57 results about "ADAR" patented technology

The present invention relates to modulators, e.g., double stranded RNA (dsRNA) agents, antisense polynucleotide agents, antibodies, guideRNAs that effect ADAR editing, or guideRNAs that effect CRISPR editing, that modulate, e.g., inhibit, the expression and / or activity of inhibin subunit beta E (INHBE). The invention also relates to methods of using such modulators to inhibit expression and / or activity of INHBE and to methods of preventing and treating an INHBE-associated disorder, e.g., metabolic disorder, e.g., metabolic syndrome, in a subject.

Methods of preventing or treating liver disease in a subject characterised as having a PNPLA3- 148M protein variant are provided. The methods include administering to the subject an oligonucleotide or a DNA molecule encoding an oligonucleotide, wherein the oligonucleotide is capable of effecting ADAR-mediated editing of a polynucleotide encoding the PNPLA3-148M protein variant, and wherein the polynucleotide encoding the PNPLA3-148M protein variant undergoes ADAR-mediated editing at a codon encoding the methionine at position 148 such that the polynucleotide encodes a PNPLA3-148V protein variant. The disclosure also provides methods of editing a target ribonucleic acid (RNA) molecule encoding a PNPLA3-148M protein variant and methods of restoring the function of the PNPLA3 protein in a subject characterised as having the PNPLA3-148M protein variant.

The present application relates to a modified guide agRNA (ADAR guide RNA, agRNA), which has a phosphorothioate modification at an agRNA site where off-target editing is present, and further has other modifications. The present application further provides a composition containing the modified agRNA composition, a method for editing RNA, and the use.

RNA editing tools for use in systems designed to measure RNA in vivo and manipulate specific cell types are disclosed herein. An RNA sensor system comprising a) a single-stranded RNA (ssRNA) sensor comprising a stop codon and a payload; optionally wherein the ssRNA sensor further comprises a normalizing gene; and b) an adenosine deaminase acting on RNA (ADAR) deaminase; wherein the sensor is capable of binding to a ssRNA target to form a double-stranded RNA (dsRNA) duplex that becomes a substrate for the ADAR deaminase; wherein the substrate comprises a mispairing within the stop codon; and wherein the mispairing is editable by the ADAR deaminase, which editing can effectively remove the stop codon so as to enable translation and expression of the payload. A method of quantifying ribonucleic acid (RNA) levels using the RNA sensor system is also disclosed.

Provided is an oligonucleotide capable of inducing the editing activity of ADAR in a cell and having excellent stability. The oligonucleotide comprises a first oligonucleotide and a second oligonucleotide linked to the 5'-side of the first oligonucleotide, and is capable of inducing the site-specific editing of target RNA. The first oligonucleotide includes: a target-corresponding nucleotide residue; an oligonucleotide that is located on the 5'-side of the target-corresponding nucleotide residue, is complementary to the target RNA, and is composed of 3 to 6 residues; and an oligonucleotide that is located on the 3'-side of the target-corresponding nucleotide residue, is complementary to the target RNA, and is composed of 10 to 24 residues. At least one of a phosphorus-containing linking group that links a first nucleoside residue to a second nucleoside residue and a phosphorus-containing linking group that links a fourth nucleoside residue to a fifth nucleoside residue is an alkylphosphonic acid residue, an alkyl phosphate residue or a substituted phosphoramide residue, wherein the nucleoside residues are numbered from the target-corresponding nucleotide in 3' direction. The second oligonucleotide has a nucleotide residue in which a nucleotide residue corresponding to the target RNA is deleted at the 3'-end or does not form a complementary pair with the target RNA. The second oligonucleotide is composed of 2 to 10 residues, wherein a nucleotide residue other than a nucleotide residue located at the 3'-end is complementary to the target RNA.

The invention relates to chemically modified oligonucleotides for use in site-directed A-to-I editing of a target RNA inside a cell with endogenous adenosine deaminase acting on RNA (ADAR), the oligonucleotide comprising a sequence capable of binding to a target sequence in a target RNA and a central base triplet (CBT) of (3) nucleotides (N-1 N0 N+1), wherein N0 is the central nucleotide directly opposite to a target adenosine in the target RNA that is to be edited, wherein the oligonucleotide comprises at least one internucleoside linkage that is a methanesulfonyl (mesyl) linkage.

