EP0393145A1 - Vesiculation spontanee de liposomes multilamellaires - Google Patents

Vesiculation spontanee de liposomes multilamellaires

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Publication number
EP0393145A1
EP0393145A1 EP89901663A EP89901663A EP0393145A1 EP 0393145 A1 EP0393145 A1 EP 0393145A1 EP 89901663 A EP89901663 A EP 89901663A EP 89901663 A EP89901663 A EP 89901663A EP 0393145 A1 EP0393145 A1 EP 0393145A1
Authority
EP
European Patent Office
Prior art keywords
liposomes
vesicles
lipid
incubated
medium
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP89901663A
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German (de)
English (en)
Other versions
EP0393145A4 (en
Inventor
Thomas D. Madden
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Elan Pharmaceuticals LLC
Original Assignee
Liposome Co Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US07/136,267 external-priority patent/US4963297A/en
Application filed by Liposome Co Inc filed Critical Liposome Co Inc
Publication of EP0393145A1 publication Critical patent/EP0393145A1/fr
Publication of EP0393145A4 publication Critical patent/EP0393145A4/en
Withdrawn legal-status Critical Current

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/127—Liposomes
    • A61K9/1277—Processes for preparing; Proliposomes

Definitions

  • the present invention is directed to a method of forming unilamellar vesicles without the use of homogenization, filtration, sonication, or extrusion techniques, or other techniques that require energy input to the system, or exposure of lipids to harsh environments.
  • harsh environments include for example detergent or extreme pH environments.
  • Liposomes are completely closed lipid bilayer membranes containing an entrapped aqueous volume. Liposomes may be unilamellar vesicles (possessing a single membrane bilayer) or multilameller vesicles (onion-like structures characterized by multiple membrane bilayers, each separated from the next by an aqueous layer).
  • the bilayer is composed of two lipid monolayers having a hydrophobic "tail” region and a hydrophilic "head” region.
  • the structure of the membrane bilayer is such that the hydrophobic (nonpolar) "tails" of the lipid monolayers orient towards the center of the bilayer while the hydrophilic "heads" orient towards the aqueous phase.
  • the original liposome preparation of Bangham et al. involves suspending phospholipids in an organic solvent which is then evaporated to dryness leaving a phospholipid film on the reaction vessel. Next, an appropriate amount of aqueous phase is added, the mixture is allowed to "swell," and the resulting liposomes which consist of multilamellar vesicles (MLVs) are dispersed by mechanical means. MLVs so formed may be used in the practice of the present invention.
  • Another class of multilamellar liposomes that may be used as the starting liposomes of this invention are those characterized as having substantially equal lamellar solute distribution.
  • This class of liposomes is denominated as stable plurilamellar vesicles (SPLV) as defined in U.S. Patent No. 4,522,803 to Lehk, et al., reverse phase evaporation vesicles (REV) as described in U.S. Patent No. 4,235,871 to Papahadjopoulos et al., monophasic vesicles as described in U.S. Patent No.
  • SPLV plurilamellar vesicles
  • REV reverse phase evaporation vesicles
  • Liposomes are comprised of lipids; the term lipid as used herein shall mean any suitable material resulting in a bilayer such that a hydrophobic portion of the lipid material orients toward the interior of the bilayer while a hydrophilic portion orients toward the aqueous phase.
  • the lipids which can be used in the liposome formulations of the present invention are the phospholipids such as phosphatidylcholine (PC) and phosphatidylglycerol (PG), more particularly dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG) .
  • Liposomes may be formed and vesiculated using DMPG, or DMPG mixed with DMPC in, for example, a 3:7 mole ratio, respectively.
  • organic solvents may be used to suspend the lipids.
  • Suitable organic solvents are those with intermediate polarities and dielectric properties, which solubilize the lipids, and include but are not limited to halogenated, aliphatic, cycloaliphatic, or aromatic-aliphatic hydrocarbons, such as benzene, chloroform, methylene chloride, or alcohols, such as methanol, ethanol, and solvent mixtures such as benzene:methanol (70:30).
  • solutions mixture ' s in which the lipids and other components are uniformly distributed throughout
  • Solvents are generally chosen on the basis of their biocompatability, low toxicity, and solubilization abilities.
  • the starting multilamellar liposomes and resulting unilamellar liposomes of the present invention may contain lipid soluble bioactive agents. Such agents are typically associated with the lipid bilayers of the liposomes.
