M, A, C, I, R, L, D, F, V, Q, N, or H; (b) X2 is I, P, E, N, D, S, A, T, M, or Q; (c) X3 is N, E, G, Y, W, M, T, I, K, Q, F, S, V, A, or L; (d) X4 is G, D, R, or E; Specification 25 / 285 pages 35 CN 121079310 A (e) X5 is H, Q, N or D; (f) X6 is D or R; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(f), for example, a conservative substitution; and (ii) [B] comprises SPHKSG (SEQ ID NO: 946).

[0225] 148. AAV particles as described in embodiment 147, wherein

[0226] (a) X1 is: T, M, A or I; (b) X2 is: E, I or D; (c) X3 is: N, Q, Y, I, M or V; (d) X4 is G; (e) X5 is H; and (f) X6 is D.

[0227] 149. AAV particles as described in embodiments 147 or 148, wherein [A] comprises: (i) TINGHD (SEQ ID NO: 5297), MPEGHD (SEQ ID NO: 5298), MEGGHD (SEQ ID NO: 5299), MEYGHD (SEQ ID NO: 5300), AEWGHD (SEQ ID NO: 5301), CEWGHD (SEQ ID NO: 5302), ANNGQD (SEQ ID NO: 5303), IPEGHD (SEQ ID NO: 5304), EGGHD (SEQ ID NO: 5305), EGGHD (SEQ ID NO: 5306), EGGHD (SEQ ID NO: 5307), EGGHD (SEQ ID NO: 5308), EGGHD (SEQ ID NO: 5309), EGGHD (SEQ ID NO: 5300), EGGHD (SEQ ID NO: 5301), EGGHD (SEQ ID NO: 5302), EGGHD (SEQ ID NO: 5303), EGGHD (SEQ ID NO: 5304), EGGHD (SEQ ID NO: 5305), EGGHD (SEQ ID NO: 5306), EGGHD (SEQ ID NO: 5307), EGGHD (SEQ ID NO: 5308), EGGHD (SEQ ID NO: 5309 ...5304)、ADMGHD (SEQ ID NO: 5305)、IEYGHD (SEQ ID NO: 5306)、ADYGHD (SEQ ID NO: 5307)、IETGHD (SEQ ID NO: 5308)、MEWGHD (SEQ ID NO: 5309)、CEYGHD (SEQ ID NO: 5310)、RINGHD (SEQ ID NO: 5311)、MEIGHD (SEQ ID NO: 5312)、LEYGHD (SEQ ID NO: 5313)、ADWGHD (SEQ ID NO: 5314)、IEIGHD (SEQ ID NO: 5315)、TIKDND (SEQ ID NO: 5316)、DIMGHD (SEQ ID NO: 5317)、FEQGHD (SEQ ID NO: 5318)、MEFGHD (SEQ ID NO: 5319)、CDQGHD (SEQ ID NO: 5320)、LPEGHD (SEQ ID NO: 5321)、IENGHD (SEQ ID NO: 5322)、MESGHD (SEQ ID NO: 5323)、AEIGHD (SEQ ID NO: 5324)、VEYGHD (SEQ ID NO: 5325)、TSNGDD (SEQ ID NO: 5326)、IEVGHD (SEQ ID NO: 5327)、MEMGHD (SEQ ID NO: 5328)、AEVGHD (SEQ ID NO: 5329)、MDAGHD (SEQ ID NO: 5330)、VEWGHD (SEQ ID NO: 5331)、AEQGHD (SEQ ID NO: 5332)、LEWGHD (SEQ ID NO: 5333)、MELGHD (SEQ ID NO: 5334)、METGHD (SEQ ID NO: 5335)、MEAGHD (SEQ ID NO: 5336)、TINRQR (SEQ ID NO: 5337)、IESGHD (SEQ ID NO: 5338)、TAKDHD (SEQ ID NO: 5339)、MEVGHD (SEQ ID NO: 5340)、CEIGHD (SEQ ID NO: 5341)、ATNGHD (SEQ ID NO: 5342)、MDGGHD (SEQ ID NO: 5343)、QEVGHD (SEQ ID NO:(ii) any part of the amino acid sequence in (i), such as any 2, 3, 4 or 5 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence of one, two or three but no more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) a sequence of one, two or three but no more than four different amino acids relative to any of the amino acid sequences in (i). Instruction manual, pages 26 / 285, 36 CN 121079310 A

[0228] 150. An AAV particle as described in any one of embodiments 147 to 149, wherein [A][B] comprises: (i) TINGHDSPHKS (SEQ ID NO: 5354), MPEGHDSPHKS (SEQ ID NO: 5355), MEGGHDSPHKS (SEQ ID NO: 5356), MEYGHDSPHKS (SEQ ID NO: 5357), AEWGHDSPHKS (SEQ ID NO: 5358), CEWGHDSPHKS (SEQ ID NO: 5359), ANNGQDSPHKS (SEQ ID NO: 5360), IPEGHDSPHKS (SEQ ID NO: 5361), ADMGHDSPHKS (SEQ ID NO: 5362), IEYGHDSPHKS (SEQ ID NO: 5363), ADYGHDSPHKS (SEQ ID NO: 5364), EGGHDSPHKS (SEQ ID NO: 5365 ... 5364), IETGHDSPHKS (SEQ ID NO: 5365), MEWGHDSPHKS (SEQ ID NO: 5366), CEYGHDSPHKS (SEQ ID NO: 5367), RINGHDSPHKS (SEQ ID NO: 5368), MEIGHDSPHKS (SEQ ID NO: 5369), LEYGHDSPHKS (SEQ ID NO: 5370)、ADWGHDSPHKS (SEQ ID NO: 5371)、IEIGHDSPHKS (SEQ ID NO: 5372)、TIKDNDSPHKS (SEQ ID NO: 5373)、DIMGHDSPHKS (SEQ ID NO: 5374)、FEQGHDSPHKS (SEQ ID NO: 5375)、 MEFGHDSPHKS (SEQ ID NO: 5376)、CDQGHDSPHKS (SEQ ID NO: 5377)、LPEGHDSPHKS (SEQ ID NO: 5378)、IENGHDSPHKS (SEQ ID NO: 5379)、MESGHDSPHKS (SEQ ID NO: 5380)、 AEIGHDSPHKS (SEQ ID NO: 5381)、VEYGHDSPHKS (SEQ ID NO: 5382)、TSNGDDSPHKS (SEQ ID NO: 5383)、IEVGHDSPHKS (SEQ ID NO: 5384)、MEMGHDSPHKS (SEQ ID NO: 5385)、 AEVGHDSPHKS (SEQ ID NO: 5386)、MDAGHDSPHKS (SEQ ID NO: 5387)、VEWGHDSPHKS (SEQ ID NO: 5388)、AEQGHDSPHKS (SEQ ID NO: 5389)、LEWGHDSPHKS (SEQ ID NO: 5390)、 MELGHDSPHKS (SEQ ID NO: 5391)、METGHDSPHKS (SEQ ID NO: 5392)、MEAGHDSPHKS (SEQ ID NO: 5393)、TINRQRSPHKS (SEQ ID NO: 5394)、IESGHDSPHKS (SEQ ID NO: 5395)、 TAKDHDSPHKS (SEQ ID NO: 5396)、MEVGHDSPHKS (SEQ ID NO: 5397)、CEIGHDSPHKS (SEQ ID NO: 5398)、ATNGHDSPHKS (SEQ ID NO: 5399)、MDGGHDSPHKS (SEQ ID NO: 5400)、 QEVGHDSPHKS (SEQ ID NO: 5401)、ADQGHDSPHKS (SEQ ID NO: 5402)、NMNGHDSPHKS (SEQ ID NO:5403), TPWEHDSPHKS (SEQ ID NO: 5404), IEMGHDSPHKS (SEQ ID NO: 5405), TANEHDSPHKS (SEQ ID NO: 5406), TINGHDSPHKS (SEQ ID NO: 5407), QQQGHDSPHKS (SEQ ID NO: 5408), TPQDHDSPHKS (SEQ ID NO: 5409), HDWGHDSPHKS (SEQ ID NO: 5410), IEGGHDSPHKS (SEQ ID NO: 5411) (ii) any portion of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence containing one, two, or three, but no more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) Relative to any of the amino acid sequences in (i), an amino acid sequence comprising one, two, or three, but not more than four, different amino acids.

[0229] 151. An AAV particle as described in any of embodiments 147 to 150, wherein the AAV capsid variant further comprises one, two, three, four, or all of the following amino acids: at position 456 other than Q (e.g., R or L), at position 457 other than N (e.g., H, K, or R), at position 458 other than Q (e.g., R or T), at position 459 other than Q (H), and / or at position 460 other than T (N or S).

[0230] 152. An AAV particle as described in any of embodiments 147 to 151, wherein the AAV capsid variant further comprises R at position 456 as described in SEQ ID NO: 138.

[0231] 153. An AAV particle as described in any one of embodiments 147 to 151, wherein the AAV capsid variant further comprises L at position 456 according to SEQ ID NO: 138 (page 27 / 285, CN 121079310 A).

[0232] 154. An AAV particle as described in any one of embodiments 147 to 153, wherein the AAV capsid variant further comprises H at position 457 and R at position 458 according to SEQ ID NO: 138.

[0233] 155. An AAV particle as described in any one of embodiments 147 to 153, wherein the AAV capsid variant further comprises L at position 456 according to SEQ ID NO: 138.138, K at position 457 and N at position 460.

[0234] 156. An AAV particle as described in any of embodiments 147 to 153, wherein the AAV capsid variant further comprises T at position 458, H at position 459 and S at position 460 according to SEQ ID NO: 138.

[0235] 157. An AAV particle as described in any of embodiments 147 to 151, wherein the AAV capsid variant further comprises R at position 456, R at position 457 and R at position 458 according to SEQ ID NO: 138.

[0236] 158. An AAV particle as described in any of embodiments 147 to 157, wherein [A][B] are present in ring IV.

[0237] 159. An AAV particle as described in any of embodiments 147 to 158, wherein [A] is present immediately after position 449 according to SEQ ID NO: 138.

[0238] 160. An AAV particle as described in any of embodiments 147 to 159, wherein [A] replaces positions 450-453 (e.g., T450, I451, N452, G453) according to SEQ ID NO: 138.

[0239] 161. An AAV particle as described in any of embodiments 147 to 160, wherein [A] is present immediately after position 449, and wherein [A] replaces positions 450-453 (e.g., T450, I451, N452, G453) according to SEQ ID NO: 138.

[0240] 162. An AAV particle as described in any one of embodiments 147 to 161, wherein [A][B] replaces positions 450-455 (e.g., T450, I451, N452, G453, S454, G455) numbered according to SEQ ID NO: 138.

[0241] 163. An AAV particle as described in any one of embodiments 147 to 162, wherein [A][B] is present immediately after position 449, and wherein [A][B] replaces positions 450-455 (e.g., T450, I451, N452, G453, S454, G455) numbered according to SEQ ID NO: 138.

[0242] 164. An AAV particle as described in any of embodiments 147 to 163, wherein [B] is present immediately after [A] and replaces positions 454 and 455 (e.g., S454 and G455), these positions being numbered according to SEQ ID NO: 138.

[0243] 165. An AAV particle as described in any of embodiments 147 to 164, wherein [B] is present immediately after position 455 numbered according to SEQ ID NO: 4, 36, 981 or 982.

[0244] 166. An AAV particle as described in any one of embodiments 147 to 165, wherein [B] is present immediately following [A].

[0245] 167. An AAV particle as described in any one of embodiments 147 to 166, wherein the AAV capsid variant comprises [A][B] from the N-terminus to the C-terminus.

[0246] 168. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence having the following formula: [N1]-[N2]-[N3] (SEQ ID NO: 6407), wherein: (i) [N1] comprises X1, X2, and X3, wherein X2 is S and X3 is G; (ii) [N2] comprises the amino acid sequence SPH; and (iii) [N3] comprises X4, X5, and X6, wherein X5 is K.

[0247] 169. An AAV particle as described in embodiment 168, wherein: (i) X4 of [N3] is S, T, N, or A; and (ii) X5 of [N3] is A, V, T, S, G, R, L, or N; Optionally, the AAV capsid variant comprises an amino acid modification, for example a conservative substitution, of any of the aforementioned amino acids in (i) or (ii).

[0248] 170. An AAV particle as described in embodiment 168 or 169, wherein X4 is S and / or X5 is A.

[0249] 171. An AAV particle as described in any one of embodiments 168 to 170, wherein [N3] comprises SK, TK, NK, AK, KA, KV, KT, KS, KG, KR, KL, or KN.

[0250] 172. An AAV particle as described in any one of embodiments 168 to 171, wherein [N3] is or includes SKA, SKV, SKT, SKS, SKG, SKR, TKA, NKA, SKL, SKN, or AKA.

[0251] 173. An AAV particle as described in any one of embodiments 168 to 172, wherein [N3] is or includes SKA.

[0252] 174. An AAV particle as described in any one of embodiments 168 to 173, wherein [N2]-[N3] includes SPHSK (SEQ ID NO: 4701), SPHTK (SEQ ID NO: 4725), SPHNK (SEQ ID NO: 4726), or SPHAK (SEQ ID NO: 4727).

[0253] 175. An AAV particle as described in any one of embodiments 168 to 174, wherein [N2]-[N3] is or comprises:(i) SPHSKA (SEQ ID NO: 941), SPHSKV (SEQ ID NO: 4737), SPHSKT (SEQ ID NO: 4731), SPHSKS (SEQ ID NO: 962), SPHSKG (SEQ ID NO: 4732), SPHSKR (SEQ ID NO: 978), SPHTKA (SEQ ID NO: 4739), SPHNKA (SEQ ID NO: 4734), SPHSKL (SEQ ID NO: 960), SPHSKN (SEQ ID NO: 4735), or SPHAKA (SEQ ID NO: 4736); (ii) any portion of the amino acid sequence in (i), such as any 2, 3, 4, or 5 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence containing one, two, or three, but no more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) Relative to any of the amino acid sequences in (i), an amino acid sequence comprising one, two, or three, but not more than four, different amino acids.

[0254] 176. An AAV particle as described in any of embodiments 168 to 175, wherein [N2]-[N3] is or comprises SPHSKA (SEQ ID NO: 941).

[0255] 177. An AAV particle as described in any of embodiments 168 to 176, wherein the AAV capsid variant comprises an amino acid other than G (e.g., M, T, I, E, S, A, N, V, L, K, H, P, R, W, or D) at position 453 according to SEQ ID NO: 138 or 981.

[0256] 178. An AAV particle as described in any of embodiments 168 to 177, wherein the AAV capsid variant comprises amino acid G at position 453 according to SEQ ID NO: 138 or 981.

[0257] 179. An AAV particle as described in any one of embodiments 168 to 178, wherein X1 of [N1] is G, M, T, I, E, S, A, N, V, L, K, H, P, R, W, or D; optionally, wherein the AAV capsid variant comprises an amino acid modification of any one of the aforementioned amino acids, for example, a conserved substitution.

[0258] 180. An AAV particle as described in any one of embodiments 168 to 179, wherein [N1] comprises SG, GS, MS, TS, IS, ES, SS, AS, NS, VS, LS, KS, HS, PS, RS, WS, or DS.

[0259] 181. An AAV particle as described in any one of embodiments 168 to 180, wherein [N1] is or comprises: GSG, MSG,TSG, ISG, ESG, SSG, ASG, NSG, VSG, LSG, KSG, HSG, PSG, RSG, WSG, or DSG.

[0260] 182. AAV particles as described in any one of embodiments 168 to 181, wherein [N1] is or contains GSG.

[0261] 183. AAV particles as described in any one of embodiments 168 to 182, wherein [N1]-[N2] contains SGSPH (SEQ ID NO: 4752). Instruction manual, pages 29 / 285, CN 121079310 A

[0262] 184. AAV particles as described in any one of embodiments 168 to 183, wherein [N1]-[N2] are or comprise: (i) GSGSPH (SEQ ID NO: 4695), MSGSPH (SEQ ID NO: 4798), TSGSPH (SEQ ID NO: 4800), ISGSPH (SEQ ID NO: 4801), ESGSPH (SEQ ID NO: 4803), SSGSPH (SEQ ID NO: 4804), ASGSPH (SEQ ID NO: 4806), NSGSPH (SEQ ID NO: 4807), VSGSPH (SEQ ID NO: 4786), LSGSPH (SEQ ID NO: 4808), KSGSPH (SEQ ID NO: 4810), HSGSPH (SEQ ID NO: 4811), PSGSPH (SEQ ID NO: 4811), (ii) any part of the amino acid sequence in (i), such as any 2, 3, 4 or 5 amino acids, such as an amino acid sequence of consecutive amino acids; (iii) an amino acid sequence containing one, two or three but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) an amino acid sequence containing one, two or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0263] 185. AAV particles as described in any one of embodiments 168 to 184, wherein [N1]-[N2]-[N3] are or comprise: (i) GSGSPHSKA (SEQ ID NO: 4697), GSGSPHSKV (SEQ ID NO: 4956), MSGSPHSKA (SEQ ID NO: 4957), TSGSPHSKA (SEQ ID NO: 4957), TSGSPHSKA (SEQ ID NO: 4957), GS ...4959), ISGSPHSKA (SEQ ID NO: 4960), GSGSPHSKT (SEQ ID NO: 4962), ESGSPHSKA (SEQ ID NO: 4963), SSGSPHSKA (SEQ ID NO: 4964), GGSPHSKS (SEQ ID NO: 4953), ASGSPHSKA (SEQ ID NO: 4966), NSGSPHSKA (SEQ ID NO: 4967), VSGSPHSKA (SEQ ID NO: 4913), LSGSPHSKA (SEQ ID NO: 4968), KSGSPHSKA (SEQ ID NO: 4970), GSGSPHSKG (SEQ ID NO: 4972), GSGSPHSKR (SEQ ID NO: 4945), HSGSPHSKA (SEQ ID NO: 4973), PSGSPHSKA (SEQ ID NO: 4975), RSGSPHSKA (SEQ ID NO: (ii) any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8 or 9 amino acids, such as a sequence of consecutive amino acids; (iii) a sequence of one, two or three but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) a sequence of one, two or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0264] 186. An AAV particle as described in any one of embodiments 168 to 185, wherein [N1]-[N2]-[N3] is or contains GSGSPHASKA (SEQ ID NO: 4697).

[0265] 187. An AAV capsid variant as described in any one of embodiments 168 to 186, comprising an amino acid other than Q (e.g., R, P, H, L, K, I, G, S, M, or E) at position 456 according to SEQ ID NO: 138, and an amino acid other than N at position 457.Amino acids other than Q (e.g., D, V, S, P, T, G, Y, W, E, R, H, K, F, A, I, L, or M), amino acids other than Q at position 458 (e.g., R, L, A, P, H, T, I, F, K, V, M, G, W, Y, S, E, N, or D), amino acids other than Q at position 459 (e.g., H, K, A, L, P, E, M, I, S, N, R, Y, C, V, T, W, D, G), and / or amino acids other than T at position 460 (e.g., I, N, S, H, R, L, D, Y, A, or Q).

[0266] 188. An AAV particle as described in any one of embodiments 168 to 187, wherein the AAV capsid variant comprises according to the specification, pages 30 / 285, 40 CN 121079310 A SEQ ID NO: Amino acids other than Q (e.g., R, P, H, L, K, I, G, S, M, or E) at position 462 of number 981, amino acids other than N (e.g., D, V, S, P, T, G, Y, W, E, R, H, K, F, A, I, L, or M) at position 464, amino acids other than Q (e.g., R, L, A, P, H, T, I, F, K, V, M, G, W, Y, S, E, N, or D) at position 465, amino acids other than Q (e.g., H, K, A, L, P, E, M, I, S, N, R, Y, C, V, T, W, D, G) at position 466, and / or amino acids other than T (e.g., I, N, S, H, R, L, D, Y, A, or Q) at position 466.

[0267] 189. An AAV particle as described in any one of embodiments 168 to 188, wherein the AAV capsid variant comprises amino acid Q at position 456, amino acid N at position 457, amino acid Q at position 458, amino acid Q at position 459, and / or amino acid T at position 460, as specified in SEQ ID NO: 138.

[0268] 190. An AAV particle as described in any one of embodiments 168 to 189, wherein the AAV capsid variant comprises amino acid Q at position 462, amino acid N at position 463, amino acid Q at position 464, amino acid Q at position 465, and / or amino acid T at position 466, as specified in SEQ ID NO: 981.

[0269] 191. An AAV particle as described in any one of embodiments 168 to 190, wherein the AAV capsid variant further comprises [N4], wherein [N4] comprises X7, X8, X9, X10, and X11, wherein: (a) X7 is Q, R, P, H, L, K, I, G, S, M, or E; (b) X8 is N, D, V, S, P, T, G, Y, W, E, R, H, K, F, A, I, L, or M; (c)X9 is Q, R, L, A, P, H, T, I, F, K, V, M, G, W, Y, S, E, N, D; (d) X10 is Q, H, K, A, L, P, E, M, I, S, N, R, Y, C, V, T, W, D, G; and (e) X11 is T, I, N, S, H, R, L, D, Y, A, Q; Optionally, the AAV capsid variant contains an amino acid modification of any of the aforementioned amino acids in (a)-(e), such as a conservative substitution.

[0270] 192. The AAV particle of embodiment 191, wherein [N4] is or comprises: (i) QNQQT (SEQ ID NO: 5412), QNRHT (SEQ ID NO: 5413), RDQQT (SEQ ID NO: 5414), PNLQT (SEQ ID NO: 5415), HVRQT (SEQ ID NO: 5416), PNQHT (SEQ ID NO: 5417), QSQQT (SEQ ID NO: 5418), QNQQI (SEQ ID NO: 5419), QPAKT (SEQ ID NO: 5420), QTQQN (SEQ ID NO: 5421), QNLAT (SEQ ID NO: 5422), QNQLT (SEQ ID NO: 5423), QGQQT (SEQ ID NO: 5424), LNRQS (SEQ ID NO: 5425), HNQQT (SEQ ID NO: 5426), QNQQT (SEQ ID NO: 5427), QNQQT (SEQ ID NO: 5428), QNQQT (SEQ ID NO: 5429), QNQQT (SEQ ID NO: 5420), QNQQT ...NQQT (SEQ ID NO: 5422), QNQQT (SEQ ID NO: 5423), NO: 5426), QNPPT (SEQ ID NO: 5427), QNLQT (SEQ ID NO: 5428), QYQQT (SEQ ID NO: 5429), QDQET (SEQ ID NO: 5430), QNHQT (SEQ ID NO: 5431), QDQQT (SEQ ID NO: 5432), HWQQT (SEQ ID NO: 5433), PNQQT (SEQ ID NO: 5434), QNQLI (SEQ ID NO: 5435), PEQQT (SEQ ID NO: 5436), QRTMT (SEQ ID NO: 5437), QNQQH (SEQ ID NO: 5438), LHQHT (SEQ ID NO: 5439), QHRIT (SEQ ID NO: 5440), QYIHT (SEQ ID NO: 5441), QKFET (SEQ ID NO: 5442), QFPST (SEQ ID NO: 5443), HNQQR (SEQ ID NO:5444), QAIKT (SEQ ID NO: 5445), QNRQT (SEQ ID NO: 5446), QYQHT (SEQ ID NO: 5447), QNPQS (SEQ ID NO: 5448), QHQLT (SEQ ID NO: 5449), QSPPT (SEQ ID NO: 5450), QAKLT (SEQ ID NO: 5451), KSQQT (SEQ ID NO: 5452), QDRPT (SEQ ID NO: 5453), QSQQL (SEQ ID NO: 5454), QAFHT (SEQ ID NO: 5455), QKQQD (SEQ ID NO: 5456), QNAQT (SEQ ID NO: 5457), HNQLT (SEQ ID NO: 5458), QNQQY (SEQ ID NO: 5459), QKLNT (SEQ ID NO: 5460), QNVQT (SEQ ID NO: 5461), QAQQT (SEQ ID NO: 5462), QNLQA (SEQ ID NO: 5463), QTPPT (SEQ ID NO: 5464), QYQHA (SEQ ID NO: Description 31 / 285 pages 41 CN 121079310 A 5465), QGQQA (SEQ ID NO: 5466), QPPAT (SEQ ID NO: 5467), QERPT (SEQ ID NO: 5468), QDLQT (SEQ ID NO: 5469), QAMHT (SEQ ID NO: 5470), LNQQT (SEQ ID NO: 5471), QHPST (SEQ ID NO: 5472), PGLQT (SEQ ID NO: 5473), QGIRT (SEQ ID NO: 5474), QAPAT (SEQ ID NO: 5475), QSQQI (SEQ ID NO: 5476), QIPPT (SEQ ID NO: 5477), QTQLT (SEQ ID NO: 5478), QAPST (SEQ ID NO: 5479), QNTYA (SEQ ID NO: 5480), QNQHI (SEQ ID NO: 5481), QNHLT (SEQ ID NO: 5482), QIGMT (SEQ ID NO: 5483), LNKQT (SEQ ID NO:5484)、QLQQT (SEQ ID NO: 5485)、QRMST (SEQ ID NO: 5486)、QGILT (SEQ ID NO: 5487)、QDRQT (SEQ ID NO: 5488)、RDWQT (SEQ ID NO: 5489)、QNTHD (SEQ ID NO: 5490)、PNLQI (SEQ ID NO: 5491)、QERST (SEQ ID NO: 5492)、QNYQT (SEQ ID NO: 5493)、QRTCT (SEQ ID NO: 5494)、QIGHT (SEQ ID NO: 5495)、QGAIT (SEQ ID NO: 5496)、QVPPT (SEQ ID NO: 5497)、QVQQI (SEQ ID NO: 5498)、LMRQT (SEQ ID NO: 5499)、QYSVT (SEQ ID NO: 5500)、QAITT (SEQ ID NO: 5501)、QKTLT (SEQ ID NO: 5502)、QNQWT (SEQ ID NO: 5503)、QLHHT (SEQ ID NO: 5504)、QNIII (SEQ ID NO: 5505)、QGHHT (SEQ ID NO: 5506)、QSKVT (SEQ ID NO: 5507)、QLPST (SEQ ID NO: 5508)、IGKQT (SEQ ID NO: 5509)、QAIHT (SEQ ID NO: 5510)、QHGLT (SEQ ID NO: 5511)、QFMCT (SEQ ID NO: 5512)、QHLQT (SEQ ID NO: 5513)、QNHQN (SEQ ID NO: 5514)、QPART (SEQ ID NO: 5515)、QSLQT (SEQ ID NO: 5516)、QSQLT (SEQ ID NO: 5517)、QDRQS (SEQ ID NO: 5518)、QMPST (SEQ ID NO: 5519)、QGSLT (SEQ ID NO: 5520)、QVPAT (SEQ ID NO: 5521)、QDKQT (SEQ ID NO: 5522)、HYQQT (SEQ ID NO: 5523)、QVPST (SEQ ID NO: 5524)、RGEQT (SEQ ID NO: 5525)、PGQQT (SEQID NO: 5526), ​​QSLQI (SEQ ID NO: 5527), LEQQT (SEQ ID NO: 5528), QNQST (SEQ ID NO: 5529), QKVIT (SEQ ID NO: 5530), QNNDQ (SEQ ID NO: 5531), QSVHT (SEQ ID NO: 5532), QPLGT (SEQ ID NO: 5533), HNQET (SEQ ID NO: 5534), QNLQI (SEQ ID NO: 5535), QIQQT (SEQ ID NO: 5536), QVRNT (SEQ ID NO: 5537), PSNQT (SEQ ID NO: 5538), QVGHT (SEQ ID NO: 5539), QRDIT (SEQ ID NO: 5540), QMPNT (SEQ ID NO: 5541), RGLQT ​​(SEQ ID NO: 5542), QKQQT (SEQ ID NO: 5543), PSLQT (SEQ ID NO: 5544), QRDQT (SEQ ID NO: 5545), QAKGT (SEQ ID NO: 5546), QSAHT (SEQ ID NO: 5547), QSTMT (SEQ ID NO: 5548), QREMT (SEQ ID NO: 5549), QYRAT (SEQ ID NO: 5550), QWQQT (SEQ ID NO: 5551), QRMNT (SEQ ID NO: 5552), GDSQT (SEQ ID NO: 5553), QKIST (SEQ ID NO: 5554), PSMQT (SEQ ID NO: 5555), SPRQT (SEQ ID NO: 5556), MEQQT (SEQ ID NO: 5557), QYQNT (SEQ ID NO: 5558), QHQQT (SEQ ID (ii) any portion of the amino acid sequence in (i), such as any 2, 3, or 4 amino acids, such as a sequence of consecutive amino acids; (iii) amino acids of the same type.Relative to any of the amino acid sequences in (i), there is an amino acid sequence containing one, two, or three but no more than four modified amino acids; or (iv) relative to any of the amino acid sequences in (i), there is an amino acid sequence containing one, two, or three but no more than four different amino acids.