The invention belongs to the field of biological medicine, and relates to a guide agRNA for regulating RNA splicing. The invention provides an application of a guide agRNA (Ribonucleic Acid) in preparation of a medicine for changing expression of a target protein by cells of a subject. The guide agRNA recruits ADAR protein to the 3 'splice site or pseudo 3' splice site to edit the A base of the 3 'splice site or pseudo 3' splice site, thereby splicing the whole exon located at the 3 'splice site or flanking the pseudo 3' splice site of the intron from the precursor mRNA, thereby changing the level of mRNA encoding the target protein, and altering the expression of the target protein in the cell. The target gene pre-mRNA selective splicing is regulated and controlled to influence the function or expression of the functional RNA and the target protein of the target gene, so that the purpose of treating diseases is achieved.

A method and compound for regulating SCN1A gene mRNA and protein expression. Provided is a use of a guide agRNA in preparation of a drug for altering expression of a target protein in a cell of a subject. The cell has a precursor mRNA encoding the target protein, wherein the precursor mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or a pseudo 3' splice site of the intron; the guide agRNA is capable of binding to the precursor mRNA to form a structure capable of recruiting ADAR in the cell, thereby forming a complex containing an ADAR protein; and the target protein is a Nav1.1 protein. By regulating alternative splicing of a target gene pre-mRNA, the functional RNA of the target gene and the function or expression of the target protein are affected, thereby achieving the objective of disease treatment.

The present application provides an engineered circular RNA capable of recruiting adenosine deaminase acting on RNA (ADAR; arRNA) and its application in treating Duchenne muscular dystrophy (DMD).

1. A method for expressing a protein in a target cell, comprising binding said target cell with a sensor RNA comprising: (ia) a first nucleotide sequence comprising a sensor nucleotide sequence that is reverse-complementary to a target RNA, the sensor nucleotide sequence comprising a stem-loop sequence comprising one or more editable codons; and / or (ib) a first nucleotide sequence comprising a sensor nucleotide sequence that is reverse-complementary to a 3'UTR of a target RNA, the sensor nucleotide sequence comprising one or more editable codons; and (ii) a second nucleotide sequence encoding a first cleavage domain, and (iii) a third nucleotide sequence encoding an output protein; where the target RNA is present in the target cell.

A method for regulating the mRNA and protein expression of an SCN1A gene, and a compound. Provided is a use of an antisense oligonucleotide guide RNA in the preparation of a drug for altering the expression of a target protein in a cell of a subject. The cell has a pre-mRNA encoding the target protein, wherein the pre-mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or pseudo 3' splice site of the intron; the antisense oligonucleotide guide RNA can bind to the pre-mRNA to form a structure capable of recruiting ADAR in a cell, thereby forming a complex containing an ADAR protein; and the target protein is an Nav1.1 protein. That is, the function or expression of the functional RNA and target protein of a target gene is affected by regulating the alternative splicing of the pre-mRNA of the target gene, thereby achieving the objective of treating diseases.

Provided is an oligonucleotide having excellent stability and capable of inducing ADAR editing activity in a cell. The oligonucleotide comprises a first oligonucleotide and a second oligonucleotide linked to the 5'side thereof, and induces site-specific editing against a target RNA. The first oligonucleotide is composed of a target-corresponding nucleotide residue, an oligonucleotide having 3 to 6 residues complementary to the target RNA on the 5'side thereof, and an oligonucleotide having 10 to 24 residues complementary to the target RNA on the 3 'side thereof. And at least one of a phosphorus-containing linking group connecting the nucleoside residues at the first position and the second position and a phosphorus-containing linking group connecting the nucleoside residues at the fourth position and the fifth position is an alkyl phosphonic acid residue, an alkyl phosphate residue or a substituted phosphamide residue. The second oligonucleotide has a nucleotide residue corresponding to the target RNA deleted at the 3'terminal, or has a nucleotide residue that does not form a complementary pair with the target RNA, the number of residues being 2 to 10, and the nucleotide residues other than the 3 'terminal being complementary to the target RNA.

