Lipid bilayer carrier for drugs or imaging agents

Abstract

Disclosed are carriers for drugs and/or MR imaging agents having a lipid bilayer shell comprising a phospholipid having two terminal alkyl chains, one being a short chain having a chain length of at most seven carbon atoms, the other being a long chain having a chain length of at least fifteen carbon atoms. The mixed long/short chain phospholipids serve to tune the release properties of the carrier. Preferred phospholipids are phosphatidylcholines.

Claims

1. A composition comprising a thermosensitive carrier having a semipermeable lipid bilayer shell, wherein the semipermeable lipid bilayer comprises a phospholipid having two terminal alkyl chains, one being a short chain having a chain length of at most five carbon atoms, the other being a long chain having a chain length of at least fifteen carbon atoms.

2. The composition of claim 1, wherein a difference in length between the short chain and the long chain is between eleven and sixteen carbon atoms.

3. The composition of claim 1, wherein the long chain has a length of at least sixteen carbon atoms.

4. The composition of claim 3, wherein the long chain has at most twenty carbon atoms.

5. The composition of claim 1, wherein the long chain has at most twenty carbon atoms.

6. The composition of claim 1, wherein the long chain has at most thirty carbon atoms.

7. The composition of claim 1, wherein the phospholipid is selected from the group consisting of phospholipids satisfying formula (I) and phospholipids satisfying formula (II), ##STR00007## wherein R is an alkyl chain of fifteen to thirty carbon atoms, and n is an integer of 1 to 4.

8. The composition of claim 7, wherein R is selected from the group consisting of C.sub.15H.sub.31 and C.sub.17H.sub.35, and n is 1 to 4.

9. The composition of claim 1, further comprising at least one drug substance.

10. The composition of claim 1, further comprising at least one magnetic resonance imaging (MRI) contrast enhancing substance.

11. The composition of claim 1, further comprising: at least one drug substance, and at least one magnetic resonance imaging (MRI) contrast enhancing substance.

12. The composition of claim 1, further comprising a substance contained within the shell, wherein the carrier is configured for the in vivo release of the substance, and wherein the substance is selected from the group consisting of drugs, MRI contrast enhancing substances, and combinations thereof.

13. The composition of claim 1, wherein a membrane of the carrier includes a paramagnetic agent which is not a chemical shift reagent.

14. The composition of claim 13, wherein the carrier is aspherical.

15. The composition of claim 13, wherein the paramagnetic agent includes an amphiphilic compound comprising a lanthanide complex and having an apolar tail.

16. The composition of claim 1, further comprising a water solution of a paramagnetic shift reagent encapsulated within an interior compartment defined by the shell.

17. A method for magnetic resonance imaging (MRI) guided delivery of a drug to a subject, the method comprising: administering to the subject a thermosensitive carrier comprising a semipermeable lipid bilayer shell, wherein the semipermeable lipid bilayer comprises a phospholipid having two terminal alkyl chains, one being a short chain having a chain length of at most five carbon atoms, the other being a long chain having a chain length of at least fifteen carbon atoms, the thermosensitive carrier carrying a drug and an MRI contrast enhancing substance; allowing the carrier to release the drug and the MRI contrast enhancing substance; and rendering a magnetic resonance image using the contrast provided by the contrast enhancing substance.

Description

DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 presents a bar diagram showing the melting phase transition temperatures of temperature sensitive liposomes (TSLs) containing either 10 mol % of Lyso-PC or 10 mol % of 1.sub.n,R in the lipid bilayer and 250 mM ProHance and doxorubicin in their aqueous lumen.

(2) FIGS. 2A and 2B shows fluorescence and longitudinal relaxivity results for TSLs containing 10 mol % acetylated PCs (1n,R: n=0, RC15H31 or C17H35) in the lipid bilayer and 250 mM ProHance and doxorubicin in their aqueous lumen.

(3) In particular, FIG. 2A depicts the fluorescence and longitudinal relaxivity during a linear temperature increase (0.5 K/min) from 300 K to 323 K for LTSLs containing 10,R (RC15H31), and FIG. 2B depicts the fluorescence and longitudinal relaxivity during a linear temperature increase (0.5 K/min) from 300 K to 323 K for LTSLs containing 10,R (RC17H35), in HBS. Meanwhile, FIG. 2C depicts the release of doxorubicin as a function of temperature for LTSLs containing 10,R (RC15H31), and FIG. 2D depicts the release of doxorubicin as a function of temperature for LTSLs containing 10,R (RC17H35), in HBS.

