NL195092C - Preparations of water-soluble peptides with a slow release. - Google Patents

Description

1 195092, Slow-release water-soluble peptides preparations

This invention relates to preparations of slow-release drugs (depot preparations), in particular of water-soluble peptides, for example somatostatin or somatostatin analogues, such as octreotide, in a biodegradable and biocompatible, polymeric carrier, for example a matrix or a coating, for example in the form of an implant or preferably a microparticle (also known as a microcapsule or a microsphere).

The invention also relates to such compositions which possess satisfactory peptide release profiles for a special period of time.

Peptide drugs often exhibit poor bioavailability in the blood after oral or parenteral administration, for example due to their short biological half-lives caused by their metabolic instability. Orally or administered through the nose, they often exhibit poor resorption through the mucous membranes. A therapeutically relevant blood level over a long period of time is difficult to achieve.

The parenteral administration of peptide drugs as a depot preparation in a biodegradable polymer, for example as microparticles or implants, has been proposed, with a delayed release after a residence time in the polymer protecting the peptide against enzymatic and hydrolytic influences of the biological media. can be released.

Although some parenteral depot preparations or peptide drugs are known in a polymer in the form of microparticles or an implant, satisfactory peptide release profiles are only obtained in practice in a few cases. Special measures must be taken to provide a continuous peptide release for a serum level of a therapeutically active medicine and, if desired, to avoid too high serum concentrations of the medicine that cause undesirable pharmacological side reactions.

The release pattern of a peptide drug depends on numerous factors, for example, the type of peptide and, for example, whether it is present in the free form or in another form thereof, for example in the salt form, which may affect its solubility in water. Another important factor is the choice of the polymer, from an extensive list of possibilities described in the literature.

Each polymer type has its characteristic biological decomposition rate. Free carboxyl groups can be formed which contribute to the pH value in the polymer and thus further influence the water solubility of the peptide and thus its release pattern.

Other factors that influence the release pattern of the depot preparation are the drug loading of its polymeric carrier, its mode of distribution into the polymer, the particle size and, in the case of an implant, moreover its shape. Furthermore, the location of the preparation in the body is of influence.

So far, no somatostatin preparation in a sustained-release form for parenteral administration has come onto the market, probably because no preparation showing a satisfactory serum level profile could be obtained.

Polymer preparations with drugs intended to provide a sustained or delayed release of the drug are known in the art.

U.S. Pat. No. 3,773,919 describes compositions from which the drug is released at a controlled rate, wherein the drug, e.g., a water-soluble peptide drug, is dispersed in a biodegradable and biocompatible linear polylactide or polylactide-co-glycolide polymer . However, no patterns for the release of the drug have been described and there is no reference to a somatostatin. U.S. Pat. No. 4,293,539 describes antibacterial preparations in a microspherical form.

U.S. Pat. No. 4,675,189 describes sustained-release preparations of the LHRH analog decapeptide nafarelin and analog LHRH relatives in polylactide co-glycolide polymers. No release pattern has been described.

50, T. Chang, J. Bioeng., Vol. 1, pages 25-32, 1976 describe the sustained release of biological materials, enzymes, and vaccines from microparticles.

Polymer copolymers of lactic acid and lactide glycolide copolymers and related compositions for use in surgical applications and for delayed release and biological decomposition are described in U.S. Pat. Nos. 3,991,776; 4,076,798 and 4,118,470.

55 European patent application 0.203.031 describes a series of somatostatin octapeptide analogs, 195092 2, for example compound RC-160 of formula, * * D-Phe-Cys-Tyr-DT rp-Lys-Val-Cys-T rp-NH2, with a bridge between the -Cys sections, in columns 15-16.

The possibility of the somatostatin to be encapsulated in a micro form with polylactide-co-glycolide polymer is mentioned in claim 18, but no indications are given how a continuous therapeutically active serum level can be obtained.

U.S. Pat. No. 4,011,312 describes that a continuous release of an antimicrobial drug, for example the water-soluble polymyxin B, from a polylactide-co-glycolide matrix with a low molecular weight (lower than 2000) and a relatively high glycolid content in in the form of an implant, can be obtained if the implant is inserted in the cow's nipple channel. The drug is released in a short period of time, due to the high glycolid content and the low molecular weight of the polymer, both of which stimulate rapid biological decomposition of the polymer and thus a corresponding rapid release of the drug. Moreover, a relatively high drug load contributes to a rapid release of the drug. No somatostatin and no drug release patterns have been described.

European Patent No. 58,481 describes that continuous release of a water-soluble peptide from a polylactide polymer implant is stimulated by reducing the molecular weight of at least a portion of the polymer molecules, by introducing glycolide units into the polymer molecule, by increasing of the polymer block character if polylactide-coglycolid molecules are used, by increasing the drug loading of the polymer matrix and by increasing the surface of the implant.

Although somatostatin has been mentioned as water-soluble peptides, no somatostatin release profiles have been described and no indication has been given as to how all of these parameters can be combined to obtain, for example, a continuous somatostatin serum level for at least a week, for example a month.

It is described in European patent 92,918 that a continuous release of peptides, preferably hydrophilic peptides, can be obtained for a long period of time if the peptide is included in a conventional hydrophobic polymer matrix, for example of a polylactide that is made more accessible to water. by introducing into its molecule a hydrophilic moiety, for example of polyethylene glycol, polyvinyl alcohol, dextran, polymethacrylamide. The hydrophilic contribution to the amphipathic polymer is given to all ethylene oxide groups in the case of a polyethylene glycol unit, by the free hydroxyl groups in the case of a polyvinyl alcohol unit or of a dextran unit and by the amide groups in the case of a polymethylacrylamide unit.