Provided herein are nucleic acid molecules, compositions, and methods for expressing a payload (e.g., protein). The nucleic acid molecules can change form and become susceptible to ADAR editing when they hybridize with a target RNA, thereby expressing the payload in the presence of the target RNA.

The present invention relates to methods and compositions for disrupting interaction of an NRF2 protein and a KEAP1 protein. The methods include contacting at least one polynucleotide selected from the group consisting of a polynucleotide encoding the NRF2 protein and a polynucleotide encoding the KEAP1 protein with a guide oligonucleotide that effects one or more (e.g., at least two) adenosine deaminase acting on RNA (ADAR)-mediated adenosine to inosine alterations in said at least one polynucleotide, wherein the adenosine to inosine alterations generate a mutant amino acid, thereby disrupting interaction of the NRF2 protein and the KEAP1 protein. The invention also relates to methods of treating a KEAP1-NRF2 pathway related disease in a subject in need thereof, the method comprising contacting, within the subject, at least one polynucleotide selected from the group consisting of a polynucleotide encoding an NRF2 protein and a polynucleotide encoding a KEAP1 protein with a guide oligonucleotide that effects an adenosine deaminase acting on RNA (ADAR)-mediated adenosine to inosine alteration in said at least one polynucleotide, wherein the adenosine to inosine alteration generates a mutant amino acid, thereby disrupting interaction of the NRF2 protein and the KEAP1 protein and treating the disease in the subject; and compositions thereof.

The invention relates to chemically modified oligonucleotides for use in site-directed A-to-I editing of a target RNA inside a cell with endogenous adenosine deaminase acting on RNA (ADAR), the oligonucleotide comprising a sequence capable of binding to a target sequence in a target RNA and a central base triplet (CBT) of 3 nucleotides (N−1N0N+1), wherein N0 is the central nucleotide directly opposite to a target adenosine in the target RNA that is to be edited, wherein the oligonucleotide comprises at least one internucleoside linkage that is a methanesulfonyl (mesyl) linkage.

Provided herein are RNA editing oligonucleotides capable of effecting ADAR-mediated deamination of a target adenosine comprised in a target RNA, wherein the oligonucleotide comprises a sequence that is capable of hybridizing with a region in the target RNA comprising said target adenosine, and wherein the oligonucleotide comprises at least one conformationally restricted nucleotide (CRN) positioned at the 5' and / or at the 3' terminus of the oligonucleotide and / or internally. In particular, the target adenosine may be part of a therapeutic target codon in the MECP2 protein-coding RNA. Provided herein are RNA editing oligonucleotides capable of effecting ADAR-mediated deamination of a target adenosine in a target RNA molecule encoding a target protein, wherein the oligonucleotide comprises a sequence that is capable of hybridizing with a region in the target RNA molecule comprising said target adenosine, and wherein the target adenosine is part of a codon encoding a posttranslational modification site in the target protein. In particular, the target adenosine may be part of a ubiquitination site in the frataxin (FXN) protein. Provided herein are RNA editing oligonucleotides capable of effecting ADAR-mediated deamination of a target adenosine in a target RNA molecule encoding a target protein, wherein the oligonucleotide comprises a sequence that is capable of hybridizing with a region in the target RNA molecule comprising said target adenosine, and wherein the target adenosine is part of a start codon in the 5' untranslated region of the target RNA molecule. In particular, the target adenosine may be part of the start codon of an untranslated ORF (uORF) in the human Sodium-dependent phosphate transporter 2 protein (SLC20A2).