  • bioactive agent is understood to include any compound having biological activity; e.g., lipid soluble drugs such as non steroidal antinflammatory drugs such as ibuprofen, ind ' omethacin, sulindac, piroxicam, and naproxen, antinoeplastic drugs such as doxorubicin, vincristine, vinblastine, methotrexate and the like, and other therapeutic agents such as anesthetics such as dibucaine, cholinergic agents such as pilocarpine, antihistimines such as benedryl, analgesics such as codeine, anticholinergic agents such as atropine, antidepressants such as imiprimine, antiarrythmic agents such as propranolol, and
  • the liposomes of the invention may be used in a liposome-drug delivery system.
  • a bioactive agent such as a drug is associated with the liposomes and then administered to the patient to be treated.
  • a bioactive agent such as a drug
  • U.S. Patent No. 3,993,754 Sears, U.S. Patent No. 4,145,410; Papahadjopoulos et al., U.S. Patent No. 4,235,871; Schnieder, U.S. Patent No. 4,224,179; Lenk et al., U.S. Patent No. 4,522,803; and Fountain et al., U.S. Patent No. 4,588,578.
  • amphotericin B is an extremely toxic antifungal polyene antibiotic with the single most reliability in the treatment of life-threatening fungal infections (Taylor et al., Am. Rev. Respir. Dis., 1982, 125:610-611). Because amphotericin B is a hydrophobic drug, it is insoluble in aqueous solution and is commercially available as a colloidal dispersion in desoxycholate, a detergent used to suspend it which in itself is toxic.
  • Amphotericin B methyl ester and amphotericin B have also been shown to be active against the HTLV-III/LAV virus, a lipid-enveloped retrovirus, shown in the etiology of acquired immuno-deficiency syndrome (AIDS)- (Schaffner et al., Biochem, Pharmacol., 1986, 35:4110-4113).
  • AIDS acquired immuno-deficiency syndrome
  • amphotericin B methyl ester ascorbic acid salt (water soluble) and amphotericin B were added to separate cultures of HTLV-III/LAV infected cells and the cells assayed for replication of the virus.
  • liposome-encapsulated amphotericin B reports of the use of liposome-encapsulated amphotericin B.
  • Such liposomes comprise phospholipid, for example dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG) in a 7:3 mole ratio, and cholesterol.
  • DMPC dimyristoylphosphatidylcholine
  • DMPG dimyristoylphosphatidylglycerol
  • LD_ n s Acute toxicity studies (LD_ n s) and in vitro assays comparing free and liposome-entrapped amphotericin B showed lower toxicities using the liposomal preparations with substantially unchanged antifungal potency.
  • Lopez-Berestein et al. J. Infect. Dis., 1986, 151:704-710 administered liposome-encapsulated amphotericin B to patients with systemic fungal infections.
  • the liposomes comprised a 7:3 mole ratio of DMPC:DMPG, and the drug was encapsulated at a greater than 90% efficiency.
  • Liposomes to buffer the toxicity of entrapped drugs with little or no decrease in efficacy is becoming increasingly well established. Therefore, there is an increasing need to be able to form liposomes of all types which have these qualities.
  • Unilamellar vesicles are clearly preferred for certain types of in vivo drug delivery over multilamellar vesicles, as well as for studies of membrane-mediated processes. As used as in vivo delivery vehicles, for example, unilamellar vesicles are cleared more slowly from the blood than are MLVs, and exhibit an enhanced distribution to the lungs and possibly bone marrow.
  • the methods known for producing these type vesicles relied upon harsh treatment of multilamellar vesicles, such as extrusion through filters, or other physically damaging processes requiring energy input such as sonication, homogenization or milling.
  • Chemical treatment techniques employing harsh detergents or solutions at high or low pH to form unilamellar vesicles have also been employed.
  • the present invention advances the art in that it allows formation of unilamellar vesicles from multilamellar vesicles without the heretofore harsh treatments required, but through the incubation of the liposomes in low ionic strength media at selected temperatures. Additionally, the unexpected simplicity of preparation of these systems is complemented by the highly defined conditions under which they may be formed. The fact that vesiculation of these lipids occurs only around about the lipid phase transition temperature
  • the present invention discloses a method for spontaneously forming unilamellar vesicles from multilamellar vesicles (MLVs).
  • MLVs comprise lipids, and unilamellar vesicles are formed by incubating the multilamellar vesicles in low ionic strength medium at neutral pH, around about the transition temperature of the 0 u lipids.
  • the lipids comprise phospholipids, specifically phosphatidylglycerol alone or in combination with phosphatidylcholine, more specifically dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol, in a 7:3 mole ratio.
