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US 11,529,427 B2Granted
Gene editing for hemophilia A with improved factor VIII expression
AssigneesCrispr Therapeutics AG, Bayer HealthCare LLC
InventorAlan Richard Brooks
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Abstract
Provided herein, in some embodiments, are materials and methods for treating hemophilia A in a subject ex vivo or in vivo. Also provided herein, in some embodiments, are materials and methods for knocking in a coding sequence encoding a synthetic FVIII having a B domain substitute into a genome.
Claims20
A system for altering a host cell DNA sequence, comprising:
- a deoxyribonucleic acid (DNA) endonuclease or a nucleic acid encoding the DNA endonuclease, wherein the DNA endonuclease is Cas9;
- a guide RNA (gRNA) comprising a spacer sequence complementary to a host cell locus or a nucleic acid encoding the gRNA;
- and
- a donor template comprising a nucleic acid sequence encoding a synthetic FVIII protein, wherein the synthetic FVIII protein comprises a B domain substitute, wherein the B domain substitute comprises the amino acid sequence of any one of SEQ ID NOs:
- 362-364, 366-369, 371, and 373.
2 more elements
A method of editing a genome in a host cell, the method comprising providing to the cell:
- (a) a gRNA comprising a spacer sequence complementary to a host cell locus or a nucleic acid encoding the gRNA;
- (b) a DNA endonuclease or a nucleic acid encoding the DNA endonuclease, wherein the DNA endonuclease is Cas9;
- and
- (c) a donor template comprising a nucleic acid sequence encoding a synthetic FVIII protein, wherein the synthetic FVIII protein comprises a B domain substitute, wherein the B domain substitute comprises the amino acid sequence of any one of SEQ ID NOs:
- 362-364, 366-369, 371, and 373.
2 more elements
A method of treating hemophilia A in a subject, the method comprising:
- providing the following to a cell in the subject:
- (a) a gRNA comprising a spacer sequence complementary to a host cell locus or a nucleic acid encoding the gRNA;
- (b) a DNA endonuclease or a nucleic acid encoding the DNA endonuclease, wherein the DNA endonuclease is Cas9;
- and
- (c) a donor template comprising a nucleic acid sequence encoding a synthetic FVIII protein, wherein the synthetic FVIII protein comprises a B domain substitute, wherein the B domain substitute comprises the amino acid sequence of any one of SEQ ID NOs:
- 362-364, 366-369, 371, and 373.
3 more elements
A method of increasing the amount of FVIII in a subject, the method comprising:
- providing the following to a cell in the subject, wherein the subject has a first serum level of FVIII:
- (a) a gRNA comprising a spacer sequence complementary to a host cell locus or a nucleic acid encoding the gRNA;
- (b) a DNA endonuclease or a nucleic acid encoding the DNA endonuclease, wherein the DNA endonuclease is Cas9;
- and
- (c) a donor template comprising a nucleic acid sequence encoding a synthetic FVIII protein, wherein the synthetic FVIII protein comprises a B domain substitute, wherein the B domain substitute comprises the amino acid sequence of any one of SEQ ID NOs:
- 362-364, 366-369, 371, and 373.
3 more elements
Dependencies are inferred from the claim text.