[0271] 193. AAV particles as described in embodiments 191 or 192, wherein [N1]-[N2]-[N3]-[N4] are or comprise: (i) an amino acid sequence of any one of SEQ ID NO: 200 or 2887-3076; (ii) an amino acid sequence comprising any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or 13 amino acids thereon, such as a sequence of consecutive amino acids; (iii) an amino acid sequence comprising one, two or three but not more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) an amino acid sequence comprising one, two or three but not more than four different amino acids relative to any one of the amino acid sequences in (i).

[0272] 194. An AAV particle as described in any one of embodiments 191 to 193, wherein [N1]-[N2]-[N3]-[N4] is or contains GSGSPHSKAQNQQT (SEQ ID NO: 200).

[0273] 195. An AAV particle as described in any one of embodiments 191 to 193, wherein [N1]-[N2]-[N3]-[N4] is or contains VSGSPHSKAQNQQT (SEQ ID NO: 903).

[0274] 196. An AAV particle as described in any one of embodiments 168 to 195, wherein the AAV capsid variant comprises an amino acid other than K (e.g., T, E, or N) at position 449, an amino acid other than T (e.g., S, E, A, N, V, Q, or G) at position 450, an amino acid other than I (e.g., F, E, V, L, D, S, C, T, A, N, H, R, G, or W) at position 451, and / or an amino acid other than N (e.g., I, P, K, R, H, S, M, Q, D, T, L, A, Y, V, F, E, W, or G) at position 452.

[0275] 197. An AAV particle as described in any one of embodiments 168 to 196, wherein the AAV capsid variant comprises amino acid K at position 449, amino acid T at position 450, amino acid I at position 451, and / or amino acid N at position 452, as specified in SEQ ID NO: 138 or 981.

[0276] 198.The AAV particle of any one of embodiments 168 to 197, wherein the AAV capsid variant further comprises [NO], wherein [NO] comprises XA, XB, XC and XD, wherein: (a) XA is K, T, E or N; (b) XB is T, S, E, A, N, V, Q or G; (c) XC is I, F, E, V, L, D, S, C, T, A, N, H, R, G or W; and (d) XD is N, I, P, K, R, H, S, M, Q, D, T, L, A, Y, V, F, E, W or G; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(d), for example, a conservative substitution.

[0277] 199. AAV particles as described in embodiment 198, wherein [NO] is or comprises: (i) KTII (SEQ ID NO: 5565), KTFP (SEQ ID NO: 5566), KTEK (SEQ ID NO: 5567), KTVN (SEQ ID NO: 5568), KTFN (SEQ ID NO: 5569), KTIN (SEQ ID NO: 5570), TTIN (SEQ ID NO: 5571), KSIN (SEQ ID NO: 5572), KTER (SEQ ID NO: 5573), KELH (SEQ ID NO: 5574), KAIN (SEQ ID NO: 5575), KTDN (SEQ ID NO: 5576), KTFH (SEQ ID NO: 5577), KTSN (SEQ ID NO: 5578), ETIN (SEQ ID NO: 5579), NTIN (SEQ ID NO: 5579), KTN (SEQ ID NO: 5570), KTN (SEQ ID NO: 5576), KTFH (SEQ ID NO: 5577), KTSN (SEQ ID NO: 5578), ETIN (SEQ ID NO: 5579), NTIN (SEQ ID NO: 5570), KTN (SEQ ID NO: 5576), KTN (SEQ ID NO: 5577), KTN (SEQ ID NO: 5578), KTN (SEQ ID NO: 5579), KTN ... NO: 5580), KTEN (SEQ ID NO: 5581), KTSS (SEQ ID NO: 5582), KTCN (SEQ ID NO: 5583), KTEH (SEQ ID NO: 5584), KAEM (SEQ ID NO: 5585), KATN (SEQ ID NO: 5586), KAIK (SEQ ID NO: 5587), KTDK (SEQ ID NO: 5588), KTFK (SEQ ID NO: 5589), KSDQ (SEQ ID NO: 5590), KTEI (SEQ ID NO: 5591), KTID (SEQ ID NO: 5592), KNTN (SEQ ID NO: Instructions 33 / 285 Page 43 CN 121079310 A 5593), KTET (SEQ IDNO: 5594)、KTEL (SEQ ID NO: 5595)、KNIN (SEQ ID NO: 5596)、 KTEA (SEQ ID NO: 5597)、KTANI (SEQ ID NO: 5598)、NTIY (SEQ ID NO: 5599)、NTIFS (SEQ ID NO: 5599) 5600)、KTES (SEQ ID NO: 5601)、KTTN (SEQ ID NO: 5602)、KTED (SEQ ID NO: 5603)、KTNN (SEQ ID NO: 5604)、KEVH (SEQ ID NO: 5605)、SEQ ID NO: 5605)、SEQ ID NO: 5605) 5606)、KTVR (SEQ ID NO: 5607)、KTDR (SEQ ID NO: 5608)、ETIK (SEQ ID NO: 5609)、 KNHI (SEQ ID NO: 5610)、KESD (SEQ ID NO: 5611)、KTVR (SEQ ID NO: 5611) (SEQ ID NO: 5613)、KTVP (SEQ ID NO: 5614)、KTVI (SEQ ID NO: 5615)、KAEH (SEQ ID NO: 5616)、KNCL (SEQ ID NO: 5617)、KTVK (SEQ ID NO: 5618)、KTVK (SEQ ID NO: 5618)、KTVK (SEQ ID NO: 5618) 5619)、KTIT (SEQ ID NO: 5620)、KNCV (SEQ ID NO: 5621)、KNAL (SEQ ID NO: 5622)、KVIN (SEQ ID NO: 5623)、KTEF (SEQ ID NO: 5624)、KTEF (SEQ ID NO: 5624)、SEQ ID NO: 5625)、KQGE (SEQ ID NO: 5626)、KSEK (SEQ ID NO: 5627)、KNVN (SEQ ID NO: 5628)、KGGE (SEQ ID NO: 5629)、KEFV (SEQ ID NO: 5630)、KEFV (SEQ ID NO: 5630) (SEQ ID NO: 5630) 5631)、KTEQ (SEQ ID NO: 5632)、KEVQ (SEQ ID NO: 5633)、KTEY (SEQ ID NO: 5634)、KNCW (SEQ ID NO: 5635)、KTDV (SEQ ID NO: 5636)、KTDV (SEQ ID NO: 5636)、SEQ ID NO:5637), KNSI (SEQ ID NO: 5638), KNSL (SEQ ID NO: 5639), KEVV (SEQ ID NO: 5640), KTEP (SEQ ID NO: 5641), KSEL (SEQ ID NO: 5642), KTWQ (SEQ ID NO: 5643), KTEV (SEQ ID NO: 5637) 5644), KAVN (SEQ ID NO: 5645), KGVL (SEQ ID NO: 5646), KTEG (SEQ ID NO: 5647), KTRD (SEQ ID NO: 5648), KTGN (SEQ ID NO: 5649), KNAI (SEQ ID NO: 5650), KAEN (SEQ ID NO: 5651), KAET (SEQ ID NO: 5652), KTVH (SEQ ID NO: 5653), KETA (SEQ ID NO: (ii) any part of the amino acid sequence in (i), such as any two or three amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) an amino acid sequence containing one, two or three but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) an amino acid sequence containing one, two or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0278] 200. AAV particles as described in embodiments 198 or 199, wherein [N0]-[N1]-[N2]-[N3]-[N4] are or comprise: (i) an amino acid sequence of any one of SEQ ID NO: 3239-3526 or 3591-3605; (ii) an amino acid sequence comprising any portion of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 or 17 amino acids therein, such as a sequence of consecutive amino acids; (iii) an amino acid sequence comprising one, two or three but not more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) an amino acid sequence comprising one, two or three but not more than four different amino acids relative to any one of the amino acid sequences in (i).

[0279] 201.AAV particles as described in any of embodiments 198 to 200, wherein [N0]-[N1]-[N2]-[N3]-[N4] are or contain KTINGSGSPHSKAQNQQT (SEQ ID NO: 5660).

[0280] 202. AAV particles as described in any of embodiments 198 to 200, wherein [N0]-[N1]-[N2]-[N3]-[N4] are or contain KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589). (See specification page 34 / 285, 44 CN 121079310 A)

[0281] 203. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence having the following formula: [N1]-[N2]-[N3] (SEQ ID NO: 6408), wherein: (i) [N1] comprises X1, X2, and X3, wherein X2 is an amino acid other than S and X3 is an amino acid other than G; (ii) [N2] comprises the amino acid sequence SPH; and (iii) [N3] comprises X4, X5, and X6, wherein X4 is K.

[0282] 204. The AAV particle of embodiment 203, wherein: (i) X5 of [N3] is S, I, T, R, H, Y, L or M; and (ii) X6 of [N3] is G, A, L, E, V, R, W, N, Q or K; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (i) or (ii), for example, a conservative substitution.

[0283] 205. The AAV particle of embodiment 203 or 204, wherein X5 is S and / or X6 is G.

[0284] 206. An AAV particle as described in any one of embodiments 203 to 205, wherein [N3] comprises KS, KI, KT, KR, KH, KY, KL, KM, SG, IG, TG, RG, SA, SL, SE, SV, SR, SW, SN, HG, YG, SQ, IV, SK, LW, MG, or MA.

[0285] 207. An AAV particle as described in any one of embodiments 203 to 206, wherein [N3] is or comprises KSG, KIG, KTG, KRG, KSA, KSL, KSE, KSV, KSR, KSW, KSN, KHG, KYG, KSQ, KIV, KSK, KLW, KMG, or KMA.

[0286] 208. An AAV particle as described in any one of embodiments 203 to 207, wherein [N3] is or comprises KSG.

[0287] 209.AAV particles as described in any of embodiments 203 to 208, wherein [N2]-[N3] include SPHKS (SEQ ID NO: 4704), SPHKI (SEQ ID NO: 4713), SPHKT (SEQ ID NO: 4711), SPHKR (SEQ ID NO: 4717), NPHKS (SEQ ID NO: 5661), SPHKH (SEQ ID NO: 4728), SPHKY (SEQ ID NO: 4715), SPHKL (SEQ ID NO: 4714), or SPHKM (SEQ ID NO: 4729).

[0288] 210. AAV particles as described in any one of embodiments 203 to 209, wherein [N2]-[N3] are or comprise: (i) SPHKSG (SEQ ID NO: 946), SPHKIG (SEQ ID NO: 958), SPHKTG (SEQ ID NO: 4738), SPHKRG (SEQ ID NO: 974), NPHKSG (SEQ ID NO: 5662), SPHKSA (SEQ ID NO: 977), SPHKSL (SEQ ID NO: 4740), SPHKSE (SEQ ID NO: 4741), SPHKSV (SEQ ID NO: 4742), SPHKSR (SEQ ID NO: 951), SPHKSW (SEQ ID NO: 4743), SPHKSN (SEQ ID NO: 4744), SPHKHG (SEQ ID NO: 4745), SPHKYG (SEQ ID NO: 966), SPHKSQ (SEQ ID NO: 9745), SPHKSQ (SEQ ID NO: 9746), SPHKSG ... (ii) any part of the amino acid sequence in (i), such as any 2, 3, 4 or 5 amino acids, such as a sequence of consecutive amino acids; (iii) a sequence of one, two or three but no more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) a sequence of one, two or three but no more than four different amino acids relative to any of the amino acid sequences in (i).

[0289] 211.AAV particles of any one of embodiments 203 to 210, wherein [N2]-[N3] is or contains SPHKSG (SEQ ID NO: 946). Specification 35 / 285 pages 45 CN 121079310 A

[0290] 212. AAV particles of any one of embodiments 203 to 211, wherein the AAV capsid variant contains an amino acid other than G (e.g., A, K, W, R, L, I, M, N, T, E, Q, Y, H, F or V) at position 453 according to SEQ ID NO: 138 or 981.

[0291] 213. AAV particles of any one of embodiments 203 to 212, wherein the AAV capsid variant contains amino acid G at position 453 according to SEQ ID NO: 138 or 981.

[0292] 214. An AAV particle as described in any one of embodiments 203 to 214, wherein: (i) X1 of [N1] is G, A, K, W, R, L, I, M, N, T, E, Q, Y, H, F, or V; (ii) X2 of [N1] is H, Y, R, Q, N, P, or D; (iii) X3 of [N1] is D, E, G, V, or N; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (i), (ii), or (iii), for example, a conservative substitution.

[0293] 215. An AAV particle as described in any one of embodiments 203 to 214, wherein X2 of [N1] is H and X3 of [N1] is D.

[0294] 216. An AAV particle as described in any one of embodiments 203 to 215, wherein X1 of [N1] is G, X2 of [N1] is H, and X3 of [N1] is D.

[0295] 217. An AAV particle as described in any one of embodiments 203 to 216, wherein [N1] comprises GH, HD, GY, GR, GQ, AH, GN, KH, GP, WH, RH, LH, IH, MH, GD, NH, TH, EH, QH, YH, HH, FH, VH, YD, HE, RG, QD, RD, ND, PD, QV, DD, HN, or NG.

[0296] 218. An AAV capsid variant as described in any one of embodiments 203 to 217, wherein [N1] is or includes GHD, GYD, GHE, GRG, GQD, GRD, AHD, GND, KHD, GPD, WHD, RHD, LHD, GQV, IHD, MHD, GDD, GHN, NHD, THD, GNG, EHD, QHD, YHD, HHD, FHD, or VHD.

[0297] 219. An AAV particle as described in any one of embodiments 203 to 218, wherein [N1] is or includes GHD.

[0298] 220.AAV particles as described in any of embodiments 203 to 219, wherein [N1]-[N2] contain HDSPH (SEQ ID NO: 4703).

[0299] 221. An AAV particle as described in any one of embodiments 203 to 220, wherein [N1]-[N2] is or comprises: (i) GHDSPH (SEQ ID NO: 4784), GYDSPH (SEQ ID NO: 4829), GHESPH (SEQ ID NO: 4793), GRGSPH (SEQ ID NO: 4788), GHDNPH (SEQ ID NO: 5664), GQDSPH (SEQ ID NO: 4785), GRDSPH (SEQ ID NO: 4831), AHDSPH (SEQ ID NO: 5665), GNDSPH (SEQ ID NO: 4832), KHDSPH (SEQ ID NO: 5666), GPDSPH (SEQ ID NO: 4833), WHDSPH (SEQ ID NO: 5667), RHDSPH (SEQ ID NO: 5668), LHDSPH (SEQ ID NO: 5668), ... 5669), GQVSPH (SEQ ID NO: 4835), IHDSPH (SEQ ID NO: 5670), MHDSPH (SEQ ID NO: 5671), GDDSPH (SEQ ID NO: 4792), GHNSPH (SEQ ID NO: 4836), NHDSPH (SEQ ID NO: 5672), THDSPH (SEQ ID NO: 5673), GNGSPH (SEQ ID NO: 4805), EHDSPH (SEQ ID NO: 5674), QHDSPH (SEQ ID NO: 5675), YHDSPH (SEQ ID NO: 5676), HHDSPH (SEQ ID NO: 5677), FHDSPH (SEQ ID NO: 5678) or VHDSPH (SEQ ID NO: 5679); (ii) (i) Any portion of the amino acid sequence in (i), such as any 2, 3, 4, or 5 amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) A sequence containing one, two, or three, but not more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or (iv) A sequence containing one, two, or three, but not more than four modified amino acid sequences relative to any one of the amino acid sequences in (i).The amino acid sequence of each different amino acid.

[0300] 222. The AAV particle of any one of embodiments 203 to 221, wherein [N1]-[N2]-[N3] is or comprises: (i) GHDSPHKSG (SEQ ID NO: 4698), GHDSPHKIG (SEQ ID NO: 4996), GYDSPHKSG (SEQ ID NO: 4997), GHESPHKSG (SEQ ID NO: 4998), GHDSPHKTG (SEQ ID NO: 4999), GRGSPHKRG (SEQ ID NO: 5000), GHDNPHKSG (SEQ ID NO: 5680), GQDSPHKSG (SEQ ID NO: 4908), GHDSPHKSA (SEQ ID NO: 4940), GHDSPHKSL (SEQ ID NO: 5001), GHDSPHKSE (SEQ ID NO: 5003), GRDSPHKSG (SEQ ID NO: 5004), AHDSPHKSG (SEQ ID NO: 5004), AHDSPHKSG (SEQ ID NO: 5005), GHDSPHKSG (SEQ ID NO: 5006), GHDSPHKSG (SEQ ID NO: 5007), GHDSPHKSG (SEQ ID NO: 4998), GHDSPHKSG (SEQ ID NO: 49 ... 5681), GNDSPHKSV (SEQ ID NO: 5005), AHDSPHKIG (SEQ ID NO: 5682), GHESPHKSA (SEQ ID NO: 4939), GQDSPHKIG (SEQ ID NO: 5006), GHDSPHKSV (SEQ ID NO: 5007), GHDSPHKSR (SEQ ID NO: 4942), KHDSPHKSG (SEQ ID NO: 5683), GPDSPHKIG (SEQ ID NO: 5008), GPDSPHKSG (SEQ ID NO: 5009), GHDSPHKSW (SEQ ID NO: 5010), WHDSPHKSG (SEQ ID NO: 5684), RHDSPHKSG (SEQ ID NO: 5685), GHDSPHKSN (SEQ ID NO: 5011), GHDSPHKRG (SEQ ID NO: 4937), GHDSPHKHG (SEQ ID NO: 5013), LHDSPHKSG (SEQ ID NO: 5686), GQVSPHKSG (SEQ ID NO: 5014), IHDSPHKSG (SEQ ID NO: 5687), MHDSPHKSG (SEQ ID NO: 5688), GDDSPHKSV (SEQ ID NO:5015), GHNSPHKSG (SEQ ID NO: 5016), NHDSPHKSG (SEQ ID NO: 5689), THDSPHKSG (SEQ ID NO: 5690), GGNSPHKRG (SEQ ID NO: 5017), EHDSPHKSG (SEQ ID NO: 5691), GHDSPHKYG (SEQ ID NO: 5018), GHDSPHKSG (SEQ ID NO: 5019), QHDSPHKSG (SEQ ID NO: 5692), RHDSPHKIV (SEQ ID NO: 5693), YHDSPHKSG (SEQ ID NO: 5694), GNDSPHKIG (SEQ ID NO: 5020), HHDSPHKSG (SEQ ID NO: 5695), GHDSPHKSK (SEQ ID NO: 5021), FHDSPHKSG (SEQ ID NO: 5696), GHDSPHKLW (SEQ ID NO: 5022), VHDSPHKSG (SEQ ID NO: 5697), GHDSPHKMG (SEQ ID NO: 5024), GHDSPHKMA (SEQ ID NO: 5025) or GDDSPHKSG (SEQ ID NO: 4938); (ii) comprising any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8 or 9 amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) comprising one, two or three but not more than four modified amino acid sequences relative to any of the amino acid sequences in (i); or (iv) comprising one, two or three but not more than four different amino acid sequences relative to any of the amino acid sequences in (i).

[0301] 223. AAV particles as in any of embodiments 203 to 222, wherein [N1]-[N2]-[N3] is or comprises GHDSPHKSG (SEQ ID NO: 4698).

[0302] 224. An AAV particle as described in any one of embodiments 203 to 223, wherein the AAV capsid variant comprises an amino acid other than Q at position 456 (e.g., R, P, H, K, L, V, A, E, or I) according to SEQ ID NO: 138, an amino acid other than N at position 457 (e.g., I, K, S, H, R, T, D, Y, L, W, F, A, Q, or M), and an amino acid other than Q at position 458.Amino acids (e.g., R, V, K, P, Y, H, L, I, E, or M), amino acids at position 459 other than Q (e.g., H, L, E, P, W, D, I, V, S, K, R, C, M, or N), and / or amino acids at position 460 other than T (e.g., A, E, K, S, I, P, G, or N).

[0303] 225. An AAV particle as described in any one of embodiments 203 to 224, wherein the AAV capsid variant comprises an amino acid other than Q (e.g., R, P, H, K, L, V, A, E, or I) at position 462 according to SEQ ID NO: 982, an amino acid other than N (e.g., I, K, S, H, R, T, D, Y, L, W, F, A, Q, or M) at position 463, an amino acid other than Q (e.g., R, V, K, P, Y, H, L, I, E, or M) at position 464, an amino acid other than Q (e.g., H, L, E, P, W, D, I, V, S, K, R, C, M, or N) at position 465, and / or an amino acid other than T (e.g., A, E, K, S, I, P, G, or N) at position 466.

[0304] 226. An AAV particle as described in any one of embodiments 203 to 225, wherein the AAV capsid variant comprises amino acid Q at position 456, amino acid N at position 457, amino acid Q at position 458, amino acid Q at position 459, and / or amino acid T at position 460, according to SEQ ID NO: 138.

[0305] 227. An AAV particle as described in any one of embodiments 203 to 226, wherein the AAV capsid variant comprises amino acid Q at position 462, amino acid N at position 463, amino acid Q at position 464, amino acid Q at position 465, and / or amino acid T at position 466, according to SEQ ID NO: 138.

[0306] 228. An AAV particle as described in any one of embodiments 203 to 227, wherein the AAV capsid variant further comprises [N4], wherein [N4] comprises X7, X8, X9, X10, and X11, wherein: (a) X7 is Q, R, P, H, L, K, I, G, S, M, or E; (b) X8 is N, D, V, S, P, T, G, Y, W, E, R, H, K, F, A, I, L, or M; (c) X9 is Q, R, L, A, P, H, T, I, F, K, V, M, G, W, Y, S, E, N, D; (d) X10 is Q, H, K, A, L, P, E, M, I, S, N, R, Y, C, V, T, W, D, G; and (e) X11 is T, I, N, S, H, R, L, D, Y, A, Q;Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(e), such as a conservative substitution.

