Methods for preparing vesicles and formulations produced therefrom
View Patent ↗The present disclosure provides methods for preparing vesicles. In some embodiments, the methods involve providing a molten mixture of vesicle forming lipids and then adding the molten mixture to an aqueous solution comprising an antigen such that antigen-containing vesicles are formed. In other embodiments, the methods involve providing a lyophilized lipid product and rehydrating the lyophilized lipid product with an aqueous solution comprising an antigen such that antigen-containing vesicles are formed. The lyophilized lipid product is prepared by melting vesicle-forming lipids to produce a molten lipid mixture and then lyophilizing the molten lipid mixture. The present disclosure also provides antigen-containing vesicle formulations prepared using these methods. The present disclosure also provides kits that include a lyophilized lipid product in a first container and an aqueous solution comprising an antigen in a second container.
1. A method of forming antigen-containing vesicles, the method comprising:
adding a molten mixture of 1-monopalmitoyl glycerol, cholesterol and dicetyl phosphate to an aqueous solution comprising an antigen, thereby forming antigen-containing vesicles, wherein the antigen-containing vesicles comprise an aqueous compartment enclosed by one or more lipid bilayers.
2. The method of claim 1 , wherein the aqueous solution comprising an antigen is temperature controlled.
3. The method of claim 2 , wherein the aqueous solution comprising an antigen is kept at a temperature of less than about 50° C. during the step of adding.
4. The method of claim 2 , wherein the aqueous solution comprising an antigen is kept at a temperature of less than about 40° C. during the step of adding.
5. The method of claim 2 , wherein the aqueous solution comprising an antigen is kept at a temperature of less than about 30° C. during the step of adding.
6. The method of claim 1 , wherein the molten mixture further comprises a transport enhancer which facilitates the transport of lipids across mucosal membranes.
7. The method of claim 1 , wherein the molten mixture does not comprise a transport enhancer which facilitates the transport of lipids across mucosal membranes.
8. The method of claim 1 , wherein the aqueous solution further comprises a lyoprotectant.
9. The method of claim 8 , wherein the lyoprotectant is sucrose.
10. The method of claim 1 , wherein the antigen is a virus.
11. The method of claim 10 , wherein the virus is an attenuated virus or an inactivated virus.
12. The method of claim 1 , wherein the antigen is selected from the group consisting of a polypeptide, a polynucleotide, and a polysaccharide.
13. The method of claim 1 , wherein the antigen is thermolabile.
14. The method of claim 1 , further comprising a step of adding an adjuvant after the antigen-containing vesicles are formed.
15. The method of claim 14 , wherein the adjuvant is a TLR-3 agonist.
16. The method of claim 15 , wherein the TLR-3 agonist is added with a lyoprotectant.
17. The method of claim 1 , wherein the molten mixture further comprises an adjuvant.
18. The method of claim 17 , wherein the adjuvant is a TLR-4 agonist.
19. The method of claim 1 , further comprising a step of lyophilizing the antigen-containing vesicles.
20. The method of claim 19 , further comprising a step of rehydrating the antigen-containing vesicles after they have been lyophilized.