Apolipoprotein nanodiscs with telodendrimer
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.
1. A nanodisc comprising:
a membrane scaffold protein;
a telodendrimer;
a lipid; and
a hydrophobic drug,
wherein the hydrophobic drug is covalently bound to a dendritic segment of the telodendrimer.
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 R 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 telodendrimer is selected from the group consisting of PEG 5k -D-CA 3 , 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) (DSPG), 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 hydrophobic drug is selected from the group consisting of amphotericin B and SN38.
11. The nanodisc of claim 1 , wherein the nanodisc is less than about 100 nm in size.
12. The nanodisc of claim 1 , wherein the nanodisc is less than about 10 nm in size.
13. The nanodisc of claim 1 , wherein the ratio of lipid to telodendrimer is from about 200:1 to about 5:1 (w/w).
14. The nanodisc of claim 1 , wherein the ratio of lipid to telodendrimer is about 9:1 (w/w).
15. 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
wherein the telodendrimer comprises a hydrophobic drug covalently bound to a dendritic segment of the telodendrimer.
16. The method of claim 15 , wherein the membrane scaffold protein is apolipoprotein.
17. The method of claim 15 , wherein the reaction mixture further comprises a lysate, a buffer, and a polymerase.
18. The method of claim 15 , wherein the lipid: telodendrimer are present in a ratio of about 99:1 (w/w).
19. The method of claim 15 , wherein the lipid: telodendrimer are present in a ratio of about 9:1 (w/w).