[0307] 229. AAV particles as described in embodiment 228, wherein [N4] is or comprises: (i) QNQQT (SEQ ID NO: 5412), QIRQT (SEQ ID NO: 5698), QNQHA (SEQ ID NO: 5699), QKQQT (SEQ ID NO: 5543), QSVQT (SEQ ID NO: 5700), RSQQT (SEQ ID NO: 5701), QNKLE (SEQ ID NO: 5702), QNQQK (SEQ ID NO: 5703), QHQQA (SEQ ID NO: 5704), QIQHT (SEQ ID NO: 5705), PRQQT (SEQ ID NO: 5706), HTQQT (SEQ ID NO: 5707), QRQHT (SEQ ID NO: 5708), QSQQT (SEQ ID NO: 5418), QNQQS (SEQ ID NO: 5418), QNQQT (SEQ ID NO: 5412), QIRQT (SEQ ID NO: 5698), QNQHA (SEQ ID NO: 5699), QKQQT (SEQ ID NO: 5543), QSVQT (SEQ ID NO: 5700), RSQQT (SEQ ID NO: 5701), QNKLE (SEQ ID NO: 5702), QNQQK (SEQ ID NO: 5703), QHQQA (SEQ ID NO: 5704), QIQHT (SEQ ID NO: 5705), PRQQT (SEQ ID NO: 5706), HTQQT (SEQ ID NO: 5707), QRQHT (SEQ ID NO: 5708), QSQQT (SEQ ID NO: 5418), QNQQS (SEQ ID NO: 5412), QIQHT (SEQ ID NO: 5543), QSQQT (SEQ ID NO: 5543), NO: 5709), RNQET (SEQ ID NO: 5710), QTQLT (SEQ ID NO: 5478), KNQQT (SEQ ID NO: 5711), QDQQT (SEQ ID NO: 5432), HNQQT (SEQ ID NO: 5426), QNQLT (SEQ ID NO: 5423), QTQQT (SEQ ID NO: 5712), QTQQI (SEQ ID NO: 5713), QSKQA (SEQ ID NO: 5714), QNQPP (SEQ ID NO: 5715), QSPQT (SEQ ID NO: 5716), QNYQT (SEQ ID NO: 5493), QNHQT (SEQ ID NO: 5431), QNRQT (SEQ ID NO: 5446), QNQQG (SEQ ID NO: 5717), QNHLT (SEQ ID NO: 5482), QYQHT (SEQ ID NO: 5447), QNQWT (SEQ ID NO: 5503), QNQHT (SEQ ID NO: 5718), QTRQT (SEQ ID NO: 5719), QNLHT (SEQ ID NO: 5720), LNQQT (SEQ ID NO: 5471), QNQET (SEQ ID NO:5721), QHLQT (SEQ ID NO: 5513), LNQPT (SEQ ID NO: 5722), QNQDT (SEQ ID NO: 5723), RNQQT (SEQ ID NO: 5724), QNLLT (SEQ ID NO: 5725), QLVIT (SEQ ID NO: 5726), RTQET (SEQ ID NO: 5727), QTHQT (SEQ ID NO: 5728), QNQPA (SEQ ID NO: 5729), QDQHT (SEQ ID NO: 5730), QSQHT (SEQ ID NO: 5731), RNQQI (SEQ ID NO: 5732), VRQQT (SEQ ID NO: 5733), QNQHS (SEQ ID NO: 5734), AWQQT (SEQ ID NO: 5735), QSVPT (SEQ ID NO: 5736), QNIQP (SEQ ID NO: 5737), QNHLN (SEQ ID NO: 5738), LDQQT (SEQ ID NO: 5739), PDQQS (SEQ ID NO: 5740), ESQQT (SEQ ID NO: 5741), QNKQT (SEQ ID NO: 5742), QRQLT (SEQ ID NO: 5743), QIIVT (SEQ ID NO: 5744), QKQST (SEQ ID NO: 5745), QSHQT (SEQ ID NO: 5746), QFVVT (SEQ ID NO: 5747), QNLQT (SEQ ID NO: 5428), QNQQI (SEQ ID NO: 5419), QSQPT (SEQ ID NO: 5748), QNEQT (SEQ ID NO: 5749), QSLQT (SEQ ID NO: 5516), RNRQT (SEQ ID NO: 5750), QSKQT (SEQ ID NO: 5751), QNPLT (SEQ ID NO: 5752), RDQKT (SEQ ID NO: 5753), HNQQN (SEQ ID NO: 5754), QWKRT (SEQ ID NO: 5755), QSQQI (SEQ ID NO: 5476), QAQQT (SEQ ID NO:5462), QNHQI (SEQ ID NO: 5756), QNQQA (SEQ ID NO: 5757), QNQLN (SEQ ID NO: 5758), QTQPT (SEQ ID NO: 5759), INQQT (SEQ ID NO: 5560), QKQLT (SEQ ID NO: 5760), RNQLA (SEQ ID NO: 5761), RNQQS (SEQ ID NO: 5762), ISIQT (SEQ ID NO: 5763), QNQQN (SEQ ID NO: 5764), QSQQS (SEQ ID NO: 5765), QTVCT (SEQ ID NO: 5766), QYQQI (SEQ ID NO: 5767), QQIMT (SEQ ID NO: 5768), QNEQS (SEQ ID NO: 5769), LNHQT (SEQ ID NO: 5770), QMIHT (ii) any part of the amino acid sequence in (i), such as any 2, 3 or 4 amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) a modified amino acid sequence containing one, two or three but not more than four modified amino acids relative to any of the amino acid sequences in (i); or (iv) an amino acid sequence containing one, two or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0308] 230. The AAV particle of embodiment 228 or 229, wherein [N1]-[N2]-[N3]-[N4] is or comprises: (i) an amino acid sequence of any one of SEQ ID NO: 201 or 3160-3237; (ii) any portion of the amino acid sequence of (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or 13 amino acids thereon, such as a sequence of consecutive amino acids; (iii) a sequence of one, two or three but not more than four modified amino acid sequences relative to any one of the amino acid sequences of (i); or (iv) a sequence of one, two or three but not more than four different amino acids relative to any one of the amino acid sequences of (i).

[0309] 231.AAV particles as described in any of embodiments 228 to 230, wherein [N1]-[N2]-[N3]-[N4] is or contains GHDSPHKSGQNQQT (SEQ ID NO: 201).

[0310] 232. AAV particles as described in any of embodiments 203 to 231, wherein the AAV capsid variant contains an amino acid other than K (e.g., T) at position 449 according to SEQ ID NO: 138 or 982, an amino acid other than T at position 450 (e.g., A, S, I, V, N, E, Y, C, G, W or Q), an amino acid other than I at position 451 (e.g., E, V, S, T, N, D, C, G, Q, L, P, A) and / or an amino acid other than N at position 452 (e.g., S, Y, I, K, F, T, D, E, G, V, L, A, M, Q, H, P or R).

[0311] 233. An AAV particle as described in any one of embodiments 203 to 232, wherein the AAV capsid variant comprises amino acid K at position 449, amino acid T at position 450, amino acid I at position 451, and / or amino acid N at position 452, as specified in SEQ ID NO: 138 or 982.

[0312] 234. An AAV particle as described in any one of embodiments 203 to 233, wherein the AAV capsid variant further comprises [NO], wherein [NO] comprises XA, XB, XC and XD, wherein: (a) XA is K or T; (b) XB is T, A, S, I, V, N, E, Y, C, G, W or Q; (c) XC is I, E, V, S, T, N, D, C, G, Q, L, P, A; and (d) XD is N, S, Y, I, K, F, T, D, E, G, V, L, A, M, Q, H, P or R; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(d), for example, a conservative substitution.

[0313] 235. AAV particles as described in embodiment 234, wherein [NO] is or comprises: (i) KAIN (SEQ ID NO: 5575), KTIN (SEQ ID NO: 5570), KTES (SEQ ID NO: 5601), TTIN (SEQ ID NO: 5571), KSIN (SEQ ID NO: 5572), KTVN (SEQ ID NO: 5568), KSIY (SEQ ID NO: 5775), KTSN (SEQ ID NO: 5578), KTTN (SEQ ID NO: 5602), KIIN (SEQ ID NO: 5568), KSIY (SEQ ID NO: 5775), KTSN (SEQ ID NO: 5578), KTTN (SEQ ID NO: 5602), KIIN (SEQ ID NO: 5570), KTIY (SEQ ID NO: 5775), KTSN (SEQ ID NO: 5578), KTTN (SEQ ID NO: 5602), KIIN (SEQ ID NO: 5570), KTIY (SEQ ID NO: 5575), KTSN (SEQ ID NO: 5578), KTTN (SEQ ID NO: 5602), KIIN (SEQ ID NO: 5570), KTIY (SEQ ID NO: 5575), KTIY (SEQ ID NO: 5570 ...5776)、KTIS (SEQ ID NO: 5606)、KAII (SEQ ID NO: 5777)、KTIK (SEQ ID NO: 5612)、KTEF (SEQ ID NO: 5624)、KTIT (SEQ ID NO: 5620)、KTIT (SEQ ID NO: 5620)、KTIN (SEQ ID NO: 5620) 5604)、KTID (SEQ ID NO: 5592)、KAIS (SEQ ID NO: 5778)、KTVD (SEQ ID NO: 5779)、KTIE (SEQ ID NO: 5780)、KTEG (SEQ ID NO: 5647)、KTEG (SEQ ID NO: 5647)、KTEG NO: 5623)、KAVN (SEQ ID NO: 5645)、KTIY (SEQ ID NO: 5781)、KTDN (SEQ ID NO: 5576)、KTCN (SEQ ID NO: 5583)、KNVV (SEQ ID NO: 5782)、KNVV (SEQ ID NO: 5782) (SEQ ID NO: 5782) 5595)、KTDA (SEQ ID NO: 5783)、KTEV (SEQ ID NO: 5644)、KSEL (SEQ ID NO: 5642)、KTEM (SEQ ID NO: 5784)、 KTEQ (SEQ ID NO: 5632)、SEQ ID NO: 5565)、KIVN (SEQ ID NO: 5785)、KTEK (SEQ ID NO: 5567)、KTEN (SEQ ID NO: 5581)、KIGN (SEQ ID NO: 5786)、KEVM (SEQ ID NO: 5787)、SEQ ID NO: 5788)、KTEA (SEQ ID NO: 5597)、KATN (SEQ ID NO: 5586)、KTEH (SEQ ID NO: 5584)、KTVE (SEQ ID NO: 5789)、KAID (SEQ ID NO: 5790)、KTIM (SEQ ID NO: 5580) 5791)、KEVG (SEQ ID NO: 5792)、KSEM (SEQ ID NO: 5793)、KAQQ (SEQ ID NO: 5794)、KCGE (SEQ ID NO: 5795)、KASN (SEQ ID NO: 5796)、SEQ ID NO: T 5594)、KTIG (SEQ ID NO:5797), KTDP (SEQ ID NO: 5798), KELV (SEQ ID NO: 5799), KELM (SEQ ID NO: 5800), KNEI (SEQ ID NO: 5801), KTPN (SEQ ID NO: 5802), KITN (SEQ ID NO: 5803), KTDI (SEQ ID NO: 5804), KTDQ (SEQ ID NO: 5805), KGIN (SEQ ID NO: 5806), KSEI (SEQ ID NO: 5807), KSEK (SEQ ID NO: 5627), KWSA (SEQ ID NO: 5808), KELA (SEQ ID NO: 5809), KQTQ (SEQ ID NO: 5804) 5810), KGAD (SEQ ID NO: 5811), KVGE (SEQ ID NO: 5812), KANE (SEQ ID NO: 5813), KTDT (SEQ ID NO: 5814), KTCI (SEQ ID NO: 5815), KELR (SEQ ID NO: 5816), KCQI (SEQ ID NO: 5817), KGVM (SEQ ID NO: 5818), KACD (SEQ ID NO: 5819), KNEL (SEQ ID NO: 5820), KAAE (SEQ ID NO: 5821), KGQN (SEQ ID NO: 5822), KNEF (SEQ ID NO: 5823), KTSI (SEQ ID NO: 5824), KAEH (SEQ ID NO: 5616), KCDQ (SEQ ID NO: 5825), KEIL (SEQ ID NO: 5826), KTER (SEQ ID NO: 5573), KNAI (SEQ ID NO: 5650), KTDK (SEQ ID NO: 5588), KTPD (ii) any part of the amino acid sequence in (i), such as any two or three amino acids therein, such as an amino acid sequence of consecutive amino acids; (iii) a sequence of one, two, or three but no more than four modified amino acid sequences relative to any one of the amino acid sequences in (i); or specification 40 / 285 pages 50 CN 121079310 A(iv) An amino acid sequence comprising one, two, or three, but not more than four different amino acids, relative to any one of the amino acid sequences in (i).

[0314] 236. The AAV particle as described in embodiment 234 or 235, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or comprises: (i) an amino acid sequence of any one of SEQ ID NO: 3606-3836; (ii) an amino acid sequence comprising any part of the amino acid sequence in (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, or 17 amino acids, such as a sequence of consecutive amino acids; (iii) an amino acid sequence comprising one, two, or three, but not more than four modified amino acid sequences, relative to any one of the amino acid sequences in (i); or (iv) an amino acid sequence comprising one, two, or three, but not more than four different amino acids, relative to any one of the amino acid sequences in (i).

[0315] 237. An AAV particle as described in any one of embodiments 234 to 236, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains KTINGHDSPHKSGQNQQT (SEQ ID NO: 5828).

[0316] 238. An AAV particle as described in any one of embodiments 234 to 236, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains KAEIGHDSPHKSGQNQQT (SEQ ID NO: 1754).

[0317] 239. An AAV particle as described in any one of embodiments 234 to 236, wherein [N0]-[N1]-[N2]-[N3]-[N4] is or contains KTEKMSGSPHSKAQNQQT (SEQ ID NO: 3241).

[0318] 240. An AAV particle as described in any one of embodiments 168 to 239, wherein [N1]-[N2]-[N3] are present, for example, in a ring IV numbered according to SEQ ID NO: 4, 36, 138, 981 or 982.

[0319] 241. An AAV particle as described in any one of embodiments 198 to 202 or 234 to 240, wherein [NO] and [N4] are present, for example, in a ring IV numbered according to SEQ ID NO: 4, 36, 138, 981 or 982.

[0320] 242. An AAV particle as described in any one of embodiments 198 to 202 or 234 to 241, wherein [NO] is present immediately after position 448 of the amino acid sequence numbered according to SEQ ID NO: 4, 36, 138, 981 or 982.

[0321] 243.AAV particles as in any of embodiments 198 to 202 or 234 to 242, wherein [NO] replaces positions 449-452 (e.g., K449, T450, I451, and N452) numbered according to SEQ ID NO: 4, 36, 138, 981, or 982.

[0322] 244. AAV particles as in any of embodiments 198 to 202 or 234 to 243, wherein [NO] is present immediately after position 448, and wherein [NO] replaces positions 449-452 (e.g., K449, T450, I451, and N452) numbered according to SEQ ID NO: 4, 36, 138, 981, or 982.

[0323] 245. An AAV particle as described in any one of embodiments 198 to 202 or 234 to 244, wherein [NO] corresponds to positions 449-452 of any one of SEQ ID NO: 4, 36, 138, 981 or 982 (e.g., K449, T450, I451 and N452).

[0324] 246. An AAV particle as described in any one of embodiments 168 to 245, wherein [N1] is present immediately after position 452 numbered according to SEQ ID NO: 4, 36, 138, 981 or 982.

[0325] 247. An AAV particle as described in any one of embodiments 168 to 246, wherein [N1] replaces positions 453-455 (e.g., G453, S454, and G455) numbered according to SEQ ID NO: 4, 36, 138, 981, or 982.

[0326] 248. An AAV particle as described in any one of embodiments 168 to 246, wherein [N1] replaces position 453 (e.g., G453) numbered according to SEQ ID NO: 4, 36, 138, 981, or 982.

[0327] 249. An AAV particle as described in any one of embodiments 168 to 177, 179 to 181, 183 to 185, 187 to 193, 195 to 200, 202 or 240 to 246, wherein: (i) X1 of [N1] replaces position 453 (e.g., G453); (ii) X2 of [N1] corresponds to position 454 (e.g., S454); and (iii) X3 of [N1] corresponds to position 455 (e.g., G455), wherein (i), (ii) and (iii) are numbered according to SEQ ID NO: 138 or SEQ ID NO: 981.

[0328] 250. An AAV particle as described in any one of embodiments 168 to 176, 178 to 201, or 240 to 246, wherein: (i) X1 of [N1] corresponds to position 453(i) X1 of [N1] corresponds to position 454 (e.g., S454); and (iii) X3 of [N1] corresponds to position 455 (e.g., G455); wherein (i), (ii) and (iii) are numbered according to SEQ ID NO: 138 or SEQ ID NO: 981.

[0329] 251. An AAV particle as described in any one of embodiments 203 to 248, wherein: (i) X1 of [N1] corresponds to position 453 (e.g., G453); (ii) X2 of [N1] replaces position 454 (e.g., S454); and (iii) X3 of [N1] replaces position 455 (e.g., G455), wherein (i), (ii) and (iii) are numbered according to SEQ ID NO: 138 or SEQ ID NO: 982.

[0330] 252. An AAV particle as described in any one of embodiments 203 to 248 or 251, wherein [N1] corresponds to positions 453-455 of SEQ ID NO: 982 (e.g., G453, H454, D455).

[0331] 253. An AAV particle as described in any one of embodiments 168 to 176, 178 to 201, 240 to 247 or 250, wherein [N1] corresponds to positions 453-455 of SEQ ID NO: 138 or 981 (e.g., G453, S454, G455).

[0332] 254. An AAV particle as described in any one of embodiments 168 to 253, wherein [N2] is present immediately after position 455 numbered according to SEQ ID NO: 4, 36, 138, 981 or 982.

[0333] 255. An AAV particle as described in any one of embodiments 168 to 254, wherein [N2] corresponds to positions 456-458 of SEQ ID NO: 981 or 982 (e.g., S456, P457, and H458).

[0334] 256. An AAV particle as described in any one of embodiments 168 to 254, wherein [N2] corresponds to positions 456-458 of any one of SEQ ID NO: 4 or 36-59 (e.g., S456, P457, and H458).

[0335] 257. An AAV particle as described in any one of embodiments 168 to 256, wherein [N2] is present immediately after [N1].

[0336] 258. AAV particles as described in any one of embodiments 168 to 202, 240 to 247, or 249 to 257, wherein [N3] corresponds to positions 459-460 of SEQ ID NO: 981 (e.g., S459, K460, A461).

[0337] 259.AAV particles of any one of embodiments 168 to 202, 240 to 247, or 249 to 257, wherein [N3] corresponds to positions 459-460 of SEQ ID NO: 36, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 57, or 59 (e.g., S459, K460, A461).

[0338] 260. AAV particles of any one of embodiments 168 to 259, wherein [N2]-[N3] are present immediately after position 455 according to any one of the numbers SEQ ID NO: 4, 36, 138, 981, or 982.

[0339] 261. An AAV particle as described in any one of embodiments 168 to 259, wherein [N2]-[N3] are present immediately after position 455 according to SEQ ID NO: 981.

[0340] 262. An AAV particle as described in any one of embodiments 168 to 261, wherein [N2]-[N3] correspond to positions 456-461 of SEQ ID NO: 981 (e.g., S456, P457, H458, S459, K460, A461).

[0341] 263. AAV particles as described in any of embodiments 168 to 262, wherein [N2]-[N3] correspond to positions 456-461 (e.g., S456, P457, H458, S459, K460, A461) of any of SEQ ID NO: 36, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 57 or 59.

[0342] 264. An AAV particle as described in any one of embodiments 203 to 257 or 259 to 261, wherein [N3] corresponds to positions 459-460 of SEQ ID NO: 982 on page 52 of CN 121079310 (e.g., K459, S460, G461).

[0343] 265. An AAV particle as described in any one of embodiments 203 to 257 or 259 to 261, wherein [N3] corresponds to positions 459-460 of SEQ ID NO: 37 (e.g., K459, S460, G461).

[0344] 266. An AAV particle as described in any one of embodiments 203 to 265, wherein [N2]-[N3] are present immediately after position 455 according to the numbering in SEQ ID NO: 982.

[0345] 267. AAV particles as described in any one of embodiments 203 to 246, 248, 252, 255, 257, 260, and 263 to 266.Particles, wherein [N3] replaces positions 454 and 455 (e.g., S454 and G455) numbered according to SEQ ID NO: 138.

[0346] 268. AAV particles as described in any one of embodiments 203 to 246, 248, 252, 255, 257, 260, 263 to 267, wherein [N3] is present immediately after [N2] and replaces positions 454 and 455 (e.g., S454 and G455), which are numbered according to SEQ ID NO: 138.

[0347] 269. An AAV particle as described in any one of embodiments 203 to 246, 248, 252, 255, 257, 260, 263 to 268, wherein [N3] is present immediately following [N1]-[N2] and replaces positions 454 and 455 (e.g., S454 and G455), these positions being numbered according to SEQ ID NO: 138.

[0348] 270. An AAV particle as described in any one of embodiments 203 to 257, 260, 264 to 269, wherein [N2]-[N3] corresponds to positions 456-461 of SEQ ID NO: 982 (e.g., S456, P457, H458, K459, S460, G461).

[0349] 271. An AAV particle as described in any one of embodiments 203 to 257, 260, 264 to 270, wherein [N2]-[N3] correspond to positions 456-461 of SEQ ID NO: 37 (e.g., S456, P457, H458, K459, S460, G461).

[0350] 272. An AAV particle as described in any one of embodiments 191 to 202 or 228 to 271, wherein [N4] is present immediately after position 455 according to the numbering in SEQ ID NO: 138.

[0351] 273. AAV particles as described in any one of embodiments 191 to 202 or 228 to 272, wherein [N4] replaces positions 456-460 (e.g., Q456, N457, Q458, Q459, and T460) according to SEQ ID NO: 138.

[0352] 274. AAV particles as described in any one of embodiments 191 to 202 or 228 to 273, wherein [N4] corresponds to positions 462-466 ​​(e.g., Q462, N463, Q464, Q465, and T466) of SEQ ID NO: 981 or 982.

[0353] 275. AAV particles as described in any one of embodiments 191 to 202 or 228 to 273, wherein [N4] corresponds to positions 462-466 ​​of any one of SEQ ID NO: 4 or 36-59.

[0354] 276277. AAV particles as described in any one of embodiments 191 to 202 or 228 to 274, wherein [N4] corresponds to positions 456-460 of SEQ ID NO: 138 (e.g., Q456, N457, Q458, Q459, and T460).

[0355] 277. AAV particles as described in any one of embodiments 191 to 202 or 228 to 276, wherein [N2]-[N3]-[N4] replace positions 456-460 (e.g., Q456, N457, Q458, Q459, and T460) numbered according to SEQ ID NO: 138.

[0356] 278. An AAV particle as described in any of embodiments 191 to 202 or 228 to 277, wherein [N2]-[N3]-[N4] is present immediately after position 455, and wherein [N2]-[N3]-[N4] replaces positions 456-460 (e.g., Q456, N457, Q458, Q459 and T460), which are numbered according to SEQ ID NO: 138.

[0357] 279. AAV particles as described in any one of embodiments 191 to 202 or 228 to 278, wherein [N1]-[N2]-[N3]-[N4] replace positions 453-460 (e.g., G453, S454, G455, Q456, N457, Q458, Q459 and T460) according to SEQ ID NO: 138.

[0358] 280. AAV particles as described in any of embodiments 191 to 202 or 228 to 279, wherein [N1]-[N2]-[N3]-[N4] are present immediately after position 452, and wherein [N1]-[N2]-[N3]-[N4] replace positions 453-460 (e.g., G453, S454, G455, Q456, N457, Q458, Q459 and T460), which are numbered according to SEQ ID NO: 138.

[0359] 281. An AAV particle as described on page 43 / 285 of the specification of any of embodiments 191 to 202, 240 to 247, 249, 250, 253 to 263, 266 or 272 to 280, wherein [N1]-[N2]-[N3]-[N4] corresponds to positions 453-466 of SEQ ID NO: 981 (e.g., G453, S454, G455, S456, P457, H458, S459, K460, A461, Q462, N463, Q464, Q465 and T466).

[0360] 282. Any of embodiments 168 to 202, 240 to 247, 249, 250, 253 to 263, 266 or 272 to 280AAV particles, wherein [N1]-[N2]-[N3] correspond to positions 453-461 of SEQ ID NO: 981 (e.g., G453, S454, G455, S456, P457, H458, S459, K460, A461).

[0361] 283. AAV particles as described in any one of embodiments 228 to 257, 260, 261, 264 to 282, wherein [N1]-[N2]-[N3]-[N4] correspond to positions 453-466 of SEQ ID NO: 982 (e.g., G453, H454, D455, S456, P457, H458, K459, S460, G461, Q462, N463, Q464, Q465, T466).

[0362] 284. AAV particles as described in any one of embodiments 203 to 257, 260, 261, 264 to 283, wherein [N1]-[N2]-[N3] correspond to positions 453-461 of SEQ ID NO: 982 (e.g., G453, H454, D455, S456, P457, H458, K459, S460, G461).

[0363] 285. AAV particles as described in any one of embodiments 228 to 257, 260, 261, 264 to 282, wherein [N1]-[N2]-[N3]-[N4] correspond to positions 453-466 of any one of SEQ ID NO: 4 or 36-59.

[0364] 286. AAV particles as described in any one of embodiments 198 to 202 or 234 to 286, wherein [N0]-[N1]-[N2]-[N3]-[N4] replace positions 449-460 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459 and T460) according to SEQ ID NO: 138.

[0365] 287. AAV particles as described in any one of embodiments 198 to 202 or 234 to 286, wherein [N0]-[N1]-[N2]-[N3]-[N4] are present immediately after position 448, and wherein [N0]-[N1]-[N2]-[N3]-[N4] replace positions 449-460 numbered according to SEQ ID NO: 138 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, and T460).

[0366] 288. AAV particles as described in embodiments 198 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, orAAV particles of any one of 287, wherein [N0]-[N1]-[N2]-[N3]-[N4] correspond to positions 449-466 of SEQ ID NO: 981 (e.g., K449, T450, I451, N452, G453, S454, G455, S456, P457, H458, S459, K460, A461, Q462, N463, Q464, Q465, T466).

[0367] 289. AAV particles as described in any one of embodiments 234 to 257, 260, 261, 264 to 284, 286 or 287, wherein [N0]-[N1]-[N2]-[N3]-[N4] corresponds to positions 449-466 of SEQ ID NO: 982 (e.g., K449, T450, I451, N452, G453, H454, D455, S456, P457, H458, K459, S460, G461, Q462, N463, Q464, Q465, T466).

[0368] 290. An AAV particle as described in any one of embodiments 234 to 257, 260, 261, 264 to 284, 286, or 287, wherein [N0]-[N1]-[N2]-[N3]-[N4] correspond to positions 449-466 of any one of SEQ ID NO: 4 or 36-59.

