EP0967998A2 - Excipient polymere - Google Patents

Excipient polymere

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Publication number
EP0967998A2
EP0967998A2 EP99900284A EP99900284A EP0967998A2 EP 0967998 A2 EP0967998 A2 EP 0967998A2 EP 99900284 A EP99900284 A EP 99900284A EP 99900284 A EP99900284 A EP 99900284A EP 0967998 A2 EP0967998 A2 EP 0967998A2
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Prior art keywords
pla
stereocomplex
polymer
bioactive
polymeric carrier
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EP99900284A
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German (de)
English (en)
Inventor
Avraham J. Domb
Zeev Zehavi
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Efrat Biopolymers Ltd
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Efrat Biopolymers Ltd
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Publication of EP0967998A2 publication Critical patent/EP0967998A2/fr
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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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1641—Organic macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyethylene glycol, poloxamers
    • A61K9/1647—Polyesters, e.g. poly(lactide-co-glycolide)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
    • A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule

Definitions

  • the present invention relates to a carrier formed of stereocomplexes of polymers for delivery of bioactive or bioreactive molecules.
  • Polymer characteristics are affected by the monomer types and composition, the polymer architecture, and the molecular weight.
  • the crystallinity of the polymer an important factor in polymer biodegradation, varies with the stereoregularity of the polymer.
  • racemic D,L poly(lactide) or poly(glycolide) is less crystalline than the D or L homopolymers.
  • Poly(lactide) (PLA) and its copolymers having less than 50% glycolic acid content are soluble in common solvents such as chlorinated hydrocarbons, tetrahydrofuran, and ethyl acetate while poly(glycolide) (PGA) is insoluble in common solvents but is soluble in hexafluoroisopropanol.
  • PLA has wide applications in medicine because of its biocompatibility and degradability to nontoxic products.
  • Micelles and particles of the AB block copolymer poly(lactide)-b-poly(ethyleneglycol) (PLA-b-PEG) have received attention for use in intravenous injectable delivery systems for extended and target drug release.
  • Gref, R. et al. Protein Delivery-Physical Systems, L. M. Sanders and H. Hendren, Eds, Plenum Press, (1997); and Gref, R. et al., Advanced Drug Delivery Reviews, 16: 215-233 (1995).
  • multiblock copolymers comprising a multifunctional compound covalently linked with one or more hydrophilic polymers and one or more hydrophobic bioerodible polymers and including at least three polymer blocks.
  • a PEG-coating on a microparticie or other polymeric device prevents the adsorption of plasma proteins and fast elimination by the reticulo endothelial system (RES).
  • RES reticulo endothelial system
  • AB-block copolymers of poly(styrene)-b-poly(acrylic acid) produce vesicle type particles which can be isolated from solution. Zhang, L. et al., Science (1995) 268, 1728. The vesicles have diameters up to 1 micron, which is much larger than that of a single micelle. This is explained by the irreversible formation of the vesicles by the fusion of micelles. However, once the micelles are associated, the high Tg of poly(styrene) (PS) freezes the structure since the PS- blocks are no longer in equilibrium with the solvent and the structure is still stable after removing the solvent. Vesicular architecture of block copolymers would appear to offer future opportunities for pharmaceutical drug devices. Nevertheless, PS and PAA cannot be used as biodegradable carriers because they are stable and do not degrade in biological mediums.
  • a polymeric carrier for delivery of bioactive or bioreactive molecules including a stereocomplex of one or more biocompatible polymers and having incorporated on or within the complex the molecules to be delivered.
  • the biocompatible stereoselective polymers are linear or branched D-PLA homo- and block-polymers, linear or branched L-PLA homo- and block-polymers, copolymers thereof, or mixtures thereof, in stereocomplexed form.
  • the polymeric carrier is complexed with a complementary stereospecific bioactive molecule.
  • the bioactive, or bioreactive is bound to the complex by ionic, hydrogen, or other non-covalent binding reactions not involving stereocomplexation, or is physically entrapped within the complex, either at the time of complex formation or when the polymeric material is formulated into particles, tablets, or other form for pharmaceutical application.
  • exemplary bioactive molecules include peptides, proteins, nucleotides, oligonucleotides, sugars, carbohydrates, and other synthetic or natural organic molecules, as well as stereoselective drugs of a molecular weight of 300 Dalton or higher.
