MXPA97003019A - Liposoma composition containing selegil - Google Patents

Abstract

The present invention relates to a liposome composition, characterized in that it contains as the active ingredient (-) - n-alpha-dimethyl-N- (2-propynylphenylethylamine) (selegiline) and / or a salt thereof.

Description

COMPOSITION OF LIPOSOMA CONTAINING SELEGILIN DESCRIPTION OF THE INVENTION The invention relates to liposomal compositions containing as an active ingredient selegiline (-) - (Na-dimethyl-N- (2-propynylphenylethylamine) and / or a salt thereof, In addition, the invention relates to its preparation, pharmaceutical compositions containing them and their therapeutic use.Today, a large number of liposomal compositions and processes for their preparation are known in the art, Such compositions are, in fact, "drug delivery systems" (DDS), as described above. described by G. Gregoriadis et al (Receiver-mediated targeting of drugs, Plenum Press, New York, 243-266, 1980.) They contain the active ingredient similarly to the encapsulated state in one or more lamellar membranes comprising lipids, i.e. The good absorption and bioavailability of the active ingredient (s) can be influenced, inter alia, by the composition and method of preparation of the liposomes, in such a way that they can supply bring the active ingredient into a specified area. In liposomal compositions, the active ingredient is comprised of one or more lamellar lipids which also serve as a carrier of the active ingredient. The ultilamellar lipid carriers (MLV) were first prepared and described by Bangham et al (J. Mol. Biol. 13, 238-252, 1965). When biologically active substances are encapsulated in small vesicles of lipids of a sheet, water-soluble substances can be encapsulated by low efficiency due to the small volume of water encapsulated in the SUV (small unilamellar lipid vesicles) (patent specification of the United States No. 4,089,801). The unilamellar lipid vesicles were prepared by other methods, i.e. by injection of ethanol [S. Batzri and E.D. Korn: Biochem. Biophys. Acta. 298: 1015-1019, (1973)] or ether [D. Deamer and A.D. Bangham: Biochem. Biophys. Acta 443, 629-634, (1976)] in such a way that the lipid solution in an organic solvent was rapidly injected into a buffer solution and in this way the unilamellar liposomes are formed spontaneously. The method is rapid and generally used, but results in diluted liposomal preparations and poor encapsulation effectiveness. Unilamellar liposomes can also be formed by the so-called detergent removal system [H. G. Weder and O. Zumbuehl: Liposome Technology, ed. G. Gregoriadis, CRC Press Inc., Boca Raton, Florida, Vol. 1. Ch 7, pp. 79-107, (1984)], in the course of which lipids and other substances are arranged together with detergents and then the detergents are removed by dialysis. The multilamellar liposomal encapsulation is carried out in accordance with United States Patent Specification No. 4,234,871 (Papahadjopoulos) by the so-called reverse phase evaporation (REV) technique and in accordance with the United States patent specification No 4,016,100 (Suzuki) by lyophilization of the lipid-aqueous dispersion of lipids and the biologically active substance. The preparation of a stable aqueous liposomal suspension which can be stored for 6 months at 40 ° C is described in the United States patent specification published No. WO 93/20934. In the course of the preparation of the above liposomal compositions, the dispersion of the lipids and the aqueous phase is carried out in contact reactions on an inert solid material, in most cases on glass beads [patent specification of United States No. 4,485,054. According to U.S. Patent Specification No. 4,761,288, which is expressly incorporated herein by reference, multi-phase systems are prepared to improve the absorption of biologically active substances that have poor water solubility, which contain the active ingredient in the form of a supersaturated solution, in solid form and encapsulated in multilamellar lipid vesicles. The vesicle, the solution and the solid form of the biologically active compound are dispersed in a hydrocolloidal gel. The hydrocolloid gel is prepared using the method for preparing the multilamellar lipid vesicles described in U.S. Patent No. 4,485,054 expressly incorporated herein by reference. Thus, liposomal compositions can be "prepared, in which the active ingredient is present in higher concentrations than what might be expected based on its water solubility and / or liposolubility. In the patent specification of the States No. 4,937,078 discloses liposomal compositions of locally applied anesthetic and analgesic active ingredients. It was found that the locally applied active ingredients are more effective in the encapsulated liposomal state than the usual ointment, cream or liquid compositions. The formation of the liposome by itself has been carried out as described in US Pat. Nos. 4,485,054 and 4,761,288.

