Title:
METHODS AND COMPOSITIONS FOR EDITING NUCLEOTIDE SEQUENCES
Document Type and Number:
WIPO Patent Application WO/2020/191248
Kind Code:
A8
Abstract:
The present disclosure provides compositions and methods for conducting prime editing of a target DNA molecule (e.g., a genome) that enables the incorporation of a nucleotide change and/or targeted mutagenesis. The nucleotide change can include a single-nucleotide change (e.g., any transition or any transversion), an insertion of one or more nucleotides, or a deletion of one or more nucleotides. More in particular, the disclosure provides fusion proteins comprising nucleic acid programmable DNA binding proteins (napDNAbp) and a polymerase (e.g., reverse transcriptase), which is guided to a specific DNA sequence by a modified guide RNA, named an PEgRNA. The PEgRNA has been altered (relative to a standard guide RNA) to comprise an extended portion that provides a DNA synthesis template sequence which encodes a single strand DNA flap, which is homologous to a strand of the targeted endogenous DNA sequence to be edited, but which contains the desired one or more nucleotide changes and which, following synthesis by the polymerase (e.g., reverse transcriptase), becomes incoporated into the target DNA molecule. Also disclosed herein are various methods that leverage prime editing, including treating trinucleotide repeat contraction diseases, installing targeted peptide tags, treating prion disease through the intallation of protection mutations, manipulating RNA-encoding genes for the installation of RNA tags for controlling the function and expression of RNA, using prime editing to construct sophisticated gene libraries, using prime editing to insert immunoepitopes into proteins, use of prime editing to insert inducible dimerization domains into protein targets, and delivery methods, among others.
Inventors:
LIU DAVID (US)
ANZALONE ANDREW (US)
NELSON JAMES (US)
ANZALONE ANDREW (US)
NELSON JAMES (US)
Application Number:
PCT/US2020/023730
Publication Date:
October 21, 2021
Filing Date:
March 19, 2020
Export Citation:
Assignee:
BROAD INST INC (US)
HARVARD COLLEGE (US)
HARVARD COLLEGE (US)
International Classes:
C12N15/11; C12N9/12; C12N9/22; C12N15/10
Attorney, Agent or Firm:
MCCOOL, Gabriel, J. et al. (US)
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