Apolipoprotein nanodiscs with telodendrimer

Patent metadata

Field	Value
Publication number	US-11713359-B2
Application number	US-202117308921-A
Country	US
Kind code	B2
Filing date	May 5, 2021
Priority date	Dec 21, 2011
Publication date	Aug 1, 2023
Grant date	Aug 1, 2023

Abstract

The present invention provides a nanodisc with a membrane scaffold protein. The nanodisc includes a membrane scaffold protein, a telodendrimer and a lipid. The membrane scaffold protein can be apolipoprotein. The telodendrimer has the general formula PEG-L-D-(R) n , wherein D is a dendritic polymer; L is a bond or a linker linked to the focal point group of the dendritic polymer; each PEG is a poly(ethylene glycol) polymer, each R is and end group of the dendritic polymer, or and end group with a covalently bound hydrophobic group, hydrophilic group, amphiphilic compound, or drug; and subscript n is an integer from 2 to 20. Cell free methods of making the nanodiscs are also provided.

First claim

What is claimed is:

1. A nanodisc comprising:
a membrane scaffold protein;
a telodendrimer;
a lipid; and
a hydrophobic drug.

2. The nanodisc of claim 1 , wherein the membrane scaffold protein is apolipoprotein.

3. The nanodisc of claim 1 , wherein the telodendrimer has the formula:
PEG - D -( R ) n
wherein
D is a dendritic polymer having a single focal group and a plurality of end groups;
PEG is polyethyleneglycol (PEG) of 1-100 kDa linked to the focal group of the dendritic polymer;
each R is independently selected from the group consisting of the end group of the dendritic polymer and an amphiphilic compound, such that when R is not an end group each A is linked to one of the end groups; and
subscript n is an integer from 2 to 20, wherein subscript n is equal to the number of end groups on the dendritic polymer, and wherein at least half the number n of R's are each an amphiphilic compound.

4. The nanodisc of claim 3 , wherein the dendritic polymer is a poly(lysine) dendritic polymer wherein each end group is hydroxy.

5. The nanodisc of claim 3 , wherein each amphiphilic compound is cholic acid (CA).

6. The nanodisc of claim 5 , wherein the telodendrirner is selected from the group consisting of PEG 5k -D-CA 8 , PEG 5k -D-CA 4 and PEG 2k -D-CA 4 , wherein each dendritic polymer D is a poly(lysine) dendritic polymer wherein each end group is hydroxy.

7. The nanodisc of claim 1 , wherein the lipid is selected from the group consisting of a phospholipid, cholesterol, a phosphatidylcholine, a phosphatidylethanolamine, a phosphatidylglycerol, a phosphatidylserine, and a phosphatidylinositol.

8. The nanodisc of claim 7 , wherein the lipid is selected from the group consisting of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-ditetradecanoyl-sn-glycero-3-phosphocholine (DMPC), 1-tetradecanoyl-2-hexadecanoyl-sn-glycero-3-phosphocholine (MPPC), 1,2-dioctadecanoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE), 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DPPE), 1,2-distearoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DSPC), 1,2-dipalmitoyl-sn-glycero-3-phospho-(1′-rac-glycerol) (DPPG), 1-tetradecanoyl-2-hexadecanoyl-sn-glycero-3-phosphoglycerol (MPPG) and cholesterol.

9. The nanodisc of claim 7 , wherein the lipid is DMPC.

10. The nanodisc of claim 1 , wherein the nanodisc is less than about 100 nm in size.

11. The nanodisc of claim 1 , wherein the nanodisc is less than about 10 nm in size.

12. The nanodisc of claim 1 , herein the ratio of lipid to telodendrimer is from about 200:1 to about 5:1 (w/w).

13. The nanodisc of claim 1 , wherein the ratio of lipid to telodendrimer is about 9:1 (w/w).

14. A cell-free method of making a nanodisc, the method comprising:
forming a vesicle comprising a telodendrimer and a lipid, wherein the ratio of lipid to telodendrimer is from about 500:1 to about 1:1 (w/w); and
forming a reaction mixture of the vesicle and a membrane scaffold protein in the absence of a cell, thereby preparing the nanodisc
the method further comprising
incorporating a hydrophobic drug in the reaction mixture and/or
loading the hydrophobic drug into the nanodisc.

15. The method of claim 14 , wherein the membrane scaffold protein is apolipoprotein.

16. The method of claim 14 , wherein the reaction mixture further comprises a lysate, a buffer, and a polymerase.

17. The method of claim 14 , wherein the lipid:telodendrimer are present in a ratio of about 99:1 (w/w).

18. The method of claim 14 , wherein the lipid:telodendrimer are present in a ratio of about 9:1 (w/w).

19. The nanodisc of claim 1 , wherein the hydrophobic drug is embedded within the nanodisc.

Assignees

• L Livermore Nat Security Llc

Inventors

• Luo Juntao
• He Wei
• Lam Kit S
• Henderson Paul
• Coleman Matthew A

Classifications

• C07K17/02 Primary

  Peptides being immobilised on, or in, an organic carrier · CPC title

• C07K14/775 Primary

  Apolipopeptides · CPC title