[0369] 291. An AAV particle as described in any one of embodiments 191 to 202 or 228 to 290, wherein [N4] is present immediately after position 461 numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0370] 292. The AAV particle as described in any one of embodiments 191 to 202 or 228 to 291, wherein [N4] replaces positions 462-466 ​​(e.g., Q462, N463, Q464, Q465, and T466) numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0371] 293. The AAV particle as described in any one of embodiments 191 to 202 or 228 to 292, wherein [N2]-[N3]-[N4] replace positions 462-466 ​​(e.g., Q462, N463, Q464, Q465, and T466) numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0372] 294. An AAV particle as described in any of embodiments 191 to 202 or 228 to 293, wherein [N2]-[N3]-[N4] is present immediately after position 455, and wherein [N2]-[N3]-[N4] replaces positions 462-466 ​​(e.g., Q462, N463, ... Specification 44 / 285 pages 54 CN 121079310 AQ464, Q465, and T466), these positions are numbered according to SEQ ID NO: 4, 36, 981, or 982.

[0373] 295. An AAV particle as described in any one of embodiments 168 to 294, wherein the AAV capsid variant comprises [N2]-[N3] from the N-terminus to the C-terminus.

[0374] 296. An AAV particle as described in any one of embodiments 168 to 295, wherein the AAV capsid variant comprises [N1]-[N2]-[N3] from the N-terminus to the C-terminus.

[0375] 297. An AAV particle as described in any one of embodiments 168 to 296, wherein the AAV capsid variant comprises [NO]-[N1]-[N2]-[N3] from the N-terminus to the C-terminus.

[0376] 298. An AAV particle as described in any one of embodiments 168 to 297, wherein the AAV capsid variant comprises [N1]-[N2]-[N3]-[N4] from the N-terminus to the C-terminus.

[0377] 299. An AAV particle as described in any one of embodiments 168 to 298, wherein the AAV capsid variant comprises [N0]-[N1]-[N2]-[N3]-[N4] from the N-terminus to the C-terminus.

[0378] 300. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding a human CDKL5 protein), wherein the AAV capsid variant comprises formula [A]-[B] (SEQ ID NO: 4696), wherein: (i) [A] comprises GSGSPH (SEQ ID NO: 4695); and (ii) [B] comprises X1, X2, X3, X4, and X5, wherein: (a) X1 is S, I, F, V, C, Y, W, R, P, L, Q, M, K, or G; (b) X2 is K, M, R, F, V, C, P, Y, L, W, G, N, S, T, I, or A; (c) X3 is A, Y, L, R, W, C, T, F, H, I, P, M, K, S, V, G, Q, or N; (d) X4 is Q, M, F, K, H, R, C, W, P, V, L, G, S, Y, I, A, T, D, N, or E; and (e) X5 is A, N, Y, R, K, L, I, M, Q, S, C, W, F, T, G, V, or P; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(e), such as a conservative substitution.

[0379] 301. AAV particles as described in embodiment 300, wherein: (a) X1 is S, L, R, V or P; (b) X2 is K, C, F, L, P, R, S or V; (c) X3 is A, C, F, I, K, L, M, P, R, T, W or Y;(d) X4 is Q, R, S, T, C, F, K, L, P or Y; and (e) X5 is N, R, S, T, K, M, Q or Y; Optionally, the AAV capsid variant comprises an amino acid modification of any of the aforementioned amino acids in (a)-(e), for example, a conservative substitution.

[0380] 302. AAV particles as described in embodiment 300 or 301, wherein [B] comprises SKA, SMY, SKL, SKR, SKW, SRC, SFT, SKF, IVW, SKY, SCH, FPW, SKI, VYY, SLY, SKP, SRF, SRM, SVK, SWA, SLW, SFR, SKK, SYA, SCS, SGA, SFP, SFF, SMC, SKT, SGK, FYR, CRV, YGI, VNC, SLA, WSY, RWL, PSC, SSW, SKG, VPW, SGC, STT, PKR, SKC, WVP, SFW, RIK, SKM, LRW, LPT, SYM, LLC, RCC, LCV, SYL, QGC, MAF, SFQ, SLC, RPW, RPR, SCP, SVR, SLP, VYH, SYT, LVY, YRY, SWL, CPA, SPP, RWT, PRK, PFV, SKS, WVA, SKV, CAL, SSC, SKN, LCT, STC, SKQ, KSG, SYY, SLT, SCQ, FPF, SVF, GRY, AQA, AQN, YMN, AFY, LKR, RHR, AQK, WRL, CRN, TCN, FFI, AQY, WQN, YFM, ARQ, HQN, IRR, YQN, YWN, AFS, FWN, AQC, MRN, KKN, APN, WKN, ARW, RPN, KVF, AFN, ACS, RLW, SRN, CPN, ACN, FRQ, PFN, FGN, CQN, LFW, TRK, KRN, RQN, VQN, IQN, AQR, PFR, Instructions 45 / 285 Page 55 CN 121079310 A AWN, RSY, LQN, WLN, RRA, AQT, GCT, RYT, TPN, ARM, CFL, PQN, WSN, FKN, KQN, APR, RYN, MIC, TQN, WKS, AAR, LTR, IRG, LVN, FQN, ACQ, WGL, ILR, QIN, ACI, ALR, AHA, CLN, AFV, AQF, RCN, MPC, KTS, PYN, AQS, TRN, LKN, AQM, CTN, PDN, RNY, ACR, CSV, ARI, LPK, SEQ, VRM, NSR, RKR, ARN, QRP, RVV,GQN, YSN, QSN, AKG, CTS, FEN, AKK, KAQ, MYM, KAF, KLK, KRH, KWR, RCR, FTC, KFF, VWQ, KYF, KAR, CHQ, PWQ, KIR, YYQ, LYW, KPK, RFW, RMR, VKK, WAP, LWK, FRP, KKV, YAF, KAC, KRL, CSR, RCP, GAC, KFR, FFG, MCQ, KLF, KTR, GKR, YRQ, RVQ, GIQ, NCQ, KPF, LAW, KRS, SYQ, WLQ, KRR, KGC, KRY, GCQ, FTP, TTC, KRQ, KCF, VPQ, FWS, KFK, IKQ, KAP, FRY, KMI, RWQ, PTQ, KWK, YMR, KAA, LCQ, CCQ, CVQ, KLT, KLC, YLV, AFQ, KWG, KIL, FQI, KAL, KAH, LCL, PRQ, CPQ, VRY, VRC, KMP, KKT, LPY, YHQ, YTR, VYQ, RYQ, WLK, PAQ, MCT, PPD, WTQ, RKQ, KCS, FVQ, KLP, KSE, VAQ, LYQ, KVR, ALQ, SCT, KNS, KRK, CTQ, TCL, YAR, KQR, KRV, SGQ, YYS, LTC, CQS, KAK, KPQ, PFQ, KCT or VFE.

[0381] 303. An AAV particle as in any one of embodiments 300 to 302, wherein [B] comprises SKAQ (SEQ ID NO: 5829), SMYM (SEQ ID NO: 5830), SKAF (SEQ ID NO: 5831), SKLK (SEQ ID NO: 5832), SKRH (SEQ ID NO: 5833), SKWR (SEQ ID NO: 5834), SRCR (SEQ ID NO: 5835), SFTC (SEQ ID NO: 5836), SKFF (SEQ ID NO: 5837), IVWQ (SEQ ID NO: 5838), SKYF (SEQ ID NO: 5839), SKAR (SEQ ID NO: 5840), SCHQ (SEQ ID NO: 5841), FPWQ (SEQ ID NO: 5842), SKIR (SEQ ID NO: 5843), VYYQ (SEQ ID NO: 5844), SLYW (SEQ ID NO: 5845), SKPK (SEQ ID NO: 5846), SRFW (SEQ ID NO:5847)、SRMR (SEQ ID NO: 5848)、SVKK (SEQ ID NO: 5849)、SWAP (SEQ ID NO: 5850)、SLWK (SEQ ID NO: 5851)、SFRP (SEQ ID NO: 5852)、SKKV (SEQ ID NO: 5853)、SYAF (SEQ ID NO: 5854)、SKAC (SEQ ID NO: 5855)、SKRL (SEQ ID NO: 5856)、SCSR (SEQ ID NO: 5857)、SRCP (SEQ ID NO: 5858)、 SGAC (SEQ ID NO: 5859)、SKFR (SEQ ID NO: 5860)、SFPF (SEQ ID NO: 5861)、SFFG (SEQ ID NO: 5862)、SMCQ (SEQ ID NO: 5863)、SKLF (SEQ ID NO: 5864)、SKTR (SEQ ID NO: 5865)、SGKR (SEQ ID NO: 5866)、FYRQ (SEQ ID NO: 5867)、CRVQ (SEQ ID NO: 5868)、YGIQ (SEQ ID NO: 5869)、VNCQ (SEQ ID NO: 5870)、SKPF (SEQ ID NO: 5871)、 SLAW (SEQ ID NO: 5872)、SKRS (SEQ ID NO: 5873)、WSYQ (SEQ ID NO: 5874)、RWLQ (SEQ ID NO: 5875)、PSCQ (SEQ ID NO: 5876)、SSWL (SEQ ID NO: 5877)、SKRR (SEQ ID NO: 5878)、SKGC (SEQ ID NO: 5879)、VPWQ (SEQ ID NO: 5880)、SKRY (SEQ ID NO: 5881)、SGCQ (SEQ ID NO: 5882)、SFTP (SEQ ID NO: 5883)、STTC (SEQ ID NO: 5884)、 PKRQ (SEQ ID NO: 5885)、SKCF (SEQ ID NO: 5886)、WVPQ (SEQ ID NO: 5887)、SFWS (SEQ ID NO: 5888)、SKFK (SEQ ID NO: 5889)、RIKQ (SEQ ID NO:5890), SKAP (SEQ ID NO: 5891), SFRY (SEQ ID NO: 5892), SKMI (SEQ ID NO: 5893), LRWQ (SEQ ID NO: 5894), LPTQ (SEQ ID NO: 5895), SKWK (SEQ ID NO: 5896), SYMR (SEQ ID NO: 5897), SKAA (SEQ ID NO: 5898), LLCQ (SEQ ID NO: 5899), RCCQ (SEQ ID NO: 5900), LCVQ (SEQ ID NO: 5901), SKLT (SEQ ID NO: 5902), SKLC (SEQ ID NO: 5903), SYLV (SEQ ID NO: 5904), QGCQ (SEQ ID NO: 5905), MAFQ (SEQ ID NO: 5906), SKWG (SEQ ID NO: 5907), SKIL (SEQ ID NO: 5908), SFQI (SEQ ID NO: 5909), SKAL (SEQ ID NO: 5910), SKAH (SEQ ID NO: 5911), SLCL (SEQ ID NO: 5912), RPWQ (SEQ ID NO: 5913), RPRQ (SEQ ID NO: 5914), SCPQ (SEQ ID NO: 5915), SVRY (SEQ ID NO: 5916), SVRC (SEQ ID NO: 5917), SKMP (SEQ ID NO: 5918), SKKT (SEQ ID NO: 5919), SLPY (SEQ ID NO: 46 / 285 pages of the specification 56 CN 121079310 A 5920), VYHQ (SEQ ID NO: 5921), SYTR (SEQ ID NO: 5922), LVYQ (SEQ ID NO: 5923), YRYQ (SEQ ID NO: 5924), SWLK (SEQ ID NO: 5925), CPAQ (SEQ ID NO: 5926), SMCT (SEQ ID NO: ...... It should be noted that the original text seems to be incomplete as there are some parts like "SLPY (SEQ ID NO: 说 明 书 46 / 285 页 56 CN 121079310 A 5920)" which are not in a proper format. This translation is based on the best understanding of the provided text.NO: 5932)、PFVQ (SEQ ID NO: 5933)、SKLP (SEQ ID NO: 5934)、SKSE (SEQ ID NO: 5935)、WVAQ (SEQ ID NO: 5936)、 SLYQ (SEQ ID NO: 5937)、SEQ ID NO: 5937) 5938)、CALQ (SEQ ID NO: 5939)、SSCT (SEQ ID NO: 5940)、SKNS (SEQ ID NO: 5941)、SKRK (SEQ ID NO: 5942)、LCTQ (SEQ ID NO: 5943)、STCCL (SEQ ID NO: 5943) (SEQ ID NO: 5945)、SKQR (SEQ ID NO: 5946)、SKRV (SEQ ID NO: 5947)、KSGQ (SEQ ID NO: 5948)、SYYS (SEQ ID NO: 5949)、 SLTC (SEQ ID NO: 5950) 5951)、SKAK (SEQ ID NO: 5952)、SKPQ (SEQ ID NO: 5953)、FPFQ (SEQ ID NO: 5954)、SKCT (SEQ ID NO: 5955)、SVFE (SEQ ID NO: 5956)、SEQ ID NO: 5956) (SEQ ID NO: 5958)、KAQN (SEQ ID NO: 5959)、MYMN (SEQ ID NO: 5960)、KAFY (SEQ ID NO: 5961)、KLKR (SEQ ID NO: 5962)、 KRHR (SEQ ID NO: 593) 5964)、KWRL (SEQ ID NO: 5965)、RCRN (SEQ ID NO: 5966)、FTCN (SEQ ID NO: 5967)、KFFI (SEQ ID NO: 5968)、KAQY (SEQ ID NO: 5969)、VW ID NO: 5970)、KYFM (SEQ ID NO: 5971)、KARQ (SEQ ID NO: 5972)、CHQN (SEQ ID NO: 5973)、PWQN (SEQ ID NO: 5974)、KIRR (SEQ ID NO:5975)、 YYQN (SEQ ID NO: 5976)、LYWN (SEQ ID NO: 5977)、KPKR (SEQ ID NO: 5978)、KAFS (SEQ ID NO: 5979)、RFWN (SEQ ID NO: 5980)、KAQC (SEQ ID NO: 5981)、RMRN (SEQ ID NO: 5982)、VKKN (SEQ ID NO: 5983)、WAPN (SEQ ID NO: 5984)、LWKN (SEQ ID NO: 5985)、KARW (SEQ ID NO: 5986)、FRPN (SEQ ID NO: 5987)、KKVF (SEQ ID NO: 5988)、 YAFN (SEQ ID NO: 5989)、KACS (SEQ ID NO: 5990)、KRLW (SEQ ID NO: 5991)、CSRN (SEQ ID NO: 5992)、RCPN (SEQ ID NO: 5993)、GACN (SEQ ID NO: 5994)、KFRQ (SEQ ID NO: 5995)、FPFN (SEQ ID NO: 5996)、FFGN (SEQ ID NO: 5997)、MCQN (SEQ ID NO: 5998)、KLFW (SEQ ID NO: 5999)、KTRK (SEQ ID NO: 6000)、GKRN (SEQ ID NO: 6001)、 YRQN (SEQ ID NO: 6002)、RVQN (SEQ ID NO: 6003)、GIQN (SEQ ID NO: 6004)、KAQR (SEQ ID NO: 6005)、NCQN (SEQ ID NO: 6006)、KPFR (SEQ ID NO: 6007)、LAWN (SEQ ID NO: 6008)、KRSY (SEQ ID NO: 6009)、SYQN (SEQ ID NO: 6010)、WLQN (SEQ ID NO: 6011)、SCQN (SEQ ID NO: 6012)、SWLN (SEQ ID NO: 6013)、KRRA (SEQ ID NO: 6014)、 KAQT (SEQ ID NO: 6015)、KGCT (SEQ ID NO: 6016)、KRYT (SEQ ID NO: 6017)、GCQN (SEQ ID NO:6018), FTPN (SEQ ID NO: 6019), TTCN (SEQ ID NO: 6020), KARM (SEQ ID NO: 6021), KRQN (SEQ ID NO: 6022), KCFL (SEQ ID NO: 6023), VPQN (SEQ ID NO: 6024), FWSN (SEQ ID NO: 6025), KFKN (SEQ ID NO: 6026), IKQN (SEQ ID NO: 6027), KAPR (SEQ ID NO: 6028), FRYN (SEQ ID NO: 6029), KMIC (SEQ ID NO: 6030), RWQN (SEQ ID NO: 6031), PTQN (SEQ ID NO: 6032), KWKS (SEQ ID NO: 6033), YMRN (SEQ ID NO: 6034), KAAR (SEQ ID NO: 6035), LCQN (SEQ ID NO: 6036), CCQN (SEQ ID NO: 6037), CVQN (SEQ ID NO: 6038), KLTR (SEQ ID NO: 6039), KLCT (SEQ ID NO: 6040), KIRG (SEQ ID NO: 6041), YLVN (SEQ ID NO: 6042), AFQN (SEQ ID NO: 6043), KACQ (SEQ ID NO: 6044), KWGL (SEQ ID NO: 6045), KILR (SEQ ID NO: 6046), FQIN (SEQ ID DESCRIPTION 47 / 285 pages 57 CN 121079310 A NO: 6047), KACI (SEQ ID NO: 6048), KALR (SEQ ID NO: 6049), KAHA (SEQ ID NO: 6050), LCLN (SEQ ID NO: 6051), KAFV (SEQ ID NO: 6052), PRQN (SEQ ID NO: 6053), CPQN (SEQ ID NO: 6054), KAQF (SEQ ID NO: 6055), VRYN (SEQ ID NO: 6056), VRCN (SEQ ID NO: 6057), KMPC (SEQ ID NO: 6058), KKTS (SEQ ID NO: 6059), LPYN (SEQ IDNO: 6060), YHQN (SEQ ID NO: 6061), KAQS (SEQ ID NO: 6062), YTRN (SEQ ID NO: 6063), VYQN (SEQ ID NO: 6064), RYQN (SEQ ID NO: 6065), WLKN (SEQ ID NO: 6066), KAQM (SEQ ID NO: 6067), PAQN (SEQ ID NO: 6068), MCTN (SEQ ID NO: 6069), PPDN (SEQ ID NO: 6070), KRNY (SEQ ID NO: 6071), WTQN (SEQ ID NO: 6072), KACR (SEQ ID NO: 6073), RKQN (SEQ ID NO: 6074), KCSV (SEQ ID NO: 6075), KARI (SEQ ID NO: 6076), FVQN (SEQ ID NO: 6077), KLPK (SEQ ID NO: 6078), KSEQ (SEQ ID NO: 6079), VAQN (SEQ ID NO: 6080), LYQN (SEQ ID NO: 6081), KVRM (SEQ ID NO: 6082), ALQN (SEQ ID NO: 6083), SCTN (SEQ ID NO: 6084), KNSR (SEQ ID NO: 6085), KRKR (SEQ ID NO: 6086), CTQN (SEQ ID NO: 6087), TCLN (SEQ ID NO: 6088), YARN (SEQ ID NO: 6089), KQRP (SEQ ID NO: 6090), KRVV (SEQ ID NO: 6091), SGQN (SEQ ID NO: 6092), YYSN (SEQ ID NO: 6093), LTCN (SEQ ID NO: 6094), CQSN (SEQ ID NO: 6095), KAKG (SEQ ID NO: 6096), KPQN (SEQ ID NO: 6097), PFQN (SEQ ID NO: 6098), KCTS (SEQ ID NO: 6099), VFEN (SEQ ID NO: 6100) or KAKK (SEQ ID NO: 6101).

