This invention provides RNA, oligoribonucleotide, and polyribonucleotide molecules comprising pseudouridine or a modified nucleoside, gene therapy vectors comprising same, methods of synthesizing same, and methods for gene replacement, gene therapy, gene transcription silencing, and the delivery of therapeutic proteins to tissue in vivo, comprising the molecules. The present invention also provides methods of reducing the immunogenicity of RNA, oligoribonucleotide, and polyribonucleotide molecules.

The invention relates to RNA editing oligonucleotides (EONs) that can bring about specific editing of a target nucleotide (adenosine) in a target RNA molecule in a eukaryotic cell, wherein said oligonucleotide is for use in the treatment of Stargardt disease, and more preferably for the deamination of target adenosines present in the ABCA4 pre-mRNA or ABCA4 mRNA.

Disclosed herein are compositions that comprise engineered polynucleotides, pharmaceutical compositions comprising the same, methods of making the same, and methods of treatment comprising the compositions that comprise the engineered polynucleotides.

The present disclosure provides adenine base editors (ABEs) that are variants of known adenine base editors. The adenosine deaminase domain of a known ABE was modified to produce adenosine deaminase variants. The deaminase variants provided herein have broader compatibility with diverse napDNAbp domains, such as Cas homologs, for base editing applications. The ABEs provided herein comprise a deaminase variant and a napDNAbp domain. The ABEs provided herein exhibit reduced off-target editing effects while retaining high on-target editing efficiencies. These ABEs exhibit reduced off-target DNA editing effects and reduced off-target editing effects in cellular mRNA. In addition, methods for targeted nucleic acid editing are provided. Further provided are pharmaceutical compositions comprising the ABEs. Also provided are vectors and kits useful for the generation and delivery of the ABEs, including vector systems for engineering the ABEs through directed evolution. Cells containing such vectors and ABEs are also provided. Further provided are methods of treatment comprising administering the ABEs.

A method for producing genetically engineered immune cells, e.g. T cells, or iPSCs which uses an RNA-scaffold mediated base editing system. The method enables precise modifications to be made to the genome whilst minimizing the possibility of off-target effects, making the method particularly suitable for therapeutic applications.

The present disclosure provides for systems, methods, and compositions for targeting nucleic acids. In particular, the invention provides Cas proteins and their use in modifying target sequences.

The disclosure provides adenosine deaminases that are capable of deaminating adenosine in DNA. 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 further comprise a nuclear localization sequence (NLS), and/or an inhibitor of base repair, such as, a nuclease dead inosine specific nuclease (dISN).

Engineered transversion base editors that enable expanded amino acid modifications and methods of using the same. Described herein, for example, are fusion proteins containing cytidine deaminases (e.g. human or rat APOBECs, pmCDA1 or AID) or adenosine deaminases (e.g. E. coli TadAs) or a combination thereof, catalytically impaired CRISPR-Cas proteins (e.g. Cas9, CasX or Cas12 nucleases), linkers, nuclear localization signals (NLSs) and a human or E. coli uracil-n-glycosylase (UNG) and/or REV1 protein that enable the CRISPR-guided programmable introduction of C-to-G and G-to-C transversions in DNA. The UNG may be fused to the deaminase-Cas fusion or not, in which case endogenous UNG may be recruited using molecular machinery that is integrated into the deaminase-Cas fusion architecture, e.g. using peptide or RNA aptamers or scFVs, sdABs or Fabs.

Provided herein is a composition comprising a fusion protein or a fragment or a variant thereof comprising an anti-PD1 antibody or a fragment/variant thereof and a TGF-β trap. Provided herein is a composition comprising a fusion protein or a fragment thereof or a variant thereof comprising an anti-PD1 antibody or a fragment/variant thereof and a ADA2 polypeptide. Also provided herein are methods of using the composition in treating cancer.

The present disclosure provides for systems, methods, and compositions for targeting nucleic acids. In particular, the invention provides small Cas proteins and their use in modifying target sequences. In one aspect, the present disclosure provides a non-naturally occurring or engineered system comprising: a Cas protein that comprises a RuvC domain and a HNH domain, and is less than 850 amino acids in size; and a guide sequence capable of forming a complex with the Cas protein and directing the complex to bind to a target sequence.