Polymer nanoparticle and DNA nanostructure compositions and methods for non-viral delivery

Patent metadata

Field	Value
Publication number	US-12109223-B2
Application number	US-202418444146-A
Country	US
Kind code	B2
Filing date	Feb 16, 2024
Priority date	Dec 3, 2020
Publication date	Oct 8, 2024
Grant date	Oct 8, 2024

Abstract

The invention relates to polymer nanoparticle and DNA nanostructure delivery compositions for non-viral delivery, and methods therefor. More particularly, the invention relates to polymer nanoparticle delivery compositions, such as reversible addition-fragmentation chain transfer (RAFT) polymer compositions, and DNA nanostructure delivery compositions, such as DNA origami compositions, for the delivery of more than one payload, or for the delivery of a nucleic acid construct payload of 3 kB or more, and methods therefor.

First claim

What is claimed is:

1. A RAFT block copolymer comprising
a. a first terminus comprising a first capping unit derived from a first chain transfer agent in a RAFT copolymerization process;
b. a first block prepared from one or more monomer units covalently attached to the first reactive functional unit, and having a molecular weight (M n ) in the range of about 20 kDa to about 80 kDa and a degree of polymerization in the range of about 20 to about 400;
c. a second block prepared from one or more monomer units covalently attached to the first block, and having a molecular weight (M n ) in the range of about 5 kDa to about 80 kDa and a degree of polymerization in the range of about 10 to about 500; and
d. a second terminus comprising a second capping unit derived from a second chain transfer agent;
wherein the RAFT block copolymer has one or more of an overall molecular weight (M n ) in the range of about 25 kDa to about 160 kDa, and overall degree of polymerization in the range of about 30 to about 900, a size in the range of about of about 10 to about 60 nm, and a maximum corona-to-core ratio (CCR) of about 1 to about 4.

2. The RAFT block copolymer of claim 1 , wherein the first block is prepared from one or more monomer units selected from the group consisting of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, and methyl methacrylate.

3. The RAFT block copolymer of claim 1 , wherein the first block is prepared from one of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, or methyl methacrylate.

4. The RAFT block copolymer of claim 1 , wherein the second block is prepared from one or more monomer units selected from the group consisting of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, and methyl methacrylate.

5. The RAFT block copolymer of claim 1 , wherein the second block is a random copolymer prepared from two different monomer units independently selected from the group consisting of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, and methyl methacrylate.

6. The RAFT block copolymer of claim 1 , wherein the second block is a random copolymer prepared from three different monomer units independently selected from the group consisting of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, and methyl methacrylate.

7. The RAFT block copolymer of claim 1 , wherein each chain transfer agent is independently selected from the group consisting of bis(carboxymethyl)trithiocarbonate, bis(2-amino-2-oxoethyl) trithiocarbonate, bis[4-(2-hydroxyethoxycarbonyl)benzyl] trithiocarbonate, 4-cyano-4-(ethylsulfanylthiocarbonyl) sulfanylvpentanoic acid, 4-cyano-4-((phenylcarbonothioyl)thio)pentanoic acid, and 4-cyano-4-[(dodecylsulfanylthiocarbonyl)sulfanyl] pentanoic acid.

8. The RAFT block copolymer of claim 1 , wherein the first capping unit is of the formula

wherein * represents a point of covalent attachment to the first block.

9. The RAFT block copolymer of claim 1 , wherein the second capping unit is of the formula

wherein * represents a point of covalent attachment to the second block, and R is —SC 2 -C 12 alkyl or C 6 H 5 .

10. A RAFT block copolymer conjugate comprising a RAFT block copolymer according to claim 1 , and further comprising a biomolecule, drug, or label covalently attached to the RAFT block copolymer through a functional group on one of the first or second chain transfer agents that is incorporated into the RAFT block copolymer.