  • the liposomes are incubated at about 22-26°C, preferably about 24°C in a medium of between about 0 mM and 25 mM salt. More preferably, the medium comprises about 0 - lOmM salt at pH of about 7.0 to about 8.0, preferably pH 7.6 and incubation time is about 15 minutes to about 24 hours.
  • FIGURE 1 demonstrates vesiculation of DMPC:DMPG (7:3) MLVs as a function of ionic strength of the incubation medium.
  • DMPC:DMPG (lOmM) was hydrated at 4°C in the media shown below and incubated at 24°C (see Examples 1 and 2).
  • Sample media were H.O (open circles); 2mM HEPES (closed squares); lOmM NaCl, 2mM HEPES, pH 7.6, (open triangles); or 25 mM NaCl, 2 mM HEPES, pH 7.6 (closed triangles) .
  • FIGURE 2 are 31 P-NMR spectra of DMPC:DMPG.
  • ' Lipid (10 mM) was hydrated in H.O at 4°C and its spectrum was recorded at 30°C (A). The same lipid mixture was then incubated at 24°C for 1 hour (B) and 12 hours (C).
  • DMPC:DMPG (7:3 mole ratio) hydrated in 150 mM NaCl, 10 mM HEPES, pH 7.6 and incubated at 24°C for 12 hours is shown in (D).
  • FIGURE 3 are P-NMR spectra for mixtures of phosphatidylcholine with phosphatidylglycerol.
  • Lipid (10 mM) was hydrated in 150 mM NaCl, 10 mM HEPES, pH 7.6 (A,B,C,) or 2 mM HEPES, pH 7.6 (D,E,F,G,H,J) and incubated at 24°C (A,B,C,G,H,J) or 10°C
  • the unilamellar liposomes of this invention are formed by the exposure of multilamellar liposomes to conditions of low ionic strength media at neutral pH, and incubation temperatures around about the gel-to-liquid crystalline transition temperature (T ) . Under such incubation conditions, MLVs vesiculate to form unilamellar vesicles.
  • T gel-to-liquid crystalline transition temperature
  • the liposomes of the present invention are preferably comprised of phospholipids, specifically dimyristoylphosphatidylglycerol (DMPG) or with dimyristoylphosphatidylcholine (DMPC).
  • DMPG dimyristoylphosphatidylglycerol
  • DMPC dimyristoylphosphatidylcholine
  • DMPC dimyristoylphosphatidylcholine
  • LUVs or SUVs from MLVs usually requires aggressive disruption, for example, by sonication (Huang, 1969, Biochemistry, 8:344) or extrusion through polycarbonate filters (Hope et al., 1985, Biochim. Biophys. Acta, 812, 55), as mentioned above.
  • incubation media of high ionic strength high than about 50 mM salt
  • vesiculation occurs at a decreased rate, or not at all. Vesiculation occurs as a function of lowering the ionic strength of the incubation medium.
  • MLVs vesiculate spontaneously when exposed to low ionic strength incubation media (about 10 mM ionic strength and less) when incubated around about the T of the lipid.
  • Any ionic species solutions may be used as incubation media, such as the salts sodium chloride, potassium chloride, and others. While a range, therefore, of about 0-25 mM salt in the incubation medium will promote vesiculation, the optimum conditions are around about 0-10 mM salt.
  • Vesiculation of MLV systems may be determined by incubating the liposomes in low ionic strength medium for 15 minutes to several hours, at around the gel-to-liquid crystalline transition temperature of the lipids used. Whether vesiculation has occurred may be measured by the size of the resulting liposomes using quasi-elastic light scattering, (unilamellar versus multilamellar), visualization of the resulting vesicles using freeze-fracture 31 electron microscopy, and P-NMR analysis of lineshape and spectrum width. For example, narrow spectrum width and isotropic signal is indicative of unilamellar vesicle structure, while a low field shoulder and high field peaks are indicative of larger vesicles.
  • Liposomes incorporating a bioactive agent, such as a drug, such as for example, amphotericin B may be formed according to the processes of the invention as follows. Amphotericin B is suspended in an aqueous solution, for example distilled water, by sonication. The suspended drug is then admixed with a suspension of lipid in aqueous solution, such as distilled water or sodium chloride solution. The mixture is incubated at or above the transition temperature of the lipid employed, with the resultant formation of vesicles.
  • a bioactive agent such as a drug, such as for example, amphotericin B
  • dimyristoylphosphatidylglycerol is used alone or in combination to form the liposomes, and when the lipid has been admixed with an aqueous solution having an ionic strength of about 0 mM to about 25 mM salt, and incubated at about the transition temperature (T ) of the lipid (i.e., at about 22-24°C), the liposomes spontaneously vesiculate, forming large unilamellar vesicles (LUVs).
  • T transition temperature
  • LUVs large unilamellar vesicles
  • DMPG can be used alone or, for example, with other lipid such as with DMPC, e.g., in a 3:7 mole ratio of DMPC:DMPG.
  • DMPC lipid
