A self-circularization RNA construct that can be expressed in a DNA vector and simultaneously circularized through a self-targeting and splicing reaction to form a circRNA is disclosed. The circRNA can consist only of a gene of interest which can be a coding, non-coding, or a combination thereof. The gene of interest has the advantage of being able to rapidly express a peptide or protein. The formed circRNA has a circular structure and has a stable and high half-life because 5′ and 3′ ends are not exposed. Accordingly, functional RNA such as miRNA, anti-miRNA, siRNA, shRNA, aptamer, functional RNA for gene/RNA editing, ADAR (adenosine deaminase acting on the RNA)-recruiting RNA, mRNA vaccine, mRNA therapeutic agent, vaccine adjuvant, and CAR-T mRNA can be produced as a stable circRNA in cells.

Circular RNA and transfer vehicles, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises group I intron fragments, spacers, an IRES, duplex forming regions, and an expression sequence. In some embodiments, the expression sequence encodes a chimeric antigen receptor (CAR). In some embodiments, circular RNA of the invention has improved expression, functional stability, immunogenicity, ease of manufacturing, and/or half-life when compared to linear RNA. In some embodiments, inventive methods and constructs result in improved circularization efficiency, splicing efficiency, and/or purity when compared to existing RNA circularization approaches.

A trans-splicing ribozyme capable of splicing a rhodopsin transcript at a target splicing site and containing a sequence that is capable of complementarily binding to a target binding site of the rhodopsin transcript is disclosed. The trans-splicing ribozyme may further containing a desired rhodopsin transcript at 3′-end. The trans-splicing ribozyme may further contains an antisense sequence that is complementary to a region downstream the target binding site of the rhodopsin transcript. A nucleotide molecule encoding the trans-splicing ribozyme is also disclosed. Delivery systems to delivery the nucleotide molecule and/or the trans-splicing ribozyme to target tissue or cells as well as uses of the trans-splicing ribozyme, the nucleotide molecule, delivery systems, or pharmaceutical compositions containing any of them are also disclosed.

Circular RNA and transfer vehicles, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises group I intron fragments, spacers, an IRES, duplex forming regions, and an expression sequence. In some embodiments, the expression sequence encodes a chimeric antigen receptor (CAR). In some embodiments, circular RNA of the invention has improved expression, functional stability, immunogenicity, ease of manufacturing, and/or half-life when compared to linear RNA. In some embodiments, inventive methods and constructs result in improved circularization efficiency, splicing efficiency, and/or purity when compared to existing RNA circularization approaches.

The present specification provides compositions and methods that are capable of directly installing an insertion or deletion of a given nucleotide at a specified genetic locus. The compositions and methods involve the novel combination of the use an engineered RNA enzyme (i.e., “ribozyme”) that is capable of site-specifically inserting or deleting a single nucleotide at a genetic locus and the use of a nucleic acid programmable DNA binding protein (napDNAbp) (e.g., Cas9) to target the engineered ribozyme to a specified genetic locus, thereby allowing for the direct installation of an insertion of deletion at the specified genetic locus by the engineered ribozyme.

Circular RNA and transfer vehicles, along with related compositions and methods are described herein. In some embodiments, the inventive circular RNA comprises group I intron fragments, spacers, an IRES, duplex forming regions, and an expression sequence. In some embodiments, the expression sequence encodes a chimeric antigen receptor (CAR). In some embodiments, circular RNA of the invention has improved expression, functional stability, immunogenicity, ease of manufacturing, and/or half-life when compared to linear RNA. In some embodiments, inventive methods and constructs result in improved circularization efficiency, splicing efficiency, and/or purity when compared to existing RNA circularization approaches.

A trans-splicing ribozyme capable of splicing a rhodopsin transcript at a target splicing site and containing a sequence that is capable of complementarily binding to a target binding site of the rhodopsin transcript is disclosed. The trans-splicing ribozyme may further containing a desired rhodopsin transcript at 3′-end. The trans-splicing ribozyme may further contains an antisense sequence that is complementary to a region downstream the target binding site of the rhodopsin transcript. A nucleotide molecule encoding the trans-splicing ribozyme is also disclosed. Delivery systems to delivery the nucleotide molecule and/or the trans-splicing ribozyme to target tissue or cells as well as uses of the trans-splicing ribozyme, the nucleotide molecule, delivery systems, or pharmaceutical compositions containing any of them are also disclosed.

Disclosed are compositions comprising: a trans-splicing nucleic acid comprising (a) one or more replacement domains that encode a therapeutic sequence; and (b) one or more intronic domains that promote RNA splicing of the replacement domain primarily in a specific tissue or cell type; and (c) one or more antisense domains that promote binding to a target RNA molecule, wherein an RNA trans-splicing reaction promotes insertion of the replacement domain into a target RNA. Methods of making and methods of using compositions of the disclosure are also provided, including but not limited to compositions of the disclosure that may be used in the treatment of a disease or disorder in a patient or subject. Example disease or disorders of the disclosure include genetic and epigenetic diseases or disorders.

A cancer-specific trans-splicing ribozyme and a use thereof are disclosed. The trans-splicing ribozyme does not act on normal tissue, but is specifically expressed in cancer tissue. Therefore, it is very safe and has excellent expression efficiency at the post-transcription level, and thus can be effectively used in treatment of cancer.

A cancer-specific trans-splicing ribozyme and a use thereof are disclosed. The trans-splicing ribozyme does not act on normal tissue, but is specifically expressed in cancer tissue. Therefore, it is very safe and has excellent expression efficiency at the post-transcription level, and thus can be effectively used in treatment of cancer.