[0382] 304.An AAV particle as in any one of embodiments 300 to 303, wherein [B] is or comprises: (i) SKAQA (SEQ ID NO: 6102), SKAQN (SEQ ID NO: 6103), SMYMN (SEQ ID NO: 6104), SKAFY (SEQ ID NO: 6105), SKLKR (SEQ ID NO: 6106), SKRHR (SEQ ID NO: 6107), SKAQK (SEQ ID NO: 6108), SKWRL (SEQ ID NO: 6109), SRCRN (SEQ ID NO: 6110), SFTCN (SEQ ID NO: 6111), SKFFI (SEQ ID NO: 6112), SKAQY (SEQ ID NO: 6113), IVWQN (SEQ ID NO: 6114), SKYFM (SEQ ID NO: 6115), SKARQ (SEQ ID NO: 6116), SCHQN (SEQ ID NO: 6117), FPWQN (SEQ ID NO: 6118), SKIRR (SEQ ID NO: 6119), VYYQN (SEQ ID NO: 6120), SLYWN (SEQ ID NO: 6121), SKPKR (SEQ ID NO: 6122), SKAFS (SEQ ID NO: 6123), SRFWN (SEQ ID NO: 6124), SKAQC (SEQ ID NO: 6125), SRMRN (SEQ ID NO: 6126), SVKKN (SEQ ID NO: 6127), SWAPN (SEQ ID NO: 6128), SLWKN (SEQ ID NO: 6129), SKARW (SEQ ID NO: 6130), SFRPN (SEQ ID NO: 6131), SKKVF (SEQ ID NO: 6132), SYAFN (SEQ ID NO: 6133), SKACS (SEQ ID NO: 6134), SKRLW (SEQ ID NO: 6135), SCSRN (SEQ ID NO: 6136), SRCPN (SEQ ID NO: 6137), SGACN (SEQ ID NO: 6138), SKFRQ (SEQ ID NO: 6139), SFPFN (SEQ ID NO: 6140), SFFGN (SEQ ID NO:6141), SMCQN (SEQ ID NO: 6142), SKLFW (SEQ ID NO: 6143), SKTRK (SEQ ID NO: 6144), SGKRN (SEQ ID NO: 6145), FYRQN (SEQ ID NO: 6146), CRVQN (SEQ ID NO: 6147), YGIQN (SEQ ID NO: 6148), SKAQR (SEQ ID NO: 6149), VNCQN (SEQ ID NO: 6150), SKPFR (SEQ ID NO: 6151), SLAWN (SEQ ID NO: 6152), SKRSY (SEQ ID NO: 6153), WSYQN (SEQ ID NO: 6154), RWLQN (SEQ ID NO: 6155), PSCQN (SEQ ID NO: 6156), SSWLN (SEQ ID NO: 6157), SKRRA (SEQ ID NO: 6158), SKAQT (SEQ ID NO: 6159), SKGCT (SEQ ID NO: 6160), VPWQN (SEQ ID NO: 6161), SKRYT (SEQ ID NO: 6162), SGCQN (SEQ ID NO: 6163), SFTPN (SEQ ID NO: Description 48 / 285 pages 58 CN 121079310 A 6164), STTCN (SEQ ID NO: 6165), SKARM (SEQ ID NO: 6166), PKRQN (SEQ ID NO: 6167), SKCFL (SEQ ID NO: 6168), WVPQN (SEQ ID NO: 6169), SFWSN (SEQ ID NO: 6170), SKFKN (SEQ ID NO: 6171), RIKQN (SEQ ID NO: 6172), SKAPR (SEQ ID NO: 6173), SFRYN (SEQ ID NO: 6174), SKMIC (SEQ ID NO: 6175), LRWQN (SEQ ID NO: 6176), LPTQN (SEQ ID NO: 6177), SKWKS (SEQ ID NO: 6178), SYMRN (SEQ ID NO: 6179), SKAAR (SEQ ID NO: 6180), LLCQN (SEQ ID NO:6181)、RCCQN (SEQ ID NO: 6182)、LCVQN (SEQ ID NO: 6183)、SKLTR (SEQ ID NO: 6184)、SKLCT (SEQ ID NO: 6185)、SKIRG (SEQ ID NO: 6186)、SYLVN (SEQ ID NO: 6187)、QGCQN (SEQ ID NO: 6188)、MAFQN (SEQ ID NO: 6189)、SKACQ (SEQ ID NO: 6190)、SKWGL (SEQ ID NO: 6191)、SKILR (SEQ ID NO: 6192)、SFQIN (SEQ ID NO: 6193)、SKACI (SEQ ID NO: 6194)、SKALR (SEQ ID NO: 6195)、SKAHA (SEQ ID NO: 6196)、SLCLN (SEQ ID NO: 6197)、SKAFV (SEQ ID NO: 6198)、RPWQN (SEQ ID NO: 6199)、RPRQN (SEQ ID NO: 6200)、SCPQN (SEQ ID NO: 6201)、SKAQF (SEQ ID NO: 6202)、SVRYN (SEQ ID NO: 6203)、SVRCN (SEQ ID NO: 6204)、SKMPC (SEQ ID NO: 6205)、SKKTS (SEQ ID NO: 6206)、SLPYN (SEQ ID NO: 6207)、VYHQN (SEQ ID NO: 6208)、SKAQS (SEQ ID NO: 6209)、SYTRN (SEQ ID NO: 6210)、LVYQN (SEQ ID NO: 6211)、YRYQN (SEQ ID NO: 6212)、SWLKN (SEQ ID NO: 6213)、SKAQM (SEQ ID NO: 6214)、CPAQN (SEQ ID NO: 6215)、SMCTN (SEQ ID NO: 6216)、SPPDN (SEQ ID NO: 6217)、SKRNY (SEQ ID NO: 6218)、RWTQN (SEQ ID NO: 6219)、SKACR (SEQ ID NO: 6220)、PRKQN (SEQ ID NO: 6221)、SKCSV (SEQ ID NO: 6222)、SKARI (SEQID NO: 6223), PFVQN (SEQ ID NO: 6224), SKLPK (SEQ ID NO: 6225), SKSEQ (SEQ ID NO: 6226), WVAQN (SEQ ID NO: 6227), SLYQN (SEQ ID NO: 6228), SKVRM (SEQ ID NO: 6229), CALQN (SEQ ID NO: 6230), SSCTN (SEQ ID NO: 6231), SKNSR (SEQ ID NO: 6232), SKRKR (SEQ ID NO: 6233), LCTQN (SEQ ID NO: 6234), STCLN (SEQ ID NO: 6235), SYARN (SEQ ID NO: 6236), SKQRP (SEQ ID NO: 6237), SKRVV (SEQ ID NO: 6238), KSGQN (SEQ ID NO: 6239), SYYSN (SEQ ID (ii) any part of the amino acid sequence in (i), such as any 2, 3, or 4 amino acids, such as a sequence of consecutive amino acids; (iii) a sequence of one, two, or three but not more than four modified amino acid sequences relative to any of the amino acid sequences in (i); or (iv) a sequence of one, two, or three but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0383] 305. AAV particles as described in any one of embodiments 300 to 304, wherein [A]-[B] are or comprise: (i) GSGSPHSKAQA (SEQ ID NO: 6250), GSGSPHSKAQN (SEQ ID NO: 6251), GSGSPHSMYMN (SEQ ID NO: 6252), GSGSPHSKAFY (SEQ ID NO: 6253), GSGSPHSKLKR (SEQ ID NO: 6254 ...6254), GSGSPHSKRHR (SEQ ID NO: 6255), GSGSPHSKAQK (SEQ ID NO: 6256), Page 49 / 285 of the specification 59 CN 121079310 A GSGSPHSKWRL (SEQ ID NO: 6257), GSGSPHSRCRN (SEQ ID NO: 6258), GSGSPHSFTCN (SEQ ID NO: 6259), GSGSPHSKFFI (SEQ ID NO: 6260), GSGSPHSKAQY (SEQ ID NO: 6261), GSGSPHIVWQN (SEQ ID NO: 6262), GSGSPHSKYFM (SEQ ID NO: 6263), GSGSPHSKARQ (SEQ ID NO: 6264), GSGSPHSCHQN (SEQ ID NO: 6265), GSGSPHFPWQN (SEQ ID NO: 6266), GSGSPHSKIRR (SEQ ID NO: 6267), GSGSPHVYYQN (SEQ ID NO: 6268), GSGSPHSLYWN (SEQ ID NO: 6269), GSGSPHSKPKR (SEQ ID NO: 6270), GSGSPHSKAFS (SEQ ID NO: 6271), GSGSPHSRFWN (SEQ ID NO: 6272), GSGSPHSKAQC (SEQ ID NO: 6273), GSGSPHSRMRN (SEQ ID NO: 6274), GSGSPHSVKKN (SEQ ID NO: 6275), GSGSPHSWAPN (SEQ ID NO: 6276), GSGSPHSLWKN (SEQ ID NO: 6277), GSGSPHSKARW (SEQ ID NO: 6278), GSGSPHSFRPN (SEQ ID NO: 6279), GSGSPHSKKVF (SEQ ID NO: 6280), GSGSPHSYAFN (SEQ ID NO: 6281), GSGSPHSKACS (SEQ ID NO: 6282), GSGSPHSKRLW (SEQ ID NO: 6283), GSGSPHSCSRN (SEQ ID NO: 6284), GSGSPHSRCPN (SEQ ID NO: ......6286)、 GSGSPHSKFRQ (SEQ ID NO: 6287)、GSGSPHSFPFN (SEQ ID NO: 6288)、GSGSPHSFFGN (SEQ ID NO: 6289)、GSGSPHSMCQN (SEQ ID NO: 6290)、GSGSPHSKLFW (SEQ ID NO: 6291)、 GSGSPHSKTRK (SEQ ID NO: 6292)、GSGSPHSGKRN (SEQ ID NO: 6293)、GSGSPHFYRQN (SEQ ID NO: 6294)、GSGSPHCRVQN (SEQ ID NO: 6295)、GSGSPHYGIQN (SEQ ID NO: 6296)、 GSGSPHSKAQR (SEQ ID NO: 6297)、GSGSPHVNCQN (SEQ ID NO: 6298)、GSGSPHSKPFR (SEQ ID NO: 6299)、GSGSPHSLAWN (SEQ ID NO: 6300)、GSGSPHSKRSY (SEQ ID NO: 6301)、GSGSPHWSYQN (SEQ ID NO: 6302)、GSGSPHRWLQN (SEQ ID NO: 6303)、GSGSPHPSCQN (SEQ ID NO: 6304)、GSGSPHSSWLN (SEQ ID NO: 6305)、GSGSPHSKRRA (SEQ ID NO: 6306)、GSGSPHSKAQT (SEQ ID NO: 6307)、GSGSPHSKGCT (SEQ ID NO: 6308). 6313)、GSGSPHSKARM (SEQ ID NO: 6314)、GSGSPHPKRQN (SEQ ID NO: 6315)、GSGSPHSKCFL (SEQ ID NO: 6316)、GSGSPHWVPQN (SEQ ID NO: 6317)、GSGSPHSFWSN (SEQ ID NO: 6318)、GSGSPHSKFKN (SEQ ID NO:6319)、GSGSPHRIKQN (SEQ ID NO: 6320)、GSGSPHSKAPR (SEQ ID NO: 6321)、 GSGSPHSFRYN (SEQ ID NO: 6322)、GSGSPHSKMIC (SEQ ID NO: 6323)、GSGSPHLRWQN (SEQ ID NO: 6324)、GSGSPHLPTQN (SEQ ID NO: 6325)、GSGSPHSKWKS (SEQ ID NO: 6326)、 GSGSPHSYMRN (SEQ ID NO: 6327)、GSGSPHSKAAR (SEQ ID NO: 6328)、GSGSPHLLCQN (SEQ ID NO: 6329)、GSGSPHRCCQN (SEQ ID NO: 6330)、GSGSPHLCVQN (SEQ ID NO: 6331)、 GSGSPHSKLTR (SEQ ID NO: 6332)、GSGSPHSKLCT (SEQ ID NO: 6333)、GSGSPHSKIRG (SEQ ID NO: 6334)、GSGSPHSYLVN (SEQ ID NO: 6335)、GSGSPHQGCQN (SEQ ID NO: 6336)、 GSGSPHMAFQN (SEQ ID NO: 6337)、GSGSPHSKACQ (SEQ ID NO: 6338)、GSGSPHSKWGL (SEQ ID NO: 6339)、GSGSPHSKILR (SEQ ID NO: 6340)、GSGSPHSFQIN (SEQ ID NO: 6341)、 GSGSPHSKACI (SEQ ID NO: 6342)、GSGSPHSKALR (SEQ ID NO: 6343)、GSGSPHSKAHA (SEQ ID NO: 6344)、GSGSPHSLCLN (SEQ ID NO: 6345)、GSGSPHSKAFV (SEQ ID NO: 6346)、 GSGSPHRPWQN (SEQ ID NO: 6347)、GSGSPHRPRQN (SEQ ID NO: 6348)、GSGSPHSCPQN (SEQ ID NO: 6349)、GSGSPHSKAQF (SEQ ID NO: 6350)、GSGSPHSVRYN (SEQ ID NO: 6351)、 GSGSPHSVRCN (SEQ ID NO:6352), GSGSPHSKMPC (SEQ ID NO: 6353), GSGSPHSKKTS (SEQ ID NO: 6354), GSGSPHSLPYN (SEQ ID NO: 6355), GSGSPHVYHQN (SEQ ID NO: 6356), GSGSPHSKAQS (SEQ ID NO: 6357), GSGSPHSYTRN (SEQ ID NO: 6358), GSGSPHLVYQN (SEQ ID NO: 6359), GSGSPHYRYQN (SEQ ID NO: 6360), GSGSPHSWLKN (SEQ ID NO: 6361), GSGSPHSKAQM (SEQ ID NO: 6362), GSGSPHCPAQN (SEQ ID NO: 6363), GSGSPHSMCTN (SEQ ID NO: 6364), GSGSPHSPPDN (SEQ ID NO: 6365), GSGSPHSKRNY (SEQ ID NO: 6366), GSGSPHRWTQN (SEQ ID NO: 6367), GSGSPHSKACR (SEQ ID NO: 6368), GSGSPHPRKQN (SEQ ID NO: 6369), GSGSPHSKCSV (SEQ ID NO: 6370), GSGSPHSKARI (SEQ ID NO: 6371), GSGSPHPFVQN (SEQ ID NO: 6372), GSGSPHSKLPK (SEQ ID NO: 6373), GSGSPHSKSEQ (SEQ ID NO: 6374), GSGSPHWVAQN (SEQ ID NO: 6375), GSGSPHSLYQN (SEQ ID NO: 6376), GSGSPHSKVRM (SEQ ID NO: 6377), GSGSPHCALQN (SEQ ID NO: 6378), GSGSPHSSCTN (SEQ ID NO:6384), GSGSPHSKQRP (SEQ ID NO: 6385), GGSSPHSKRVV (SEQ ID NO: 6386), GSGSPHKSGQN (SEQ ID NO: 6387), GSGSPHSYYSN (SEQ ID NO: 6388), GGSSPHSLTCN (SEQ ID NO: 6389), GSGSPHSCQSN (SEQ ID NO: 6390), GGSSPHSKAKG (SEQ ID NO: 6391), GGSSPHSKPQN (SEQ ID NO: 6392), GGSSPHSPFQN (SEQ ID NO: 6393), GGSPHSKCTS (SEQ ID NO: 6394), GGSSPHSVFEN (SEQ ID NO: 6395), GGSSPHSKAKK (SEQ ID NO: 6396) or GGSSPHGRYQN (SEQ ID NO: 6397); (ii) (i) any portion of the amino acid sequence, such as any 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids therein, such as a sequence of consecutive amino acids; (iii) a sequence of one, two, or three, but not more than four modified amino acid sequences relative to any of the amino acid sequences in (i); or (iv) a sequence of one, two, or three, but not more than four different amino acids relative to any of the amino acid sequences in (i).

[0384] 306. An AAV particle as described in any of embodiments 300 to 305, wherein [A]-[B] does not contain the amino acid sequence of GSGSPHSKAQN (SEQ ID NO: 6251).

[0385] 307. An AAV particle as described in any one of embodiments 300 to 306, wherein the AAV capsid variant comprises one, two, or all of the following amino acids according to SEQ ID NO: 138: position 458 other than Q (e.g., R, C, S, W, L, F, Y, H, I, V, A, or P), position 459 other than Q (e.g., K, I, R, L, or S), and / or position 460 other than T (e.g., R).

[0386] 308. An AAV particle as described in any one of embodiments 300 to 307, wherein the AAV capsid variant comprises: (i) amino acid R at position 458; (ii) amino acid W at position 458; (iii) amino acid Y at position 458; (iv) amino acid F at position 458; (v) amino acid S at position 458; (vi) amino acid C at position 458; (vii) amino acid C at position 458;Amino acid I at position 458; (viii) Amino acid L at position 458; (ix) Amino acid P at position 458; (x) Amino acid I at position 459; (xi) Amino acid H at position 458; or (xii) Amino acid V at position 458; wherein (i)-(xii) are numbered according to SEQ ID NO: 138.

[0387] 309. An AAV particle as described in any one of embodiments 300 to 307, wherein the AAV capsid variant comprises: (i) amino acid R at position 458 and amino acid K at position 459; (ii) amino acid C at position 458 and amino acid I at position 459; (iii) amino acid S at position 458 and amino acid R at position 459; (iv) amino acid L at position 458 and amino acid K at position 459; (v) amino acid F at position 458 and amino acid K at position 459; (vi) amino acid C at position 458 and amino acid R at position 459; (vii) amino acid H at position 458 and amino acid R at position 459; (viii) amino acid I at position 458 and amino acid L at position 459; (ix) amino acid V at position 458 and amino acid R at position 459; (x) amino acid A at position 458 and amino acid K at position 459; (xi) Amino acid I at position 458 and amino acid K at position 459; (xii) amino acid C at position 458 and amino acid S at position 459; or (xiii) amino acid C at position 458 and amino acid L at position 459, wherein (i)-(xiii) are numbered according to SEQ ID NO: 138.

[0388] 310. An AAV particle as described in any one of embodiments 300 to 307, wherein the AAV capsid variant comprises amino acid F at position 458, amino acid K at position 459, and amino acid R at position 460, as numbered according to SEQ ID NO: 138.

[0389] 311. An AAV particle as described in any one of embodiments 300 to 310, wherein the AAV capsid variant comprises one, two, or all of the amino acids other than T (e.g., Y, P, W, R, K, S, or F) at position 450, the amino acids other than I (e.g., R, S, Y, L, V, H, P, A, or F) at position 451, and / or the amino acids other than N (e.g., V, W, A, T, F, Y, L, R, H, S, or M) at position 452 according to SEQ ID NO: 138.

[0390] 312. An AAV particle as described in any one of embodiments 300 to 311, wherein the AAV capsid variant comprises one, two, or all of the amino acids other than T (e.g., Y, P, W, R, K, S, or F) at position 450, the amino acids other than I (e.g., R, S, Y, L, V, H, S, or M) at position 452 according to SEQ ID NO: 138.313. The AAV particle of any one of embodiments 300 to 312, wherein the AAV capsid variant comprises amino acid Y at position 450 and amino acid V at position 452 according to SEQ ID NO: 138.

[0392] 314. The AAV particle of any one of embodiments 300 to 312, wherein the AAV capsid variant comprises amino acid R at position 450 and amino acid Y at position 451 according to SEQ ID NO: 138.

[0393] 315. An AAV particle as described in any one of embodiments 300 to 311, wherein the AAV capsid variant comprises: (i) amino acid P at position 450, amino acid R at position 451, and amino acid W at position 452; (ii) amino acid Y at position 450, amino acid S at position 451, and amino acid A at position 452; (iii) amino acid Y at position 450, amino acid Y at position 451, and amino acid T at position 452; (iv) amino acid P at position 450, amino acid R at position 451, and amino acid F at position 452; (v) amino acid W at position 450, amino acid L at position 451, and amino acid T at position 452; (vi) amino acid R at position 450, amino acid S at position 451, and amino acid Y at position 452; (vii) amino acid R at position 450, amino acid S at position 451, and amino acid Y at position 452; (viii) Amino acid Y at position 450, amino acid V at position 451, and amino acid F at position 452; (ix) Amino acid P at position 450, amino acid P at position 451, and amino acid L at position 452; (x) Amino acid P at position 450, amino acid A at position 451, and amino acid R at position 452; (xi) Amino acid S at position 450, amino acid R at position 451, and amino acid R at position 452; (xii) Amino acid F at position 450, amino acid F at position 451, and amino acid H at position 452; (xiii) Amino acid R at position 450, amino acid F at position 451, and amino acid S at position 452; (xiv) Amino acid Y at position 450, amino acid S at position 451, and amino acid M at position 452; or (xv) Amino acid P at position 450, amino acid F at position 451, and amino acid L at position 452; wherein (i)-(xv) are numbered according to SEQ ID NO: 138.

[0394] 316.The AAV particle of any one of embodiments 300 to 315, wherein the AAV capsid variant comprises: (i) an amino acid sequence of any one of SEQ ID NO: 3849-3982, 2984-4010, 4681-4693; (ii) an amino acid sequence comprising any part of the amino acid sequence of (i), such as any 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16 or 17 amino acids, such as a sequence of consecutive amino acids; (iii) an amino acid sequence comprising one, two or three but not more than four modified amino acid sequences relative to any one of the amino acid sequences of (i); or (iv) an amino acid sequence comprising one, two or three but not more than four different amino acids relative to any one of the amino acid sequences of (i).

[0395] 317. An AAV particle as described in any one of embodiments 300 to 316, wherein the AAV capsid variant does not contain the amino acid sequence of GSGSPHSKAQNQQ (SEQ ID NO: 1801) or GSGSPHSKAQNQQT (SEQ ID NO: 200).

[0396] 318. An AAV particle as described in any one of embodiments 300 to 317, wherein [A]-[B] are present in ring IV.

[0397] 319. An AAV particle as described in any one of embodiments 300 to 318, wherein [A] is present immediately after position 452 according to SEQ ID NO: 138 or 981.

[0398] 320. An AAV particle as described in any one of embodiments 300 to 319, wherein [A] replaces positions 453-455 (e.g., G453, S454, G455) numbered according to SEQ ID NO: 138 or 981.

[0399] 321. An AAV particle as described in any one of embodiments 300 to 320, wherein [A] is present immediately after position 452, and wherein [A] replaces positions 453-455 (e.g., G453, S454, G455) numbered according to SEQ ID NO: 138 or 981.

[0400] 322. An AAV particle as described in any one of embodiments 300 to 321, wherein [B] is present immediately after [A].

[0401] 323. An AAV particle as described in any one of embodiments 300 to 322, wherein [B] replaces positions 456 and 457 (e.g., Q456, N457) according to SEQ ID NO: 138.

[0402] 324. An AAV particle as described in any one of embodiments 300 to 323, wherein [A]-[B] replaces positions 453-457 according to SEQ ID NO: 138.(e.g., G453, S454, G455, Q456, N457).

[0403] 325. An AAV particle as described in any of embodiments 300 to 324, wherein [A]-[B] are present immediately after position 452, and wherein [A]-[B] replace positions 453-457 (e.g., G453, S454, G455, Q456, N457), which are numbered according to SEQ ID NO: 138.

[0404] 326. An AAV particle as described in any of embodiments 300 to 325, wherein the AAV capsid variant comprises [A][B] from the N-terminus to the C-terminus. Specification 53 / 285 pages 63 CN 121079310 A

[0405] 327. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises at least one, at least two, at least three, or at least four (e.g., 1-4 to 1-5) charged amino acid residues (e.g., acidic and / or basic amino acid residues) relative to SEQ ID NO: 138, which are present at the N-terminus of the amino acid sequence of SPH (e.g., within 1, 2, 3, 4, 5, or 6 amino acids starting from the amino acid sequence of SPH (e.g., within positions 450-455 according to SEQ ID NO: 138)), optionally, wherein the amino acid sequence of SPH is present at positions 456-458 according to any one of SEQ ID NO: 36-59, 981, or 982.

[0406] 328. An AAV particle as described in embodiment 327, wherein the amino acid sequence of SPH is present at positions 456-458 according to any one of the numberings in SEQ ID NO: 36-59, 981, or 982.

[0407] 329. An AAV particle as described in embodiment 327 or 328, wherein the AAV capsid variant comprises fewer than four, fewer than three, or fewer than two (e.g., two or one) charged amino acid residues (e.g., acidic and / or basic amino acid residues) relative to SEQ ID NO: 138.

[0408] 330. An AAV particle as described in any one of embodiments 327 to 329, wherein the AAV capsid variant comprises one charged amino acid residue (e.g., acidic or basic amino acid residue) relative to SEQ ID NO: 138, optionally at any one of the positions 450-455 relative to the numbering in SEQ ID NO: 138.

[0409] 331. An AAV particle as described in any one of embodiments 327 to 330, wherein the charged amino acid residue is an acidic amino acid (e.g., D or E).

[0410] 332. An AAV particle as described in any one of embodiments 327 to 331, wherein the charged amino acid residue is a negatively charged amino acid (e.g., D or E).

[0411] 333. An AAV particle of any one of embodiments 327 to 332, wherein the charged amino acid residue is D.

[0412] 334. An AAV particle of any one of embodiments 327 to 333, wherein the charged amino acid residue is E.

[0413] 335. An AAV particle of any one of embodiments 327 to 334, wherein the charged amino acid residue is a basic amino acid (e.g., K, R, or H).

[0414] 336. An AAV particle of any one of embodiments 327 to 335, wherein the charged amino acid residue is a positively charged amino acid (e.g., K, R, or H).

[0415] 337. An AAV particle of any one of embodiments 327 to 336, wherein the charged amino acid residue is H.

[0416] 338. An AAV particle of any one of embodiments 327 to 337, wherein the charged amino acid residue is R.

[0417] 339. An AAV particle of any one of embodiments 327 to 338, wherein the charged amino acid residue is K.

[0418] 340. An AAV particle of any one of embodiments 327 to 339, wherein the AAV capsid variant comprises an acidic amino acid (e.g., E or D) and a basic amino acid (e.g., R, K or H).

[0419] 341. An AAV particle of any one of embodiments 327 to 340, wherein at least one, two, three or four charged amino acid residues are present in 1, 2, 3, 4, 5 or 6 (e.g., 1-6) amino acids starting from the SPH amino acid sequence.

[0420] 342. An AAV particle as described in any one of embodiments 327 to 341, wherein the AAV capsid variant comprises two charged amino acid residues immediately preceding the amino acid sequence of SPH (e.g., at positions 454 and 455 numbered according to SEQ ID NO: 138 or SEQ ID NO: 982).

[0421] 343. An AAV particle as described in any one of embodiments 327 to 342, wherein the AAV capsid variant comprises charged amino acid residues (e.g., E) within 1, 2, 3, 4, 5 (e.g., 5) amino acids starting from the amino acid sequence of SPH.

[0422] 344. An AAV particle as described in any one of embodiments 327 to 343, wherein the AAV capsid variant comprises a charged amino acid residue (e.g., E) at position 451 numbered according to any one of SEQ ID NO: 138, 981, or 982.

[0423] 345. An AAV particle as described in any one of embodiments 327 to 344, wherein the AAV capsid variant comprises E at position 451 of any of the numbers listed on page 54 / 285 of the specification, CN 121079310 A SEQ ID NO: 138, 981, or 982.

[0424] 346. An AAV particle of any one of embodiments 327 to 345, wherein the AAV capsid variant comprises a charged amino acid residue (e.g., R or K) at position 452 numbered according to any one of SEQ ID NO: 138, 981, or 982.

[0425] 347. An AAV particle of any one of embodiments 327 to 346, wherein the AAV capsid variant comprises R at position 452 numbered according to SEQ ID NO: 138 or SEQ ID NO: 982.

[0426] 348. An AAV particle of any one of embodiments 327 to 347, wherein the AAV capsid variant comprises E at position 451 numbered according to SEQ ID NO: 138 or SEQ ID NO: 982 and R at position 452.

[0427] 349. An AAV particle as described in any one of embodiments 327 to 348, wherein the AAV capsid variant has a reduced tropism to hepatocytes or tissues relative to the tropism of an AAV capsid containing the amino acid sequence of SEQ ID NO: 138 or SEQ ID NO: 981.

[0428] 350. An AAV particle as described in any of the preceding embodiments, wherein the AAV capsid variant comprises at least one, at least two, at least three, or at least four (e.g., 1-4 to 1-5) charged amino acid residues (e.g., basic amino acid residues) relative to SEQ ID NO: 138, which are present at the C-terminus of the amino acid sequence of SPH (e.g., within 1, 2, 3, 4, 5, 6, or 7 amino acids from the end of the amino acid sequence of SPH (e.g., within position 459-465 according to any of the numbers in SEQ ID NO: 36-59 or 981)), optionally, wherein the amino acid sequence of SPH is present at position 456-458 according to any of the numbers in SEQ ID NO: 36-59, 981, or 982.

[0429] 351. An AAV particle as described in embodiment 350, wherein the amino acid sequence of SPH is present at positions 456-458 according to any one of the numberings in SEQ ID NO: 36-59, 981, or 982.

[0430] 352. An AAV particle as described in embodiment 350 or 351, wherein the AAV capsid variant comprises fewer than four, fewer than three, or fewer than two (e.g., two or one) charged amino acid residues (e.g., basic amino acid residues) relative to SEQ ID NO: 138.

[0431] 353. An AAV particle as described in any one of embodiments 350 to 352, wherein the AAV capsid variant comprises one charged amino acid residue (e.g., basic amino acid residue) relative to SEQ ID NO: 138, which optionally is located at positions 456-458 according to SEQ ID NO:138 at any of positions 456-460, or at positions 462-466 ​​according to any of the numbers SEQ ID NO: 36-59, 981, or 982.

[0432] 354. An AAV particle as described in any of embodiments 350 to 353, wherein the charged amino acid residue is a basic amino acid (e.g., R or K).

[0433] 355. An AAV particle as described in any of embodiments 350 to 354, wherein the charged amino acid residue is a positively charged amino acid (e.g., R or K).

[0434] 356. An AAV particle as described in any of embodiments 350 to 355, wherein the charged amino acid residue is R.

[0435] 357. An AAV particle as described in any of embodiments 350 to 355, wherein the charged amino acid residue is K.

[0436] 358. An AAV particle of any one of embodiments 350 to 357, wherein at least one, two, three, or four charged amino acid residues are present in the first, second, third, fourth, fifth, sixth, or seventh (e.g., 1-7) amino acids following the end of the SPH amino acid sequence.

[0437] 359. An AAV particle of any one of embodiments 350 to 358, wherein the AAV capsid variant comprises a charged amino acid residue (e.g., K or R) immediately following the SPH sequence (e.g., at position 459 according to SEQ ID NO: 981).

[0438] 360. An AAV particle of any one of embodiments 350 to 359, wherein the AAV capsid variant comprises a charged amino acid residue (e.g., K or R) at position 459 according to SEQ ID NO: 138 or SEQ ID NO: 982. Specification 55 / 285 pages 65 CN 121079310 A

[0439] 361. An AAV particle as described in any one of embodiments 350 to 360, wherein the AAV capsid variant comprises K at position 459 according to SEQ ID NO: 981.

[0440] 362. An AAV particle as described in any one of embodiments 350 to 360, wherein the AAV capsid variant comprises R at position 459 according to SEQ ID NO: 981.

[0441] 363. An AAV particle as described in any one of embodiments 350 to 362, wherein the AAV capsid variant comprises charged amino acid residues (e.g., R or K) at one, two, three, four, five, or all of positions 460, 461, 462, 463, 464, and / or 465 according to SEQ ID NO: 138 or 981.

[0442] 364. AAV particles as described in any one of embodiments 300 to 326 or 350 to 363, wherein the AAV particles are associated with the inclusion of SEQ ID NO:The AAV capsid with amino acid sequence 138 exhibits tropism towards liver cells or tissues, wherein the AAV capsid variant has increased tropism towards liver cells or tissues.

[0443] 365. AAV particles as described in any one of embodiments 300 to 326 or 350 to 364, wherein, when measured, for example, by means of the analysis described in Example 4, the AAV capsid variant is enriched in the liver at least 10 times, at least 15 times, at least 20 times, at least 25 times, at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 55 times, at least 60 times, at least 65 times, at least 70 times, at least 75 times, at least 80 times, at least 85 times, at least 90 times, at least 95 times, at least 100 times, at least 105 times, at least 110 times, at least 115 times, at least 120 times. At least 125 times, at least 130 times, at least 135 times, at least 140 times, at least 150 times, at least 160 times, at least 170 times, at least 180 times, at least 190 times, or at least 200 times.

[0444] 366. An AAV particle as described in any one of embodiments 300 to 326, 364, or 365, wherein the AAV capsid variant has a reduced tropism to CNS cells or tissues, such as brain cells, brain tissue, spinal cord cells, or spinal cord tissue, relative to the tropism of the AAV capsid containing the amino acid sequence SEQ ID NO: 138.