  • These lipids can be co-lyophilized from a 70:30 v/v solution of benzene:methanol, and stored at -20°C until use.
  • MLVs are prepared by hydrating the lipid (for example, a total of lipid of 13.5 umoles/ml) in aqueous solution such as distilled water or buffer at 4°C.
  • the lipid When formation of amphotericin B-containing LUVs is desired, the lipid is hydrated in an aqueous solution of ionic strength of about ) mM to about 25 mM salt, and incubated at aout 23°C. Amphotericin B, dispersed in distilled water by bath sonication, at a concentration of about 0.98 umoles/ml is then added to the hydrated lipid and incubated at about 23°C for about one hour, resulting in LUVs containing amphotericin B. These proportions of lipid and amphotericin B result in about a 7 mole % ratio of amphotericin B.
  • the lipids of the present invention may be hydrated to form liposomes using any available aqueous solutions, for example, distilled water, saline, or aqueous buffers.
  • buffers include but are not limited to buffered salines such as phosphate buffered saline (“PBS”), tris-(hydroxymethyl)-aminomethane hydrochloride (“tris”) buffers, and preferably N-2-hydroxyethyl piperazine-N-2-ethane sulfonic acid (“HEPES”) buffer.
  • PBS phosphate buffered saline
  • tris tris-(hydroxymethyl)-aminomethane hydrochloride
  • HEPES N-2-hydroxyethyl piperazine-N-2-ethane sulfonic acid
  • buffers are preferably used at pH of about 7.0 to about 8.0, preferably about pH 7.6. If required, the ionic strength of the medium may be adjusted to physiological values following the vesiculation procedure
  • the liposomes of the present invention may be dehydrated either prior to or following vesiculation, thereby enabling storage for extended periods of time until use.
  • Standard freeze-drying equipment or equivalent apparatus may be used to lyophilize the liposomes.
  • Liposomes may also be dehydrated simply by placing them under reduced pressure and allowing the suspending solution to evaporate. Alternatively, the liposomes and their surrounding medium may be frozen prior to dehydration.
  • Such dehydration may be performed in the presence of one or more protectants such as protective sugars, according to the process of Janoff et al. , PCT 86/01103, published February 27, 1986, and incorporated herein by reference.
  • the liposomes resulting from the processes of the present invention can be used therapeutically in mammals, including man, in the treatment of infections or conditions which benefit from the employment of liposomes which give for example, sustained release, reduced toxicity, and other qualities which deliver the drug in its bioactive form.
  • the mode of administration of the preparation may determine the sites and cells in the organism to which the compound will be delivered.
  • the liposomes of the present invention can be administered alone but will generally be administered in admixture with a pharmaceutical carrier selected with regard to the intended route of administration and standard pharmaceutical practice.
  • the preparations may be injected parenterally, for example, intra-arterially or intravenously.
  • the preparations may also be administered via oral, subcutaneous, or intramuscular routes.
  • parenteral administration they can be used, for example, in the form of a sterile aqueous solution which may contain other solutes, for example, enough salts or glucose to make the solution isotonic.
  • Other uses, depending upon the particular properties of the preparation may be envisioned by those skilled in the art.
  • the liposomes of the present invention may be incorporated into dosage forms such as gels, oils, emulsions, and the like. Such preparations may be administered by direct application as a cream, paste, ointment, gel, lotion or the like.
  • the liposomes of this invention encapsulating a bioactive agent can be used in the form of tablets, capsules, losenges, troches, powders, syrups, elixirs, aqueous solutions and suspensions, and the like.
  • carriers which can be used include lactose, sodium citrate and salts of phosphoric acid.
  • Various disintegrants such as starch, and lubricating agents, such as magnesium stearate, sodium lauryl sulfate and talc, are commonly used in tablets.
  • useful diluents are lactose and high molecular weight polyethylene glycols.
  • the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening and/or flavoring agents can be added.
  • DMPC:DMPG (7:3 M ratio) was lyophilized from benzene:methanol (70:30 v/v).
  • the lipid was hydrated to 10 mM with distilled water pH 7.6, at 4°C, forming MLVs.
  • the suspension was then incubated at 24°C for 15 minutes.
  • QELS studies showed the resulting liposomes to be about 200 nm in diameter, corresponding to LUVs.
  • Example 1 The procedures and materials of Example 1 were employed using 150 mM NaCl, 2 mM HEPES buffer as the hydrating solution. QELS measurements revealed no change in liposome size (no vesiculation) after incubation.