Due to the presence of the hydrophilic unit in the polymer molecules, the implant will acquire hydrogel properties after the absorption of water. Somatostatin is mentioned as a hydrophilic peptide, but no release profile has been described nor an indication given which polymer type is recommended for this peptide and what molecular weight and how many hydrophilic groups it will have.

It is described in U.S. Pat. No. GB 2,145,422 B that a sustained release of 40 different types of drugs, e.g. vitamins, enzymes, antibiotics and antigens, can be obtained over a prolonged period of time if the drug is inserted into an implant, e.g. of the size of microparticles made from a polymer of a polyol, for example glucose or mannitol, with one or more, preferably at least 3, polylactide ester groups. The polylactide ester groups preferably contain, for example, glycolide units. No 45 peptides, for example somatostatin, are mentioned as drugs and no serum drug levels are described.

This invention relates to sustained-release preparations, for example preparations of microparticles of a medicament, in particular of a hormonally active, water-soluble, somatostatin or a somatostatin analogue, such as octreotide, to provide a satisfactory drug plasma level and, for example, in a biologically decomposable, biocompatible, polymer, e.g., in an encapsulating polymer matrix. The polymer matrix can be a synthetic or a natural polymer.

The microparticles of this invention can be obtained by any conventional method, for example an organic phase separation method, a spray drying technique or a triple emulsion method, wherein the polymer is precipitated together with the drug, followed by curing of the resulting product, if the phase separation or triple emulsion method.

If desired, the sustained-release preparations may be in the form of an implant.

It has been found that a new salt of octreotide is the pamoate, which is very stable in such compositions.

The present invention accordingly provides (i) octreotide pamoate and (ii) a process for preparing octreotide pamoate, which comprises the reaction of octreotide with embonic acid (or a reactive derivative thereof).

A particularly useful modification of a phase separation method for preparing microparticles of each drug is thus a method for preparing or manufacturing a microparticle containing a drug in a biologically decomposable, biocompatible carrier, which comprises the following steps: a) dissolving the polymeric carrier material in a suitable solvent in which the drug compound is not soluble; b) addition and dispersion of a solution of the drug compound in a suitable solvent, e.g. an alcohol that is not a solvent for the polymer, in the solution of step a), c) addition of a phase inducing agent to the dispersion of step b), to induce the formation of microparticles, d) addition of an oil-in-water emulsion to the mixture of step c), for curing from the microparticle, and e) recovering the microparticle.

Also a particularly useful modification is that of the triple emulsion method for preparing microparticles of each drug, which comprises a method for preparing microparticles of (i) intimately mixing a water-in-oil emulsion formed from an aqueous medium and an immiscible water-soluble organic solvent containing in one phase the drug and in the other phase a biologically decomposable, biocompatible polymer, with an excess of an aqueous medium containing an emulsifying material or a protective colloid to form an aqueous contains in-oil-in-water emulsion, without adding any material containing the drug to the water-in-oil emulsion or applying any intermediate, viscosity-increasing step, (ii) desorbing its organic solvent, (iii) isolating and drying the resulting microparticles.

According to these methods, microparticles are provided.

Also provided is: a) a sustained-release preparation that a peptide drug compound contains a 40/60 to 60/40 polylactide-co-glycolide ester of a polyol, wherein the polyol unit is selected from the group of an alcohol with a carbon chain of 3 -6 carbon atoms and 3-6 hydroxyl groups and a mono- or disaccharide and the esterified polyol contains at least 3 polylactide-co-glycolide chains.

b) a sustained-release preparation comprising a peptide drug compound selected from the group of a calcitonin, lypressin or somatostatin in a 40/60 to 60/40 polylactide-co-glycolide polymer with linear chains with a molecular weight Mw of between 25,000 and 100,000, a polydispersibility between 1.2 and 2 in a concentration of 0.2, preferably 2-10 wt. % of the peptide drug-40 part compound therein.

c) a sustained-release preparation containing octreotide or a salt or a derivative thereof in a biodegradable, biocompatible, polymeric carrier.

The naturally occurring somatostatin is a tetradecapeptide of the formula Ala-Gly-Cys-Lys-Asn-Phe-Phe-T rp 45 I

Cys-Ser-Thr-Phe-Thr-Lys.

This hormone is produced by the hypothalamic gland as well as other organs, for example the G1 system and mediates, together with GRF, q.v. the neuroregula system and mediates, together with GRF, q.V. the neuroregulation of the release of the pituitary growth hormone. In addition to the inhibition of the release of GH by the pituitary gland, somatostatin is a potent inhibitor of a number of systems, including the central and peripheral neural, gastrointestinal and vascular smooth muscle. It also inhibits the release of insulin and glucagon.

The term "somatostatin" includes the analogs or derivatives thereof. Derivatives and analogues are understood to be polypeptides with a straight chain, a bridge or cyclic polypeptides in which one or a number of the amino acid units have been omitted and / or have been replaced by one or more other amino group (s) and / or in which one or a number of the functional groups have been replaced by one or more other functional groups and / or one or a number of groups have been replaced by one or more 195092 4 other isosteric groups. In general, the term includes all modified derivatives of a v biologically active peptide that exhibits a qualitatively similar effect to that of the unmodified somatostatin peptide.

Agonist analogues of somatostatin are thus useful as substitutes for natural somatostatin in its effect on the regulation of physiological functions.

