Protein enriched microvesicles and methods of making and using the same

Patent metadata

Field	Value
Publication number	US-12319934-B2
Application number	US-202318129292-A
Country	US
Kind code	B2
Filing date	Mar 31, 2023
Priority date	Jun 11, 2013
Publication date	Jun 3, 2025
Grant date	Jun 3, 2025

Abstract

Protein enriched micro-vesicles and methods of making and using the same are provided. Aspects of the methods include maintaining a cell having a membrane-associated protein comprising a first dimerization domain and a target protein having a second dimerization domain under conditions sufficient to produce a micro-vesicle from the cell, wherein the micro-vesicle includes the target protein. Also provided are cells, reagents and kits that find use in making the micro-vesicles, as well as methods of using the micro-vesicles, e.g., in research and therapeutic applications.

First claim

What is claimed is:

1. A method of introducing a target protein into a cell, the method comprising:
contacting the cell with a micro-vesicle comprising:
(a) a chimeric membrane-associated protein comprising a membrane-associated domain and a first dimerization domain, wherein the membrane-associated domain and the first dimerization domain are heterologous; and
(b) a chimeric target protein comprising a second dimerization domain, wherein the target protein is not membrane-associated in the absence of the chimeric membrane-associated protein;
wherein the first and second dimerization domains are specifically bound to each other in a dimerized complex or the first and second dimerization domains are bound to each other by a single dimerization mediator;

under conditions sufficient for the micro-vesicle to fuse with the cell and introduce the target protein into the cell.

2. The method according to claim 1 , wherein the chimeric membrane-associated protein and chimeric the target protein are present in a dimerized complex.

3. The method according to claim 2 , wherein the first and second dimerization domains are specifically bound to each other in the dimerized complex.

4. The method according to claim 2 , wherein first and second dimerization domains are bound to each other by a dimerization mediator.

5. The method according to claim 4 , wherein the dimerization mediator is a modifiable dimerization mediator.

6. The method according to claim 5 , wherein the method further comprises applying a stimulus to the micro-vesicle to modify the dimerization mediator to dissociate the complex of the chimeric target protein and the chimeric membrane-associated protein.

7. The method according to claim 2 , further comprising contacting the micro-vesicle with a dimerization disruptor to dissociate the chimeric target protein from the chimeric membrane-associated protein.

8. The method according to claim 1 , wherein the micro-vesicle further comprises a second chimeric target protein comprising a third dimerization domain.

9. The method according to claim 1 , wherein the chimeric target protein comprises a therapeutic protein.

10. The method according to claim 1 , wherein the micro-vesicle exhibits tropism for the cell.

11. The method according to claim 1 , wherein the cell is a human cell.

12. The method according to claim 1 , wherein the cell is in vitro.

13. The method according to claim 1 , wherein the cell is in vivo.

Assignees

• Takara Bio Usa Inc

Inventors

• Haugwitz Michael
• Quinn Thomas Patrick
• Farmer Andrew Alan
• Fernández Montserrat Morell

Classifications

• C12Y105/01003

  Dihydrofolate reductase (1.5.1.3) · CPC title

• C12N2760/00022

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• C12N2501/998

  Proteins not provided for elsewhere · CPC title

• C12N9/1241

  Nucleotidyltransferases (2.7.7) · CPC title

• C12N9/003

  Dihydrofolate reductase [DHFR] (1.5.1.3) · CPC title