  • Figure 1 demonstrates vesiculation by plotting the vesicle diameter (obtained by quasi elastic light scattering, QELS) as an indication of MLV or LUV against time of incubation, and shows that the rate of vesiculation at 24°C is directly related to the ionic strength of the hydration medium.
  • Figure 2 demonstrates the
  • DMPG 10 mM
  • DMPG 10 mM
  • HEPES 4°C
  • pH 7.6 a pH 7.6
  • MLVs MLVs were incubated at 24°C for 15 minutes, and the sample analyzed by QELS.
  • the resulting liposomes were unilamellar (LUVs).
  • Example 13 This Example may be compared with Example 13, where liposomes made of a 3:7 M ratio of DMPC:DMPG incubated in lOmM NaCl (Example 13) only approach the 200 nm diameter vesicles of Example 3 after 5 hours incubation.
  • Example 1 A 7:3 M ratio of dry DMPC:DMPG was equilibrated at 32°C in a water-saturated atmosphere for 60 minutes, and then the procedures and materials of Example 1 were followed to make MLVs (10 mM lipid), using 2 mM HEPES as hydration medium and an incubation temperature of 32°C. After 6 hours incubation, no vesiculation had occurred as QELS measurements revealed the liposomes had a mean diameter of greater than 2 microns.
  • This Example is a control for the incubation of the liposome systems around about the T of the lipid; it shows this incubatic parameter is an important requirement of the invention.
  • Example 4 The procedures and materials of Example 4 were employed using 2 M HEPES as the hydration medium and an incubation temperature of 15°C. After 6 hours incubation, no vesiculation had occurred as QELS measurements revealed the liposomes had a mean diameter greater than 2 microns.
  • This Example serves as a further control for T being an important incubation parameter. No vesiculation occured at this incubation temperature. However, when this system was incubated at 24°C, the liposomes rapidly vesiculated.
  • Example 1 The procedures and materials of Example 1 were employed, using a 7:3 M ratio of D0PC:DMPG.
  • the lipid was hydrated with 2 mM HEPES and incubated at 24°C for 16 hours.
  • Example 7 The procedures and materials of Example 7 were employed, using a 7:3 M ratio of DMPC:D0PG.
  • the lipid was hydrated with 2 mM HEPES and incubated at 24°C for 16 hours.
  • P-NMR spectroscopy revealed little or no vesiculation.
  • Example 7 The procedures and materials of Example 7 were employed, using a 7:7:3:3 M ratio of DOPC:DMPC:D0PG:DMPG.
  • the lipid was hydrated with 2 mM HEPES and incubated at 24°C for 16 hours.
  • 31 3 (A-J) demonstrates the P-NMR spectra for such samples incubated at either 10°C or 24°C. All spectra are characteristic of large vesicles in the bilayer phase (MLVs); the samples did not vesiculate.
  • Example 7 The procedures and materials of Example 7 were employed, using a 7:3 M ratio of D0PC:DMPG.
  • the lipid was hydrated with 150 ' mM NaCl, mM HEPES and incubated for 16 hours at 24° C .
  • Example 7 The procedures and materials of Example 7 were employed, using a :3 M ratio of DMPC:D0PG.
  • the lipid was hydrated with 150 mM NaCl, mM HEPES and incubated for 16 hours at 24°C.
  • Example 7 The procedures and materials of Example 7 were employed, using a 7:7:3:3 M ratio of D0PC:DMPC:D0PG:DMPG.
  • the lipid was hydrated with 150 mM NaCl, 2 mM HEPES and incubated for 16 hours at 24°C.
  • Example 3 The procedures and materials of Example 3 were employed, using a 3:7 M ratio of DMPC:DMPG.
  • the lipid was hydrated in 10 mM NaCl, 2 mM HEPES at pH 7.6 at 4°C, forming MLVs.
  • the suspension was then incubated for 1 hour at 24°C.
  • QELS measurements revealed that vesiculation of the MLVs had formed LUVs.
  • Lipid (14.8 umol/ml, 7:3 mol ratio of DMPC:DMPG) was hydrated in distilled water and incubated at 4°C. The resulting MLVs were extruded through two stacked polycarbonate filters ten times using the LUVET procedure.
  • Amphotericin B was dispersed in distilled water using a bath sonicator at a concentration of 10.8 umol/ml.
  • the amphotericin B disperson was added to the lipid suspension to a final lipid and amphotericin B concentration of 13.5 umol/ml and 0.98 umol/ml, respectively.
  • 20 ml of the sample were centrifuged at 15,000 X g for 30 minutes in a Ti60 or SW 27 rotor (Beckman) at 22°C in a Beckman L8-60 ultracentrifuge. The supernatant free amphotericin B was removed without disturbing the liposome pellet.
  • the resulting liposomes were measured by quasi-elastic light scattering to be larger than 1.0 urn in diameter.
  • Example 14 The materials and procedures of Example 14 were employed, but wherein the lipid suspended in distilled water was incubated with the amphotericin B at 22°C. The resulting liposomes were unilamellar and measured at about 0.1 - 0.2 urn in mean diameter by quasi elastic light scattering.

