HK1058366B - Method for producing peptide salts, their use, and pharmaceutical preparations containing these peptide salts - Google Patents

Description

Method for producing peptide salts, use of peptide salts and pharmaceutical preparations comprising peptide salts

The invention relates to a novel method for producing peptide salts, in particular poorly soluble peptide salts, and to the use thereof for producing medicaments. The invention further relates to pharmaceutical preparations containing at least one peptide salt produced according to the invention and to the production thereof.

A process for the preparation of poorly soluble peptides by precipitation of poorly soluble acid addition salts of the peptide by reacting an aqueous solution of an acid salt with an acetic acid solution of a basic peptide is described in International patent application No. PCT/EP 94/03904. For example, a process for the preparation of the LHRH (luteinizing hormone releasing hormone) antagonist cetrorelix pamoate (cetrorelix emonate) is described.

The invention relates to a novel process for the preparation of peptide salts, characterized in that an acid addition salt of a basic peptide (starting peptide salt) (1) is reacted with an ion exchanger of mixed bed or a mixture of acid and base ion exchangers in the presence of a suitable diluent to form a free basic peptide, the ion exchanger is subsequently separated off, the free basic peptide is then reacted with an inorganic or organic acid to form the desired acid addition salt of the peptide (final peptide salt) (2), and the diluent is subsequently removed.

Here, the term "basic peptide" refers to a poly (amino acid) containing a basic amino acid such as arginine, pyridylalanine or lysine, or to the N-terminus of the peptide or just at least one basic group, even in a substructure sense within a larger overall structure.

Preferred peptides are the LHRH antagonists antide, A-75998, ganirelix, NaI-Glu antagonists, cetrorelix, teverelix (Antarelix *) and antagonists according to US 5,942,493 and DE19911771.3, the contents of which are incorporated herein by reference. Additional peptides are abarelix (abarelix), azaline B, degeprelix (demirelix), ramorex (ramorelix) (Stoeckemann and Sandow, j. cancer res. clin. oncol.1993, 119, 457) and RS-68439. The structure of these peptides can be found in Behre et al, "GnRH antagonists: review "(Proceedings of the 2)^ndWorld Conference on emulsification introduction (The second World Ovulation Induction Conference corpus), The part Publishing Group Ltd.); an article by Kutscher et al (angelw. chem. (applied chemistry) 1997, 109, 2240).

The acid addition salts of the peptides used as starting materials are preferably readily soluble salts such as acetates, hydrochlorides, sulfates.

According to the novel process, the starting peptide salt is completely or partially dissolved in a diluent or suspended therein. The diluent is then added. The solvent and diluent may be the same or different. Possible solvents and diluents are, for example: water, ethanol, methanol, propanol, isopropanol, butanol, acetone, dimethyl ketone, methyl ethyl ketone, dimethyl acetamide, dimethylformamide, N-methylpyrrolidone, acetonitrile, pentane, hexane, heptane, and mixtures thereof. Ethanol, isopropanol or acetone are preferred. A water content of 1 to 60%, preferably 5 to 50%, is likewise preferred.

The mixed bed ion exchanger, i.e. the mixture of acidic and basic ion exchangers, is added to the solution or suspension of the starting peptide salt. One possible ion exchanger is for example Amberlite *.

The amount of ion exchanger used depends on the number of basic groups per peptide. This amount is determined by the method added until a certain constant pH is reached. For example, 1 gram of cetrorelix requires 10 grams of Amberlite MB-3.

In the preparation of the base, the pH of the base solution is adjusted to 7.5 to 13, depending on the active compound salt used, in particular in the case of peptide salts containing amino acids which react in a basic manner, in the case of salts of LHRH antagonists, such as cetrorelix, D-63153, abarelix, ganirelix, ramorex, which may be present, for example, as acetate.

To avoid decomposition of the peptide, the temperature should not exceed 25-30 ℃. The reaction time to prepare the free base is typically carried out within a few minutes, for example, 20 minutes starting with cetrorelix acetate. It may also be longer, for example, about 1 hour from the start of the reaction with cetrorelix pamoate. After the pH has reached a constant value, the reaction should be terminated, since, for example, decomposition products may form owing to the alkalinity of the solution.

The ion exchanger is then removed from the reaction mixture. The removal can be carried out by sieving, filtering off, centrifugation or column filtration.

Solutions of the free peptide base are unstable, become clear or cloudy, and should be reacted with the acid as quickly as possible to form the desired acid addition salt. The acid may be added as a solid or as a solution or suspension. The free peptide base solution can be converted to an acid in exactly the same manner.

The reaction time is in the range of several minutes to several hours. For example, for the formation of cetrorelix pamoate, the reaction time is 1.5 hours.

The reaction solution, which is usually clear, is then subjected to sterile filtration. The solvent is then removed to yield the pure peptide salt. Alternatively, excipients (excipients), additives or carriers (vehicles) may be added to the solution prior to removal of the solvent. Excipients may be added in solid form prior to sterile filtration or in the form of a sterile filtered solution after sterile filtration.

Suitable excipients are, for example, mannitol, sorbitol, xylitol, soluble starch.

According to the invention, the following salts can be prepared by adding suitable acids: acetate, adipate, ascorbate, alginate, benzoate, benzenesulfonate, bromide, carbonate, citrate, chloride, dibutyl phosphate, dihydrogen citrate, dioctyl phosphate, dihexadecyl phosphate, fumarate, gluconate, glucuronate, glutamate, bicarbonate, bitartrate, hydrochloride, bicarbonate, iodide, lactate, alpha-lipoic acid, malate, maleate, malonate, pamoate, (pamoate), palmitate, phosphate, salicylate, stearate, succinate, sulfate, tartrate, tannate, oleate, octyl phosphate.

