# US 12,600,989 B2 · Adeno-associated virus (AAV) vector and uses therefor > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 12,600,989 B2 - Document ID: US12600989B2 - Status: Granted - Kind: B2 - Application number: 18/248,967 - Filed: 2021-10-18 - Published: 2023-11-30 - Granted: 2026-04-14 - Earliest claimed: 2020-10-18 - Assignees: The Trustees of the University of Pennsylvania - Applicants: The Trustees of the University of Pennsylvania - Inventors: Qiang Wang - Web page: https://patentgrep.com/patent/US12600989B2 ## CPC - C12N 5/10: Cells modified by introduction of foreign genetic material - C12N 15/86: Viral vectors - C07K 14/005: from viruses - C12N 2510/00: Genetically modified cells - C12N 2511/00: Cells for large scale production - C12N 2750/14121: Viruses as such, e.g. new isolates, mutants or their genomic sequences - C12N 2750/14122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes - C12N 2750/14142: virus or viral particle as vehicle, e.g. encapsulating small organic molecule - C12N 2750/14143: viral genome or elements thereof as genetic vector - C12N 2750/14151: Methods of production or purification of viral material - C12N 2750/14152: relating to complementing cells and packaging systems for producing virus or viral particles - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) ## Abstract Provided herein are engineered nucleic acid sequences encoding AAVhu68 which provide improved yields of packaged AAVhu68 viral particles. Also provided is a packaging host cell comprising the engineered nucleotide sequence and methods of using same to product recombinant AAVhu68 vectors useful for gene delivery. Further provided are production cell supernatants comprising improved yields of recombinant AAV vectors having AAVhu68 capsids produced using the nucleotide sequences provided herein. ## Claims ### Claim 1 A production host cell for generating a recombinant adeno-associated virus (rAAV) having an AAVhu68 capsid comprising: (a) a nucleic acid sequence of SEQ ID NO: 1 or a sequence at least 99% identical thereto encoding the AAVhu68 vp1 capsid protein operably linked to expression control sequences which direct expression of the AAVhu68 vp1 capsid protein in the host cell; (b) a nucleic acid molecule for packaging into the AAVhu68 capsid, said nucleic acid molecule comprising at least one AAV inverted terminal repeat (ITR) and a non-AAV nucleic acid sequence encoding a gene product operably linked to sequences which direct expression of the product in a host cell; and (c) sufficient AAV rep functions and helper functions to permit packaging of the nucleic acid molecule into the AAVhu68 capsid. ### Claim 2 (depends on 1) The production host cell according to claim 1, wherein the host cell further comprises a nucleic acid sequence of about nucleotide 607 to about nucleotide 2211 of SEQ ID NO: 1 or a sequence at least 99% identical thereto encoding the AAVhu68 vp3 of about amino acid 203 to about amino acid 736 of SEQ ID NO: 2. ### Claim 3 (depends on 1) The production host cell according to claim 1, wherein the host cell is a human cell line or an insect cell line. ### Claim 4 (depends on 1) The production host cell according to claim 1, wherein the host cells is a HEK293 cell, HuH-7 cell, BHK cell, or Vero cell. ### Claim 5 (depends on 1) The production host cell according to claim 1, wherein the helper functions are provided by adenovirus, baculovirus, or herpes simplex virus proteins. ### Claim 6 (depends on 1) The production host cell according to claim 1, wherein the AAV rep is from a different AAV. ### Claim 7 (depends on 6) The production host cell according to claim 6, wherein the AAV rep is from AAV2. ### Claim 8 (depends on 1) The production host cell according to claim 1, wherein the AAV rep coding sequence and cap genes are on the same nucleic acid molecule, wherein there is optionally a spacer between the rep sequence and cap gene. ### Claim 9 (depends on 8) The production host cell according to claim 8, wherein the spacer is SEQ ID NO: 9. ### Claim 10 (depends on 1) The production host cell according to claim 1, wherein the AAV rep is AAVhu68rep characterized by the amino acid sequence of SEQ ID NO: 6, or a functional fragment thereof. ### Claim 11 (depends on 10) The production host cell according to claim 10, wherein the AAV rep is encoded by the nucleic acid sequence of SEQ ID NO: 5. ### Claim 12 (depends on 1) A suspension comprising production host cells according to claim 1. ### Claim 13 (depends on 12) The suspension according to claim 12, wherein the helper functions are herpes simplex virus helper functions. ### Claim 14 (depends on 12) The suspension according to claim 12, wherein the helper functions are baculovirus helper functions. ### Claim 15 (depends on 1) A cell culture comprising the production host cell according to claim 1. ### Claim 16 (depends on 15) The cell culture according to claim 15, wherein helper functions are adenovirus helper functions. ### Claim 17 (depends on 1) A substrate comprising an adherent cell line comprising the production host cell according to claim 1. ### Claim 18 (depends on 12) The suspension according to claim 12, wherein the host cells can produce an amount of rAAVhu68 particles that is at least 20% higher than the amount produced by AAVhu68 of SEQ ID NO: 3.