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  • Health & Medical Sciences (AREA)
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Abstract

Nouveau procédé permettant de produire des vésicules unilamellaires à partir de vésicules multilamellaires. On forme lesdits vésicules sans avoir recours à des procédés de rupture physique ou chimique, connus dans l'art et permettant de former des visicules unilamellaires. On met en incubation les liposomes à un pH neutre, à ou proche de la température de transition des lipides utilisés, dans un milieu à faible résistance ionique tel que de l'eau distillée. Les liposomes peuvent comprendre des agents bioactifs, tels que les produits pharmaceutiques hydrophobes toxiques comme l'amphotéricine B antibiotique de polyène, et peuvent être utilisés in vivo ou in vitro. Les compositions de lipides sont de préférence une combinaison des phospholipides dimyristoylphosphatidylcholine (DMPC) et dimyristoylphosphatidylglycérol (DMPG) dans un rapport de mole d'environ 7:3.
EP19890901663 1987-12-22 1988-12-22 Spontaneous vesiculation of multilamellar liposomes Withdrawn EP0393145A4 (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US07/136,267 US4963297A (en) 1987-12-22 1987-12-22 Spontaneous vesticulation of multilamellar liposomes
US23670088A 1988-08-25 1988-08-25
US236700 1988-08-25
US136267 1998-08-19