Known somatostatin are: * * a) (D) Phe-Cys-Phe- (D) Trp-Lys-Thr-Cys-Thr-ol (generic name octreotide) b) (D) Phe-Cys-Tyr- (D ) Trp-Lys-Val-Cys-Ti rNH2 * * c) (D) Phe-Cys-Tys- (D) Trp-Lys-Val-Cys-TrpNH2 * * 15 d) (D) Trp-Cys-Phe- (D) Trp-Lys-Thr-Cys-ThrNH2 * * e) (D) Phe-Cys-Phe- (D) Trp-Lys-Thr-Cys-ThrNH2 * * f) 3- (2- (naphthyl) - (D) -Ala-Cys-Tyr- (D) Trp-Lys-Val-Cys-ThrNH2 20 g) (D) Phe-Cys-Tyr- (D) Trp-Lys-Val-Cys- | 3-Na1- NH 2 * * h) 3- (2-naphthyl) -Ala-Cys-Tyr- {D) Trp-Lys-Val-Cys-p-Na 1 -NH 2 * * i) (D) Phr-Cys-p-Na 1 - (D) Trp-Lys-Val-Cys-Thr-NH 2 wherein in each of the compounds a) to i) there is a bridge between the amino acids labeled with an * as indicated by the following formula.

Other somatostatin are: * ______________________________________

30 I I

H-Cys-Phe-Phe- (D) Trp-Lys-Thr-Phe-Cys-OH

(see Vale et al. Metabolism, 27, Supp. 1, 139 (1978)).

* *

Asn-Phe-Phe- (D) Trp-Lys-Thr-Phe-Gaba 35 (see European Patent Publication No. 1295 and Application No. 78100994.9).

* *

Me Ala-T yr- (D) T rp-Lys-Val-Phe (see Verber et al., Life Sciences, 34, 1371-1378 (1984) and European Patent Application 82106205.6 (published as No. 70021)) also known as (N-Me-Ala-Tyr-D-Trp-Lys-Val-Phe).

40 NMePhe-His (D) T rp-Lys-Val-Ala (see RF, Nutt and med., Klin. Wochenschr. (1986) 64 (suppl.VII) * *

H-Cys-His-His-Phe-Phe (D) Trp-Lys-Thr-Phe-Thr-Ser-Cys-OH

45 (see EP-A-200,188).

* * X-Cys-Phe-D-Trp-Lys-Thr-Cys-Thr-NH2 and * * 50 X-Cys-Phe-DT rp-Lys-Thr-Cys-Thr-ol wherein X is a cationic anchor, in particular * "

Ac-hArg (Et2) -Gly-Cys-Phe-D-Trp-Lys-Thr-Cys-NH2 (see EP 0.363.589 A2) in which in the aforementioned amino acids there is a bridge between the amino acids marked with an a * 55 is present.

The term "derivative" also includes the corresponding derivatives with a sugar residue.

If somatostatin contains a sugar moiety, it is preferably coupled to an N-terminal 195092, amino group and / or to at least one amino group present in a peptide side chain, more particularly to an N-terminal amino group. Such compounds and their preparation are described, for example, in WO 88/02756.

The term octreotide derivatives includes those containing a 5-D-Phe-Cys-Phe-DTrp-Lys-Thr-Cys group with a bridge between the Cys residues. Particularly recommended derivatives are N "- [α-glucosyl- (1-4-deoxyfructosyl] -DPhr = -Cys-Phe-DTrp-Lys-Thr-Cys-Thr-ol and Ν" [β -deoxyfructosyl-DPhe-Cys- Phe-DTrp-Lys-Thr-Cys-Thr-ol, each with a bridge between the -Cys portions, in particular in acetate salt form and described in Examples 2 and 1 respectively of the aforementioned application.

The somatostatin may, for example, exist in free form, salt form or in the form of complexes thereof. Acid addition salts can be formed with, for example, organic acids, polymeric acids and inorganic acids. Acid addition salts include, for example, the hydrochloride and acetates. For example, complexes are formed from somatostatin by the addition of inorganic materials, for example inorganic salts or hydroxides, such as calcium and zinc salts and / or addition of polymeric organic materials.

The acetate salt is a recommended salt for such preparations, in particular for microparticles that lead to a reduced explosion of drug at the start. The present invention also provides the pamoate salt, which is useful, in particular for implants and a method for its preparation.

The pamoate can be obtained in a conventional manner, for example by reacting embonic acid (pamoic acid) with octreotide, for example in the form of a free base. The reaction can be carried out in a polar solvent, for example at room temperature.

The somatostatin are indicated for use in the treatment of diseases where long-term administration of the drug must be taken into account, for example diseases with an etiology comprising or associated with an excess of GH secretion, for example treatment of acromegaly, for use in the treatment of gastrointestinal disorders, for example the treatment or prevention of gastric ulcer, enterocutaneous and pancreaticocutaneous fistulas, irritable gut syndrome, dumping syndrome, aqueous diarrhea syndrome, acute pancreatitis and gastroenteropathic endocrine tumors (for example vipomas, GRFomas, glucagonomas, insulinomas, gastinomas and carcinoid tumors), as well as bleeding in the gastrointestinal tract, breast cancer and complications associated with diabetes.

The polymeric carrier can be obtained from biocompatible and biologically decomposable polymers, such as linear polyesters, branched polyesters, which are linear chains that radiate from a polyol portion, e.g., glucose. Other esters are those of polylactic acid, polyglycolic acid, polyhydroxy butyric acid, polycaprolactone, polyalkylene oxalate, polyalkylene glycol esters of acids from the Kreb's cycle, for example the citric acid cycle, etc. and copolymers thereof.

The recommended polymers are the linear polyesters and polyesters with a branched chain.

The linear polyesters can be prepared from the alpha-hydroxy carboxylic acids, for example lactic acid and glycolic acid, by condensation of the lactone dimers, see, for example, U.S. Pat. No. 3,773,919.

