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WO2009033588A2 - Système d'administration auto-contrôlée d'insuline - Google Patents

Système d'administration auto-contrôlée d'insuline Download PDF

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Publication number
WO2009033588A2
WO2009033588A2 PCT/EP2008/007149 EP2008007149W WO2009033588A2 WO 2009033588 A2 WO2009033588 A2 WO 2009033588A2 EP 2008007149 W EP2008007149 W EP 2008007149W WO 2009033588 A2 WO2009033588 A2 WO 2009033588A2
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WO
WIPO (PCT)
Prior art keywords
vesicle
insulin
liposome
vesicle according
glucose
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.)
Ceased
Application number
PCT/EP2008/007149
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English (en)
Other versions
WO2009033588A3 (fr
Inventor
Wolfgang Burkhardt
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.)
F Hoffmann La Roche AG
Original Assignee
F Hoffmann La Roche AG
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
Application filed by F Hoffmann La Roche AG filed Critical F Hoffmann La Roche AG
Publication of WO2009033588A2 publication Critical patent/WO2009033588A2/fr
Anticipated expiration legal-status Critical
Publication of WO2009033588A3 publication Critical patent/WO2009033588A3/fr
Ceased 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—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/28—Insulins

Definitions

  • the present invention relates to a self-controlled insulin delivery system.
  • Type 1 diabetics need to control their glucose-level continually to keep it within the range of 80-120 mg/dl.
  • measures have to be taken, e.g. intake of carbohydrates in the case of hypoglycemia, or injection of insulin in the case of hyperglycemia. This means that in order to maintain blood glucose it is necessary to adjust them.
  • the present invention provides liposomes which can be used in a method to automatically adjust insulin levels in a glucose dependent manner.
  • glucokinase converts glucose to glucose-6-phosphate. This mediates the release of insulin from vesicles by exocytosis.
  • glucokinase is the glucose sensor for insulin release.
  • the present invention relates in particular to vesicles comprising a glucose sensor integrated into the membrane of said vesicle, which further comprises glucokinase, insulin or analogs thereof, Mg++ and ATP.
  • a glucose sensor integrated into the membrane of said vesicle, which further comprises glucokinase, insulin or analogs thereof, Mg++ and ATP.
  • insulin refers to both insulin and to analogs of insulin.
  • said vesicle is a liposome.
  • vesicle relates to a small and enclosed compartment, which comprises at least one membrane enclosing the compartment.
  • compartment relates to the core of the vesicle.
  • the membrane separates the contents of the core from the outside environment of the vesicle.
  • membrane as used herein refers to a lipid bilayer enclosing a compartment.
  • liposome as used herein relates to a spherical vesicle with a membrane comprising a phospholipid and a cholesterol bilayer.
  • lipid as used herein relates to an amphiphilic class of hydrocarbon-containing organic compounds.
  • insulin is encapsulated in a vesicle comprising insulin independent glucose sensors.
  • said insulin is human insulin (Seq ID No. 4) or porcine insulin (Seq ID No. 5) or an analog thereof.
  • Preferred analogs are Glulysine insulin (human insulin Lys(B3)-Glu(B29)); Lispro insulin (human insulin Lys(B28)-Pro(B29)); Aspart insulin (human insulin Asp(B28)); Glargine insulin (human insulin Gly(A21)-L- Arg(B31, B32)) or Detemir insulin (Thr(B30) is deleted, Lys(B29) is ⁇ -myristoylated) or combinations thereof.
  • said glucose sensor which is integrated into the membrane of the liposome is GLUTl, GLUT2 or GLUT3. More preferably, said GLUTl, GLUT2 or GLUT3 are human GLUTl, human GLUT2 or human GLUT3. Human Glucose transporter
  • GLUT2 is defined by Seq ID No. 6.
  • Human Glucose transporter GLUT2 is defined by Seq ID No. 1.
  • Human Glucose transporter GLUT3 is defined by Seq ID No. 2.
  • the liposome may further comprise Zn++.
  • said glucokinase is human glucokinase (hexokinase 4, EC 2.7.1.2) (Seq ID No. 3).
  • the concentration of Mg++ is equal or higher than the dissociation constant of glucokinase.
  • said concentration is between 0.5 to 15 mM, more preferably between 2 to 10 mM.