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EP0393145A1 true EP0393145A1 (fr) 1990-10-24
EP0393145A4 EP0393145A4 (en) 1991-09-11

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WO (1) WO1989005636A1 (fr)

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DE4107153A1 (de) * 1991-03-06 1992-09-10 Gregor Cevc Praeparat zur wirkstoffapplikation in kleinsttroepfchenform
DE4107152C2 (de) * 1991-03-06 1994-03-24 Gregor Cevc Präparate zur nichtinvasiven Verabreichung von Antidiabetica
IS1685B (is) * 1990-12-11 1998-02-24 Bracco International B.V. Aðferð við að búa til fitukúlur (liposomes) sem eru gæddar auknum hæfileika til að draga í sig og halda í sér aðskotaefnum
FR2754272B1 (fr) * 1996-10-08 1998-11-13 Rhone Poulenc Rorer Sa Procede de preparation de compositions pour le transfert d'acides nucleiques
DE10255285A1 (de) * 2002-11-26 2004-06-03 Mcs Micro Carrier Systems Gmbh Selbst formende Phospholipid-Gele
EP2344198B1 (fr) * 2008-09-27 2020-11-04 Jina Pharmaceuticals Inc. Préparations pharmaceutiques à base de lipide(s) à usage topique
AU2016256979B2 (en) 2015-05-04 2021-01-28 Versantis AG Method for preparing transmembrane pH-gradient vesicles
CN112545994B (zh) * 2020-11-19 2022-10-14 河南科技大学 一种小粒径、高稳定性多室及单室脂质体的同步制备方法

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US4310505A (en) * 1979-11-08 1982-01-12 California Institute Of Technology Lipid vesicles bearing carbohydrate surfaces as lymphatic directed vehicles for therapeutic and diagnostic substances
DE3374837D1 (en) * 1982-02-17 1988-01-21 Ciba Geigy Ag Lipids in the aqueous phase
US4515736A (en) * 1983-05-12 1985-05-07 The Regents Of The University Of California Method for encapsulating materials into liposomes
JPS6150912A (ja) * 1984-08-16 1986-03-13 Shionogi & Co Ltd リポソ−ム製剤の製造法
JPS6176414A (ja) * 1984-09-21 1986-04-18 Shionogi & Co Ltd リポソーム製剤の製法

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Publication number Priority date Publication date Assignee Title
US4663167A (en) * 1984-04-16 1987-05-05 The Board Of Regents Of The University Of Texas System Composition and method for treatment of disseminated fungal infections in mammals

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Title
See also references of WO8905636A1 *

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CA1330199C (fr) 1994-06-14
JPH03501732A (ja) 1991-04-18
EP0393145A4 (en) 1991-09-11
WO1989005636A1 (fr) 1989-06-29

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