  • Stereocomplexation of macromolecules to biodegradable polymers is a new approach in the delivery of macromolecules.
  • the interaction at the molecular level between the polymer carrier and the bioactive macromolecule provides stability, which allows for extended release and for easy access to the target cell or tissue with minimal toxicity.
  • Stereocomplexes including a racemic packing of enantiomorphic poly(L-lactide) [L- PLA] and poly(D-Lactide) [D-PLA] have a melting point 60°C higher than chiral crystals with the packing of isomorphic PLA's. As a consequence of the different packing, the chiral and racemic single crystals exhibit different morphologies.
  • the stereocomplex forms lamella triangular or rounded hedrite type crystals instead of lozenge shaped crystals.
  • PLA-b-PEG aggregates to supra molecular assemblies like flat or tubular rods of hundreds of nanometers wide and a few microns long. Powder-diffraction patterns indicate that the crystallization of both blocks are the driving force for the formation of the mesoscopic suprastructures. The crystallized blocks are not in equilibrium with the solvent for very long, which explains the stability of the structures after removing the solvent.
  • vesicle type particles emerge from dioxane and acetonitrile solutions.
  • the racemic particles of PEG-b-L-PLA/PEG-b-D-PLA should have a similar assembly to the PS- b-PAA particles.
  • the racemic particles of PEG-b-PLA have a much higher potential as a drug carrier system because of the safety of the polymer and its degradation products.
  • the hydrophobic content within the vesicles should support the encapsulation of non-polar drugs.
  • the hydrophobic/hydrophilic content may provide a type of target mechanism to pass through lipid membranes.
  • PLA is a polyester and peptides are polyamides. Esters cannot form hydrogen bonds to each other since they lack hydroxyl groups. In organic solvents poly(amino acids) do not form hydrogen bonds. Thus the crystallization and packing of poly(amino acids) is comparable to PLA. Enantiomorphic poly(alanine) and PLA were crystallized into a racemic lattice to better understand the specific interactions between peptides and synthetic polymers. Until now only microparticles of the statistical copolymer poly(lactide)-b-poly(glycolide) containing LHRH showed the delayed liberation of the hormone.
  • Polymeric carriers are provided for delivery of bioactive or bioreactive molecules, which include at least one biocompatible stereoselective polymer.
  • useful polymers include polyhydroxy acids, polyhydroxyalkyls, polyalkylene oxides, polyesters, polycarbonates, and polyanhydrides.
  • the carrier is formed from linear or branched D-PLA homo- or block-polymers, linear or branched L-PLA homo- or block-polymers, copolymers thereof, and mixtures thereof, where the copolymers are linear or branched D-PLA or L-PLA block copolymer copolymerized with a component such as a poly(hydroxyalkyl acid), polycarbonate or polyanhydride.
  • Polymeric carriers can also be formed of, or include, a stereocomplex of homo- or block-copolymers of D-lactide or homo- or block-copolymers of L-lactide with inert polyamino acids (such as polyalanine or polylysine), polypeptides or polysaccharides.
  • the block length of the enantiomeric segment is typically equivalent to ten lactide units or more.
  • the general synthetic procedures for the synthesis of the polymers is as follows. Polymers are dissolved in a suitable solvent and appropriate polymerization catalysts such as short alcohols, polyethylene glycol (PEG), fatty alcohol or polyalcohol, added in a range such as 0.1 to 3 mole percent per lactide. The solvent is removed after polymerization is initiated, for example, by solvent evaporation. The molar ratio between the monomer and the catalyst determine the polymer block molecular weight. The length and number of blocks is controlled by the number of and amount of each monomer units added and the amount of catalyst used.
  • PEG polyethylene glycol
  • polyalcohol polyethylene glycol
  • the molar ratio between the monomer and the catalyst determine the polymer block molecular weight. The length and number of blocks is controlled by the number of and amount of each monomer units added and the amount of catalyst used.
  • pre-prepared polymer blocks with hydroxyi and carboxylic acid can be conjugated via an ester, phosphate, anhydride, or carbonate bond.