  • ATP adenosine triphosphate
  • the ATP concentration is preferably >10 mM, more preferably >30 mM.
  • the vesicles hereinbefore described may further comprise multiple layers, each of which comprises the ingredients listed above.
  • Such liposomes are also known as multi- lamellar vesicles (MLVs).
  • MLVs multi- lamellar vesicles
  • Such vesicles in which protein drugs are encapsulated are e.g. disclosed in Anderson et al. 1994, CYTOKINE 6, p.92-101.
  • the term multi-lamellar liposome (MLV) as used herein relates to a liposome with a multiple layer structure wherein said layers are separated by aqueous medium.
  • the vesicles may comprise a phospholipid and cholesterol bilayer.
  • Said phospholipid is selected from one or more phospholipids of the group comprising DMPC, DOTAP, DOPC, MOPC, egg phosphatidylethanolamine, DOPE, egglecithin, dicetylphosphate, dipalmitoyl lecithin, digalactosyl diglyceride, sphingosine, lecithin, egg phosphatidylcholine, dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylcholine and/or long-chain or intermediate-chain phosphatidylcholine.
  • the vesicles may, as a non-limiting example, comprise DMPC, DOTAP, DOPC, MOPC, egg phosphatidylethanolamine,DOPE and/or cholesterol.
  • DMPC is dimyristoyl phosphatidyl choline
  • DOTAP is l,2-Dioleoyl-3-Trimethylammonium- Propane
  • DOPC is l ⁇ -di-oleoyl-sn-glycero-S-phosphocholine
  • DOPE dioleylphosphatidylethanolamine
  • MOPC is l-myristoyl-2-oleyl-phosphatidylcholine
  • Cholesterol is also known as 10,13-dimethyl-17-(6-methylheptan-2-yl)- 2,3,4,7,8,9,1 l ⁇ .U.lSJ ⁇ . ⁇ -dodecahydrolH-cyclopentataJphenanthren-S-ol.
  • the vesicles may also comprise a mixture of lipids.
  • the mixture comprises egglecithin, dicetylphosphate and cholesterol.
  • the mixture comprises cholesterol, dipalmitoyl lecithin, digalactosyl diglyceride and sphingosine.
  • the vesicle comprises lecithin and cholesterol, preferably in a molar ratio of 9:2.
  • the vesicle comprises egg phosphatidylcholine and cholesterol or dipalmitoylphosphatidylcholine and cholesterol, preferably dipalmitoylphosphatidylcholine and cholesterol.
  • the vesicle comprises phospholipid, cholesterol and triglyceride.
  • the vesicle may comprise long-chain phosphatidylcholine or intermediate- chain phosphatidylcholine.
  • the vesicles may additionally also comprise a neutral lipid.
  • Said neutral lipid may be a triglyceride.
  • Such a triglyceride may be a short-chain triglyceride.
  • all embodiments of the vesicles described above may additionally comprise PEG (polyethylene glycol) on the surface of the liposome.
  • PEG polyethylene glycol
  • insulin preferably human insulin
  • expression and purification of glucokinase is also well known in the art (e.g. in Lange et al, Biochem. J. (1991), 277, 159-163).
  • GLUTl maybe isolated from erythrocyte membranes or expressed in a suitable expression system as described below for GLUT2.
  • GLUT2 can be expressed in a suitable expression system, e.g. in RINm5F cells (Eisner et al., Diabetologia), and purified, e.g. by affinity chromatography using a GLUT2-specific antibody.
  • a suitable expression system e.g. in RINm5F cells (Eisner et al., Diabetologia)
  • purified e.g. by affinity chromatography using a GLUT2-specific antibody.
  • GLUT3 may be isolated from neuron membranes or expressed in a suitable expression system as described above for GLUT2.
  • liposomes for encapsulation of drugs is well known (Katre et al., Am J
  • the present invention also relates to a method of preparing the vesicles hereinbefore described comprising: adding an aqueous solution comprising a glucose sensor integrated into the membrane of a vesicle, glucokinase, insulin or analogs thereof, Mg++ and ATP to a lipid- organic phase to obtain an emulsion followed by removing the organic.
  • the present invention also relates to a method of preparing a vesicle comprising the steps of (a) sterilizing powdered lipid, (b) adding sterile powdered lipid to aqueous protein, (c) rapidly mixing the suspension of b), (d) bath sonicating said mixture, (e) freezing said mixture, (f) thawing said mixture, (g) repeating steps c) to f) two times.
  • lipid powder or solution 15000 to 150 000 rads of gamma irradiation are used, preferably using a 137Cs source.
  • the protein mixture is present in a 0.9 % saline buffer comprising Mg++, Zn++ and ATP as described above.
