Composition and preparation of extracellular vesicles

Abstract

In various aspects and embodiments, compositions comprising extracellular vesicles (including modified extracellular vesicles), methods for preparing formulations of extracellular vesicles, methods for modifying extracellular vesicles, and methods for using modified and / or unmodified extracellular vesicles are provided.

Claims

1. A composition comprising extracellular vesicles, wherein the extracellular vesicles comprise 0.1 to 2 μM of ATP or an ATP analog.

2. The composition according to claim 1, wherein the extracellular vesicles contain 0.5 to 1.5 μM of ATP or an ATP analog.

3. The composition according to claim 2, wherein the extracellular vesicles contain 0.5 to 1.5 μM of ATP or an ATP analog.

4. The composition according to claim 3, wherein the extracellular vesicle is an extramammary cell vesicle.

5. The composition according to claim 4, wherein the extracellular vesicle is a bovine milk extracellular vesicle.

6. The composition according to claim 5, wherein the average particle size of the extracellular vesicles is 100 to 200 nm.

7. The composition according to claim 6, wherein 99% of the extracellular vesicles have a particle size greater than 20 nm and less than 600 nm.

8. A method for preparing the composition according to claim 7, comprising contacting bovine milk cell extracellular vesicles with ATP.

9. The composition according to claim 8, wherein the extracellular vesicle carries one or more cargo molecules.

10. The composition according to claim 9, wherein the cargo molecule comprises one or more molecular types selected from the group consisting of nucleic acids, polypeptides, carbohydrates, and steroids.

11. A method comprising administering the composition described in claim 10 to a subject.

12. The method according to claim 11, wherein the composition is administered to the subject orally, intranasally, parenterally, or intravenously.

13. A method for isolating extracellular vesicles from a milk sample, comprising: (i) contacting the milk with tryptophan or a tryptophan analog at a concentration of 50 μM to 300 μM, at a pH of 2.0 to 8.0, at a temperature of 10°C to 80°C for 10 to 60 minutes; and (ii) separating the extracellular vesicles from other components of the milk by filtration and / or centrifugation.

14. The method according to claim 13, wherein the milk used in step (i) is defat.

15. The method according to claim 14, wherein the milk used in step (i) is either raw milk or pasteurized milk.

16. The method according to claim 15, further comprising subjecting the extracellular vesicles obtained from step (ii) to a filtration to remove extracellular vesicles having a particle size of less than 20 nm, and another filtration to remove vesicles having a particle size of more than 600 nm.

17. A composition comprising extracellular vesicles obtained by the method described in claim 16.

18. The composition according to claim 17, wherein the composition is freeze-dried by either rapid freezing or controlled freezing, followed by primary drying, secondary drying, and vacuum capping of the sample.

19. A composition comprising lyophilized extracellular vesicles obtained from the composition according to claim 18, to which 100 μM tryptophan and 50 mM trehalose have been added.

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