SiRNAs with vinylphosphonate at the 5′ end of the antisense strand
Inventors
WEINGÄRTNER, Adrien • Bethge, Lucas
Assignees
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Publication Number
US-11560563-B2
Publication Date
2023-01-24
Expiration Date
Abstract
The present invention relates to nucleic acids for inhibiting expression of a target gene in a cell, comprising at least one duplex region that comprises at least a portion of a first strand and at least a portion of a second strand that is at least partially complementary to the first strand, wherein said first strand is at least partially complementary to at least a portion of RNA transcribed from said target gene to be inhibited. The first strand of the nucleic acid has a terminal 5′ (E)-vinylphosphonate nucleotide that is linked to the second nucleotide in the first strand by a phosphodiester linkage.
Core Innovation
The patent provides a nucleic acid for inhibiting expression of a target gene in a cell. The nucleic acid comprises at least one duplex region including a first strand and a second strand that is at least partially complementary to the first strand, and the first strand is at least partially complementary to RNA transcribed from the target gene to be inhibited. The first strand has a terminal 5 (E)-vinylphosphonate nucleotide linked to the second nucleotide in the first strand by a phosphodiester linkage.
The chemically modified pattern of nucleotides is defined such that all even-numbered nucleotides of the first strand are modified by a 2-F modification, and all odd-numbered nucleotides of the first strand are modified by a 2-OMe modification. The second strand has a corresponding position-specific modification pattern relative to nucleotides 11-13 of the first strand, with 2-F modifications at corresponding positions and 2-OMe modifications at other corresponding positions.
The document also describes GaINAc-conjugated nucleic acid designs for gene inhibition, including triantennary and related multi-ligand GaINAc-conjugated nucleic acid designs with ligand-linker chemistry. Embodiments include a nucleic acid conjugated to a triantennary ligand via a phosphate or modified phosphate linker, with GaINAc affinity for hepatic ASGP-R receptors, and embodiments with terminal 5 (E)-vinylphosphonate nucleotide linkages.
Claims Coverage
The claim coverage centers on a duplex nucleic acid for inhibiting expression of a target gene in a cell, with a terminal 5 (E)-vinylphosphonate nucleotide and a defined positional modification scheme on both strands. The claim features repeatedly identify the same core architecture: duplex complementary to target gene RNA, phosphodiester linkage at the 5 terminal vinylphosphonate, and 2-F/2-OMe patterning by nucleotide position.
Duplex region complementary to target gene RNA
A nucleic acid for inhibiting expression of a target gene in a cell comprising at least one duplex region including a first strand and a second strand that is at least partially complementary to the first strand, where the first strand is at least partially complementary to RNA transcribed from the target gene to be inhibited.
Terminal 5 (E)-vinylphosphonate nucleotide linked by phosphodiester
The first strand has a terminal 5 (E)-vinylphosphonate nucleotide linked to the second nucleotide in the first strand by a phosphodiester linkage.
Even/odd position-specific 2-F and 2-OMe modifications on first strand
All even-numbered nucleotides of the first strand are modified by a 2-F modification, and all odd-numbered nucleotides of the first strand are modified by a 2-OMe modification.
Position-specific 2-F and 2-OMe modifications on second strand relative to first strand nucleotides 11-13
Nucleotides of the second strand in positions corresponding to nucleotides 11-13 of the first strand are modified by a 2-F modification, and all nucleotides of the second strand other than the nucleotides corresponding to nucleotides 11-13 of the first strand are modified by 2-OMe modification.
GaINAc ligand conjugation to the 5 end of the second strand
A conjugate whose ligand portion includes one or more GaINAc ligands, one or more GaINAc ligand derivatives, or a GaINAc moiety conjugated to the 5 end of the second strand.
Composition including the nucleic acid or conjugate with physiologically acceptable excipient
A composition including a nucleic acid from the claims or a conjugate from the claims together with a physiologically acceptable excipient.
The claims are centered on a chemically modified duplex nucleic acid that inhibits expression of a target gene by targeting RNA transcribed from that gene, with a terminal 5 (E)-vinylphosphonate nucleotide, a defined phosphodiester linkage, and specific 2-F/2-OMe modification rules on both strands. Additional claim coverage includes GaINAc-containing conjugates and compositions formulated with a physiologically acceptable excipient.
Stated Advantages
Improved target mRNA knockdown.
In vitro stability in acidic tritosome lysate.
In vivo duration/efficacy.
Improved potency and in vivo duration for thiophosphate-containing conjugates.
Inhibiting expression of a target gene in a cell.
Documented Applications
Inhibiting expression of a target gene in a cell.
GalNAc-conjugated siRNA/vinylphosphonate constructs for inhibition of target mRNA.
Gene inhibition using triantennary and related multi-ligand GaINAc-conjugated nucleic acid designs with GaINAc targeting to hepatic ASGP-R receptors.
Formulation and delivery contexts including lipid-nucleic acid complexes and pharmaceutical compositions.
Disease treatment by administering the nucleic acid or conjugate to a subject.
A conjugate in which a ligand portion includes GaINAc ligands, GaINAc ligand derivatives, or a GaINAc moiety conjugated to the 5 end of the second strand.
A composition including the nucleic acid or conjugate together with a physiologically acceptable excipient.
Knockdown of TMPRSS6 using GalNAc siRNA conjugates in primary murine hepatocytes.
Serum stability assessment and analytical purity and identity assessments in the example context.
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