Linear polylactide co-glycolides suitably have a molecular weight between 25,000 and 100,000 and a polydispersibility Mw / Mn of, for example, between 1.2 and 2.

The branched polyesters that can be used can be obtained using 45 polyhydroxy compounds, e.g. polyol, e.g. glucose or mannitol, as an initiator. These polyol esters are known and described in British Patent Specification GB 2,145,422 B. The polyol contains at least 3 hydroxyl groups and has a molecular weight of at most 20,000, with at least 1, preferably at least 2, for example an average of 3 of the hydroxyl groups of the polyol in the form of ester groups containing polylactide or copolylactide chains. Typically, 0.2% glucose is used to initiate the polymerization. The structure of the branched polyester is star-shaped. The preferred polyester chains in the linear and star-shaped polymer compounds that are preferably used according to the invention are copolymers of the alpha-carboxylic acid portions, lactic acid and glycolic acid, or of the lactone dimers. The molar ratios of lactide: glycolide is about 75:25 to 25:75, for example 60:40 to 40:60, with 55:45 to 45:55, for example 55:45 to 50:50, being the most recommended.

The star-shaped polymers can be obtained by reacting a polyol with a lactide and preferably also a glycolide, at an elevated temperature and in the presence of a catalyst which makes ring-polymerization feasible.

195092 6

An advantage of the star-type polymer in the compositions is that its molecular weight can be relatively high, thereby imparting physical stability, e.g., a certain hardness, to implants and microparticles, thereby avoiding their sticking together, although relatively short polylactide chains are present, leading to a controllable biological decomposition rate of the polymer ranging from a few weeks to 1 or 2 months and to a correspondingly delayed release of the peptide, which makes a depot preparation thereof suitable for, for example, a release during a month.

The star-shaped polymers preferably have an average molecular weight Mw in the range of about 10,000 to 200,000, preferably 25,000 to 100,000, in particular 35,000 to 60,000, and a polydispersibility of, for example, 1.7 to 3.0, for example 2.0 to 2.5. The intrinsic viscosities of star-shaped polymers with Mw 35,000 and Mw 60,000 are 0.36 and 0.51 dl / g in chloroform, respectively.

A star-shaped polymer with an Mw 52,000 has a viscosity of 0.474 dl / g in chloroform.

The designations "microsphere, microcapsule and microparticles" are interchangeable with regard to the invention and indicate the encapsulation of the peptides by the polymer, preferably with the peptide divided by the polymer, which is then a matrix for the peptide. In that case, the term "microsphere" or more generally "microparticle" is preferably used.

Using the phase separation method, the compositions can be prepared, for example, by dissolving the polymeric carrier material in a solvent that is not a solvent for the peptide, followed by addition and dispersion of a solution of the peptide in the polymer-solvent product. A phase inducer, e.g., a silicone fluid, is then added to induce encapsulation of the peptide by the polymer.

The effect of the explosive release of the drug can be significantly precipitated in situ from the ultrafine drug particles, by adding a drug solution to the polymer solution, before the phase separation. The prior art method involves adding dry particles directly to the polymer solution.

The therapeutic duration of the peptide release can be extended by curing the microparticles with an emulsion of buffer heptane. The method known from the prior art involves a hardening step after which no washing is carried out or which is followed by a separate aqueous washing step.

An oil-in-water type {= 0 / w) emulsion can be used for washing and curing the microspheres and for removing non-encapsulated peptide. The wash promotes the removal of the non-encapsulated peptide from the surface of the microspheres. Removing the excess peptide from the microspheres reduces the initial drug explosion, which is characteristic of many conventional encapsulation preparations. Thus, with the present microsphere compositions, a more consistent release of the drug over a period of time is possible.

The emulsion also promotes removal of residual polymer solvent and silica liquid.

The emulsion can be added to the polymer peptide mixture or the mixture can be added to the emulsion. It is recommended that the polymer peptide mixture be added to the emulsion.

The O / W emulsion can be prepared using an emulsifying agent, such as sorbitan monooleate (Span 80 ICI Corp.) or the like, to form a stable emulsion. The emulsion can be buffered with a buffer that is not harmful to the peptide and the polymer matrix material. The buffer can have a pH between 2 and 8, with a pH of 4 being recommended. The buffer can be prepared from acidic buffers, such as a phosphate buffer, acetate buffer, etc. Water can replace the buffer.

Heptane, hexane, etc. can be used as the organic phase of the buffer.

The emulsion may contain dispersants, such as silicone oil.

A recommended emulsion may contain heptane, pH 4 phosphate buffer, silicone oil, and sorbitan monooleate.

If drug release is initially desirable, a single cure step without solvent can be used instead of the emulsion cure. As the solvent, heptane, hexane, etc. can be used.

Other alternatives to the O / W emulsion can also be used for curing the microcapsules, such as solvent plus emulsifier for curing the microcapsules without washing and solvent plus emulsifier for curing followed by a separate washing step.

The O / W emulsion can be used without dispersant. However, the dispersant avoids aggregation of the dry particles of the microcapsules due to static electricity and promotes the reduction of the residual solvent content.

Examples of the solvent for the polymer matrix material include methylene chloride, chloroform, benzene, ethyl acetate, etc. The peptide is preferably dissolved in an alcoholic solvent, e.g. methanol, miscible with the polymer solvent.

The phase inducers (coacervatives) are solvents that are miscible with the polymer drug mixture and that cause the embryonic microcapsules to form before curing; silicone oils are the recommended phase inducers.

The O / W emulsion can be prepared in a conventional manner using heptane, hexane, etc. as an organic phase.