  • the hydroxyi end groups are reacted with either a diacid chloride (i.e. adipoyl chloride, sebacoyl chloride), alkyl phosphodichloridate, or phosgene to form ester, phosphate or carbonate block conjugates, respectively.
  • Anhydride copolymers are prepared by activating the PLA carboxylic acid end group with acetic anhydride and copolymerized with sebacic acid prepolymer according to Domb et al., J. Poly. Sci. 25:3373 (1987).
  • Multiblock copolymers of PLA are prepared by using a polyalcohol such as pentaerythritol, or glycerol in the catalyst mixture.
  • a polyalcohol such as pentaerythritol, or glycerol
  • the structures and block length can be determined by H-NMR and GPC.
  • Typical MW of polymers is in the range of 5,000 to 100,000.
  • homopolymers of PLA were synthesized by dissolving D-lactide or L-lactide in dry toluene at 100°C and adding a solution of stannous octoate and alcohol as polymerization catalyst (5% solution in toluene, 0.1 to 3 mole % per lactide). After 3 hours the solvent was evaporated to dryness and the viscous residue was left at 130°C for additional 2 hours to yield the polymer.
  • the first block i.e. L-lactide
  • Block copolymers with cyclic hydroxy alkyl acids and cyclic carbonates are prepared in a similar manner, but the second portion is the desired cyclic monomer (caprolactone, trimethylene carbonate, glycolide), instead of lactide.
  • a tetrablock copolymer consisting of two D-PLA chains and two L-PLA chains was prepared by polymerizing D-lactide with dibenzyl tartarate using stannous octoate as catalyst. After polymerization at 130°C as described above, the benzyl protecting groups were removed by hydrolysis or hydrogenation. The free acid groups were esterified with hydroxyi terminated L-PLA in chloroform solution and DCC as coupling agent. The length of the blocks varied depending on the amount of D-lactide used for polymerization and the L-PLA chain length. This polymer formed an intra and inter-stereocomplexation.
  • Other multiblock polymers contained blocks of either or both D-PLA and L-PLA and other biodegradable polymers or poly(oxyalkanes) or contained other branching molecules like mucic acid, pentaerythritol, citric acid and malonic acid 2-methanol.
  • polymer is either dissolved in a solvent in which the polymers are soluble or the polymer components melted together.
  • the polymer mixture is stored under conditions at which the polymers will complex and precipitate out of solution, or cool.
  • the mixture can be formed into a desired shape as it forms, or processed after stereocomplex formation.
  • Solvents which can be used to dissolve polymers for formation of stereocomplexes include dioxane, chloroform, tetrahydrofuran, ethyl acetate, acetone, N-methylpyrrolidone, ethyl and methyl lactate, ethyl acetate and mixtures of these solvents, and other solvents, such as water, short chain alcohols and carboxylic acids (C5 or below).
  • the particle size of the precipitate is controlled by the selected solvent, the drug, and polymer concentrations, and the reaction conditions (temperature, mixing, volume etc.).
  • L-PLA blocks of 20 to 100 lactide units were conjugated to a biodegradable polyanhydride, polycaprolactone and polyhydroxybutyrate, or to the hydrophilic poly(ethylene glycol) or polypropylene glycol).
  • the stereocomplexation of these copolymers with short and long chain polymers of D-lactide at different solutions and conditions was characterized by atomic force microscopy (AFM) and related surface characterization methods (SEM, TEM, XPS).
  • X ester, carbonate, ether, phosphate, anhydnde, orthoester, or a branching molecule b.
  • the stereocomplexes can be used to deliver any of a variety of molecules which may broadly be classed as bioactive or bioreactive molecules, for therapeutic, prophylactic or diagnostic applications, referred to generally herein as ⁇ bioactive molecules!].
  • bioactive molecules can be any of the broad chemical classes of peptides, proteins, sugars, carbohydrate, lipids, nucleotides, oligonucleotides, and combinations thereof such as glycoproteins.
  • preferred bioactive molecules include hormones, clotting factors, proteases, growth factors; and vaccines.
  • peptides include hormones such as LHRH, GNRH, enkephalin, ACTH, a-MSH, somatostatin, caicitonin, insulin and their analogs.