The invention discloses a modified linear arRNA (Ribonucleic Acid) or cyclized arRNA and an application of the modified linear arRNA or cyclized arRNA in RNA (Ribonucleic Acid) editing. The invention provides an RNA (referred to as a modified arRNA), comprising n copies of arRNA truncated bodies; and n is a natural number greater than 2. The modified arRNA can be a linear RNA (Ribonucleic Acid) or a circular RNA (Ribonucleic Acid). To minimize the effect of bystanders without reducing the editing efficiency. The RNA sequence structure is modified by minimizing the specific sequence, the recruitment structural domain is combined, and the cyclization mechanism is optimized. Compared with the existing ADAR technology, the method and the device have the advantage that the editing efficiency is remarkably improved.

A method for expressing a protein in a target cell, the method including: combining the target cell with a sensor RNA including the following: (i) a first nucleotide sequence including a sensor nucleotide sequence including: (a) a first region that is reverse complementary to the target RNA, (b) a second region including a stem-loop sequence including one or more editable codons wherein a first portion of the stem-loop sequence is reverse complementary to a sequence in the sensor RNA, and (c) a third region that is reverse complementary to the target RNA, and (ii) a second nucleotide sequence encoding a first cleavage domain, and (iii) a third nucleotide sequence encoding an output protein; wherein the target RNA is present in the target cell.

The present invention relates to a single base editor using adenosine deaminase acting on RNA (ADAR enzyme) and a single base editing method using same. Specifically, the present invention relates to a single base editor using an ADAR enzyme and a single base editing method using same, wherein the single base editor solves the problem of not only a target single base but also all single bases around the target single base being replaced, which is the biggest limitation of conventional base editing tools, and can selectively replace just a single base.

The present disclosure provides a method for screening a candidate agent for activity in altering a target RNA in a cell, the method including: a) contacting the candidate agent with the cell containing a sensor RNA, wherein the sensor RNA contains: (i) a first nucleotide sequence containing a sensor nucleotide sequence that hybridizes to the target RNA, wherein the sensor nucleotide sequence contains one or more editable codons, (ii) a second nucleotide sequence encoding a first cleavage domain, and (iii) a third nucleotide sequence encoding an output protein; and b) assaying for the presence of the output protein to assess the activity of the candidate agent to alter the target RNA.

The disclosure relates to the field of chronic pain and the treatment thereof. The disclosure involves antisense oligonucleotides and the use thereof in pre-mRNA and mRNA modulation, in one aspect related to RNA editing using endogenous ADAR enzymes, in targeting an adenosine in a (pre-) mRNA for human Nav1.7, preferably to change from a lysine residue to an arginine residue in the protein at position 1406 (K1406R), thereby impairing the ability of the Nav1.7 protein to act as a sodium ion channel. In another aspect the disclosure relates to exon skipping wherein antisense oligonucleotides are used to induce skipping of exon 23 in the generation of human SCN9A mRNA thereby generating a shortened Nav1.7 protein with an impaired ability to act as a sodium ion channel. The disclosure relates to methods and means to alter the Nav1.7 protein in cells of the peripheral nervous system, preferably large nociceptor cells (type Aα / Aβ), to provide a treatment of chronic pain.

A readrRNA (RNA sensing by Endogenous ADAR) molecule is a modular RNA molecule that facilitates sensing and detection of a cell type or cell status of a cell, including a cell of a mammalian nervous system, such as a neuronal or non-neuronal cell of the mammalian central and / or peripheral nervous system, and / or facilitates delivery of an effector protein to the selected cell. A composition that includes such a modular RNA molecule and another nucleic acid (linked or unlinked to the modular RNA molecule) is a CellREADR (Cell access through RNA sensing by Endogenous ADAR). CellREADR senses the presence of a selected cell RNA in a cell of a mammalian nervous system via readrRNA and leverages RNA editing mediated by ADAR (adenosine deaminase acting on RNA) for coupling the detection of a cell-defining RNA with translation of one or more effector proteins in a cell of a mammalian nervous system.

Systems and methods for predicting deamination efficiency or specificity associated with a guide RNA (gRNA) are provided. A nucleic acid sequence for the gRNA is received. Responsive to inputting a data structure into a model, a metric for an efficiency or specificity of deamination by a first Adenosine Deaminase Acting on RNA (ADAR) protein of a target nucleotide position in mRNA transcribed from a target gene is obtained as output from the model. The data structure includes an encoding of the nucleic acid sequence for the gRNA.