The preparation of all liposomal compositions has been directed to active ingredients which have little solubility in water first of all to increase absorption capacity and local concentration and / or for programmed absorption. The invention relates to the preparation of liposomal compositions of selegiline or its salts, which are very soluble in water and solvents (1 g / 3 ml in water, 1 g / 5 ml in chloroform or 1 g / 3 ml in methanol) , in addition to its oral, parenteral or local therapeutic applications, optionally in controlled release, transdermal compositions. Selegiline is a known pharmaceutical composition, widely sold under the name of Jumex, Deprenyl, Eldepryl or L-Deprenyl, which are widely effective, for example in the treatment of tuberculosis or immune regulation [A. Dow: The Deprenyl Story, Toronto: Stoddart (1990); Inhibitors of Monoamine Oxidase B, edited by I. Szelenyi, Birkhuser Verlag, Basel-Boston-Berlin 237-358 (1993)]. One of its important effects is its antidepressant and psychoestamulant and its effect that inhibits MAO, more intimately, its selective effect of inhibiting MAO-B. Several processes are known for their preparation, see for example Hungarian patent specifications Nos. 151,090, 154,655 and 187,775. L-Deprenyl was approved by the FDA in 1989 as an agent for the treatment of Parkinson's disease. The liposomal composition according to the present invention preferably contains from 0.1 to 40% by weight of selegiline (-) - [Na-dimethyl-N- (2-propynylphenyl-ethylamine] and / or a salt thereof, 2 to 40% by weight of lipids, preferably phospholipids, from 0 to 10% by weight of cholesterol, from 0 to 20% by weight of an alcohol, from 0 to 25% by weight of a glycol, from 0 to 3% by weight of an antioxidant, from 0 to 3% by weight of a preservative, from 0 to 2% by weight of an agent influencing the viscosity, from 0 to 50% by weight of cyclodextrin or a derivative of the cyclodextrin and from 30 to 90% by weight of water The liposome composition according to the present invention preferably contains from 0.1 to 20, more preferably from 0.1 to 10% by weight of selegiline and / or a salt thereof and by at least 10% by weight of this amount in a uni- and / or multilamellar lipid vesicle and the remaining amount necessary for 100% by weight in free state and / or as a saturated solution. The liposomal composition according to the invention contains as a lipid preferably a phospholipid, preferably phosphatidylcholine and / or lysophosphatidylcholine and / or phosphatidylserine and / or phosphatidyl-ethanolamine and / or phosphatidylinositol; as a preferred alcohol, ethanol or isopropanol; as a glycol, preferably a propylene glycol or polyethylene glycol; as an antioxidant preferably tocopherol or BHA (butylhydroxyanisole); as a preservative agent of preference germaben (International Specialty Product, Vienna, Austria); as an agent influencing the viscosity preferably a hydrocarbon or a cellulose derivative, preferably carbopol (Carbomer, Goodrich, Cleveland); and as a cyclodextrin and / or a cyclodextrin derivative preferably a, β or gamma-cyclodextrin, a water soluble cyclodextrin polymer, a methylated, hydroxypropylated, or succinylmethylated cyclodextrin derivative or any mixture thereof. The compositions according to the invention may contain the liposome composition, if desired, together with fillers, diluents or auxiliary agents generally used. They can be administered orally preferably, parenterally or transdermally. When preparing a transdermal formulation, the liposome composition can be applied on a carrier surface, preferably a thin sheet, a film or a plaster. The liposomal compositions can be prepared as described in US Pat. Nos. 4,485,054 and 4,761,288, the organic solvent is evaporated from an organic solvent mixture containing fat-soluble components, comprising at least one lipid and selegiline, then it is combined with an aqueous solution of the water-soluble components under stirring. As an organic solvent mixture preferably a mixture of chloroform and methanol is used. The liposomal compositions according to the invention can preferably be used for the treatment of Alzheimer's disease, Parkinson's disease, depression, shock, movement disease or myelitis. The liposomal composition * itself contains the active ingredient in a multiple phase, unilamellar and / or multilamellar vesicle, in a free state and in its saturated solution, i.e. it is a multiple phase liposomal drug delivery system. The liposomal system thus obtained is stable and can be freely diluted with water. Their rheological characteristics can be varied from the diluted liquid to the gelatinous state. From the pharmacological point of view, it is intended to prepare selegiline containing liposomal compositions, which are controlled release drug delivery systems and thus allow the administration of exact doses during transdermal treatment even at a dose regimen of "once a week". The mode of administration and dose depends, among others, on the disease to be treated (Alzheimer's disease, Parkinson's disease, depression, shock, movement disease or myelitis), in its severity, the patient's general condition , etc. In vivo pharmacological and pharmacokinetic examination of the liposomal compositions has been carried out in albino guinea pigs weighing 300-350 g (Charless-River line, SPF: specific pathogen-free quality) by local treatment, in groups comprising three animals randomly selected for the test animals. The second phase of the experiments has been carried out in pigs from 20 to 22 kg, treating three animals with a single dose of one of the formulas. The animals are kept separately in boxes filled with wool waste at an average temperature of 23 ° C, fed with the same feed and water. As the test compositions, the products according to Examples 1, 2, 3 and 6 were used. Deprenyl liposomal preparations were applied on the unshaven back (guinea pigs) or on the skin of the neck (pigs) a surface of 1.5 x 1.5 and 3 x 3 cm2, respectively, and after drying for a few minutes held by Tegaderm (produced by 3M, USA). As a control, the pigs are treated orally with selegiline tablets once a day.