The microparticles can also be obtained according to the generally known spray-drying method. According to this method, the somatostatin or a solution of the peptide in an organic solvent, for example methanol, in water or in a buffer, for example pH 3-8 and a solution of the polymer in an organic solvent, which is immiscible with the the former, for example methylene chloride, intimately mixed.

The first-mentioned solution, suspension or emulsion, is then sprayed into a stream of air, preferably warm air. The resulting microparticles are collected, for example with a cyclone and, if desired, washed, for example in a buffer solution with a pH of for example between 3.0 and 8.0, preferably of 4.0, or distilled water and dried under reduced pressure, for example at a temperature of 20-40 ° C. The washing step can be used if the particles give a drug explosion in vivo and the extent of this explosion is undesirable. An acetate buffer can be used as the buffer.

As a result, microparticles can be obtained that exhibit a better somatostatin release profile in vivo.

A sustained-release preparation is provided prepared by mixing a somatostatin or a solution of a somatostatin in methanol or water or a pH 3-8 buffer and a solution of the polylactide co-glycolide in methylene chloride and the solution formed, emulsion or suspension of somatostatin, spray a stream of warm air into the polymer solution, collect the microspheres and wash them in a buffer solution with pH 3.0-8.0 or distilled water and place them under reduced pressure at a temperature of 20 to 40 ° C to dry. Compared to microparticles prepared by the phase separation method, they do not contain silicone oil, even in traces, since no silicone oil is used in the spray-drying process.

The compositions can also be prepared by a triple emulsion method. According to a typical method, peptide, e.g. octreotide, is dissolved in a suitable solvent, e.g. water, and intensively in a solution of the polymer, e.g. 50/50 poly (D, L-lactide-co-glycolide) glucose in a solvent which does not solvent for the peptide is emulsified, for example, in methylene chloride. Examples of the solvent for the polymer matrix material include.

Methylene chloride, chloroform, benzene, ethyl acetate, etc. The resulting water-oil (w / o) emulsion is further dissolved in an excess of water containing an emulsifying material, for example an anionic or non-ionic surfactant or lecithin or a protective colloid for example gelatin, dextrin, carboxymethylcellulose, polyvinylpyrrolidone, polyvinyl alcohol, emulsified to give continuous formation of the triple (w / o / w) emulsion. The microparticles are formed by spontaneous precipitation of the polymer and cured by evaporation of the organic solvent. Gelatin serves to prevent agglomeration of the microspheres. After sedimentation of the microspheres, the supernatant is decanted off and the microparticles are washed with water and then with acetate buffer. Finally, the microparticles are filtered and dried.

The peptide can also be dispersed directly in the polymer solution, after which the resulting suspension is mixed with the gelatin-containing aqueous phase.

The triple emulsion method is known from U.S. Pat. No. 4,652,441. According to this patent, in a first step, a solution of a drug (1), for example somatostatin in water (column 2, lines 31-32), is intimately mixed with an excess of a polylactide-co-glycolide solution (2) in another solvent, in which the first solvent is insoluble, for example methylene chloride, whereby a water-in-oil type (w / 0) emulsion (3) of fine, drug-containing droplets of (1) in solution ( 2) is obtained. In solution (1), a so-called drug-containing material (column 1, line 31), for example gelatin, albumin, pectin or agar, is also dissolved.

In a second step, the viscosity of the internal phase (1) is increased by applying suitable measures such as heating, cooling, pH change, addition of metal ions or crosslinking of, for example, gelatin with aldehyde.

According to a third step, an excess of water is thoroughly mixed with the w / 0 emulsion (3) (column 7, lines 52-54), which leads to a w / 0 ftt-type temary layer emulsion. The excess water can, if desired, contain a so-called emulsifier (column 7, line 56), selected from the group consisting of, for example, an anionic or non-ionic surfactant, or, for example, polyvinylpyrrolidone, polyvinyl alcohol or gelatin.

In a fourth step, the "" / "/" emulsion is subjected to "in-water drying" (line 52). This means that the organic solvent in the oil layer is desorbed to develop micro particles. The desorption is carried out in a manner known per se (column 8, lines 3-5), for example by reducing the pressure with stirring (column 8, lines 5-7) or, for example, by nitrogen gas through the oil layer (e.g. methylene chloride) (line 19).

The microparticles formed are recovered by centrifugation or filtration (lines 26-27) and the components not included in the polymer are removed by washing with water (line 29). If desired, the microparticles are heated under reduced pressure to obtain better removal of water and solvent (e.g., methylene chloride from the wall of microparticles (lines 30-32).

Although the above-described method is satisfactory for the preparation of the compositions, the so-called drug-containing material mentioned above, for example gelatin, albumin, pectin or agar, is still present in the obtained microparticles.

If the addition of the drug-containing material (= in solution (1)) and the step of increasing the viscosity of the internal phase are avoided and in the excess of water of the ternary w / o / w emulsion the measure of while adding an emulsifying material or a protective colloid, such as gelatin, satisfactory microparticles can nevertheless be obtained, while, moreover, the microparticles no longer contain any drug-containing material and only a very small amount of methylene chloride.

