Engineered cell lines for increased production of recombinant adeno-associated virus (raav)
AssigneeUltragenyx Pharmaceutical Inc.
InventorsNicholas Richards, Justin Tedeschi, Caroline Adams, Charles Pugh, Matthew Scott Fuller
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Abstract
This application relates to recombinant adeno-associated virus (rAAV) packaging and/or producer cell lines which have been engineered to reduce expression and/or activity of one or more genes and/or proteins to increase rAAV titers. The methods of generating the engineered cell lines have also been described herein.
Claims78
A recombinant adeno-associated virus (rAAV) packaging and/or producer cell line comprising cells in which the expression of a gene selected from the group consisting of RIG-1 (DDX58), IFIT3, MDA5 (IFIH1), CGAS (cGAS), CHUK (IKK-α), DDX41, DHX58 (LGP2), IFI6, IKBKB (IKK-β), IRF3, IRF7, MAVS, MYD88, NFKB1, NFKB2, TBK1, TRIF, and TRIM25, and any combination thereof is reduced compared to that in control parental cells.
A cell comprising:
- (a) AAV rep and cap genes and
- (b) an siRNA comprising a nucleotide sequence selected from any one of SEQ ID NOs:
- 1-36.
A cell comprising:
- (a) AAV rep and cap genes and
- (b) (i) a guide RNA (gRNA) comprising a nucleotide sequence selected from SEQ ID Nos:
- 37-99;
- and/or
- (ii) a gRNA which targets a DNA sequence selected from SEQ ID NOs:
- 100-162.
A cell comprising:
- (a) AAV rep and cap genes and
- (b) a nuclease selected from the group consisting of Zinc Finger nuclease (ZFN), a meganuclease, or a transcription activator-like effector nuclease (TALEN), wherein the nuclease targets a gene selected from the group RIG-1 (DDX58), IFIT3, MDA5 (IFIH1), CGAS (cGAS), CHUK (IKK-α), DDX41, DHX58 (LGP2), IFI6, IKBKB (IKK-β), IRF3, IRF7, MAVS, MYD88, NFKB1, NFKB2, TBK1, TRIF, TRIM25, and combinations thereof.
A cell comprising:
- (a) AAV rep and cap genes and
- (b) a knockout version of a gene selected from the group consisting of RIG-1 (DDX58), IFIT3, MDA5 (IFIH1), CGAS (cGAS), CHUK (IKK-α), DDX41, DHX58 (LGP2), IFI6, IKBKB (IKK-β), IRF3, IRF7, MAVS, MYD88, NFKB1, NFKB2, TBK1, TRIF, TRIM25, and
- combinations thereof.
A recombinant adeno-associated virus (rAAV) packaging and/or producer cell line comprising cells which have been engineered to reduce the expression and/or activity of a gene product expressed from a gene selected from the group consisting of RIG-1 (DDX58), IFIT3, MDA5 (IFIH1), CGAS (cGAS), CHUK (IKK-α), DDX41, DHX58 (LGP2), IFI6, IFIT3, IKBKB (IKK-β), IRF3, IRF7, MAVS, MDA5 (IFIH1), MYD88, NFKB1, NFKB2, RIG-1 (DDX58), TBK1, TRIF, and TRIM25, and any combination thereof as compared to corresponding control parental cells.
A recombinant adeno-associated virus (rAAV) packaging and/or producer cell line, wherein said cell line exhibits reduced expression and/or activity of a polypeptide or polyribonucleotide expressed from a gene selected from the group consisting of RIG-1 (DDX58), IFIT3, MDA5 (IFIH1), CGAS (cGAS), CHUK (IKK-α), DDX41, DHX58 (LGP2), IFI6, IFIT3, IKBKB (IKK-β), IRF3, IRF7, MAVS, MDA5 (IFIH1), MYD88, NFKB1, NFKB2, RIG-1 (DDX58), TBK1, TRIF, and TRIM25, and any combination thereof as compared to that of a control parental cell line.
Dependencies are inferred from the claim text.