11. The RAFT block copolymer complex of claim 10 , wherein the payload comprises nucleic acids.

12. The RAFT block copolymer complex of claim 10 , wherein the nucleic acids comprise DNA or RNA.

13. The RAFT block copolymer complex of claim 10 , wherein the nucleic acids comprise a ribonucleoprotein.

14. The RAFT block copolymer complex of claim 10 , wherein the payload is a nucleic acid and the nucleic acid payload is of a size selected from the group consisting of 3 kB or more, 3.5 kB or more, 4 kB or more, 4.5 kB or more, 5 kB or more, 5.5 kB or more, 6 kB or more, 6.5 kB or more, 7 kB or more, 7.5 kB or more, 8 kB or more, and 8.5 kB or more.

15. A RAFT block copolymer complex comprising a RAFT block copolymer according to claim 1 , and further comprising a payload complexed to the RAFT block copolymer through electrostatic interaction.

16. The RAFT block copolymer complex of claim 15 , wherein the payload comprises nucleic acids.

17. The RAFT block copolymer complex of claim 15 , wherein the nucleic acids comprise DNA or RNA.

18. The RAFT block copolymer complex of claim 15 , wherein the payload is a nucleic acid and the nucleic acid payload is of a size selected from the group consisting of 3 kB or more, 3.5 kB or more, 4 kB or more, 4.5 kB or more, 5 kB or more, 5.5 kB or more, 6 kB or more, 6.5 kB or more, 7 kB or more, 7.5 kB or more, 8 kB or more, and 8.5 kB or more.

19. A method of preparing a RAFT block copolymer comprising:
i. contacting a first chain transfer agent, a first initiator and one or more monomer units to provide a first block;
ii. contacting a second chain transfer agent, a second initiator and one or more monomer units to provide a second block
iii. contacting the first block and the second block under conditions capable of coupling the blocks to provide the RAFT block copolymer.

20. The method of claim 19 , wherein the first chain transfer agent is a diamino or dihydroxy chain transfer agent, and the second chain transfer agent is a dicarboxylic acid chain transfer agent.

21. The method of claim 19 , wherein the first chain transfer agent is bis(2-amino-2-oxoethyl) trithiocarbonate or bis[4-(2-hydroxyethoxycarbonyl)benzyl] trithiocarbonate, and the second chain transfer agent is bis(carboxymethyl)trithiocarbonate.

22. The method of claim 19 , wherein the first chain transfer agent is a dicarboxylic acid chain transfer agent, and the second chain transfer agent is a diamino or dihydroxy chain transfer agent.

23. The method of claim 19 , wherein the first chain transfer agent is bis(carboxymethyl)trithiocarbonate, and the second chain transfer agent is bis(2-amino-2-oxoethyl) trithiocarbonate or bis[4-(2-hydroxyethoxycarbonyl)benzyl] trithiocarbonate.

24. The method of claim 19 , wherein the one or more monomer units in step (i) are independently selected from the group consisting of 2-dimethylaminoethyl acrylate, 2-(diethylamino) ethyl methacrylate, 2-(diisopropylamino) ethyl methacrylate, butyl methacrylate, ethyl acrylic acid, propyl acrylic acid, (hydroxyethyl)methacrylate, and methyl methacrylate.

25. The method o

Assignees

• Battelle Memorial Institute

Inventors

• Duong Anthony D
• Gupta Cherry
• Sims Jr Kenneth R
• Huk Danielle
• Lilly Jacob
• Kute Stephanie M
• Koeris Mike

Classifications

• C08F2438/03

  Use of a di- or tri-thiocarbonylthio compound, e.g. di- or tri-thioester, di- or tri-thiocarbamate, or a xanthate as chain transfer agent, e.g . Reversible Addition Fragmentation chain Transfer [RAFT] or Macromolecular Design via Interchange of Xanthates [MADIX] · CPC title

• C08F293/005

  using free radical "living" or "controlled" polymerisation, e.g. using a complexing agent · CPC title

• A61K47/6455 Primary

  Polycationic oligopeptides, polypeptides or polyamino acids, e.g. for complexing nucleic acids · CPC title

• B82Y5/00

  Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery · CPC title

• C12N2310/11

  Antisense · CPC title