A method for preparing microparticles is thus intimately mixing: a) a solution of a drug, preferably a somatostatin, in particular octreotide, in an aqueous medium, preferably water or a buffer, preferably in a wt. /full. ratio of 0.8-4.0 g / 1-120 ml, in particular 2.5 / 10 and in a buffer with pH 3-8, in particular an acetate buffer, and b) a solution of a polymer, at preferably a polylactide co-glycolide, as mentioned above, in an organic solvent that is immiscible with the aqueous medium, for example methylene chloride, preferably in a weight percent. ratio of 40 g / 90 to 400 ml, in particular 40/100, preferably in such a way that the wt / wt. the ratio of the drug to the polymer is between 1/10 to 50, in particular 1/16 and the volume / volume ratio of the aqueous medium / organic solvent is 1 / 1.5 to 30, in particular 1 / 10, intimately mixing the w / c emulsion of a) in b) together with c) an excess of an aqueous medium, preferably water or a buffer, for example an acetate or phosphate buffer, preferably at a pH of 3 8, which contains an emulsifying material or a protective colloid 35, preferably in a concentration of 0.01 to 15.0%, in particular gelatin, especially in a concentration of 0.1 to 3%, especially 0.5 wt. %, preferably in a full / full. mixing speed ratio of ab) / c) from 1/10 to 100, in particular 1/40, without adding any drug-containing material to the water-in-oil emulsion or by using any intermediate viscosity-increasing step, 40 - curing the embryonic microparticles in the formed w / o / w emulsion by desorption, preferably by evaporation, of the organic solvent, preferably methylene chloride, and - by isolation, optionally washing and drying the resulting microparticles.

There is also a process variant in which the drug is directly dispersed in the polymer solution, after which the resulting dispersion is mixed with the gelatin-containing aqueous phase.

Like microparticles obtained by the spray drying method, the microparticles obtained by the above method do not contain silicone oil. Compared to the microparticles prepared according to the known triple emulsion process type, they do not contain a protective colloid.

The sustained-release preparations may also be prepared by other methods known per se, e.g. 50 - if the peptide is sufficiently stable to produce an implant, by heating microparticles containing the peptide, e.g. a somatostatin in a polylactide co -glycolide, in particular as described above or a mixture thereof obtained by mixing the peptide and the polymer, at a temperature of, for example, 70 to 100 ° C and extruding and cooling the compact mass, after which the extrudate is cut and optionally washed and dried.

Suitably, the compositions are prepared or prepared under aseptic conditions.

The compositions can be used in depot form, for example as injectable microspheres or implants.

They can be administered in a conventional manner, for example subcutaneously or by intramuscular injection, for example for indications known for the medicine contained therein.

The sustained-release preparations containing octreotide can be administered for all known indications of the octreotide or derivatives thereof, for example those described in British Pat. No. 2,199,829 A, pages 89-96, as well as for acromegaly and breast cancer.

The microparticles can have a size ranging in diameter from about 1 to 250 microns, preferably from 10 to 200, especially 10 to 130 microns, e.g. from 10 to 90 microns. Implants can, for example, have a size of approximately 1 to 10 m3. The amount of drug, i.e., peptide present in the composition, depends on the desired daily dose being delivered and thus on the biological decomposition rate of the encapsulating polymer. The precise amount of peptide can be determined by bioavailability studies. The compositions can be peptide in an amount of at least 0.2, preferably 0.5 to 20% by weight, relative to the polymer matrix, preferably 2.0 to 10, in particular 3.0 to 6,% by weight contain.

The release time of the peptide from the microparticle can be 1 to 2 weeks to about 2 months.

Suitably, the sustained-release preparation contains a somatostatin, for example octreotide, in a biologically decomposable, biocompatible polymeric carrier which, administered subcutaneously at a dose of 10 mg of somatostatin to a body weight of the animal to a rat, has a concentration of a somatostatin in the blood plasma of at least 0.3 ng / ml and preferably less than 20 ng / ml over a period of 30 days or suitably a period of 60 days.

It is also possible that the sustained-release preparation contains a somatostatin, for example octreotide, in a biologically decomposable, biocompatible, polymeric carrier, which, intramuscularly administered to a rabbit at a dose of 5 mg / kg body weight, has a concentration of somatostatin of at least 0.3 ng / ml over a period of 50 days and suitably a concentration of at most 20 ng / ml.

Other recommended properties of the somatostatin obtained, for example octreotide-containing depot preparations, depend on the preparation methods used:

Phase separation method 30

Rabbit 5 mg somatostatin / kg, intramuscular delay (CM2 days) 76% average plasma level (Cp ideal) (0-42 days) 4 ng / ml AUC (0-42 days) 170 ng / ml x days 35

Spray drying method

Rat 10 mg somatostatin / kg, Subcutaneous delay (0-42 days)> 75% average plasma level (Cp ideal) (0-42 days) 4-6 ng / m AUC (0-42 days) 170-210 ng / ml x days 40

Rabbit 5 mg somatostatin / kg, intramuscular delay (0-43 days)> 75% average plasma level (Cp.ideaa,) (0-43 days) 4-6 ng / m „AUC (0-43 days) 200-240 ng / ml x days 45

Tripel emulsion method

Rat 10 mg somatostatin / kg subcutaneous delay (0-42 days)> 75% average plasma level (Cp.ideaai) (0-42 days) 4-6.5 ng / ml AUC (0-42 days) 170-230 ng / ml x days

Rabbit 5 mg somatostatin / kg intramuscular delay (0-42 / 43 days)> 74% ψ 195092 10 average plasma level (Cp ideal) (0-42 / 43 days) 3.5-6.5 ng / ml 'AUC ( 0-42/43 days) 160-270 ng / ml x days

Thus, somatostatin, preferably octreotide and octreotide analogous preparations, having the following properties are provided: 1. a delay of at least 70%, preferably at least 74%, for example at least 75%, 80%, 88% or at least 89% over a period of 0 to 42 or 43 days, and / or 2. an average plasma level (Cp ideal) of 2.5-6.5, preferably 4-6.5 ng / ml over a period of 0 to 42 days, in rats, if 10 mg of somatostatin is administered subcutaneously and / or a noise-reduced plasma level of 3.5-6.5, for example 4-6.5 ng / ml over a period of 0 to 42 or 43 days at a rabbit if 5 mg somatostatin is administered intramuscularly, and / or 3. an AUC over a period of 0 to 42 days of at least 160, preferably of 170-230 ng / ml x days for a rat, if 10 mg of somatostatin subcutaneously is administered and / or an AUC over a period of 0 to 42 or 43 days of at least 160, preferably from 180-275, for example from 20 0 1 g to 275 mg / ml x days for a rabbit, if 5 mg of somatostatin is administered intramuscularly.