  • proteins include erythropoeitin, t- PA, Factor VIII, growth hormone, growth factors like FGF, BMP, and EGF; and vaccines against bacterial or viral pathogens such as vaccines containing as antigens staphylococcal enterotoxin B toxoid, HGC-DT, diphtheria toxoid and ribonuclease A.
  • preferred oligonucleotides include antisense, genes, plasmids and viral vectors.
  • the bioactive molecules can be incorporated onto or into the stereocomplexes. They can be coupled to the stereocomplexes by ionic, hydrogen or other types of bond formation, including covalent bond formation.
  • the bioactive molecules can be incorporated into the stereocomplexes when the polymers are mixed together in solution or melted, so that the molecules are entrapped within the polymer complex as it precipitates or cools. They can also physically be mixed with the stereocomplexes as these are formulated into tablets, molds, or particles, as described below.
  • the bioactive molecules can be coupled to the stereocomplexes after formation of the complexes or the formulations containing the complexes.
  • stereocomplexes are typically formed into devices for drug delivery using standard polymer processing techniques to form particles including nanoparticies, microspheres, and microcapsules, pellets, tablets, films, rods, or beads.
  • the polymers can be formulated into a paste, ointment, cream, gel, or transdermal patch.
  • the polymers can also be lyophilized and then formulated into an aqueous suspension in a range of microgram/ml to 100 mg/ml prior to use.
  • Suitable vehicles include water, saline, and phosphate buffered saline.
  • the bioactive or bioreactive component can be administered once, or may be divided into a number of smaller doses to be administered at varying intervals of time, depending on the release rate of the carrier and the desired dosage.
  • the above formulation principles are well known in the art.
  • the formulations can be administered to a patient in a variety of routes, for example, orally, parenterally, intravenously, intradermally, subcutaneously, or topically, in liquid, cream, gel, or solid form, as appropriate for the drug to be delivered.
  • the carrier should contain the substance to be delivered in an amount sufficient to deliver to a patient a therapeutically effective amount of compound.
  • the desired concentration of active compound with the carrier will depend on absorption, inactivation, and excretion rates of the drug, as well as the delivery rate of the compound from the carrier.
  • dosage values will also vary with the severity of the condition to be alleviated. It is to be further understood that, for any particular subject, specific dosage regimens should be adjusted over time according to the individual need and the professional judgment of the person administering or supervising the administration of the compositions.
  • Atomic-force-Microscopy A scanning probe microscope 'Nanoscope III' (Digital Instruments Inc.) was used. Rectangular Si cantilevers (Nano-probes) were applied for the tapping mode experiments. Simultaneous registration was performed in the tapping mode for height and amplitude images.
  • the specimen for AFM were prepared by depositing one drop of 0.1% solution of the AB-block copolymer on mica and evaporating the solvent. Stereocomplexes of enantiomorphic PLA-blocks were prepared by mixing equimolar solutions and stirring on a low speed for 1 week.
  • Stereocomplexes were prepared as follows. Table 2 illustrates the differences between the melting points of homopolymers of the starting materials and the melting points of the stereocomplexes. a. L-PLA and D-PLA in acetonitrile
  • the clear solution became turbid after about 4-5 hours, and after two days at 60°C, a heavy white solid precipitated. After three days, the solution was filtered, and the stereocomplex was collected and dried in vacuum over night.
  • the precipitate was analyzed for particle size and shape by particle size analyzer (Coulter) and AFM.
  • the average particle size was 2.1 microns with various particle shapes, mostly consisting of flat discs.
  • the melting point of the complex was 234.8°C.
  • a stereocomplex of D-PLA-PEG and L-PLA-PEG was prepared according to the procedure described in example b. The melting point was determined to be 221°C.
  • Table 2 Stereocomplexes melting points (DS :
  • PLA-PEG and L-PLA-PEG (CHClj). 221
  • the center of the discs was indented.
  • the block copolymers PEG-b-L-PLA-2/ PEG-b-D-PLA-2 formed monodisperse discs with diameters of approximately 1 mm and heights of between 90 and 120 nm.
  • the dimensions of the discs exceeded the size of a single block copolymer micelle, which has a diameter of between 60 and 80 nm. Preparation of discs with the same dimensions and shapes was reproducible. In contrast, the rods and coiled coils emerged randomly and irregularly, with different dimensions, and in small amounts.