During the evaluation the inhibition of MAO in the blood, brain, liver and intestines was measured, as well as the concentration of the active ingredient and its metabolites in the blood. The accumulation of the active ingredient and its metabolites in organs other than pigs (blood, brain, heart, liver, kidney, lung and spleen) were examined after treatment with selegiline-liposomes labeled with - * H-isotope by radioactive detection. Blood samples are taken to determine the serum concentration before treatment and then 6, 24, 48, 72, 96, 120 144 and 168 hours after administration. The amount of selegiline and its metabolites, in addition to the MAO-B activity of the platelets was measured. Tagederm bras are removed after 6 hours and the remaining alcoholic extract, the unabsorbed amount of selegiline was measured. After finishing the MAO activity experiments were also determined from the brain, liver and intestines isolated from the sacrificed animals. In determining the activity of the MAO-A and MAO-B enzymes, the selectivity of the inhibition of the enzyme was also examined. The combined concentration of Deprenyl and its metabolites was measured in the blood and in the brain, lungs, spleen, liver, heart, stomach, small intestine and large intestine and the kidney isolated from the slaughtered animals, as well as on the surface of the skin, where the liposomal treatment was carried out. Blood samples are taken at certain intervals from the guts of the eye of the guinea pigs and the major vein of the throat of pigs. At the end of the seventh day the guinea pigs are sacrificed and a blood sample is drawn immediately from the heart. In the case of blood samples of the pigs are taken before sacrificing them as described in the above. The removed organs and tissues are measured and homogenized with a volume of 4 times of physiological sodium chloride solution. Aliquots of 3 x 5 μl are pipetted into cuvettes containing 2 ml of Solune 320 and 0.5 ml of isopropanol. The samples are further processed as described above for the blood samples. The radioactivity of the organs was determined by liquid scintillation counting, the data obtained gives the total amount of the measurable Deprenyl and the metabolites. The amount of the drug that remains in the parafilm and the Tegaderm is also determined by the radioactive technique. The specific radioactivity is determined from the aliquot samples of the original liposomal preparations, which were used to calculate the value that relates to the organs and tissues. The activity of MAO in the brain was determined by the method of Wurtman and Axelrod [Biochem. Pharmacol. 12 1414-1419 (1963)] and the protein content of the homogenates was determined by the method of Lowry et al [J. Biol. Chem. 193, 265-275 (1951)]. The MAO activity of the platelets was examined by the method of Willberg and Oreland [Med. Biol. 54, 137-144 (1976)]. The concentration of selegiline and in addition to its metabolites that of amphetamine, jethamphetamine and demethyl-selegiline was determined by gas chromatography. The biological pharmacological results are clarified in the following figures. Figures 1 and 2 show the blood level data of the major metabolite of selegiline, ie methamphetamine, after applying once transdermally to the selegiline-liposomal composition. When the unilamellar liposomal composition of smaller vesicle distribution is mainly applied according to Example 6 absorption and metabolism is rapid, a dose-dependent blood level is measured for 24 hours (Figure 1). The multilamellar liposomes mainly fall under the range of larger particle size distribution (Examples 1 to 3) produces a delayed, slow blood level of the components having a low lamellar number, ensures a relatively high blood level for 72 hours and the multilamellar composition ensures a measurable blood level even after 168 hours (Figure 2). In the case of the same compositions, it can also be established that the composition that ensures rapid absorption caused an inhibition of stable MAO, while the slow absorption causes inhibition that regenerates relatively quickly in the blood (Figures 3 and 4) . Similar data were also measured in the brain (Figures 5 and 6). Similar blood levels were measured in the radioactive tests in guinea pigs (selegiline without change and their metabolites together) (Figure 7). In the case of liposomes of significant levels of low absorption, it could be measured in organs despite the decrease in blood level at 168 * hours, both in the brain which is important with respect to the effect, both playing an important role in the liver in the metabolism and in all the lymphoid organs (Figure 8 and Table 1). These organs can serve as a reservoir that releases the active ingredient later. The intestines contain a relatively small amount of the substance, thus supporting the inhibition data and the lack of inhibition of MAO-A (Figure 5). Figure 9 (Table 1) compares the levels measured in the organs with the amount of substances accumulated in the skin. It can also be seen that although significant, values above 10 ng / g can be measured in the organs, which are greater than the known postmortem human data (dotted line in Figure) there are still significant preservations in the skin, which It can ensure a supply of active ingredient for a long time. The different doses, 10 and 140 mg, cause a significantly greater difference in the blood (approximately 100 / times) than in the brain (only 2 to 3 times) (compare Figures 7 and 8). The exams support the following advantages of liposomal compositions containing selegiline: The active ingredient prevents the gastrointestinal tract, thus causing a significant decrease in the inhibition of the MAO-A enzyme there and which plays an important role in the metabolism of tyramine, because he does not find the active ingredient. By any other route of administration apart from oral (intravenous, intramuscular, eye drops, nasal spray, evaporator, etc), the active ingredient prevents the portal vein and thus the first step of metabolism. Thus, it causes higher levels of selegiline and decreased metabolite levels, decreases the inhibition of MAO-A. The "cheese" effect is less likely to occur. Higher doses can be applied without side effects and for example the antidepressant indication can be extended. Metabolites are stimulants and cause drowsiness, their lower level means the decrease of side effects. When administered transdermally, the skin stores the amount of the active ingredient necessary for continuous absorption, in a non-limiting manner of its function. The absorption of the liposome composition (depending on the formulation) takes approximately 1 to 20 minutes, after any restraint or coverage (patch, Tegaderm) is superfluous, can not be washed, does not prevent washing, its removal can not be feared yet in dementia. When changing the formulation (composition or particle size) an optimal delay (1 day, 1 week, several weeks) can be achieved. By changing the ratio of the encapsulate and the free active ingredient a stable ingredient: metabolite ratio can be achieved, which is optimal with respect to neuroprotective and neurorescue effects. The invention is clarified by the following non-limiting examples.

