Disclosed herein are engineered ADAR systems for gene editing.

The present invention relates to methods and compositions for editing an ASL polynucleotide, e.g., an ASL polynucleotide comprising a SNP associated with argininosuccinate lyase deficiency. The invention also relates to methods and compositions for treating or preventing argininosuccinate lyase deficiency in a subject.

The present invention relates to methods and compositions for editing an LRRK2 polynucleotide, e.g., an LRRK2 polynucleotide comprising a SNP associated with Parkinson's Disease. The invention also relates to methods and compositions for treating or preventing Parkinson's Disease in a subject.

The present invention relates to compositions comprising a Cas12i polypeptide, a deaminase polypeptide, and an RNA guide, processes for characterizing the compositions, cells comprising the compositions, Cas12i fusion proteins, Cas12i complexes, and methods of using the compositions.

The present invention provides novel systems, methods and compositions for making and using a recombinantly engineered novel Cas9 optimized for human cells, for nucleic acid targeting and manipulation. The present invention is based on the discovery of a novel Cas9 species from Lachnospira bacterium that was codon-optimized and recombinantly produced for use in human ceils. In some embodiments, the novel Cas9 can be used in a base editor. In some embodiments, the novel engineered Cas9 is used to treat human diseases.

The present invention features compositions and methods for editing deleterious mutations associated with hemoglobinopathies, such as sickle cell disease (SCD). In particular embodiments, the invention provides methods for correcting mutations in a beta globin polynucleotide using modified adenosine base editors termed “ABE8” having unprecedented levels (e.g., >60-70%) of efficiency.

The disclosure provides methods and compositions for treating blood diseases/disorders, such as sickle cell disease, hemochromatosis, hemophilia, and beta-thalassemia. For example the disclosure provides therapeutic guide RNAs that target the promotor of HBG1/2 to generate point mutations that increase expression of fetal hemoglobin. As another example, the disclosure provides therapeutic guide RNAs that target mutations in HBB, Factor VIII, and HFE to treat sickle cell disease, beta-thalassemia, hemophilia and hemochromatosis. The disclosure also provides fusion proteins comprising a Cas9 (e.g., a Cas9 nickase) domain and adenosine deaminases that deaminate adenosine in DNA. In some embodiments, the fusion proteins are in complex with nucleic acids, such as guide RNAs (gRNAs), which target the fusion proteins to a DNA sequence (e.g., an HBG1 or HBG2 protmoter sequence, or an HFE, GBB, or F8 gene sequence). Such complexes may be useful for increasing expression of fetal hemoglobin or correcting a poing mutation (e.g., C282Y) in HFE.

The present application provides methods for editing RNA by introducing a deaminase-recruiting RNA in a host cell for deamination of an adenosine in a target RNA. The present application further provides deaminase-recruiting RNAs used in the RNA editing methods and compositions and kits comprising the same.

The present disclosure provides for endonuclease enzymes having distinguishing domain features, as well as methods of using such enzymes or variants thereof.

Aspects of the disclosure relate to compositions (e.g., isolated nucleic acids, rAAV vectors, rAAVs, etc.) and methods for gene editing. The disclosure is based, in part, on isolated nucleic acids encoding combinations of gene editing proteins (e.g., Cas proteins) and base editors (e.g., Adenosine Deaminase Acting on RNA deaminase domains) with certain regulatory sequences that are amenable to packaging in recombinant adeno-associated viruses (rAAVs). In some embodiments, compositions described by the disclosure are useful for treating certain diseases in a subject in need thereof.