  • the sterile lipid is added.
  • a lipid to aqueous ratio of 10 to 1000 mg lipid / ml aqueous protein solution, preferably 100 to 500, more preferably 200 to 400, containing 0.1 to 10 mg protein (insulin, insulin-independent glucose transporter, prepferably GLUTl, GLUT2 or GLUT3, and glucokinase) per ml, preferably 0.5 to 2 mg is used.
  • the suspension is mixed rapidly, bath sonicated, frozen, thawed, and this cycle is repeated twice.
  • a method of preparing a self-controlled delivery system using multivesicular liposomes is also disclosed.
  • An insulin-containing aqueous phase is emulsified into a lipid- organic phase to form a water-in-oil emulsion.
  • the organic phase (solvent) preferably comprises chloroform or methylene chloride.
  • the lipid solution preferably comprises one or two phosphatidylcholines (more preferably mono-unsaturated), a phosphatidylglycerol, cholesterol and one or two triglycerides. Then, this water-in-oil emulsion is further emulsified into an aqueous solution to obtain a water-in-oil-in-water emulsion.
  • the solvent is then removed to convert the water-in-oil-in-water emulsion into a multivesicular liposome particle.
  • the emulsion is flushed with nitrogen to remove the solvent.
  • the particles are concentrated.
  • the potency is then adjusted to the final product concentration.
  • the resulting multivesicular liposomes will comprise insulin, Mg++, Zn++, insulin-independent glucose transporter, preferably GLUTl, GLUT2 or GLUT3 and glucokinase as disclosed above (Katre et al., Am J Drug Deliv 2004, 2 (4), p 213-227).
  • the invention also relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a vesicle as hereinbefore described and a pharmaceutically acceptable excipient.
  • the invention also relates to a kit comprising a pharmaceutical composition hereinbefore described.
  • the liposome described above is administered to a patient suffering from Type 1 diabetes or a patient suffering from Type 2 diabetes who is treated with insulin.
  • the present invention also relates to the use of the liposomes described above for the preparation of a medicament for treatment of diabetic patients.
  • said patients are Type I diabetes patients.
  • interstitial glucose diffusion into the vesicles can occur mediated by an insulin-dependent glucose transporter depending on glucose concentration.
  • glucose is converted to glucose-6-phosphate by glucokinase.
  • Glucose-6-phosphate can no longer diffuse out of the liposome.
  • the osmotic pressure increases in a glucose-dependent manner, leading to an influx of water from the interstitium.
  • the liposomes are swelling and leaks will occur which allow a release of ingredients, including insulin. An increase of the surface of 2% to 3% may lead to such leaks.
  • the liposomes will burst if the osmotic pressure rises further, and the ingredients are released completely.
  • the liposome When the liposome comprises multiple layers, the outermost layer will become unstable first, optionally followed by the next layer depending on glucose concentration.
  • the depot function of multi-layer liposomes is well known (Katre et al., Am J Drug Deliv 2004, 2 (4), p 213-227).
  • Unused liposomes will be phagocytosed after 12 to 24 hours. Thus, no accumulation of unactivated liposomes occurs. Phagocytosis may be controlled by lipid composition (Nanobiotechnologie II - für in der Medizin und Pharmazie, 2004, V. Wagner and D. Wechsler, Vol. 50, Ed: VDI Technologie scholar GmbH imset des BMBF) especially by pegylation of the surface of the liposomes, which can increase the half life to about 20 h.
  • Lipid powder or solution is irradiated with 100000 rads of gamma irradiation (e.g. using a 137Cs source).
  • Sterile filtration of protein solutions can be done using any suitable method. Such methods are well known to the skilled person.
  • Aqueous carrier free protein in 0.9 % saline with 7.5 mM MgCl2 and 5 mM ATP is added to sterile, powdered lipid.
  • the suspension is mixed rapidly, bath sonicated, frozen, thawed, and this cycle is repeated twice.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Endocrinology (AREA)
  • Diabetes (AREA)
  • Dispersion Chemistry (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Immunology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicinal Preparation (AREA)