  • crystallization forms triangular or rounded crystals depends on the crystallization conditions.
  • the rounded crystals preferentially form from concentrated acetonitrile solutions (1 %).
  • Rounded crystals were obtained by fast cooling of a heated 0.1% acetonitrile solution.
  • the rounded crystals could be hedrites which are a transition from lamellae single to spherulitic crystal growth.
  • the diameter of the hedrites was 5 microns and the height was 200 nm; further they exhibited a deep hole in the middle.
  • the hole may be the result of a disordered nucleatio ⁇ , such that when the nucleus has reached a critical size, the respective sides of the crystal grow faster.
  • the similarity of the block copolymer and homopolymer discs demonstrates that the rounded shape of the block copolymer discs may be primarily due to the racemic packed enantiomorphic PLA-blocks.
  • Devices were prepared by compression molding of these nanoscale structures with drugs. This compression molding resulted in strong, dense devices with high uniformity and reproducibility.
  • the disintegration time and the release of the incorporated drug in buffer solutions was slower than the compression molded devices of the corresponding regular polymers.
  • the regular polymer powders are difficult to obtain and their properties are erratic, since they are prepared by precipitation in non-aqueous solvent and grinding at a temperature below the glass transition temperature which provides irregular particle size and shape.
  • the particles formed by stereocomplex formation are of well-defined flat crystallites that are easy to compression mold.
  • Lidocaine was released constantly for 30 days when placed in buffer solution p H7.4 at 37°C.
  • Example 5 Formulation and Release of Methotrexate (MTX).
  • the drug in the form of a powder was mixed with the stereocomplex in the form of a powder, and the mixture compression molded to form tablets.
  • the stereocomplex and MTX were dissolved in dichloromethane, and solvent evaporated to yield a fine powder.
  • LHRH Leutinizing Hormone Releasing Hormone
  • the white precipitate was filtered and dried over a vacuum over night; it yielded 460 mg of LHRH loaded PLA stereocomplex.
  • the average particle size of the powder was 2.4 microns, and the particles were in the shape of discs.
  • the LHRH content in the acetonitrile solutions was determined in the following manner: 3 ml of the acetonitrile solution (from the procedures described above) were evaporated to dryness and the residue was dissolved in 2 ml of phosphate buffer. LHRH concentration in these solutions was determined by the HPLC method described above. No LHRH peak was shown on the chromatograms for solutions a to e, while more than 80% of the LHRH was found in the solution of formulation f, and the precipitate was almost pure LHRH. LHRH release:
  • the powder of the LHRH in the stereocomplex is very thin, ail the attempts to sink it at the release medium failed.
  • the release was conducted in the following way: 100 mg of the powder in 5 ml phosphate buffer were added to a 10 ml syringe. The edge of the syringe was sealed and constantly shaken at a rate of 150 RPM and a temperature of 37°C. At each interval of time, the syringe was inverted so that the solid floated at the top of the buffer and all of the buffer was pushed out of the syringe in a dropwise fashion. Using this technique, the powders of LHRH in PLA stereocomplex (formulations a-e) were analyzed against a control blank of PLA stereocomplex (no drug).
  • LHRH was constantly released from these powders for a period of about three months with a similar release profile for formulations a and b. No LHRH was detected in the blank polymer.
  • Formulation e prepared from low molecular weight PLA, released the drug faster, within 30 to 40 days, than the other formulations.
  • the rod formulations released the LHRH for longer time periods than their respective powder formulations.
  • the oligonucleotide ISIS3521 was incorporated into the stereocomplexes in one of the three ways described for LHRH.
  • ISIS3521 concentration in the acetonitrile solutions was determined by UV absorption at 260 nm Release of ISIS
  • reaction a 0.04 mg of ISIS 3521 was detected, which is less than 1 % of the initial amount added.
  • the acetonitrile phase from reaction b did not contain ISIS at all and the small precipitate from reaction c was 100% ISIS3521.
  • TRH is a tripeptide with the following structure: pL-Glu-L-His-L-Pro_NH2.
  • ISIS an antisense oligonucleotide
  • LHRH an antisense oligonucleotide
  • methotrexate Three representative bioactive molecules, ISIS (an antisense oligonucleotide), LHRH, and methotrexate were incorporated into the polymers either during the crystallization and complex formation, or by mixing the blank stereocomplex particles with the drug and compressing the powder into tablets.