[0445] 367. An AAV particle as described in any one of embodiments 300 to 326 or 364 to 366, wherein the AAV capsid variant shows preferential transduction in the liver region relative to transduction in the brain and / or dorsal root ganglion (DRG).

[0446] 368. An AAV particle as described in any one of embodiments 300 to 326 or 364 to 367, wherein the AAV capsid variant exhibits preferential transduction in the liver region relative to transduction in the heart and / or muscles (e.g., quadriceps femoris).

[0447] 369. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., the AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding the human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence of any one of the sequences provided in Tables 1, 2A, 2B, or Tables 20-26; (b) an amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, or at least 17 consecutive amino acids from any one of the sequences provided in Tables 1, 2A, 2B, or Tables 20-26; or (c)(d) An amino acid sequence containing at least one, at least two, or at least three but no more than four different amino acids relative to any one of the sequences provided in Tables 1, 2A, 2B, or 20-26; or (d) An amino acid sequence containing at least one, at least two, or at least three but no more than four modified amino acid sequences relative to any one of the sequences provided in Tables 1, 2A, 2B, or 20-26.

[0448] 370. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence of any one of SEQ ID NO: 945-980 or 985-986; (b) an amino acid sequence comprising at least 3, at least 4, or at least 5 consecutive amino acids from any one of SEQ ID NO: 945-980 or 985-986; or (c) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of any one of SEQ ID NO: 945-980 or 985-986; (d) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of any one of SEQ ID NO: The amino acid sequence of any one of 945-980 or 985-986 contains at least one, at least two, or at least three but no more than four modified amino acid sequences.

[0449] 371. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., the AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence of any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404 or 903-909; (b) an amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12 or at least 13 consecutive amino acids from any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404 or 903-909; (c) The amino acid sequence comprising, relative to any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404 or 903-909, an amino acid sequence containing at least one, at least two, or at least three but no more than four distinct amino acids; or(d) The amino acid sequence relative to any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404 or 903-909 contains at least one, at least two or at least three but no more than four modified amino acid sequences.

[0450] 372. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence of any one of SEQ ID NO: 3849-4051 or 4681-4693; (b) an amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, or at least 17 consecutive amino acids from any one of SEQ ID NO: 3849-4051 or 4681-4693; (c) relative to SEQ ID NO: The amino acid sequence of any one of SEQ ID NO: 3849-4051 or 4681-4693 contains at least one, at least two, or at least three but no more than four different amino acid sequences; or (d) relative to the amino acid sequence of any one of SEQ ID NO: 3849-4051 or 4681-4693, contains at least one, at least two, or at least three but no more than four modified amino acid sequences.

[0451] 373. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence of any one of SEQ ID NO: 4052-4092; (b) an amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, or at least 17 consecutive amino acids from any one of SEQ ID NO: 4052-4092; (c) relative to SEQ ID NO: The amino acid sequence of any one of 4052-4092, comprising at least one, at least two, or at least three but no more than four different amino acids; or the amino acid sequence of CN 121079310 A (d) on page 57 / 285 of the specification, relative to SEQ ID NO:The amino acid sequence of any one of 4052-4092 contains at least one, at least two, or at least three but no more than four modified amino acid sequences.

[0452] 374. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence (e.g., encoding human CDKL5 protein), wherein the AAV capsid variant comprises: (a) an amino acid sequence comprising any one of SEQ ID NO: 4056, 4058, 4059, 4062-4064, 4066, 4067, 4080, 4084, 4090 or 4095-4097; (b) an amino acid sequence comprising from SEQ ID NO: 4056, 4058, 4059, 4062-4064, 4066, 4067, 4080, 4084, (c) An amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, or at least 17 consecutive amino acids relative to any one of SEQ ID NO: 4056, 4058, 4059, 4062-4064, 4066, 4067, 4080, 4084, 4090, or 4095-4097, comprising at least 1, at least 2, or at least 3 but not more than 4 distinct amino acids; or (d) An amino acid sequence comprising at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, or at least 17 consecutive amino acids relative to any one of SEQ ID NO: 4056, 4058, 4059, 4062-4064, 4066, 4067, 4080, 4084, or 4095-4097. The amino acid sequence of any one of 4090 or 4095-4097, comprising at least one, at least two, or at least three but no more than four modified amino acid sequences.

[0453] 375. The AAV particle of embodiment 369 or 371, wherein the AAV capsid variant comprises an amino acid sequence containing at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, or at least 13 consecutive amino acids from any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404, or 903-909.

[0454] 376. The AAV particle of any one of embodiments 369 to 374, wherein at least 3 consecutive amino acids comprise SPH.

[0455] 377. An AAV particle as described in any one of embodiments 369 to 371 or 376, wherein at least four consecutive amino acids comprise SPHS (SEQ ID NO: 4700).

[0456] 378. An AAV particle of any one of embodiments 369 to 371, 376, or 377, wherein at least 5 consecutive amino acids comprise SPHSK (SEQ ID NO: 4701).

[0457] 379. An AAV particle of any one of embodiments 369 to 371 or 376 to 378, wherein at least 6 consecutive amino acids comprise SPHSKA (SEQ ID NO: 941).

[0458] 380. An AAV particle of embodiments 369 to 371, wherein at least 3 consecutive amino acids comprise HDS.

[0459] 381. An AAV particle of any one of embodiments 369 to 371 or 380, wherein at least 4 consecutive amino acids comprise HDSP (SEQ ID NO: 4702).

[0460] 382. An AAV particle as described in any one of embodiments 369 to 371, 380 or 381, wherein at least 5 consecutive amino acids comprise HDSPH (SEQ ID NO: 4703).

[0461] 383. An AAV particle as described in any one of embodiments 369 to 371 or 380 to 382, ​​wherein at least 6 consecutive amino acids comprise HDSPHK (SEQ ID NO: 2).

[0462] 384. An AAV particle as described in any one of embodiments 369 to 371, wherein: (i) at least 3 consecutive amino acids contain SPH; (ii) at least 4 consecutive amino acids contain SPHK (SEQ ID NO: 6398); (iii) at least 5 consecutive amino acids contain SPHKY (SEQ ID NO: 4715); and / or (iv) at least 6 consecutive amino acids contain SPHKYG (SEQ ID NO: 966).

[0463] 385. An AAV particle as described in embodiment 369 or 371, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four modifications relative to any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404, or 903-909.

[0464] 386. An AAV particle as described in any one of embodiments 369, 371, or 385, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four modifications relative to the SPHSKA amino acid sequence (SEQ ID NO: 941).

[0465] 387. An AAV particle as described in any one of embodiments 369, 371, or 385, wherein the AAV capsid variant comprisesAn amino acid sequence comprising at least one, at least two, or at least three but no more than four modifications relative to the amino acid sequence of HDSPHK (SEQ ID NO: 2).

[0466] 388. An AAV particle as described in any one of embodiments 369 to 371, 384, or 385, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four modifications relative to the amino acid sequence of SPHKYG (SEQ ID NO: 966).

[0467] 389. The AAV particle of embodiment 370, wherein the AAV capsid variant comprises: (i) an amino acid sequence comprising at least one, at least two, or at least three but not more than four modified amino acid sequences relative to the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589); (ii) an amino acid sequence comprising at least one, at least two, or at least three but not more than four modified amino acid sequences relative to the amino acid sequence of KAEIGHDSPHKSGQNQQT (SEQ ID NO: 1754); (iii) an amino acid sequence comprising at least one, at least two, or at least three but not more than four modified amino acid sequences relative to the amino acid sequence of KTEKMSGSPHSKAQNQQT (SEQ ID NO: 3241); (iv) an amino acid sequence comprising at least one, at least two, or at least three but not more than four modified amino acid sequences relative to the amino acid sequence of KTINGHDSPHSKAQNLQT (SEQ ID NO: 4100); or (v) an amino acid sequence comprising at least one, at least two, or at least three but not more than four modified amino acid sequences relative to the amino acid sequence of KTVNGHDSPHSKAQNQQT (SEQ ID NO: 1754). 4062), comprising at least one, at least two, or at least three but no more than four modified amino acid sequences.

[0468] 390. AAV particles as described in embodiment 369 or 371, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four different amino acids relative to any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404, or 903-909.

[0469] 391. AAV particles as described in embodiment 369, 371, or 390, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four different amino acids relative to the amino acid sequence of SPHSKA (SEQ ID NO: 941).

[0470] 392. An AAV particle as described in any one of embodiments 369, 371 or 390, wherein the AAV capsid variant comprisesAn amino acid sequence comprising at least one, at least two, or at least three but no more than four different amino acids relative to the amino acid sequence of HDSPHK (SEQ ID NO: 2).

[0471] 393. An AAV particle as described in any one of embodiments 369, 371, 384, or 390, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than four different amino acids relative to the amino acid sequence of SPHKYG (SEQ ID NO: 966).

[0472] 394. The AAV particle of embodiment 369, wherein the AAV capsid variant comprises: (i) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589); (ii) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of KAEIGHDSPHKSGQNQQT (SEQ ID NO: 1754); (iii) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of KTEKMSGSPHSKAQNQQT (SEQ ID NO: 3241); (iv) an amino acid sequence comprising at least one, at least two, or at least three but not more than four different amino acids relative to the amino acid sequence of KTINGHDSPHSKAQNLQT (SEQ ID NO: 3589). 4100), comprising an amino acid sequence containing at least one, at least two, or at least three but not more than four different amino acids; or (v) relative to the amino acid sequence of KTVNGHDSPHSKAQNQQT (SEQ ID NO: 4062), comprising an amino acid sequence containing at least one, at least two, or at least three but not more than four different amino acids.

[0473] 395. An AAV particle as described in any one of embodiments 1 to 129, 269, 271, 375 to 388, or 390 to 394, wherein the AAV capsid variant comprises an amino acid sequence containing any one of SEQ ID NO: 2, 200, 201, 941, 943, 204, 208, 404, or 903-909.

[0474] 396. An AAV particle as described in any one of embodiments 295 to 297, 301 to 305, 313, 314, 318, 319 or 323, wherein the AAV capsid variant comprises the amino acid sequence of ERVGSPHSKA (SEQ ID NO: 6399), optionally wherein the amino acid sequence is present immediately after position 450 and replaces positions 451-455.(e.g., I451, N542, G453, S454, G455), these positions are numbered according to SEQ ID NO: 138.

[0475] 397. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 385, 386, 389 to 391 or 394 to 396, wherein the AAV capsid variant comprises the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589), optionally wherein the amino acid sequence is present immediately after position 448 and replaces positions 449-460 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, T460), these positions are numbered according to SEQ ID NO: 138.

[0476] 398. An AAV particle as described in any one of embodiments 269 to 371, 375, 380 to 383, 385, 387, 389, 390, 393 or 394, wherein the AAV capsid variant comprises the amino acid sequence AEIGHDSPHKSG (SEQ ID NO: 6400), optionally wherein the amino acid sequence is present immediately after position 449 and replaces positions 450-455 (e.g., T450, I451, N452, G453, S454, G455), these positions being numbered according to SEQ ID NO: 138.

[0477] 399. An AAV particle as described in any one of embodiments 369 to 371, 375, 380 to 383, 385, 387, 389, 390, 393, 394 or 398, wherein the AAV capsid variant comprises the amino acid sequence KAEIGHDSPHKSGQNQQT (SEQ ID NO: 1754), optionally wherein the amino acid sequence is present immediately after position 448 and replaces positions 449-460 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, T460), these positions being numbered according to SEQ ID NO: 138.

[0478] 400. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 390, 391, or 395, wherein the AAV capsid variant comprises the amino acid sequence of EKMSGSPHSKA (SEQ ID NO: 6401), optionally wherein the amino acid sequence is present immediately after position 450 and substitutes positions 451-455 (e.g., I451, N452, G453, S454, G455), these positions being numbered according to SEQ ID NO: 138.

[0479] 401.AAV particles as described in any one of embodiments 369 to 371, 375 to 379, 390, 391 or 395, wherein the AAV capsid variant comprises the amino acid sequence KTEKMSGSPHSKAQNQQT (SEQ ID NO: 3241), optionally wherein the amino acid sequence is present immediately after position 448 and replaces positions 449-460 (e.g. K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, T460), these positions being numbered according to SEQ ID NO: 138.

[0480] 402. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 390, 391 or 395, wherein the AAV capsid variant of this specification (page 60 / 285, CN 121079310 A) comprises the amino acid sequence HDSPHSKAQNL (SEQ ID NO: 6402), optionally wherein the amino acid sequence is present immediately after position 453 and replaces positions 456-458 (e.g., Q456, N457, Q458), which are numbered according to SEQ ID NO: 138.

[0481] 403. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 390, 391 or 395, wherein the AAV capsid variant comprises the amino acid sequence KTINGHDSPHSKAQNLQT (SEQ ID NO: 4100), optionally wherein the amino acid sequence is present immediately after position 448 and replaces positions 449-460 (e.g. K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, T460), these positions being numbered according to SEQ ID NO: 138.

[0482] 404. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 390, 391, or 395, wherein the AAV capsid variant comprises the amino acid sequence VNGHDSPHSKA (SEQ ID NO: 6403), optionally wherein the amino acid sequence is present immediately after position 450 and replaces positions 451-455 (e.g., I451, N452, G453, S454, G455), these positions being numbered according to SEQ ID NO: 138.

[0483] 405. An AAV particle as described in any one of embodiments 369 to 371, 375 to 379, 390, 391, or 395, wherein the AAV capsid variant comprises the amino acid sequence KTVNGHDSPHSKAQNQQT (SEQ ID NO: 4062), optionally whereinThe amino acid sequence immediately following position 448 exists and replaces positions 449–460 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459, T460), these positions are numbered according to SEQ ID NO: 138.

[0484] 406. An AAV particle as described in any one of embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391 or 395, wherein the AAV capsid variant comprises an amino acid sequence encoded by: the nucleotide sequence of SEQ ID NO: 942; comprising relative to SEQ ID NO: The nucleotide sequence of SEQ ID NO: 942 may contain at least one, two, three, four, five, six, or seven modifications, such as substitutions (e.g., conserved substitutions), but not more than ten modifications, such as substitutions (e.g., conserved substitutions); or a nucleotide sequence containing at least one, two, three, four, five, six, or seven, but not more than ten different nucleotides relative to the nucleotide sequence of SEQ ID NO: 942.

[0485] 407. An AAV particle as described in any one of embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, or 395, wherein the AAV capsid variant comprises an amino acid sequence encoded by: SEQ ID NO: The nucleotide sequence of SEQ ID NO: 3; a nucleotide sequence containing at least one, two, three, four, five, six or seven modifications, such as substitutions (e.g., conserved substitutions), but not more than ten modifications, such as substitutions (e.g., conserved substitutions), relative to the nucleotide sequence of SEQ ID NO: 3; or a nucleotide sequence containing at least one, two, three, four, five, six or seven, but not more than ten different nucleotides relative to the nucleotide sequence of SEQ ID NO: 3.

[0486] 408.AAV particles according to any one of embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, or 406, wherein the nucleotide sequence encoding the capsid variant comprises the nucleotide sequence of SEQ ID NO: 942; comprising relative to SEQ ID NO: The nucleotide sequence of SEQ ID NO: 942 may contain at least one, two, three, four, five, six or seven modifications, such as substitutions (e.g., conserved substitutions), but not more than ten modifications, such as substitutions (e.g., conserved substitutions); or a nucleotide sequence containing at least one, two, three, four, five, six or seven, but not more than ten different nucleotides relative to the nucleotide sequence of SEQ ID NO: 942. Instruction manual, pages 61 / 285, CN 121079310 A

[0487] 409. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particle of any one of embodiments 395 or 407, wherein the nucleotide sequence encoding the capsid variant comprises the nucleotide sequence of SEQ ID NO: 3; a nucleotide sequence comprising at least one, two, three, four, five, six or seven modifications, such as substitutions (e.g., conserved substitutions) but not more than ten modifications, such as substitutions (e.g., conserved substitutions), relative to the nucleotide sequence of SEQ ID NO: 3; or a nucleotide sequence comprising at least one, two, three, four, five, six or seven but not more than ten different nucleotides relative to the nucleotide sequence of SEQ ID NO: 3.

[0488] 410. AAV particle of any one of embodiments 369 to 409, wherein, for example, relative to the amino acid sequence of SEQ ID NO: 138, the amino acid sequence is present in ring IV.

[0489] 411. AAV particle of any one of embodiments 369 to 410, wherein the amino acid sequence immediately follows the sequence according to SEQ ID NO:The position of SEQ ID NO: 138 is present after 448, 449, 450, 451, 452, 453, 454 or 455.

[0490] 412. AAV particles as described in any one of embodiments 369 to 411, wherein the amino acid sequence substitutions are amino acids 449, 450, 451, 452, 453, 454, 455, 456, 457, 458, 459 and / or 460 (e.g., K449, T450, I451, N452, G453, S454, G455, Q456, N457, Q458, Q459 and / or T460) according to SEQ ID NO: 138.

[0491] 413. An AAV particle as described in any one of embodiments 369 to 412, wherein the amino acid sequence is present immediately after position 455 according to SEQ ID NO: 138.

[0492] 414. An AAV particle as described in any one of embodiments 369 to 413, wherein the amino acid sequence is present immediately after position 453 according to SEQ ID NO: 138.

[0493] 415. An AAV particle as described in any one of embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 02, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, or 410 to 413, wherein the AAV capsid variant comprises the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after position 455 encoded according to SEQ ID NO: 138.

[0494] 416. An AAV particle as described in any one of embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 02, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, 410 to 413 or 415, wherein the AAV capsid variant comprises the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence immediately follows the sequence according to SEQ ID NO:The position 455 of number 981 exists.

[0495] 417. The AAV particle of embodiment 415 or 416, wherein the AAV capsid variant further comprises an amino acid other than I at position 451, an amino acid other than N at position 452, and an amino acid other than G at position 453, according to any one of the numbers SEQ ID NO: 36, 138, or 981.

[0496] 418. The AAV particle of any one of embodiments 415 to 417, wherein the AAV capsid variant further comprises E at position 451, R at position 452, and V at position 453, according to any one of the numbers SEQ ID NO: 36, 138, or 981.

[0497] 419. An AAV particle as described in any one of embodiments 415 to 418, wherein the AAV capsid variant further comprises substitutions I451E, N452R, and G453V according to any one of SEQ ID NO: 36, 138, or 981.

[0498] 420. An AAV particle as described in any one of embodiments 415 to 419, wherein the AAV capsid variant comprises: (i) E at position 451, R at position 452, and V at position 453 according to any one of SEQ ID NO: 36, 138, or 981; and (ii) an amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after position 455 according to any one of SEQ ID NO: 36, 138, or 981.

[0499] 421. The AAV particle of embodiment 415 or 416, wherein the AAV capsid variant further comprises an amino acid other than I at position 451, an amino acid other than N at position 452, and / or G at position 453, as specified in SEQ ID NO: 39 or 138.

[0500] 422. The AAV particle of any one of embodiments 415, 416, or 421, wherein the AAV capsid variant further comprises E at position 451, K at position 452, and / or M at position 453, as specified in SEQ ID NO: 138 or 39.

[0501] 423. The AAV particle of any one of embodiments 415, 416, 421, or 422, wherein the AAV capsid variant further comprises substitutions I451E, N452K, and G453M, as specified in SEQ ID NO: 39 or 138.

[0502] 424. An AAV particle as described in any one of embodiments 415, 416, or 421 to 423, wherein the AAV capsid variant comprises: (i) according to SEQ ID NO:E at position 451, K at position 452, and M at position 453, as specified in SEQ ID NO: 39 or 138; and (ii) the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately following position 455 according to SEQ ID NO: 39 or 138.

[0503] 425. The AAV particle of embodiment 415 or 416, wherein the AAV capsid variant further comprises an amino acid other than S at position 454, an amino acid other than G at position 455, and / or Q at position 458, as specified in SEQ ID NO: 138.

[0504] 426. The AAV particle of any one of embodiments 415, 416, or 425, wherein the AAV capsid variant further comprises H at position 454, D at position 455, and / or L at position 458, as specified in SEQ ID NO: 138.

[0505] 427. An AAV particle as described in any one of embodiments 415, 416, 425 or 426, wherein the AAV capsid variant further comprises the substitutions S454H, G455D and Q458L as numbered according to SEQ ID NO: 138.

[0506] 428. An AAV particle as described in any one of embodiments 415, 416 or 425 to 427, wherein the AAV capsid variant comprises: (i) H at position 454, D at position 455 and / or L at position 458 as numbered according to SEQ ID NO: 138; and (ii) an amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after position 455 as numbered according to SEQ ID NO: 138.

[0507] 429. The AAV particle of embodiment 415 or 416, wherein the AAV capsid variant further comprises an amino acid other than I at position 451, an amino acid other than S at position 454, and / or an amino acid other than G at position 455, as specified in SEQ ID NO: 52 or 138.

[0508] 430. The AAV particle of any one of embodiments 415, 416, or 429, wherein the AAV capsid variant further comprises V at position 451, H at position 454, and / or D at position 455, as specified in SEQ ID NO: 52 or 138.

[0509] 431. The AAV particle of any one of embodiments 415, 416, 429, or 430, wherein the AAV capsid variant further comprises substitutions I451V, S454H, and / or G455D, as specified in SEQ ID NO: 52 or 138.

[0510] 432.AAV particles as described in any of embodiments 415, 416, or 429 to 431, wherein the AAV capsid variant comprises: (i) V at position 451, H at position 454, and / or D at position 455 according to SEQ ID NO: 52 or 138; and (ii) the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately following position 455 according to SEQ ID NO: 52 or 138.

[0511] 433. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412 or 414, wherein the AAV capsid variant contains the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the amino acid sequence is present immediately after position 453 of the amino acid sequence number according to SEQ ID NO: 138.

[0512] 434. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414 or 433, wherein the AAV capsid variant comprises the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the amino acid sequence is present immediately after position 453 of the amino acid sequence number according to SEQ ID NO: 982.

[0513] 435. As in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 toAAV particles of any one of 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, 395, 407, 409 to 412, 414, 433 or 434, wherein the AAV capsid variant comprises the amino acid sequence of SPHKSG (SEQ ID NO: 946), wherein the amino acid sequence is present immediately after position 455 of the amino acid sequence number according to SEQ ID NO: 982.

[0514] 436. An AAV particle of any one of embodiments 369 to 435, wherein the AAV capsid variant comprises: (i) an amino acid sequence of HDSPHSKA (SEQ ID NO: 4486) immediately following position 453; and (ii) a deletion of amino acid SG at positions 454 and 455; wherein (i) and (ii) are numbered according to SEQ ID NO: 138.

[0515] 437. An AAV particle of any one of embodiments 369 to 436, wherein the AAV capsid variant comprises amino acid HD at positions 454 and 455 and further comprises an amino acid sequence of SPHSKA (SEQ ID NO: 941) immediately following position 455, these positions being numbered according to SEQ ID NO: 138.

[0516] 438. An AAV particle of any one of embodiments 433 to 435, wherein the AAV capsid variant further comprises an amino acid other than T at position 450, an amino acid other than I at position 451, and an amino acid other than N at position 452, as specified in SEQ ID NO: 138 or 982.

[0517] 439. An AAV particle of any one of embodiments 433 to 435 or 438, wherein the AAV capsid variant further comprises A at position 450, E at position 451, and I at position 452, as specified in SEQ ID NO: 138 or 982.

[0518] 440. An AAV particle of any one of embodiments 433 to 435, 438, or 439, wherein the AAV capsid variant further comprises substituted T450A, I451E, and N452I, as specified in SEQ ID NO: 138 or 982.

[0519] 441. An AAV particle as described in any one of embodiments 433, 434, or 438 to 440, wherein the AAV capsid variant comprises: (i) according to SEQ ID NO:The amino acid sequence of HDSPHK (SEQ ID NO: 2) is present at position 450, position 451, and position 452 of specification page 64 / 285, 74 CN 121079310 A; and (ii) the amino acid sequence of HDSPHK immediately following position 453 of SEQ ID NO: 138 or 982.

[0520] 442. An AAV particle as described in any one of embodiments 1 to 22, 25 to 27, 31, 34 to 42, 45 to 50, 53 to 63, 69, 79, 83 to 86, 91 to 98, 102, 103, 110, 111, 118 to 129, 369 to 371, 384, 385, 390, 393, 395, 410 to 413, wherein the AAV capsid variant comprises the amino acid sequence of SPHKYG (SEQ ID NO: 966), wherein the amino acid sequence is present immediately after position 455 of the amino acid sequence number according to SEQ ID NO: 138.

[0521] 443. An adeno-associated virus (AAV) particle comprising an AAV capsid variant comprising an amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the amino acid sequence is present immediately after position 453 of the amino acid sequence number according to SEQ ID NO: 982, wherein the AAV particle further comprises nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein).

[0522] 444. An adeno-associated virus (AAV) particle comprising an AAV capsid variant and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after position 455 of the amino acid sequence number according to SEQ ID NO: 981.

[0523] 445. An adeno-associated virus (AAV) particle comprising an AAV capsid variant containing the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the amino acid sequence is present immediately following position 453 of the amino acid sequence numbered according to SEQ ID NO: 37, and optionally further comprising: (i) one, both or all of the following amino acids, excluding T, at position 450, excluding I, at position 541, and / or excluding N, at position 452; (ii) one, both or all of the following amino acids, excluding A, at position 450, E, and / or I, at position 452, according to SEQ ID NO: 138 or 37.The AAV particle further comprises nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein).

[0524] 446. An adeno-associated virus (AAV) particle comprising an AAV capsid variant comprising an amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately following position 455 of an amino acid sequence numbered according to any one of SEQ ID NO: 36, 38-55, 57 or 59, wherein the AAV particle further comprises nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein).

[0525] 447. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant further comprises: (i) modifications in rings I, II, VI and / or VIII; and / or (ii) a substitution at position K449 according to SEQ ID NO: 138, for example, a K449R substitution.