Abstract

La présente invention concerne un liposome sensible au glucose (de préférence un liposome multilamellaire) comprenant un transporteur de glucose indépendant de l'insuline intégré dans la membrane du liposome, qui comporte également une glucokinase, de l'insuline, Mg++ et ATP. Le glucose se diffuse dans le liposome dépendant de sa concentration et sera converti en glucose-6-phosphate. L'accumulation de glucose-6-phosphate entraîne éventuellement la rupture du liposome. Ainsi, ces liposomes permettent un ajustement automatique de niveaux d'insuline d'une manière dépendante de glucose et sont donc utiles pour le traitement de patients diabétiques.
PCT/EP2008/007149 2007-09-11 2008-09-02 Système d'administration auto-contrôlée d'insuline Ceased WO2009033588A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
EP07116131.9 2007-09-11
EP07116131 2007-09-11

Publications (2)

Publication Number Publication Date
WO2009033588A2 true WO2009033588A2 (fr) 2009-03-19
WO2009033588A3 WO2009033588A3 (fr) 2010-05-20

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Cited By (13)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US8569231B2 (en) 2009-03-20 2013-10-29 Smartcells, Inc. Soluble non-depot insulin conjugates and uses thereof
US8623345B2 (en) 2009-03-20 2014-01-07 Smartcells Terminally-functionalized conjugates and uses thereof
US8846103B2 (en) 2009-01-28 2014-09-30 Smartcells, Inc. Exogenously triggered controlled release materials and uses thereof
US8906850B2 (en) 2009-01-28 2014-12-09 Smartcells, Inc. Crystalline insulin-conjugates
US8933207B2 (en) 2010-07-28 2015-01-13 Smartcells, Inc. Drug-ligand conjugates, synthesis thereof, and intermediates thereto
US8940690B2 (en) 2009-01-28 2015-01-27 National Institutes Of Health (Nih) Synthetic conjugates and uses thereof
US9050370B2 (en) 2009-01-28 2015-06-09 Smartcells, Inc. Conjugate based systems for controlled drug delivery
US9068013B2 (en) 2010-07-28 2015-06-30 Smart Cells, Inc. Recombinant lectins, binding-site modified lectins and uses thereof
US9074015B2 (en) 2010-07-28 2015-07-07 Smartcells, Inc. Recombinantly expressed insulin polypeptides and uses thereof
US9427475B2 (en) 2013-10-04 2016-08-30 Merck Sharp & Dohme Corp. Glucose-responsive insulin conjugates
WO2022036270A1 (fr) * 2020-08-13 2022-02-17 The Regents Of The University Of Colorado A Body Corporate Bicouches phospholipidiques qui favorisent catalytiquement le repliement de protéines, inhibent et inversent la formation d'agrégats de protéines, et méthodes de traitement de maladies neurodégénératives mettant en œuvre de telles bicouches phospholipidiques
US12233177B2 (en) 2019-09-16 2025-02-25 Amgen Inc. Method for external sterilization of drug delivery device
US12257352B2 (en) * 2016-06-20 2025-03-25 The Regents Of The University Of Michigan Compositions and methods for delivery of biomacromolecule agents

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060193905A1 (en) * 2002-05-14 2006-08-31 University Of Louisville Research Foundation, Inc. Direct cellular energy delivery system

Cited By (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9463249B2 (en) 2009-01-28 2016-10-11 Smartcells, Inc. Crystalline insulin-conjugates
US8846103B2 (en) 2009-01-28 2014-09-30 Smartcells, Inc. Exogenously triggered controlled release materials and uses thereof
US8906850B2 (en) 2009-01-28 2014-12-09 Smartcells, Inc. Crystalline insulin-conjugates
US8940690B2 (en) 2009-01-28 2015-01-27 National Institutes Of Health (Nih) Synthetic conjugates and uses thereof
US9050370B2 (en) 2009-01-28 2015-06-09 Smartcells, Inc. Conjugate based systems for controlled drug delivery
US10398781B2 (en) 2009-01-28 2019-09-03 Smartcells, Inc. Conjugate based systems for controlled drug delivery
US9579391B2 (en) 2009-01-28 2017-02-28 Smartcells, Inc. Conjugate based systems for controlled drug delivery
US9114176B2 (en) 2009-01-28 2015-08-25 Smartcells, Inc. Exogenously triggered controlled release materials and uses thereof
US8623345B2 (en) 2009-03-20 2014-01-07 Smartcells Terminally-functionalized conjugates and uses thereof
US8569231B2 (en) 2009-03-20 2013-10-29 Smartcells, Inc. Soluble non-depot insulin conjugates and uses thereof
US8933207B2 (en) 2010-07-28 2015-01-13 Smartcells, Inc. Drug-ligand conjugates, synthesis thereof, and intermediates thereto
US9074015B2 (en) 2010-07-28 2015-07-07 Smartcells, Inc. Recombinantly expressed insulin polypeptides and uses thereof
US9068013B2 (en) 2010-07-28 2015-06-30 Smart Cells, Inc. Recombinant lectins, binding-site modified lectins and uses thereof
US9427475B2 (en) 2013-10-04 2016-08-30 Merck Sharp & Dohme Corp. Glucose-responsive insulin conjugates
US9884125B2 (en) 2013-10-04 2018-02-06 Merck Sharp & Dohme Corp. Glucose-responsive insulin conjugates
US9889205B2 (en) 2013-10-04 2018-02-13 Merck Sharp & Dohme Corp. Glucose-responsive insulin conjugates
US12257352B2 (en) * 2016-06-20 2025-03-25 The Regents Of The University Of Michigan Compositions and methods for delivery of biomacromolecule agents
US12233177B2 (en) 2019-09-16 2025-02-25 Amgen Inc. Method for external sterilization of drug delivery device
WO2022036270A1 (fr) * 2020-08-13 2022-02-17 The Regents Of The University Of Colorado A Body Corporate Bicouches phospholipidiques qui favorisent catalytiquement le repliement de protéines, inhibent et inversent la formation d'agrégats de protéines, et méthodes de traitement de maladies neurodégénératives mettant en œuvre de telles bicouches phospholipidiques

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