  • the solution was allowed to mix at 37°C over night to form a white precipitate which was separated by filtration.
  • the particles were of a disc shape with an average particle size of between 1 and 3 microns.
  • Analysis of the drug content in the solution and the precipitate showed an encapsulation yield of over 80%. Methotrexate was encapsulated in poor yield by this method.
  • Ibuprofen concentration in the acetonitrile solution was determined by UV at 254 nm. It was found that over 95% of the drug was in the acetonitrile solution, which indicates no complex formation occurred for any of the ibuprofen isomers.
  • L-phenylalanine was reacted with D-PLA or a mixture of D- PLA and L-PLA in acetonitrile, and no complex was formed.
  • testosterone blood levels was determined by a standard kit for the determination of testosterone in human blood. As demonstrated, testosterone levels remained low for the 40 days of the study which is similar to the data obtained for the clinically used formulation, Lupron, as described in the literature. This experiment demonstrates the effectiveness of and the use of these stereocomplexes for peptide delivery.
  • Example 14 Stereoselective polymers with cationic residues for gene delivery: a. PEI-D-PLA polymers:
  • Block copolymers of polyethylenimine and D-PLA are prepared by the attachment of polyD-lactic acid on polyethylenimine using DCC as a coupling agent.
  • DCC dicyclohexylcarbodimide
  • DMAP N,N-dimethylamino pyridine
  • the residue was purified by precipitation in ether: petroleum ether mixture from a concentrated dichloromethane solution.
  • the resulting polymer contained both the D-PLA and PEI as determined by NMR and nitrogen analysis.
  • Copolymers of the ratio PEI:D-PLA ranging from 3:1 to 1 :3 are prepared.
  • D-PLA of molecular weights, 2,000, 10,000 are used for these copolymers.
  • random copolymers are prepared by a transamidation reaction between PEI and D-PLA in solution or in melt.
  • D-PLA of a molecular weight 55,000 was dissolved in dry dioxane and PEI-600 (2:1 w/w) was added to the solution.
  • the reaction solution was heated to reflux for 5 hours and the solution was added to deionized water solution to precipitate the polymer.
  • the precipitated polymer was dissolved in dichloromethane, dried over MgSO and evaported to dryness.
  • the resulted polymer contained PEI residues as determined by NMR and nitrogen analysis.
  • D-PLA with carboxylic acid end groups that can react with amino groups to form amide bonds are prepared by either polycondensation of D-lactic acid (Prepared by hydrolysis of the commercially D-lactide available from Purac) in toluene or by partial hydrolysis of poly(lactide) in a base solution or by enzyme cleavage.
  • Low molecular weight PEI are synthesized by acid-catalyzed ring-opening polymerization of aziridine in aqueous solution.
  • Short chain PEI, up to 2,000 Daltons, are preferred for gene delivery as they have shown to be less toxic compared to longer chains [D. Fischer, T. Bieber, Y. Li, H.P. Elsasser, T. Kisse.
  • Polyethylenimine Synthesis and in vitro cytotoxicity of a low molecular weight polycation for gene transfer. Eur. J. Cell Biol. 75 (1998) 107; T. Bieber, D. Fischer, T. Kissel, H.P. Elsasser. A low molecular weight polyethylenimine with striking qualities for gene delivery. Eur. J. Cell Biol. 75 (1998) 108]. c. Synthesis of polylysine-D-PLA copolymers
  • Carboxylic acid terminated D-PLA was conjugated to polylysine to form a steroselective polymer with cationic capacity.
  • the preferred polylysine is of D- configuration and of low molecular weight which is considered as less toxic and easy to being eliminated from the body.
  • the D-configuration will be aligned with the D-PLA and increase the sterocomplexing capacity.
  • the methods for the formation of linear the graft block copolymers of polylysine-PLA are similar to the methods described above for PEI. Random and graft copolymers of D-PLA and lysine can be prepared using the methods described by Langer [Langer et al.
  • the preferred lysine configuration for these copolymers is the D-configuration which forms a uniform D- configuration polymer chain with the D-lactide residues.
