Polynucleotide constructs
Disclosed are polynucleotide constructs having a strand linked to a moiety carrying one or more auxiliary moieties. Also disclosed are polynucleotide constructs interrupted with a sugar analogue, and polynucleotide constructs with stereochemical{circumflex over ( )} enriched phosphorothioates. The polynucleotide constructs may be provided as hybridized polynucleotide constructs. Also featured are methods of delivery a polynucleotide construct to a cell and methods of reducing the expression of a protein in a cell by contacting the cell with the disclosed polynucleotide construct or hybridized polynucleotide construct.
1. A polynucleotide construct or a salt thereof, comprising a linker conjugated to a polynucleotide and to a plurality of targeting moieties, wherein prior to conjugation of the targeting moieties, the linker has the structure X2,
and wherein conjugation of the targeting moieties to the linker occurs through one or more of the alkyne moieties of X2, and
wherein each of the targeting moieties comprises an N-acetyl galactosamine (GalNAc).
2. The polynucleotide construct of claim 1 , wherein prior to conjugation of the targeting moieties to the linker X2, the polynucleotide construct comprises a structure of
3. The polynucleotide construct of claim 1 , wherein the targeting moieties are selected from NAG26, NAG37, NAG38, and NAG39.
4. The polynucleotide construct of claim 1 , wherein prior to conjugation to the linker X2, each of the targeting moieties has the structure of
5. The polynucleotide construct of claim 1 , wherein the polynucleotide comprises a passenger strand and a guide strand.
6. The polynucleotide construct of claim 5 , wherein the X2 linker is bound to the passenger strand.
7. The polynucleotide construct of claim 6 , wherein the X2 linker is bound to the 3′-terminus of the passenger strand.
8. The polynucleotide construct of claim 5 , wherein the passenger strand is 15 to 32 nucleotides in length.
9. The polynucleotide construct of claim 5 , wherein the guide strand is 15 to 32 nucleotides in length.
10. The polynucleotide construct of claim 1 , wherein the X2 linker is conjugated with the targeting moieties by click reaction.
11. The polynucleotide construct of claim 1 , wherein the X2 linker comprises a terminal phosphotriester.
12. The polynucleotide construct of claim 11 , wherein the X2 linker is bound to a 3′ C 3 —OH group.
13. The polynucleotide construct of claim 1 , wherein the polynucleotide construct comprises one X2 linker.