Disclosed herein is a readrRNA (RNA sensing by Endogenous ADAR) molecule comprising a modular RNA molecule that facilitates sensing and detection of a cell type or its status, including a cell of a mammalian nervous system, including neurons and / or neuronal cells of the area postrema of the mammalian brain, and / or facilitates delivery of an effector protein to the selected cell. A composition that includes such a modular RNA molecule and another nucleic acid (linked or unlinked to the modular RNA molecule) is a CellREADR (Cell access through RNA sensing by Endogenous ADAR). CellREADR senses the presence of a selected cell RNA in a cell of a mammalian nervous system via readrRNA and leverages RNA editing mediated by ADAR (adenosine deaminase acting on RNA) for coupling the detection of a cell-defining RNA with translation of one or more effector proteins in a cell of a mammalian nervous system.

The technology described herein is directed to compositions, kits, systems and methods related to an engineered, inducible adenosine deaminase (iAD) enzymes, including but not limited to, an engineered inducible adenosine deaminase acting on RNA (ADAR) enzyme, which can be activated in the presence of an inducer. Also described are synthetic RNA molecules, to which the iAD can be specifically recruited to edit at least one target codon, leading to decreased or increased translation of the RNA molecules depending on the specific construct. The technology described herein is also directed to systems comprising the iAD and synthetic RNA molecule, nucleic acids and vectors encoding the iAD and synthetic RNA molecule, and methods of using such systems, nucleic acids, and vectors.

The invention relates to a target gene transcriptional activity determination method based on RNA (Ribonucleic Acid) editing, which is characterized in that the method is based on mRNA (Messenger Ribonucleic Acid) editing and reporter gene technologies, a sensor reporter gene vector comprises an upstream internal reference reporter gene, a T2A (Transcription 2A), a sensor sequence, an F2A (Factor 2A) and a downstream main reporter gene, and the upstream reporter gene and the downstream reporter gene encode two different luciferases. Wherein the sensor sequence can specifically recognize mRNA expressed by a target gene. The sensor reporter gene vector is transfected into a cell with high ADAR expression, and a sensor reporter gene cell is constructed. After a sample to be detected acts on the cells, the expression of the main reporter gene can be activated in a dose-dependent manner. The method provided by the invention can be used for determining the transcriptional activity of target genes of different vectors such as plasmids, mRNA, lentiviruses, adeno-associated viruses and the like.

The present disclosure provides methods, compositions, systems, and kits for expressing a protein in a target cell.

To provide new oligonucleotides capable of utilizing the adenosine deaminase (ADAR) proteins to selectively edit target RNAs in a therapeutically effective manner.SOLUTION: The present invention provides an oligonucleotide comprising a structure: [Am]-X1-X2-X3-[Bn] wherein each of A and B is a nucleotide; m and n are each, independently, an integer from 1 to 50; X1, X2, and X 3 are each, independently, a nucleotide, wherein each of X1, X2, and X3 has a specific structure.SELECTED DRAWING: None

The invention belongs to the technical field of gene editing, and particularly relates to a low off-target RNA editing system and a construction method thereof. The fusion protein provided by the invention sequentially comprises an NES nuclear signal peptide, ADAR2 deaminase and a tDeg tag with degradation activity from an N terminal to a C terminal. The fusion protein and a guide RNA (Pepper-gRNA) containing a Pepper RNA (Ribonucleic Acid) aptamer jointly form an RNA (Ribonucleic Acid) editing system. When the target RNA exists, the Pepper aptamer masks the tDeg tag, the ADAR is stabilized, and A-to-I editing is triggered; when there is no target, the tDeg tag is exposed and mediates protein degradation, thereby inhibiting non-specific editing. According to the system, the off-target effect is remarkably reduced while the editing efficiency is maintained, and the system has high specificity, dynamic regulation and control and good safety and is suitable for precise repair of genetic disease related RNA mutation.