Company: PRME
Filing Date: 2025-02-28
Form Type: 10-K
Source: 0001628280-25-008884
Chunk: 23

Company: Prime Medicine, Inc.
Filing Date: 2025-02-28
Form: 10-K
Item: Item 1
Chunk 23
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 Prime Editor proteins contain two protein domains. The first domain is a programmable DNA binding domain, often a CRISPR-Cas protein domain, or Cas domain. Cas domains enable targeting of specific DNA sequences, and they have been adapted and engineered to target desired genomic locations in human cells with high specificity. In Prime Editors, programmable DNA binding domains, such as Cas domains, are typically modified such that they do not cause a double-stranded break in the DNA. The second protein domain of Prime Editors is a DNA polymerase domain, often a reverse transcriptase enzyme domain, or RT domain. Reverse transcriptases are DNA polymerase enzymes that write new DNA sequences by copying from an RNA template. In Prime Editing, the RT domain copies the edited DNA sequence directly into the target genomic site where the edit is made.

The other main component in Prime Editing is the pegRNA. The pegRNA contains a search sequence, also known as a spacer, which provides a target genomic address for the Prime Editor. This enables the Prime Editor to specifically target a desired gene sequence. The pegRNA also contains a second sequence unique to Prime Editing, a replace sequence, or edit template, which provides a blueprint for the edit that will be made to the target DNA sequence.

As shown in the second panel in the figure below, a Prime Editor protein, exemplified using a fusion protein containing a Cas domain and an RT protein domain, and the pegRNA locate the DNA target site using the pegRNA’s search sequence. As exemplified in the figure, when the correct DNA target is found (referred to as “edit check 1,” as described below), the Prime Editor’s Cas domain cleaves, or nicks, one of the two DNA strands, creating a single-stranded 3’ flap. The other DNA strand remains intact and is not cleaved by the Prime Editor, thus avoiding the formation of double-stranded DNA breaks. As shown in the third panel of the figure, the 3’ flap binds to a region of the replace sequence in the pegRNA (“edit check 2”) and “primes” the DNA synthesis. The Prime Editor’s RT domain copies a portion of the replace sequence, which incorporates the desired edit, as shown in the fourth panel. After the desired edit is copied, cellular DNA repair preferentially incorporates the corrected 3’ flap into the gene (“edit check 3”) while removing the excess original DNA sequence. The complementary DNA strand is also corrected through cellular DNA repair, using the Prime-Edited DNA strand as