  • AB block copolymers of polylysine-D-PLA the g-amino side groups of the polylysine are protected by benzoxycarbonyl protecting groups and then attach the D-PLA chain to the free carboxylic end group by an ester bond.
  • Complexation with DNA Tese polymers were complexed with Herring DNA by mixing the DNA and polymer in buffer solutions or mixtures of buffers and acetonitrile or DMSO to form a complex.
  • a steroselective structure with cationic sites is provided so that a plasmid can be complexed by diasteriomer formation and by electrostatic complexation.
  • Such polymers are: D-PLA graft and block copolymers with short polyamine such as polyethylene imine (Mw ⁇ 2,000), poly(lysine), spermine, spermidine and copolymers of D-lactide and D-lysine.
  • the present invention also provides a stereoselective polymer with cationic residues including lysine, polylysine, sperime, spermidine and polyethyleneimine which allow the complexation to DNA for gene delivery.

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  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
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Abstract

L'invention concerne un excipient polymère destiné à l'administration d'une molécule bioactive ou bioréactive, l'excipient comprenant un stéréocomplexe d'au moins un polymère stéréosélectif biocompatible et d'une molécule bioactive ou bioréactive.
EP99900284A 1998-01-14 1999-01-14 Excipient polymere Withdrawn EP0967998A2 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
IL12293398 1998-01-14
IL12293398A IL122933A (en) 1998-01-14 1998-01-14 Polymeric carrier for delivery of a bioactive molecule
PCT/IL1999/000023 WO1999036100A2 (fr) 1998-01-14 1999-01-14 Excipient polymere

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EP0967998A2 true EP0967998A2 (fr) 2000-01-05

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JP2002537343A (ja) * 1999-02-23 2002-11-05 アイシス・ファーマシューティカルス・インコーポレーテッド 多重粒子製剤
WO2002083166A1 (fr) * 2001-04-10 2002-10-24 Santen Pharmaceutical Co., Ltd. Complexes de polymeres d'interferons et leur utilisation medicinale
DE10154924A1 (de) * 2001-11-08 2003-05-28 Medinnova Ges Med Innovationen Bioabbaubare Kopolymere
EA008244B1 (ru) * 2002-05-03 2007-04-27 Янссен Фармацевтика Н.В. Полимерные микроэмульсии
JP4877225B2 (ja) * 2005-02-18 2012-02-15 日油株式会社 ポリオキシアルキレン誘導体
EP1902084A2 (fr) * 2005-06-01 2008-03-26 Yissum Research Development Company, of The Hebrew University of Jerusalem Polymeres biodegradables a chiralite predeterminee
US7794838B2 (en) * 2006-02-03 2010-09-14 National University Corporation Hokkaido University Nanodisk comprising block copolymer
PL2135887T3 (pl) 2008-06-18 2011-05-31 Inst Biopolimerow I Wlokien Chemicznych Sposób wytwarzania stereokompleksu polikwasu mlekowego w postaci proszku
WO2012005376A1 (fr) * 2010-07-09 2012-01-12 国立大学法人 東京大学 Composition d'administration d'acides nucléiques, composition d'excipient, composition pharmaceutique contenant la composition d'administration d'acides nucléiques ou la composition d'excipient et méthode d'administration d'acides nucléiques
CN108559084B (zh) * 2018-04-13 2020-12-04 华东理工大学 一种聚乳酸基疏水薄膜的制备方法
CN109908106A (zh) * 2019-03-04 2019-06-21 北京化工大学 一种可供静脉注射的甲氨蝶呤纳米胶囊及其制备方法
JP7050734B2 (ja) * 2019-09-30 2022-04-08 鳥居薬品株式会社 舌下アレルゲン免疫療法におけるフィルム製剤
CN115785471B (zh) * 2022-12-16 2023-08-08 华中科技大学 一种二氧化碳基超分子聚合物及其制备方法和应用

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CA2283648C (fr) 2007-04-17
IL122933A (en) 2005-03-20
JP2001515522A (ja) 2001-09-18
IL122933A0 (en) 1998-08-16
WO1999036100A2 (fr) 1999-07-22
CA2283648A1 (fr) 1999-07-22
WO1999036100A3 (fr) 1999-09-23
AU1888999A (en) 1999-08-02

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