Use of prime editing for treating sickle cell disease
AssigneesThe Broad Institute, Inc., President and Fellows of Harvard College, St. Jude Children's Research Hospital
InventorsDavid R. Liu, Kelcee Everette, Gregory Newby, Jonathan S. Yen, Mitchell J. Weiss
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Abstract
The present disclosure provides methods for treating sickle cell disease using prime editing. The present disclosure also provides epegRNAs targeting the β-globin (HBB) gene, which may be useful for treating sickle cell disease. Also provided herein are prime editor complexes, polynucleotides, vectors, pharmaceutical compositions, kits, and cells useful for performing the methods described herein.
Claims81
A method of treating sickle cell disease comprising contacting a target nucleotide sequence with a prime editor and an engineered prime editing guide RNA (epegRNA), wherein the epegRNA comprises the structure 5′-[spacer sequence]-[single guide RNA (sgRNA) scaffold sequence]-[extension arm sequence]-[optional engineered linker sequence]-[3′ structured motif]-3′, wherein each instance of]-[comprises an optional linker sequence.
A method of treating sickle cell disease comprising contacting a target nucleotide sequence with a prime editor and a prime editing guide RNA (pegRNA), wherein the prime editor is PE3max or PE3bmax, and wherein the pegRNA comprises the structure 5′-[spacer sequence]-[single guide RNA (sgRNA) scaffold sequence]-[extension arm sequence]-3′, wherein each instance of]-[comprises an optional linker sequence.
A method of treating sickle cell disease comprising contacting a target nucleotide sequence with a prime editor, a prime editing guide RNA (pegRNA), and a nicking single guide RNA (sgRNA), wherein the pegRNA comprises the structure 5′-[spacer sequence]-[sgRNA scaffold sequence]-[extension arm sequence]-3′, wherein each instance of]-[comprises an optional linker sequence, and wherein the spacer sequence of the nicking sgRNA comprises the nucleotide sequence 5′-GTAACGGCAGACTTCTCTTC-3′ (SEQ ID NO:
- 129), 5′-CACGTTCACCTTGCCCCACA-3′ (SEQ ID NO:
- 130), or 5′-TCCACATGCCCAGTTTCTAT-3′ (SEQ ID NO:
- 131), or a sequence at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the sequence 5′-GTAACGGCAGACTTCTCTTC-3′ (SEQ ID NO:
- 129), 5′-CACGTTCACCTTGCCCCACA-3′ (SEQ ID NO:
- 130), or 5′-TCCACATGCCCAGTTTCTAT-3′ (SEQ ID NO:
- 131) or a fragment thereof.
An engineered prime editing guide RNA (epegRNA) targeting the β-globin gene (HBB), wherein the epegRNA comprises the structure 5′-[spacer sequence]-[single guide RNA (sgRNA) scaffold sequence]-[extension arm sequence]-[optional engineered linker sequence]-[3′ structured motif]-3′, wherein each instance of]-[comprises an optional linker sequence.
A cell comprising DNA comprising the sequence 5′-ATGGTGCACCTGACTCCTGA AGAGAAG-3′ (SEQ ID NO:
- 78), or a sequence at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the sequence 5′-ATGGTGCACCTGACTCCTGAAGAGAAG-3′ (SEQ ID NO:
- 78), wherein the underlined nucleotide is an A.
Dependencies are inferred from the claim text.