For the quantitative characterization of the above-described sustained-release preparations, use is made of the method of surface deviation (AD) of F. Nimmerfall and J. Rosenthaler;

Internal. J. Pharmaceut. 32, 1-6 (1986). Briefly, according to the AD method, the surface deviations of the experimental plasma profile of an ideal profile that have a constant average plasma level (= Cp-idealai) are obtained by converting the experimental surface under the plasma-mirror-time curve (AUC) into a rectangle with same area calculated. The percentage of delay is calculated from the percentage surface deviation (indicated by AUC) in the following way:% delay = 100 x (1 - AD / AUC).

According to this method, the complete plasma profile is determined for a preselected period of time characterized by a single number index.

In Proc.Natl.Acad.Sci.USA 85 (1988) 5688-5692, Figure 4 shows a plasma mirror profile of the octapeptide analogous to somatostatin of formula * * D-Phe-Cys-Tyr-D-Trp-Lys-Val- Cys-Trp-NH 2 in rats.

However, a clear comparison cannot be made with the plasma level values of the compositions according to the invention in rats, as mentioned above, since the plasma level profile described is based on a different method of administration (intramuscular injection) and, more importantly, the loading level of the microcapsules (between 2 and 6%) and the dosage amount for administration (25 to 50 mg portions of microcapsules for 30 days, although determinations were made for at least 45 days) are not accurately indicated. In addition, the type of poly (DI-lactide-co-glycolide) used is not accurately described.

The descriptive value of the publication is therefore too low to be regarded as a pre-publication which adversely affects the invention.

The following examples illustrate the invention.

The Mw of the polymers is the average molecular weight determined according to GLPC using polystyrene as standard.

Example 1 45

Octreotide pamoate

10.19 g (10 mM) of octreotide free base and 3.88 g (10 mM) of embonic acid were dissolved in 1 l of a mixture of equal parts of water and dioxane. The reaction mixture was filtered and lyophilized, whereby a yellow powder [α] 20 D = + 7.5 ° (C = 0.35, in DMF) of octreotide pamoate hydrate was obtained. The 50 factor is 1.4, wherein factor is the weight of lyophilisate weight of the octreotide present therein.

The pamoate can replace the octreotide acetate present in the microparticles of Examples 2-10 and has excellent stability.

Example 2 55

1 g of poly (D, L-lactide-co-glycolide) 50/50 molar, Mw 45,000 was dissolved; polydispersity ca. 1.7) with magnetic stirring in 15 ml methylene chloride, after which 75 mg octreotide acetate dissolved in 0.5 ml 11 195092 methanol was added. Then 15 ml of silicone oil (Dow 360 Medical Fluid 1000cs brand) (silicone fluid) was added to the polymer peptide mixture. The resulting mixture was added to a stirred emulsion containing 400 ml of n-heptane, 100 ml of pH 4 phosphate buffer, 40 ml of Dow 360 Medical Fluid, 350 cs and 2 ml of Span 80 (emulsifier). Stirring was then continued for a minimum of 10 minutes. The resulting microparticles were recovered by filtration under reduced pressure and were dried overnight in a vacuum oven. The yield was about 90% of microparticles with a size of 10-40 microns.

The microparticles were suspended in a carrier and administered intramuscularly at a dose of 4 mg octreotide to New Zealand rabbits. Blood samples were taken periodically with plasma levels of 0.5 to 1.0 ng / ml measured for 30 days, measured by radioimmunoassay (RIA) analysis.

Example 3

1 g of poly (D, L-lactide-co-glycolide) glucose (Mw - 45,000) (55/45 molar) obtained according to the method of British Patent Specification 2,145,422 B, polydispersibility of about 1.7, was prepared (0.2% glucose) with magnetic stirring in 25 ml of ethyl acetate, after which 75 mg of octreotide dissolved in 3 ml of methanol were added. Then 25 ml of silicone oil (Dow 360 Medical Fluid brand, 1000 cs) was added to the polymer-polypeptide mixture. The resulting mixture was added to the emulsion described in Example 2. The mixture was then stirred for a minimum of 10 minutes. The resulting microparticles were recovered by vacuum filtration and dried overnight in a vacuum oven. The yield was greater than 80% microparticles in the range of 10-40 microns.

The microparticles were suspended in a carrier and administered intramuscularly at a dose of 4 mg octreotide to white New Zealand rabbits. Blood samples were taken periodically, showing that the plasma levels were between 0.5 and 2 ng / ml for 21 days, measured with RIA.

25

Example 4

A solution of 1.5 g of octreotide acetate in 20 ml of methanol was added with stirring to a solution of 18.5 g of poly (D, L-lactide-co-glycolide) glucose (50:50 molar, molecular weight) 45,000) in 500 ml of methylene chloride. Phase separation was performed by adding 500 ml of Dow 360 Medical Fluid (1000 cs) and 800 ml of Dow 360 Medical Fluid (350 cs) to the peptide polymer suspension. The resulting mixture was added to a stirred emulsion consisting of 1800 ml of n-heptane, 2000 ml of sterile water and 40 ml of Span 80. After stirring for 10 minutes, the microspheres were collected by filtration under reduced pressure.