(4) FIG. 3A shows fluorescence and longitudinal relaxivity during a linear temperature increase (0.5 K/min) from 300 K to 323 K for LTSLs containing 13,R (RC15H31) and FIG. 3B shows fluorescence and longitudinal relaxivity during a linear temperature increase (0.5 K/min) from 300 K to 323 K for LTSLs containing 13,R (RC17H35), in HBS. FIG. 3C depicts the release of doxorubicin as a function of temperature for LTSLs containing 13,R (RC15H31) and FIG. 3D depicts the release of doxorubicin as a function of temperature for LTSLs containing 13,R (RC17H35), in HBS.

(5) FIG. 4 shows the release of doxorubicin as a function of temperature for LTSLs containing of DPPC:DPPE-PEG2000, showing no quantitative release of doxorubicin.

(6) FIG. 5 shows the longitudinal relaxivity (rl) of temperature-sensitive liposomes containing 250 mM ProHance and doxorubicin during a linear temperature increase (0.5 K/min). At 310 K, the longitudinal relaxivity of paramagnetic liposomes containing 1.sub.0,R (n=0 and RC.sub.17H.sub.35) displays the lowest value. For the sake of legibility, a legend is contained in the Figure.

(7) FIG. 6 shows the longitudinal relaxivity of liposomes (TTSL and NTSL) containing doxorubicin and 250 mM [Gd(hpdo3a)(H.sub.2O)]. For the sake of legibility, a legend is contained in the Figure.

(8) FIG. 7 shows the difference in the longitudinal relaxivity between 310 K and 315 K for liposomes encapsulating 250 mM ProHance and doxorubicin. Reference experiments have been performed with non-temperature sensitive liposomes (NTSL; HSPC:Cho1:DPPE-PEG2000=75:20:3) and traditional temperature-sensitive liposomes (TTSLs; DPPC:HSPC:Chol:DPPE-PEG2000=50:25:15:3).

EXAMPLE 1

(9) A compound of the above-identified type 1.sub.n,R having n=0, and RC.sub.17H.sub.35, i.e. 1(n=0, RC.sub.17H.sub.35), was prepared as follows. A solution of 4-dimethylaminopyridine (149.1 mg, 1.22 mmol) in dichloromethane (8 mL) dried on molecular sieves was added to 1-myristoyl-2-stearoyl-sn-glycero-3-phosphocholine, abbreviated as MSPC (211.8 mg, 0.404 mmol). Subsequently, acetic anhydride (115 L, 1.23 mmol) was added and the mixture was stirred for 30 hours at room temperature under nitrogen atmosphere to yield a colorless solution. Methanol (8 mL) was added and the solvent was removed at room temperature under reduced pressure. The crude mixture was dissolved in chloroform (8 mL) and the organic layer was extracted three times with a solution of MeOH (8 mL) and 0.1 M HCl (8 mL). The mixture was centrifuged (30 minutes, 4000 rpm) to induce fast phase separation. The remaining organic layer was filtered, and then concentrated on a rotary evaporator at room temperature under reduced pressure. The crude product was dissolved in acetone (30 mL) and the solution was cooled to 20 C. to induce precipitation. To obtain the solid the mixture was centrifuged (30 minutes, 4000 rpm, 19 C.) and the solvent was decanted. The obtained product was washed with acetone, centrifuged (30 minutes, 4000 rpm, 19 C.) and dried with the aid of a nitrogen stream to obtain 1(n=0, RC.sub.17H.sub.35 (0.132 g) 58% Yield. The product was analyzed using .sup.1H- and .sup.13C NMR spectroscopy

EXAMPLE 2

(10) The compound 1(n=3, RC.sub.17H.sub.35) was obtained similar to 1(n=0, RC.sub.17H.sub.35) of Example 1, in 70% yield (0.172 g) Instead of acetic anhydride, valeric anhydride (240 L, 1.22 mmol) was used. This product did not dissolve completely in acetone therefore purification was done by stirring the crude product in acetone before cooling to 20 C. The product was analyzed using .sup.1H- and .sup.13C NMR spectroscopy.