[0526] 448. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three modifications, such as substitutions (e.g., conservative substitutions), but not exceeding 30, 20, or 10 modifications, such as substitutions (e.g., conservative substitutions), relative to the amino acid sequence of SEQ ID NO: 138.

[0527] 449. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three, but not exceeding 30, 20, or 10 different amino acids, relative to the amino acid sequence of SEQ ID NO: 138.

[0528] 450. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with SEQ ID NO: 138.

[0529] 451. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises an amino acid sequence having at least 98% identity with SEQ ID NO: 138.

[0530] 452. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%) sequence identity with SEQ ID NO: 137.453. An AAV particle as described in any of the foregoing embodiments, wherein the nucleotide sequence encoding the capsid variant comprises a sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with SEQ ID NO: 137.

[0532] 454. An AAV particle as described in any of the foregoing embodiments, wherein the AAV capsid variant comprises VP1 protein, VP2 protein, VP3 protein, or a combination thereof.

[0533] 455. An AAV particle as described in any one of embodiments 1 to 454, wherein the AAV capsid variant comprises an amino acid sequence corresponding to positions 138-742 (e.g., VP2) of SEQ ID NO: 981, 982, 36, or 4, or a sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0534] 456. An AAV particle as described in any one of embodiments 1 to 455, wherein the AAV capsid variant comprises an amino acid sequence corresponding to positions 203-742 (e.g., VP3) of SEQ ID NO: 981, 982, 36, or 4, or a sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0535] 457. An AAV particle as described in any one of embodiments 1 to 456, wherein the AAV capsid variant comprises an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with SEQ ID NO: 138.

[0536] 458. An AAV particle as described in any one of embodiments 1 to 457, wherein the AAV capsid variant comprises an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with SEQ ID NO: 138.

[0537] 459. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 toAAV particles of any one of 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, 410 to 413, 415 to 432, 444, or 446 to 458, wherein the AAV capsid variant comprises an amino acid sequence comprising at least 3, at least 4, at least 5, or at least 6 consecutive amino acids from the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein: (i) at least 3 consecutive amino acids comprise SPH; (ii) at least 4 consecutive amino acids comprise SPHS (SEQ ID NO: 4700); (iii) at least 5 consecutive amino acids comprise SPHSK (SEQ ID NO: 4701); or (iv) at least 6 consecutive amino acids comprise SPHSKA (SEQ ID NO: 4700). 941); wherein the AAV capsid variant comprises: (a) a VP1 protein containing the amino acid sequence of SEQ ID NO: 981; (b) a VP2 protein containing the amino acid sequence of positions 138-742 of SEQ ID NO: 981; (c) a VP3 protein containing the amino acid sequence of positions 203-742 of SEQ ID NO: 981; or (d) an amino acid sequence having at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with any of the amino acid sequences in (a)-(c).

[0538] 460. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, AAV particles of any one of 410 to 413, 415 to 432, 444, or 446 to 459, wherein the AAV capsid variant comprises an amino acid sequence comprising at least 3, at least 4, at least 5, or at least 6 consecutive amino acids from the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein: (i) at least 3 consecutive amino acids comprise SPH; (ii) at least 4 consecutive amino acids comprise SPHS (SEQ ID NO: 4700);(iii) At least 5 consecutive amino acids contain SPHSK (SEQ ID NO: 4701); or (iv) at least 6 consecutive amino acids contain SPHSKA (SEQ ID NO: 941); wherein the AAV capsid variant contains at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) of the amino acid sequence of SEQ ID NO: 981.

[0539] 461. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, AAV particles of any one of 410 to 413, 415 to 432, 444, or 446 to 460, wherein the AAV capsid variant comprises one or two, but no more than three, substitutions relative to the amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the AAV capsid variant comprises: (a) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 981; (b) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 981; (c) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 981; or (d) an amino acid sequence having at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with any one of the amino acid sequences in (a)-(c).

[0540] 462. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, AAV particles of any one of 410 to 413, 415 to 432, 444, or 446 to 461, wherein the AAV capsid variant contains an amino acid sequence relative to SPHSKA (SEQ ID NO:941) One or two, but no more than three, substitutions, wherein the AAV capsid variant comprises at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) amino acid sequence identical to the amino acid sequence of SEQ ID NO: 138 or SEQ ID NO: 981.

[0541] 463. An AAV particle as described in any one of embodiments 459 to 462, wherein the amino acid sequence is present immediately following position 455 according to the numbering in SEQ ID NO: 138 or 981.

[0542] 464. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, or 447 to 458, wherein the AAV capsid variant comprises an amino acid sequence comprising at least 3, at least 4, at least 5, or at least 6 consecutive amino acids from the amino acid sequence of HDSPHK (SEQ ID NO: 2, page 67 / 285, CN 121079310 A NO: 2), wherein: (i) at least 3 consecutive amino acids comprise HDS; (ii) at least 4 consecutive amino acids comprise HDSP (SEQ ID NO: 4702); (iii) at least 5 consecutive amino acids comprise HDSPH (SEQ ID NO: 4703); or (iv) at least 6 consecutive amino acids comprise HDSPHK (SEQ ID NO: 2); wherein the AAV capsid variant comprises: (a) a VP1 protein comprising SEQ ID NO: (a) an amino acid sequence of SEQ ID NO: 982; (b) a VP2 protein comprising the amino acid sequence at positions 138-742 of SEQ ID NO: 982; (c) a VP3 protein comprising the amino acid sequence at positions 203-742 of SEQ ID NO: 982; or (d) an amino acid sequence having at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with any of the amino acid sequences in (a)-(c).

[0543] 465.Implementation plans 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392. AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, or 464, wherein the AAV capsid variant comprises an amino acid sequence comprising at least 3, at least 4, at least 5, or at least 6 consecutive amino acids from the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein: (i) at least 3 consecutive amino acids comprise HDS; (ii) at least 4 consecutive amino acids comprise HDSP (SEQ ID NO: 4702); (iii) at least 5 consecutive amino acids comprise HDSPH (SEQ ID NO: 4703); or (iv) at least 6 consecutive amino acids comprise HDSPHK (SEQ ID NO: 2); wherein the AAV capsid variant comprises at least 90% of the amino acid sequence of SEQ ID NO: 982. (For example, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) the same amino acid sequence.

[0544] 466. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, 464, or 465, wherein the AAV capsid variant comprises one or two, but no more than three, substitutions relative to the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the AAV capsid variant comprises: (a) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982; (b) a VP2 protein comprising the amino acid sequence of SEQ ID NO: 982.(a) The amino acid sequence at positions 138 to 742 of SEQ ID NO: 982; (c) The VP3 protein comprising the amino acid sequence at positions 203 to 742 of SEQ ID NO: 982; or (d) An amino acid sequence having at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with any of the amino acid sequences in (a) to (c).

[0545] 467. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458 or 464 to 466, wherein the AAV capsid variant comprises one or two but no more than three substitutions relative to the amino acid sequence of HDSPHK (SEQ ID NO: 2), wherein the AAV capsid variant comprises at least 90% (e.g., at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) the same amino acid sequence as SEQ ID NO: 982.

[0546] 468. AAV particles of any one of embodiments 464 to 468, wherein the amino acid sequence is present immediately after position 453 according to the numbering of SEQ ID NO: 138 or 982.

[0547] 469. An AAV particle of any one of embodiments 1 to 468, wherein the AAV capsid variant comprises an amino acid sequence of SEQ ID NO: 981 or 982 or an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0548] 470. An AAV particle of any one of embodiments 1 to 469, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three modifications relative to the amino acid sequence of SEQ ID NO: 981 or 982, such as substitutions (e.g., conservative substitutions) but not more than 30, not more than 20, or not more than 10 modifications, such as substitutions (e.g., conservative substitutions).

[0549] 471. An AAV particle as described in any one of embodiments 1 to 470, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but no more than 30, no more than 20, or no more than 10 different amino acids relative to the amino acid sequence of SEQ ID NO: 981 or 982.

[0550] 472. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, AAV particles of any one of 410 to 413, 415 to 432, 444, 446 to 463 or 469 to 471, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 981 or an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0551] 473. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, AAV particles of any one of 410 to 413, 415 to 432, 444, 446 to 463, or 469 to 472, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three modifications, such as substitutions (e.g., conservative substitutions), but not more than 30, 20, or 10 modifications, such as substitutions (e.g., conservative substitutions), relative to the amino acid sequence of SEQ ID NO: 981.

[0552] 474. As in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99,AAV particles of any one of 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 240 to 247, 249, 250, 253 to 263, 266, 272 to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, 410 to 413, 415 to 432, 444, 446 to 463, or 469 to 473, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two, or at least three but not more than 30, not more than 20, or not more than 10 different amino acids relative to the amino acid sequence of SEQ ID NO: 981.

[0553] 475. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458 or 464 to 471, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it. Instruction manual, pages 69 / 285, CN 121079310 A

[0554] 476. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, Any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, 464 to 471 or 475AAV particles, wherein the AAV capsid variant comprises an amino acid sequence containing at least one, at least two, or at least three modifications, such as substitutions (e.g., conservative substitutions), relative to the amino acid sequence of SEQ ID NO: 982, but not more than 30, not more than 20, or not more than 10 modifications, such as substitutions (e.g., conservative substitutions).

[0555] 477. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, 464 to 471, 475 or 476, wherein the AAV capsid variant comprises an amino acid sequence comprising at least one, at least two or at least three but no more than 30, no more than 20 or no more than 10 different amino acids relative to the amino acid sequence of SEQ ID NO: 982.

[0556] 478. AAV particles of any one of embodiments 1 to 477, wherein the AAV capsid variant comprises an amino acid sequence encoded by a nucleotide sequence of SEQ ID NO: 983 or 984 or a nucleotide sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%) sequence identity with it.

[0557] 479. An AAV particle as described in any one of embodiments 1 to 478, wherein the nucleotide sequence encoding the capsid variant comprises the nucleotide sequence of SEQ ID NO: 983 or 984, or a nucleotide sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0558] 480. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 02, 240 to 247, 249, 250, 253 to 263, 266, 272AAV particles of any one of SEQ ID NO: 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406, 408, 410 to 413, 415 to 432, 444, 446 to 463, 469 to 474, 478, or 479, wherein the nucleotide sequence encoding the capsid variant comprises the nucleotide sequence of SEQ ID NO: 983 or a nucleotide sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0559] 481. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, 464 to 471 or 475 to 479, wherein the nucleotide sequence encoding the capsid variant comprises the nucleotide sequence of SEQ ID NO: 984 or a nucleotide sequence having at least 80% (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%) sequence identity with it.

[0560] 482. AAV particles of any of the foregoing embodiments, wherein the nucleotide sequence encoding the capsid variant is codon-optimized.

[0561] 483. As described in embodiments 1 to 23, 26 to 29, 32, 35 to 43, 46 to 51, 54 to 72, 69 to 89, 91, 94 to 99, 102 to 104, 107, 110 to 112, 115 to 129, 168 to 202, 02, 240 to 247, 249, 250, 253 to 263, 266, 272, specification 70 / 285 pages 80 CN 121079310 A to 281, 286, 288, 291 to 299, 327 to 363, 369 to 371, 375 to 379, 385, 386, 390, 391, 395, 406,408, 410 to 413, 415 to 432, 444, 446 to 463, 469 to 474, 478 to 480 or 482, and further comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 981.

[0562] 484. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein), wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 981.

[0563] 485. An AAV particle as described in embodiment 483 or 484, wherein the nucleotide sequence encoding the AAV capsid variant comprises the nucleotide sequence of SEQ ID NO: 983 or a nucleotide sequence that is at least 90%, at least 95%, or at least 99% identical thereto.

[0564] 486. As described in embodiments 1 to 9, 11, 12 to 22, 24, 26, 28, 30, 33, 35 to 42, 44, 46 to 50, 52, 54 to 77, 83 to 88, 90 to 97, 100 to 103, 105, 110, 111, 113 to 129, 203 to 248, 251, 252, 254 to 257, 260, 261, 264 to 280, 283 to 287, 289 to 299, 327 to 363, 369, 369, 371, 380 to 383, 385, 386, 390, 392, AAV particles of any one of 395, 407, 409 to 412, 414, 433 to 435, 438 to 441, 443, 445, 447 to 458, 464 to 471, 475 to 479, 481 or 482, and further comprising an amino acid sequence that is at least 95% identical to that of SEQ ID NO: 982.

[0565] 487. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein), wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 982.

[0566] 488. An AAV particle as described in embodiment 486 or 487, wherein the nucleotide sequence encoding an AAV capsid variant comprises the nucleotide sequence of SEQ ID NO: 984 or a nucleotide sequence that is at least 90%, at least 95%, or at least 99% identical thereto.

[0567] 489. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., an AAV9 capsid variant) and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein), wherein the AAV capsid...The variant comprises an amino acid sequence encoded by the nucleotide sequence of SEQ ID NO: 983 or 984 or a nucleotide sequence that is at least 95% identical thereto.

[0568] 490. An adeno-associated virus (AAV) particle comprising an AAV capsid variant (e.g., the AAV9 capsid variant) and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., the human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence of any one of SEQ ID NO: 4 or 36-59, optionally wherein the AAV capsid variant comprises an amino acid sequence of SEQ ID NO: 4 or 36.

[0569] 491. An adeno-associated virus (AAV) particle comprising an AAV capsid variant and nucleic acid encoding a cyclin-dependent kinase-like 5 (CDKL5) protein (e.g., human CDKL5 protein), wherein the AAV capsid variant comprises an amino acid sequence encoded by a nucleotide sequence of any one of SEQ ID NO: 12-35 or a nucleotide sequence that is at least 95% identical thereto.

[0570] 492. The AAV particle of embodiment 490 or 491, wherein the nucleotide sequence encoding the AAV capsid variant comprises a nucleotide sequence of any one of SEQ ID NO: 12-35 or a nucleotide sequence that is at least 95% identical thereto.

[0571] 493. The AAV particle of any one of embodiments 1 to 299, 369 to 371 or 375 to 492, having increased tropism to CNS cells or tissues, such as brain cells, brain tissue, spinal cord cells or spinal cord tissue, relative to the tropism of AAV particles comprising a capsid containing the amino acid sequence of SEQ ID NO: 138.

[0572] 494. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371 or 375 to 493, which transduce brain regions, such as midbrain regions (e.g., hippocampus or thalamus) or brainstem, optionally wherein, for example when measured by means of an assay, such as immunohistochemistry or qPCR, such as as described in Example 2, the transduction level is at least 5 times, at least 10 times, at least 15 times, at least 20 times, at least 25 times, at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 55 times, at least 60 times or at least 65 times that of AAV particles containing the capsid of SEQ ID NO: 138. Instruction manual, pages 71 / 285, CN 121079310 A

[0573] 495. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371 or 375 to 494, which transduce brain regions, such as midbrain regions (e.g., hippocampus or thalamus) or brainstem, optionally, wherein, for example, when measured, for exampleWhen measured by immunohistochemical assay or qPCR assay, for example as described in Example 2, the transduction level is at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 55 times, at least 60 times, or at least 65 times that of AAV particles containing the capsid of SEQ ID NO: 138.

[0574] 496. AAV particles as described in any one of Embodiments 1 to 129, 168 to 299, 369 to 371, or 375 to 495, when measured by the assay described in Example 1, for example, the AAV particles are enriched in the brain at at least 3 times, at least 4 times, at least 5 times, at least 6 times, at least 7 times, at least 8 times, at least 9 times, or at least 10 times that of AAV particles containing the capsid of SEQ ID NO: 138.

[0575] 497. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371 or 375 to 496, for example, when measured by means of the determination described in Example 1, are enriched in the brain at least 20 times, at least 25 times, at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 55 times, at least 60 times, at least 65 times, at least 70 times, at least 75 times, at least 80 times or at least 85 times.

[0576] 498. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371 or 375 to 497, for example, compared to AAV particles containing the capsid of SEQ ID NO: 138, are enriched in the brains of at least two to at least three species (e.g., non-human primates and rodents (e.g., mice)).

[0577] 499. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371, or 375 to 498, when measured by means of the determination described in Example 1 or 5, wherein the AAV particles are enriched in the brains of at least two to at least three species (e.g., non-human primates and rodents (e.g., mice)) at at least 10 times, at least 15 times, at least 20 times, at least 25 times, at least 30 times, at least 35 times, at least 40 times, at least 45 times, at least 50 times, at least 55 times, at least 60 times, at least 65 times, at least 70 times, at least 75 times, at least 80 times, at least 85 times, at least 90 times, at least 95 times, at least 100 times, at least 105 times, at least 115 times, at least 120 times, at least 125 times, at least 130 times, or at least 130 times the amount of AAV particles containing the capsid of SEQ ID NO: 138. At least 135 times, at least 140 times, at least 145 times, at least 150 times, at least 155 times, at least 160 times, at least 165 times, at least170 times, at least 175 times, at least 180 times, at least 190 times, at least 200 times, at least 205 times, or at least 210 times.

[0578] 500. AAV particles as described in embodiment 498 or 499, wherein at least two to at least three species are cynomolgus monkeys (Macaca fascicularis), green monkeys (Chlorocebus sabaeus), common marmosets (Callithrix jacchus) and / or mice (e.g., BALB / c mice, C57Bl / 6 mice, and / or CD-1 distantly related hybrid mice).

[0579] 501. AAV particles as described in any one of embodiments 130 to 146, 369, 410 to 414, 447 to 454, 457, 458, 482 or 493, for example, when measured by means of the determination described in Example 3, are enriched in the brain at at least 2 times, at least 2.5 times, at least 3 times, at least 3.5 times, at least 4 times, at least 4.5 times, at least 5 times, at least 5.5 times, at least 6 times, at least 6.5 times, at least 7 times, at least 7.5 times or at least 8 times.

[0580] 502. AAV particles as described in any one of embodiments 147 to 167, 369, 410 to 414, 447 to 454, 457, 458, 482 or 493, for example, when measured by means of the determination described in Example 3, are enriched in the brain at at least 2 times, at least 2.5 times, at least 3 times, at least 3.5 times, at least 4 times, at least 4.5 times, at least 5 times or at least 5.5 times the amount of AAV particles containing the capsid of SEQ ID NO: 982.

[0581] 503. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371 or 375 to 500, when measured by assay, such as qRT-PCR or qPCR assay (e.g., as described in Example 2 or 8), deliver an increased level of payload to the brain region, optionally wherein the level of payload is increased by at least 10-fold, at least 12-fold, at least 15-fold, at least 20-fold, at least 25-fold, at least 30-fold, at least 35-fold, at least 40-fold, at least 45-fold, at least 50-fold, at least 55-fold, at least 60-fold, at least 65-fold or at least 70-fold.

[0582] 504. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500 or 503The AAV particle, when measured by assays such as qRT-PCR or qPCR assays (e.g., as described in Examples 2 or 8), delivers an increased level of viral genome to a brain region, optionally wherein the level of viral genome is increased by at least 5-fold, at least 10-fold, at least 15-fold, at least 17-fold, at least 18-fold, at least 19-fold, at least 20-fold, at least 25-fold, at least 30-fold, at least 35-fold, at least 40-fold, at least 45-fold, or at least 50-fold compared to an AAV particle containing the capsid of SEQ ID NO: 138.

[0583] 505. The AAV particle of embodiment 503 or 504, wherein the brain region is a midbrain region (e.g., hippocampus or thalamus), frontal cortex, temporal cortex, motor cortex, cerebral cortex, caudate nucleus, putamen, dentate nucleus, substantia nigra, or brainstem.

[0584] 506. AAV particles as described in any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500 or 503 to 505, for example, when measured by means of the determination described in Example 1 or 8, are enriched in the spinal cord at at least 4 times, at least 5 times, at least 10 times, at least 15 times, at least 20 times, at least 25 times, at least 30 times or at least 35 times the amount of AAV particles containing the capsid of SEQ ID NO: 138, optionally, wherein the region of the spinal cord is the thoracic spinal cord region, the cervical spinal cord region, the C5 ventral horn region, the lumbar spinal cord region or the L5 ventral horn region.

[0585] 507. AAV particles of any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500, or 503 to 506, showing preferential transduction in a brain region relative to transduction in the dorsal root ganglion (DRG).

[0586] 508. AAV particles of any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500, or 503 to 507, showing preferential transduction in a brain region relative to the liver.

[0587] 509. AAV particles of any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500, or 503 to 508, showing preferential transduction in a brain region relative to transduction in the heart.

[0588] 510. AAV particles of any one of embodiments 1 to 129, 168 to 299, 369 to 371, 375 to 500, or 503 to 509, which exhibit preferential transduction in brain regions relative to transduction in the dorsal root ganglion (DRG) and the heart.

[0589] 511. AAV particles of any one of the foregoing embodiments, which are capable of transducing non-neuronal cells, such as glial cells (e.g., oligodendrocytes or astrocytes).

[0590] 512.AAV particles as described in embodiment 511, wherein the non-neuronal cells comprise glial cells, oligodendrocytes (e.g., Olig2-positive oligodendrocytes), or astrocytes (e.g., Olig2-positive astrocytes).

[0591] 513. AAV particles as described in any of the preceding embodiments, which are capable of transducing Olig2-positive cells, such as Olig2-positive astrocytes or Olig2-positive oligodendrocytes.

[0592] 514. AAV particles as described in any of embodiments 369, 373, 447 to 454, 457, 458, or 482, which have increased tropism toward cardiac cells or tissues, such as ventricles or atria, relative to the tropism of AAV particles comprising the capsid of SEQ ID NO: 138.

[0593] 515. AAV particles as described in any one of embodiments 369, 373, 447 to 454, 457, 458, 482 or 514, when measured by means of the determination described in Example 4, are enriched in the heart by at least 4 times, at least 5 times, at least 8 times, at least 10 times, at least 11 times, at least 12 times, at least 13 times, at least 14 times, at least 18 times, at least 19 times, at least 20 times, at least 21 times, at least 22 times, at least 24 times, at least 25 times, at least 27 times, at least 31 times, at least 33 times or at least 34 times.

[0594] 516. The AAV particle of any one of embodiments 369, 374, 447 to 454, 457, 458, 482, having increased tropism to muscle cells or tissues (e.g., quadriceps femoris cells or quadriceps femoris tissue) relative to the tropism of AAV particles comprising a capsid containing the amino acid sequence of SEQ ID NO: 138. Instruction manual 73 / 285 pages 83 CN 121079310 A

[0595] 517. AAV particles as described in any one of embodiments 369, 374, 447 to 454, 457, 458, 482, for example, when measured by means of the determination described in Example 4, the AAV particles are enriched in muscle at least 4 times, at least 5 times, at least 8 times, at least 12 times, at least 17 times, at least 18 times, at least 20 times, at least 26 times, at least 27 times, at least 28 times, at least 30 times, or at least 36 times.

[0596] 518. AAV particles as described in embodiment 516 or 517, wherein the muscle cells or tissue are cardiac muscle (e.g., ventricle or atrium or both), quadriceps femoris or both.

[0597] 519. AAV particles as described in any of the foregoing embodiments, which are then separated and / or recombined.

[0598] [Implementation schemes 520 to 585 are intentionally absent.]

[0599] 586. An AAV particle as described in any of the foregoing implementation schemes, wherein the viral genome contains a promoter effectively linked to the CDKL5 coding sequence.

[0600] 587. The AAV particle of embodiment 586, wherein the promoter is a human elongation factor 1α-subunit (EF1α) promoter, a cytomegalovirus (CMV) immediate early enhancer and / or promoter, a chicken β-actin (CBA) promoter, a CAG promoter, a CAG derivative promoter, a β-glucuronidase (GUSB) promoter, a ubiquitin C (UBC) promoter, a neuron-specific enolase (NSE) promoter, a platelet-derived growth factor (PDGF) promoter, a platelet-derived growth factor B chain (PDGF-β) promoter, an intercellular adhesion molecule 2 (ICAM-2) promoter, a synaptic protein (Syn) promoter, a methyl-CpG binding protein 2 (MeCP2) promoter, a Ca2+ / calmodulin-dependent protein kinase II (CaMKII) promoter, or a metabolite glutamate receptor 2. (mGluR2) promoter, neurofilament light chain (NFL) or heavy chain (NFH) promoter, β-globin small gene nβ2 promoter, proenkephalinogenin (PPE) promoter, enkephalin (Enk) and excitatory amino acid transporter 2 (EAAT2) promoter, glial fibrillary acidic protein (GFAP) promoter, myelin basic protein (MBP) promoter, cardiovascular promoter (e.g., αMHC, cTnT and CMV-MLC2k), liver promoter (e.g., hAAT, TBG), skeletal muscle promoter (e.g., desmin, MCK, C512) or fragments thereof (e.g., truncated) or functional variants thereof.

[0601] [Embodiments 588 and 589 are intentionally absent.]

[0602] 590. An AAV particle as described in any one of embodiments 586 to 589, wherein the viral genome further comprises a polyadenylated (polyadenylated) sequence.

[0603] 591. An AAV particle as described in any one of embodiments 586 to 590, wherein the viral genome further comprises an inverted terminal repeat (ITR) sequence.

[0604] 592. An AAV particle as described in any one of embodiments 586 to 591, wherein the viral genome comprises an ITR sequence located at the 5' relative to a CDKL5 coding sequence (e.g., encoding a human CDKL5 protein).

[0605] 593. An AAV particle as described in any one of embodiments 586 to 592, wherein the viral genome comprises an ITR sequence located at the 3' relative to a CDKL5 coding sequence (e.g., encoding a human CDKL5 protein).

[0606] 594. An AAV particle as described in any one of embodiments 586 to 593, wherein the viral genome comprises an ITR sequence located at the 3' relative to a CDKL5 coding sequence (e.g., encoding a human CDKL5 protein).The 5' ITR sequence of the CDKL5 coding sequence (e.g., encoding the human CDKL5 protein) and the 3' ITR sequence relative to the CDKL5 coding sequence (e.g., encoding the human CDKL5 protein).