The invention is illustrated by the following examples, without thereby being limited thereto.

Example 1

46.47 g of D-20761 were added in portions to 1193 g of water and dissolved with stirring (solution 1). Solution 1 was then diluted with 3261 g of 96% ethanol under stirring (solution 2). After dilution, the solution 2 was filtered through a glass fiber prefilter, and the filtrate was mixed with 390 g of Amberlite MB3 (mixed bed ion exchanger formed of strongly acidic cation and anion exchangers) with stirring (to give a mixture 1). 316.8 g of mannitol were dissolved with stirring in 1267 g of water (solution 3). After stirring for 15 minutes, the pH of the supernatant of mixture 1 was measured and after stirring for a further 5 minutes the pH was measured again. Subsequently, after the pH had reached 12.5, the solution was separated from Amberlite MB3 using a fine mesh screen (to give solution 4).

4162 g of solution 4 were mixed with 5.34 g of pamoic acid (embonic acid) with stirring. The mixture was stirred vigorously for an additional 1.5 hours, and the slightly cloudy solution was then filtered through a glass fiber prefilter. The pH of this solution was 8.4 (to give solution 5). The measured pH values were all measured by a ground glass electrode using a viscous electrolyte liquid. These pH values are only to be regarded as relative values, since the measured solutions or suspensions contain ethanol and therefore show significantly higher values.

In the reaction apparatus, 3333 g of solution 5 adjusted to room temperature was sterile-filtered, and 528 g of solution 3 adjusted to room temperature was sterile-filtered to incorporate solution 5 (to give solution 6) with stirring.

The solution was heated to 6 to 40 ℃ and then the ethanol/water mixture was distilled off in vacuo, reducing the solution to less than 1931 g (to give suspension 1). This cetrorelix pamoate suspension 1 was cooled to room temperature and diluted to 3000 g with sterile filtered water for injection with stirring (suspension 2). The ready-to-use suspension 2, adjusted to room temperature, was then dispensed in 3.0 g portions into 10 ml injection vials, fitted with freeze-drying stoppers and transferred to a freeze-drying apparatus.

These injection vials were frozen in a freeze-drying apparatus at a plate temperature (plate temperature) -40 ℃. Drying is accomplished by a programmed drying process that raises the plate temperature from-40 ℃ to 20 ℃. The Freeze Drying (FD) unit was filled with sterile filtered nitrogen, the injection vials in the unit were sealed, and the crimp cap was capped and rolled.

After freeze-drying, these sealed injection vials were sterilized by gamma irradiation at 12kGy (minimum) -15 kGy. The latter is optional.

1 injection vial of 140.0(7) mg was filled with 34.07 mg of cetrorelix pamoate, corresponding to 30mg of cetrorelix and 106 mg of mannitol. For reconstitution, 2 ml of water for injection was used. The resulting suspension may be administered by i.m. (intramuscular) or s.c. (subcutaneous) administration.

Biological effects:

cetrorelix pamoate (2: 1) lyophilisate (30mg) obtained according to example 1 is resuspended in 2 ml of water for injection and can then be administered parenterally, preferably subcutaneously (s.c.) or intramuscularly (i.m.).

In the case of subcutaneous administration, cetrorelix pamoate (2: 1) can achieve a bioavailability of about 30-50% (100% being defined as the bioavailability by intravenous administration of cetrorelix acetate). A particular advantage of the lyophilisate of cetrorelix pamoate (2: 1) is that it has little or no "breakthrough effect" on the patient. The duration of action is dose-dependent and at doses of 30-150 mg, is 2-8 weeks or longer. The lyophilisate of cetrorelix pamoate (2: 1) according to the invention has been studied in the human clinical phase I trial.

Figure 1 illustrates the progression of cetrorelix plasma concentration as a function of time (in hours) starting from the intramuscular administration of 60mg of cetrorelix pamoate (2: 1) lyophilisate described in example 1 to humans. No burst effect was measured (about 100ng/ml (ng/ml)). The duration of action is over 700 hours. 150 hours after administration, a constant plasma level of about 2ng/ml was achieved. The bioavailability is about 40%.

The fields of application of the peptide salts according to the invention are, for example, BPH, myoma and endometriosis.

Claims (6)

1. A process for the preparation of a pharmaceutical formulation useful for parenteral administration to a mammal, characterized in that a readily soluble acid addition salt of the basic peptide LHRH antagonist cetrorelix is reacted with a mixed bed ion exchanger or a mixture of acidic and basic ion exchangers in the presence of a suitable diluent to form a free basic peptide, the ion exchanger is subsequently separated off, the free basic peptide is subsequently reacted with an inorganic or organic acid to form the desired sparingly soluble acid addition salt of the peptide, suitable pharmaceutical excipients, carriers and/or fillers are subsequently added, and the diluent is removed.

2. A process according to claim 1, characterized in that the acid is pamoic acid, stearic acid or salicylic acid.

3. A process according to claim 1 or 2, characterized in that the molar ratio of cetrorelix to pamoic acid is 2: 1.

4. A method according to claim 1 or 2, characterized in that the diluent is removed by freeze drying.

5. The method according to claim 1, characterized in that the diluent is selected from the group consisting of: water, ethanol, methanol, propanol, isopropanol, butanol, acetone, dimethyl ketone, methyl ethyl ketone, dimethyl acetamide, dimethylformamide, N-methylpyrrolidone, acetonitrile, pentane, hexane, heptane, and mixtures thereof.

6. A process according to claim 1, characterized in that said excipients are selected from: mannitol, sorbitol, xylitol, soluble starch.