EXAMPLE 3

(11) Liposomes were formed by the lipid film hydration technique coupled with sequential extrusion. The phospholipids were dissolved in a solution of CHCl3/MeOH (4:1 v/v). The solvents were gently removed under reduced pressure and a thin lipidic film was obtained. The lipidic film was hydrated in a solution of 250 mM [Gd(hpdo3a)(H2O)] in 120 mM ammonium acetate buffer. The dispersion was extruded several times through polycarbonate membrane filters with pore diameters of 400, 200 and 100 nm, subsequently. After extrusion, the extraliposomal buffer was replaced by HEPES Buffered Saline (HBS), pH 7.4 (20 mM HEPES, 137 mM NaCl) by gel filtration through a PD-10 column (GE Healthcare). Subsequently, a doxorubicin solution in HBS (5 mg/mL) was added to the liposomes at a 20:1 phospholipid to doxorubicin weight ratio and incubated for 90 min at 37 C. Finally, the liposomes were passed through another PD-10 column to remove traces of non-encapsulated doxorubicin and free [Gd(hpdo3a)(H2O)].

(12) The composition of the phospholipid bilayer is given in Table 1 below.

(13) TABLE-US-00001 TABLE 1 Mixed short/long chain PC DPPC DPPE-PEG2000 Formulation (all 10 mol %) (mol %) (mol %) A 1.sub.n, R (n = 0, R = C.sub.15H.sub.31) 86 4 B 1.sub.n, R (n = 0, R = C.sub.17H.sub.35) 86 4 C 1.sub.n, R (n = 1, R = C.sub.17H.sub.35) 86 4 D 1.sub.n, R (n = 2, R = C.sub.17H.sub.35) 86 4 E 1.sub.n, R (n = 3, R = C.sub.15H.sub.31) 86 4 F 1.sub.n, R (n = 3, R = C.sub.17H.sub.35) 86 4 G 0 96 4

EXAMPLE 4

(14) The liposomes of Example 3 were loaded with doxorubicin and 250 mM [Gd(hpdo3a)(H2O)]. The Tm and the hydrodynamic diameter of the liposomes (formulation A-E) were determined by differential scanning calorimetry (DSC) and dynamic light scattering (DLS), respectively. As shown in Table 2 and FIG. 1, the Tm of the phospholipid bilayer can be modulated by the incorporation of 1n,R.

(15) TABLE-US-00002 TABLE 2 Formulation n R T.sub.m (K) Diameter (nm) A 0 C.sub.15H.sub.31 313.1 196 B 0 C.sub.17H.sub.35 313.7 197 C 1 C.sub.17H.sub.35 313.0 118 D 2 C.sub.17H.sub.35 312.9 110 E 3 C.sub.15H.sub.31 311.8 132 F 3 C.sub.17H.sub.35 312.6 136 G 313.8 109

EXAMPLE 5

(16) The release of doxorubicin and [Gd(hpdo3a)(H2O)] from the aqueous lumen of the liposomes was studied as a function of temperature probing the fluorescence at 590 nm and the longitudinal relaxivity (rl), respectively (FIGS. 2-6).

(17) For all studied systems, the fluorescence of the encapsulated doxorubicin crystals is quenched at 300 K. However, at temperatures close to the Tm and upon the release from the liposome, a steep increase in the fluorescence signal is observed. As long as the MRI agent stays intraliposomal, the longitudinal relaxivity is limited by the transmembrane water exchange rate. When the temperature is raised the agent is released and MR contrast enhancement is observed. The incorporation of mixed short/long chain PCs in the bilayer allows one to tune the membrane properties that relate to drug release and the transmembrane water exchange rate and thereby the MR contrast enhancement (FIG. 5-7). By tuning the transmembrane water exchange rate, the properties of temperature-sensitive liposomes can be optimized for applications in MR image-guided drug delivery. Moreover, the co-encapsulation of doxorubicin and a T1-agent in a temperature-sensitive liposome consisting of mixed short/long chain PCs offers the opportunity to monitor the drug release process.