[0607] 595. An AAV particle as described in any one of embodiments 586 to 594, wherein the viral genome further comprises enhancers, Kozak sequences, intron regions and / or exon regions.

[0608] [Embodiments 596 to 615 are intentionally absent.]

[0609] 616. An AAV particle as described in any one of embodiments 586 to 595, wherein the viral genome further comprises a nucleotide sequence encoding a Rep protein, such as a non-structural protein, wherein the Rep protein comprises Rep78 protein, Rep68 protein, Rep52 protein and / or Rep40 protein (e.g., Rep78 and Rep52 proteins). Specification 74 / 285 pages 84 CN 121079310 A

[0610] 617. An AAV particle as described in embodiment 616, wherein the AAV particle further comprises a nucleotide sequence encoding a Rep protein, such as a non-structural protein, wherein the Rep protein comprises Rep78 protein, Rep68 protein, Rep52 protein and / or Rep40 protein (e.g., Rep78 and Rep52 proteins).

[0611] 618. An AAV particle as described in embodiment 616 or 617, wherein the Rep78 protein, Rep68 protein, Rep52 protein and / or Rep40 protein is encoded by at least one Rep gene.

[0612] 619. An AAV particle as described in any one of embodiments 586 to 618, wherein the viral genome further comprises a nucleic acid sequence encoding an AAV capsid variant of an AAV particle as described in any one of embodiments 1 to 519, 566 or 574.

[0613] 620. AAV particles as described in any one of embodiments 575 to 619, wherein the AAV particles are isolated and / or recombined AAV particles.

[0614] [Embodiment 621 is intentionally absent.]

[0615] 622. A cell, such as a host cell, comprising AAV particles as described in any of the preceding embodiments.

[0616] 623. A cell as described in embodiment 622, wherein the cell is a mammalian cell or an insect cell.

[0617] 624. A cell as described in embodiment 622 or 623, wherein the cell is a cell of a brain region or spinal cord region, optionally a cell of the brainstem, hippocampus, or thalamus.

[0618] 625. A cell as described in any one of embodiments 622 to 624, wherein the cell is a neuron, sensory neuron, motor neuron, astrocyte, glial cell, oligodendrocyte, or muscle cell (e.g., cells of the heart, diaphragm, or quadriceps femoris muscle).

[0619] [Scheme 626 is intentionally absent.]

[0620] 627. A method for preparing adeno-associated virus (AAV) particles, comprising

[0621] (i) providing a host cell containing a viral genome containing a CDKL5 coding sequence; and

[0622] (ii) culturing the host cell under conditions suitable for encapsulating the viral genome in an AAV capsid variant as described in any one of Schemes 1 to 620; thereby preparing the AAV particles.

[0623] 628. The method of Scheme 627, further comprising introducing a first nucleic acid molecule containing the viral genome into the host cell prior to step (i).

[0624] 629. The method of Scheme 628, wherein the host cell contains a second nucleic acid encoding a capsid variant.

[0625] 630. The method of Scheme 629, wherein the second nucleic acid molecule is introduced into the host cell before, simultaneously with, or after the first nucleic acid molecule.

[0626] 631. A pharmaceutical composition comprising AAV particles as described in any one of embodiments 1 to 620 and a pharmaceutically acceptable excipient.

[0627] 632. A method of delivering a payload to cells or tissues (e.g., CNS cells or CNS tissues) comprising administering an effective amount of the pharmaceutical composition as described in embodiment 631 or AAV particles as described in any one of embodiments 1 to 620.

[0628] 633. The method of embodiment 632, wherein the cells are cells of a brain region or spinal cord region, optionally cells of the frontal cortex, sensory cortex, motor cortex, caudate nucleus, cerebellar cortex, cerebral cortex, brainstem, hippocampus, or thalamus.

[0629] 634. The method of embodiment 632 or 633, wherein the cells are neurons, sensory neurons, motor neurons, astrocytes, glial cells, or oligodendrocytes.

[0630] [Embodiment 635 is intentionally absent.]

[0631] 636. The method of any one of embodiments 632 to 634, wherein the cell or tissue is within the subject.

[0632] 637. The method of embodiment 636, wherein the subject has, has been diagnosed with, a genetic condition, such as a monogenic or polygenic disease, or is at risk of developing a genetic condition, such as a monogenic or polygenic disease.

[0633] 638. The method of embodiment 636 or 637, wherein the subject has, has been diagnosed with, a neurological condition, such as a neurodegenerative disease, or is at risk of developing a neurological condition, such as a neurodegenerative disease.

[0634] [Embodiment 639 is intentionally absent.]

[0635] 640.The method ...

Claims

1. An adeno-associated virus (AAV) particle comprising: a) an AAV capsid variant comprising an amino acid sequence having the formula: [N1]-[N2]-[N3], wherein: (i) optionally, [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G; (ii) [N2] comprises the amino acid sequence of SPH; and (iii) [N3] comprises X4, X5, and X6, wherein at least one of X4, X5, or X6 is a basic amino acid; and b) a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence.

2. The AAV particle of claim 1, wherein the amino acid sequence [N1]-[N2]-[N3] is located in hypervariable loop IV of the AAV capsid variant.

3. The AAV particle of claim 1 or claim 2, wherein the AAV capsid variant is an AAV9 capsid variant.

4. The AAV particle of any one of claims 1 to 3, wherein [N1] comprises X1, X2, and X3, wherein at least one of X1, X2, or X3 is G.

5. The AAV particle of any one of claims 1 to 4, wherein [N2]-[N3] comprises the amino acid sequence of SPHSKA (SEQ ID NO: 941).

6. An adeno-associated virus (AAV) particle comprising a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence and an AAV9 capsid variant comprising the amino acid sequence of SPHSKA (SEQ ID NO: 941).

7. The AAV particle of claim 6, wherein the amino acid sequence of SPHSKA (SEQ ID NO: 941) is located in hypervariable loop IV of the AAV9 capsid variant.

8. The AAV particle of claim 6 or claim 7, wherein the amino acid sequence of SPHSKA (SEQ ID NO: 941) is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 4 or SEQ ID NO:

36.

9. The AAV particle of any one of claims 6 to 8, wherein the AAV9 capsid variant further comprises one, two, or all of: N at the amino acid position corresponding to position 452 of SEQ ID NO: 4, E at the amino acid position corresponding to position 451 of SEQ ID NO: 4, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO:

4.

10. The AAV particle of any one of claims 6 to 9, wherein the AAV9 capsid variant comprises the amino acid sequence of KTENVSGSPHSKAQNQQT (SEQ ID NO: 3272).

11. The AAV particle of any one of claims 6 to 10, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 90% identical to SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence that is at least 90% identical to positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 90% identical to positions 203-742 of SEQ ID NO:

4.

12. The AAV particle of any one of claims 6-11, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence that is at least 95% identical to positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 95% identical to positions 203-742 of SEQ ID NO:

4.

13. The AAV particle of any one of claims 6-12, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 99% identical to SEQ ID NO: 4; (ii) a VP2 protein comprising an amino acid sequence that is at least 99% identical to positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 99% identical to positions 203-742 of SEQ ID NO:

4.

14. The AAV particle of any one of claims 6-13, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

4.

15. The AAV particle of any one of claims 6-13, wherein the AAV9 capsid variant comprises: (i) an amino acid sequence of SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 4; (ii) an E at the amino acid position corresponding to position 451 of SEQ ID NO: 4 and a V at the amino acid position corresponding to position 453 of SEQ ID NO: 4; and (iii) no other modifications relative to wild-type AAV9.

16. The AAV particle of any one of claims 6-8, wherein the AAV9 capsid variant further comprises one, two, or all of: E at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and / or V at the amino acid position corresponding to position 453 of SEQ ID NO:

36.

17. The AAV particle of any one of claims 6-8 and 16, wherein the AAV9 capsid variant comprises the amino acid sequence of KTERVSGSPHSKAQNQQT (SEQ ID NO: 3589).

18. The AAV particle of any one of claims 6-8, 16, and 17, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 90% identical to SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence that is at least 90% identical to positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 90% identical to positions 203-742 of SEQ ID NO:

36.

19. The AAV particle of any one of claims 6-8 and 16-18, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 95% identical to SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence that is at least 95% identical to positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 95% identical to positions 203-742 of SEQ ID NO:

36.

20. The AAV particle of any one of claims 6-8 and 16-19, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence that is at least 99% identical to SEQ ID NO: 36; (ii) a VP2 protein comprising an amino acid sequence that is at least 99% identical to positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising an amino acid sequence that is at least 99% identical to positions 203-742 of SEQ ID NO:

36.

21. The AAV particle of any one of claims 6-8 and 16-20, wherein the AAV9 capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

36. (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

36.

22. The AAV particle of any one of claims 6-8 and 16-20, wherein the AAV9 capsid variant comprises: (i) the amino acid sequence SPHSKA (SEQ ID NO: 941), wherein the amino acid sequence is present immediately after the amino acid position corresponding to position 455 of SEQ ID NO: 36; (ii) E at the amino acid position corresponding to position 451 of SEQ ID NO: 36, R at the amino acid position corresponding to position 452 of SEQ ID NO: 36, and V at the amino acid position corresponding to position 453 of SEQ ID NO: 36; and (iii) no other modifications relative to wild-type AAV9.

23. The AAV particle of any one of claims 1-4, wherein [N1]-[N2]-[N3] is present immediately after the position corresponding to amino acid position 452 of SEQ ID NO: 982; and wherein the AAV capsid variant comprises an amino acid sequence that is at least 90% identical, e.g., at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical to the amino acid sequence of SEQ ID NO: 982, e.g., to positions 203-742 of SEQ ID NO:

982.

24. The AAV particle of claim 23, wherein [N1] comprises GHD.

25. The AAV particle of claim 23 or claim 24, wherein [N1] comprises the amino acid G at the position corresponding to position 453 of SEQ ID NO: 138 or SEQ ID NO: 982, the amino acid H at position 454 of SEQ ID NO: 138 or SEQ ID NO: 982, and the amino acid D at position 455.

26. The AAV particle of any one of claims 23-25, wherein [N3] comprises KSG.

27. The AAV particle of any one of claims 23-26, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence that is at least 90% identical to SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982 or an amino acid sequence that is at least 90% identical to positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 982 or an amino acid sequence that is at least 90% identical to positions 203-742 of SEQ ID NO:

982.

28. The AAV particle of any one of claims 23-27, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence at least 95% identical to SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982 or an amino acid sequence at least 95% identical to positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 982 or an amino acid sequence at least 95% identical to positions 203-742 of SEQ ID NO:

982.

29. The AAV particle of any one of claims 23-28, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982 or an amino acid sequence at least 99% identical to SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982 or an amino acid sequence at least 99% identical to positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 982 or an amino acid sequence at least 99% identical to positions 203-742 of SEQ ID NO:

982.

30. The AAV particle of any one of claims 23-29, wherein the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 982; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

982.

31. The AAV particle of any one of claims 1-30, wherein the viral genome encodes a wild-type CDKL5 protein or a fragment thereof.

32. The AAV particle of any one of claims 1-31, wherein the viral genome encodes a human CDKL5 protein.

33. The AAV particle of claim 31 or claim 32, wherein the CDKL5 protein comprises the amino acid sequence of SEQ ID NO: 6413.

34. The AAV particle of any one of claims 1 to 33, wherein the CDKL5-encoding sequence comprises a nucleotide sequence that is at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identical) to SEQ ID NO: 6414.

35. The AAV particle of claim 34, wherein the CDKL5-encoding sequence comprises a nucleotide sequence that is at least 95% identical to SEQ ID NO: 6414.

36. The AAV particle of claim 35, wherein the CDKL5-encoding sequence comprises a nucleotide sequence that is at least 99% identical to SEQ ID NO: 6414.

37. The AAV particle of claim 36, wherein the CDKL5-encoding sequence comprises the nucleotide sequence of SEQ ID NO: 6414.

38. The AAV particle of claim 37, wherein the CDKL5-encoding sequence consists of the nucleotide sequence of SEQ ID NO: 6414.

39. The AAV particle of any one of claims 1 to 38, wherein the viral genome comprises a promoter operatively linked to the CDKL5-encoding sequence.

40. The AAV particle of claim 39, wherein the promoter is a human elongation factor 1 alpha-subunit (EF1a), cytomegalovirus (CMV) immediate early enhancer and / or promoter, chicken beta-actin (CBA) promoter, CAG promoter, CAG derivative promoter, beta glucuronidase (GUSB) promoter, ubiquitin C (UBC) promoter, neuron-specific enolase (NSE) promoter, platelet-derived growth factor (PDGF) promoter, platelet-derived growth factor B chain (PDGF-beta) promoter, intercellular adhesion molecule 2 (ICAM-2) promoter, synapsin (Syn) promoter, synapsin 1 (Synl) promoter, methyl-CpG binding protein 2 (MeCP2) promoter, Ca2+ / calmodulin-dependent protein kinase II (CaMKII) promoter, metabotropic glutamate receptor 2 (mGluR2) promoter, neurofilament light chain (NFL) or heavy chain (NFH) promoter, beta-globin minigene nB2 promoter, preproenkephalin (PPE) promoter, enkephalin (Enk) and excitatory amino acid transporter 2 (EAAT2) promoter, glial fibrillary acidic protein (GFAP) promoter, myelin basic protein (MBP) promoter, cardiovascular promoters (e.g., aMHC, cTnT, and CMV-MLC2k), liver promoters (e.g., hAAT, TBG), skeletal muscle promoters (e.g., desmin, MCK, C512), or fragments thereof, e.g., truncations or functional variants.

41. The AAV particle of any one of claims 1 to 40, wherein the viral genome further comprises an inverted terminal repeat (ITR) sequence.

42. The AAV particle of claim 41, wherein the viral genome comprises an ITR sequence 5’ relative to the CDKL5 coding sequence.

43. The AAV particle of claim 41 or claim 42, wherein the viral genome comprises an ITR sequence 3’ relative to the CDKL5 coding sequence.

44. The AAV particle of any one of claims 41 to 43, wherein the viral genome comprises an ITR sequence 5’ relative to the CDKL5 coding sequence, and an ITR sequence 3’ relative to the CDKL5 coding sequence.

45. An adeno-associated virus (AAV) particle comprising a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence, and an AAV capsid variant, the AAV capsid variant comprising: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 4; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

4.

46. An adeno-associated virus (AAV) particle comprising a viral genome comprising a cyclin-dependent kinase-like 5 (CDKL5) coding sequence, and an AAV capsid variant, the AAV capsid variant comprising: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36; and / or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO:

36.

47. A cell comprising the AAV particle of any one of claims 1 to 46, optionally wherein the cell is a mammalian cell (e.g., a HEK293 cell), an insect cell (e.g., a Sf9 cell), or a bacterial cell.

48. A method of making the AAV particle of any one of claims 1 to 46, the method comprising: (i) providing a cell comprising a viral genome comprising a CDKL5 coding sequence and a nucleic acid encoding the AAV capsid variant; and (ii) incubating the cell under conditions suitable for encapsidation of the viral genome in the AAV capsid variant; thereby making the AAV particle.

49. The method of claim 48, wherein: (a) the viral genome comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) thereto; and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 4 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 4; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 4 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 138-742 of SEQ ID NO: 4; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 4 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 203-742 of SEQ ID NO:

4.

50. The method of claim 49, wherein the AAV capsid variant comprises the amino acid sequence of SEQ ID NO: 4, the amino acid sequence of positions 138-742 of SEQ ID NO: 4, and / or the amino acid sequence of positions 203-742 of SEQ ID NO:

4.

51. The method of claim 48, wherein: (a) the viral genome comprises the nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) thereto; and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising the amino acid sequence of SEQ ID NO: 36 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 36; (ii) a VP2 protein comprising the amino acid sequence of positions 138-742 of SEQ ID NO: 36 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 138-742 of SEQ ID NO: 36; or (iii) a VP3 protein comprising the amino acid sequence of positions 203-742 of SEQ ID NO: 36 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 203-742 of SEQ ID NO:

36. (iii) a VP3 protein comprising an amino acid sequence of positions 203-742 of SEQ ID NO: 36 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 203-742 of SEQ ID NO:

36.

52. The method of claim 51, wherein the AAV capsid variant comprises an amino acid sequence of SEQ ID NO: 36, an amino acid sequence of positions 138-742 of SEQ ID NO: 36, and / or an amino acid sequence of positions 203-742 of SEQ ID NO:

36.

53. The method of claim 48, wherein: (a) the viral genome comprises a nucleic acid sequence of SEQ ID NO: 6414 or a nucleic acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) thereto; and (b) the AAV capsid variant comprises: (i) a VP1 protein comprising an amino acid sequence of SEQ ID NO: 982 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to SEQ ID NO: 982; (ii) a VP2 protein comprising an amino acid sequence of positions 138-742 of SEQ ID NO: 982 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 138-742 of SEQ ID NO: 982; or (iii) a VP3 protein comprising an amino acid sequence of positions 203-742 of SEQ ID NO: 982 or an amino acid sequence at least 90% identical (e.g., at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical) to positions 203-742 of SEQ ID NO:

982.

54. The method of claim 53, wherein the AAV capsid variant comprises an amino acid sequence of SEQ ID NO: 982, an amino acid sequence of positions 138-742 of SEQ ID NO: 982, and / or an amino acid sequence of positions 203-742 of SEQ ID NO:

982.

55. The method of any one of claims 48-54, further comprising introducing a first nucleic acid molecule comprising the viral genome into the cell prior to step (i).

56. The method of any one of claims 48-55, wherein the cell comprises a second nucleic acid molecule encoding the AAV capsid variant, optionally wherein the method further comprises introducing the second nucleic acid molecule into the cell prior to step (i).

57. The method of any one of claims 48-56, wherein the cell comprises a mammalian cell (e.g., a HEK293 cell), an insect cell (e.g., a Sf9 cell), or a bacterial cell.

58. A pharmaceutical composition comprising the AAV particle of any one of claims 1-46 and a pharmaceutically acceptable excipient.

59. A pharmaceutical composition comprising the AAV particle of any one of claims 5-22 and a pharmaceutically acceptable excipient.

60. A pharmaceutical composition comprising the AAV particle of any one of claims 9-15 and 45 and a pharmaceutically acceptable excipient.

61. A pharmaceutical composition comprising the AAV particle of any one of claims 16-22 and 46 and a pharmaceutically acceptable excipient.

62. A method of delivering an AAV particle encoding a CDKL5 protein to a subject, comprising administering to the subject an effective amount of the pharmaceutical composition of any one of claims 58-61 or the AAV particle of any one of claims 1-46.

63. The method of claim 62, wherein the subject has, has been diagnosed with, or is at risk of developing a CDKL5-associated disorder, optionally wherein the CDKL5-associated disorder is a CDKL5-associated neurodegenerative or neuromuscular disorder.

64. The method of claim 62 or claim 63, wherein the subject has, has been diagnosed with, or is at risk of developing CDD, developmental and epileptic encephalopathy 2, or atypical Rett syndrome.

65. A method of treating a subject having or diagnosed with a CDKL5-associated disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of any one of claims 58-61 or the AAV particle of any one of claims 1-46.

66. A method of treating a subject having or diagnosed with a CDKL5-associated disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 60 or the AAV particle of any one of claims 9-15 and 45.

67. A method of treating a subject having or diagnosed with a CDKL5-associated disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 61 or the AAV particle of any one of claims 16-22 and 46.

68. A method of treating a subject having or diagnosed with a CDKL5-associated disorder, comprising administering to the subject an effective amount of the pharmaceutical composition of claim 59 or the AAV particle of any one of claims 5-22.

69. The method of claim 35, wherein the CDKL5-associated disorder is a CDKL5-associated neurodegenerative or neuromuscular disorder.

70. The method of claim 69, wherein the CDKL5-associated neurodegenerative or neuromuscular disorder is CDD, developmental and epileptic encephalopathy 2, or atypical Rett syndrome.

71. A method of treating a subject having CDKL5 deficiency disorder (CDD) or diagnosed with CDD, comprising administering to the subject an effective amount of the pharmaceutical composition of any one of claims 58-61 or the AAV particle of any one of claims 1-46.

72. A method of treating a subject having CDKL5 deficiency disorder (CDD) or diagnosed with CDKL5 deficiency disorder (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition of claim 60 or the AAV particle of any one of claims 9-15 and 45.

73. A method of treating a subject having CDKL5 deficiency disorder (CDD) or diagnosed with CDKL5 deficiency disorder (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition of claim 61 or the AAV particle of any one of claims 16-22 and 46.

74. A method of treating a subject having CDKL5 deficiency disorder (CDD) or diagnosed with CDKL5 deficiency disorder (CDD), comprising administering to the subject an effective amount of the pharmaceutical composition of claim 59 or the AAV particle of any one of claims 5-22.

75. The method of any one of claims 32-38, wherein the subject has one or more mutations in the CDKL5 gene.

76. The method of any one of claims 62-75, wherein the subject has a reduced level of CDKL5 activity compared to a reference level in a subject not having a CDKL5-associated disorder.

77. The method of any one of claims 65-76, wherein the administration arrests progression of the subject’s disorder.

78. The method of any one of claims 65-77, wherein the administration ameliorates at least one symptom of the disorder and / or alters one or more biomarkers of the disorder.

79. The method of claim 78, wherein the one or more biomarkers comprise a marker of neurofilament light chain or CDKL5 activity, e.g., as measured by phosphorylation level of a substrate protein, e.g., MECP2, or as measured by mass spectrometry.

80. The method of any one of claims 65 to 77, wherein the treatment improves at least one symptom, optionally, wherein the at least one symptom includes epilepsy (e.g., early-onset epilepsy), autism, cognitive deficits, motor skill limitations, sleep difficulties, visual impairments, hypotonia, gastrointestinal reflux, behavioral symptoms (including unexplained laughter or crying, touch sensitivity, and sleep disturbances), facial appearance changes (including microcephaly, high and wide forehead, large and deep-set eyes, a smaller-than-normal distance between the nose and upper lip, upturned nose, full lips, and wide-set teeth), difficulty standing and walking, small and cold feet, lack of or reduced eye contact, frequent strabismus, cortical visual impairment or cortical blindness, bruxism, speech limitations or aphasia, difficulty eating, stereotypes, limited fine motor skills, gastroesophageal reflux, constipation, or a combination thereof.

81. The method of any one of claims 62 to 80, wherein the subject is a human.

82. The method of any one of claims 62 to 81, wherein the AAV particles are delivered to cells, tissues, or regions of the CNS, such as regions of the brain or spinal cord, such as the parenchyma, cortex, substantia nigra, caudate nucleus, cerebellum, striatum, corpus callosum, cerebellum, brainstem caudate nucleus-putamen, thalamus, superior colliculus, spinal cord, or combinations thereof, and / or delivered to neurons, or combinations thereof.

83. The method of any one of claims 62 to 82, further comprising assessing, for example, measuring, the level of CDKL5 expression in the subject, such as the level of CDKL5 gene expression, the level of CDKL5 mRNA expression, and / or the level of CDKL5 protein expression in the subject's cells, tissues, or body fluids.

84. The method of claim 83, wherein the CDKL5 protein expression level is measured by ELISA, Western blotting, or immunohistochemistry.

85. The method of claim 83 or claim 84, wherein the CDKL5 expression level is assessed before and after application of the AAV particle, optionally wherein the CDKL5 expression level before application is compared with the CDKL5 expression level after application.

86. The method of any one of claims 83 to 85, comprising assessing the expression level of CDKL5 in cells or tissues of the central nervous system (e.g., parenchyma).

87. The method of any one of claims 83 to 86, wherein the CDKL5 protein expression level of the subject increases after administration relative to the subject's CDKL5 protein expression level before administration.

88. The method of any one of claims 62 to 87, further comprising evaluating, for example, measuring, the level of CDKL5 activity in the subject, such as in the subject's cells or tissues.

89. The method of any one of claims 65 to 88, wherein the application results in the following increase: (i) CDKL5 activity in the subject’s cells, tissues (e.g., cells or tissues of the CNS, e.g., cortex, striatum, thalamus, cerebellum, and / or brainstem), and / or bodily fluids (e.g., CSF and / or serum) relative to CDKL5 activity in the subject prior to the administration; (ii) viral genome (VG) level per cell in the subject’s CNS tissue (e.g., cortex, striatum, thalamus, cerebellum, brainstem, and / or spinal cord) relative to VG level per cell in the subject’s peripheral tissue prior to the administration; and / or (iii) CDKL5 mRNA expression in the subject’s cells or tissues (e.g., cells or tissues of the CNS, e.g., cortex, thalamus, and / or brainstem) relative to CDKL5 mRNA expression in the subject prior to the administration.

90. The method of any one of claims 65-89, further comprising administering to the subject an additional agent suitable for treating or preventing a CDKL5-associated disorder; optionally, wherein the additional agent comprises one or more anti-epileptic drugs (e.g., valproic acid, levetiracetam, clobazam, lamotrigine, ganaxolone, topiramate, or a combination thereof).

91. The method of any one of claims 65-90, further comprising administering to the subject an immunosuppressive agent.

92. The method of claim 91, wherein the immunosuppressive agent comprises a corticosteroid (e.g., prednisone, prednisolone, methylprednisolone, and / or dexamethasone), rapamycin, mycophenolate mofetil, tacrolimus, rituximab, and / or eculizumab hydroxychloroquine.

93. The pharmaceutical composition of any one of claims 58-61 or the AAV particle of any one of claims 1-46 for use in treating a CDKL5-associated disorder; optionally, wherein the CDKL5-associated disorder is CDKL5 Deficiency Disorder (CDD), developmental and epileptic encephalopathy 2, or atypical Rett syndrome.

94. Use of the pharmaceutical composition of any one of claims 58-61 or the AAV particle of any one of claims 1-46 in the manufacture of a medicament for treating a CDKL5-associated disorder; optionally, wherein the CDKL5-associated disorder is CDKL5 Deficiency Disorder (CDD), developmental and epileptic encephalopathy 2, or atypical Rett syndrome.

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