Half of the product was dried in a vacuum oven at 37 ° C.

The level of residual methylene chloride was 1.2%.

The other half of the product was washed by stirring with 1000 ml of ethanol containing 1 mg of Span 80. After stirring for 1 hour, the ethanol was decanted and the microparticles were stirred for 1 hour with 1000 ml of n-heptane containing 1 ml of Span 80. After stirring for 1 hour, the microparticles were collected by filtration under reduced pressure and dried overnight at 37 ° C in a vacuum oven. The level of the remaining methylene chloride from the microparticles washed in this way was lowered from 1.2% to 0.12%.

The combined yield of the product was 18.2 g (91%) of microparticles containing 5.6% octreotide, with an average diameter of 24 microns and 1.5% residual heptane.

The microparticles were suspended in a carrier and administered intramuscularly at a dose of 5 mg / kg dose of octreotide to white rabbits. Blood samples were taken periodically, showing that plasma levels were between 0.3 and 7.7 ng / ml for 49 days, measured with RIA.

Example 5 50

1 g of poly (D, L-lactide-co-glycolide) glucose, molecular weight 46,000 (50:50) molar was prepared according to the method of British Patent Specification 2,145,422 B, polydispersibility about 1.7, prepared from 0.2 % glucose) with magnetic stirring in 10 ml of methylene chloride, after which 75 mg of octreotide dissolved in 0.133 ml of methanol were added. The mixture was intimately mixed for 1 minute, for example with the aid of an Ultra-Turax at 20,000 revolutions per minute, whereby a suspension of very small crystals of octreotide in the polymer solution was obtained.

The suspension was sprayed with the aid of a high-speed turbine (Niro Atomizer) 195092 12 and the small droplets were dried in a stream of warm air, whereby microparticles became.

obtained. These microparticles were collected with a "zyklon" and dried overnight in a vacuum oven at room temperature.

The microparticles were washed for 5 minutes with 1/15 molar acetate buffer, pH 4.0, and were dried again in a vacuum oven at room temperature. After 72 hours, the microparticles were sieved (0.125 mm mesh), whereby the final product was obtained.

The microparticles were suspended in a carrier and administered intramuscularly at a dose of 5 mg / kg of octreotide to white rabbits (chinchilla bastard) and subcutaneously at a dose of 10 mg / kg of male rats. Blood samples were taken periodically, showing that plasma levels were between 0.3 and 10.0 ng / ml (5 mg dose) in rabbits and between 0.5 and 7.0 ng / m! for 42 days in rats, measured by radioimmunoassay (RIA) analysis.

Example 6 Microparticles were prepared by spray drying in the same manner as described for Example 5, with the only change that octreotide was suspended directly in the polymer solution, without using methanol.

The microparticles were suspended in a carrier and administered subcutaneously to male rats at a dose of 10 mg / kg octreotide. Blood levels were taken periodically, showing that plasma levels were between 0.5 and 10.0 ng / ml for 42 days, measured by radioimmunoassay (RIA) analyzes.

Example 7 1 g of poly (D, L-lactide-co-glycolide) glucose, molecular weight 4600 (50:50 molar) prepared according to the method of British Patent Specification 2,145,422 B, polydispersibility about 1.7 prepared from (0.2% glucose) in 2.5 ml of methylene chloride, after which 75 mg of octreotide dissolved in 0.125 ml of deionized water was added. The mixture was intimately mixed for 1 minute, for example with an Ultra-Turax at 20,000 revolutions per minute (internal W / O phase).

1 g of gelatin A was dissolved in 200 ml of deionized water at 50 ° C and the resulting solution was cooled to 20 ° C (outer W phase). The W / O and W phases were intimately mixed. Thereby, the W / O / inner phase was separated into small droplets, which were homogeneously dispersed in the W outer phase. The resulting triple emulsion was stirred slowly for 1 hour. The methylene chloride was evaporated and the microcapsules were cured from the droplets of the inner phase. After the microparticles were deposited, the supernatant was filtered off with suction and the microparticles were recovered by filtration under reduced pressure and rinsed with water to remove gelatin.

Drying, sieving, washing and re-drying of the microparticles was carried out in the manner described in Example 5. The microparticles were suspended in a carrier and administered intramuscularly to white rabbits (chinchilla bastard) at a dose of 5 mg / kg octreotide and administered at a dose of 40 10 mg / kg subcutaneously to male rats. Blood samples were taken periodically to show that plasma levels were between 0.3 and 15.0 ng / ml (5 mg dose) in rabbits and between 0.5 and 8.0 ng / ml in rats for 42 days, measured with radioimmunoassay (RIA) analyzes.

Example 8 45

Microparticles were prepared by the triple-emulsion method, in the same manner as described for Example 7 with three changes: 1. 0.25 ml of acetate buffer, pH 4.0 instead of 0.125 ml of water was used to prepare the W / O inner phase.

2. rinsing after collecting the microparticles was performed with a 1/45 molar acetate buffer, pH 4.0 instead of with water.

3. the further washing of the microparticles was omitted.

Example 9 55

Microparticles were prepared according to the triple emulsion method, in the same manner as described for Example 8, with the only change that the W / O inner phase was obtained by using water

Claims (2)

13, 195092, containing 0.7% (w / v) sodium chloride instead of acetate buffer. Example 10 Microparticles were prepared in the same manner as described in Example 7 with the only difference that the drug compound was directly suspended in the polymer solution, after which the resulting dispersion was mixed with the gelatin-containing aqueous phase. 10 Conclusions 1. Octreotide pamoate. A method for preparing octreotide pamoate, wherein octreotide is reacted with embonic acid or a reactive derivative thereof.