DNA-cell conjugates

Patent metadata

Field	Value
Publication number	US-9969970-B2
Application number	US-201615057604-A
Country	US
Kind code	B2
Filing date	Mar 1, 2016
Priority date	Apr 8, 2009
Publication date	May 15, 2018
Grant date	May 15, 2018

Abstract

The present invention provides conjugates of DNA and cells by linking the DNA to a native functional group on the cell surface. The cells can be without cell walls or can have cell walls. The modified cells can be linked to a substrate surface and used in assay or bioreactors.

First claim

What is claimed is:

1. A method of preparing a conjugate of a live cell and a nucleic acid moiety, wherein (i) the cell has a surface comprising a native functional group comprising an amino acid selected from the group consisting of lysine, cysteine, tyrosine, threonine, serine, aspartic acid, glutamic acid and tryptophan, and wherein the native functional group is not an azide-modified sugar, and (ii) the cell has no cell wall, the method comprising:
contacting the cell with an activated nucleic acid moiety under conditions such that the activated nucleic acid moiety reacts with the native functional group to covalently link the nucleic acid directly to the native functional group.

2. The method of claim 1 , wherein the cell is a primary cell.

3. The method of claim 1 , wherein the cell is a mammalian cell.

4. The method of claim 1 , wherein the cell is a stem cell.

5. The method of claim 1 , wherein the native functional group comprises lysine.

6. The method of claim 1 , wherein the nucleic acid moiety comprises a member selected from the group consisting of an oligonucleotide, DNA, RNA, PNA and an aptamer.

7. The method of claim 1 , wherein the nucleic acid moiety comprises single-stranded DNA.

8. The method of claim 1 , wherein the nucleic acid moiety comprises from about 10 to about 200 nucleic acids.

9. The method of claim 1 , wherein the nucleic acid moiety comprises an aptamer.

10. The method of claim 1 , wherein the nucleic acid moiety comprises a linker.

11. The method of claim 1 , wherein the conjugate comprises a mammalian cell comprising lysine on the cell surface; and a single-stranded deoxy-nucleic acid covalently linked to the lysine via an amide.

12. A method of preparing a conjugate of a live cell and a nucleic acid moiety, wherein (i) the cell has a surface comprising a native functional group selected from the group consisting of lysine and tryptophan, and (ii) the cell has no cell wall, the method comprising contacting the cell with an activated nucleic acid moiety under conditions such that the activated nucleic acid moiety reacts with a primary amine of the native functional group to form a covalent bond.

13. The method of claim 12 , wherein the cell is a primary cell.

14. The method of claim 12 , wherein the cell is a mammalian cell.

15. The method of claim 12 , wherein the cell is a stem cell.

16. The method of claim 12 , wherein the cell has not undergone metabolic engineering to introduce an azide modified sugar.

17. The method of claim 12 , wherein the native functional group comprises lysine.

18. The method of claim 12 , wherein the nucleic acid moiety comprises a member selected from the group consisting of an oligonucleotide, DNA, RNA, PNA and an aptamer.

19. The method of claim 12 , wherein the nucleic acid moiety comprises single-stranded DNA.

20. The method of claim 12 , wherein the nucleic acid moiety comprises from about 10 to about 200 nucleic acids.

21. The method of claim 12 , wherein the nucleic acid moiety comprises an aptamer.

22. The method of claim 12 , wherein the nucleic acid moiety comprises a linker.

23. The method of claim 12 , wherein the conjugate comprises a mammalian cell comprising lysine on the cell surface; and a single-stranded deoxy-nucleic acid covalently linked to the lysine via an amide.

24. A method of preparing a conjugate of a live cell and a nucleic acid moiety, wherein the cell has a surface comprising a native functional group comprising an amino acid selected from the group consisting of lysine, cysteine, tyrosine, threonine, serine, aspartic acid, glutamic acid, and tryptophan, and wherein the cell has no cell wall, the method comprising: contacting the cell with an activated nucleic acid moiety under conditions such that the nucleic acid moiety reacts with the amino acid to form a covalent bond.

25. The method of claim 24 , wherein the cell is a primary cell.

26. The method of claim 24 , wherein the cell is a mammalian cell.

27. The method of claim 24 , wherein the cell is a stem cell.

28. The method of claim 24 , wherein the cell has not undergone metabolic engineering to introduce an azide modified sugar.

29. The method of claim 24 , wherein the native function group comprises lysine.

30. The method of claim 24 , wherein the nucleic acid moiety comprises a member selected from the group consisting of an oligonucleotide, DNA, RNA, PNA and an aptamer.

31. The method of claim 24 , wherein the nucleic acid moiety comprises single-stranded DNA.

32. The method of claim 24 , wherein the nucleic acid moiety comprises from about 10 to about 200 nucleic acids.

33. The method of claim 24 , wherein the nucleic acid moiety comprises an aptamer.

34. The method of claim 24 , wherein the nucleic acid moiety comprises a linker.

Assignees

• Univ California

Inventors

• Hsiao Shih-Chia
• Francis Matthew B
• Bertozzi Carolyn
• Mathies Richard
• Chandra Ravi
• Douglas Erik
• Twite Amy
• Toriello Nicholas
• Onoe Hiroaki

Classifications

• A61K47/6901

  Conjugates being cells, cell fragments, viruses, ghosts, red blood cells or viral vectors · CPC title

• C12N5/0006 Primary

  Modification of the membrane of cells, e.g. cell decoration · CPC title