Example 1 to 6 Preparation of Deprenyl liposomal Compositions Example 1 g of phospholipon 90-G (unsaturated phospholipid) and 10 g of selegiline hydrochloride are dissolved in a round bottom flask in 20 ml of a 2: 1 mixture of chloroform and methanol at a temperature of 40 ° C. 100 g of small glass beads are added to the solution. The solvent is evaporated in vacuo in a Rotavapor and during this process the thin film is formed on the wall of the glass flask and the surface of the glass beads. 70 g of a 1: 3 mixture of ethanol and water heated to 40 ° C is added to it. The contents of the flask are well stirred and after further stirring at an average speed of 200 rotations / minute for 30 minutes at 35 ° C. The glass beads are removed by filtration through a Büchner funnel without filter paper. The filtrate is allowed to stand for one hour at room temperature to allow the liposome system to form. The formation of liposomes is confirmed by optical microscopic examination. The total weight of this liposomal product (CH-L-1) is 100 g. The liposomal structure is tested by microscopic examination and measurement of the particle size distribution.

Example 2 A liposomal selegiline composition is prepared as described in Example 1 with the difference that Phospholipon 90 -H was used as the phospholipid. 100 g of the product (CH-L-2) was also obtained. Its liposomal structure is tested by microscopic examination and measurement of the particle size distribution.

Example 3 The procedure is as described in Example 1 with the difference that the phospholipid in an amount of less than 1 g and the cholesterol in an amount of 1 g is dissolved in the organic solvent mixture. The product obtained was called (CH-L-3).

Example 4 The procedure is as described in Example 1 with the difference that 20 g of selegiline and 30 g of phospholipid were used. It is proved that 118 g of the product is a liposomal composition by its microscopic examination and the measurement of particle size distribution.

Example 5 The procedure is as described in Example 2 but 30 g of selegilna, 40 g of a phospholipid and 40 ml of a mixture of chloroform in methanol were used. 135 g of the product were obtained, which proved to be a liposomal composition by its microscopic examination and the measurement of the particle size distribution.

Example 6 The procedure is as described in Example 1, with the difference that 16 g of Fosfolipon 90-H, 4 g of cholesterol, 10 g of selegiline hydrochloride, 50 g of distilled water and 5 g of propylene glycol were used and stirring was carried out at a speed of 250 rotation / minute. The product obtained (CH-L-10) proved to be a liposomal composition by its microscopic examination and the measurement of the particle size distribution. Its average particle size is smaller than that of the products obtained according to Examples 1 to 5.

Table 1 The radioactivity content of different organs (in μgeq / g of tissue) of the guinea pigs one week after 140 mg of 3H deprenyl-formulated liposome (containing 14 μg of deprenyl, approximately 42 mg / kg). The first shaded data were measured after 24 hours, not included in the average value).

Figure 1: Average plasma levels of metamfeta ina "(± SD, n = 3) in the pig, after oral and transdermal treatment with (-) Deprenyl (CHL-10) Figure 2: Levels of plasma of average metamfetamine ( SD, n = 3) in the pig after transdermal treatment with a liposomal composition containing (-) Deprenyl Figure 3: Inhibition of platelet MAO-B after transdermal administration of a liposome composition (CHL- 10) and oral administration of traditional tablets Figure 4: Inhibition of MAO-B in platelets 168 hours after transdermal administration of a multilamellar liposomal composition Figure 5: Inhibition of MAO-B in the brain, liver and small intestine 168 hours after the transdermal administration of the CHL-2 liposomal composition. Figure 6: Inhibition of MAO-B in the brain and liver 168 hours after the transdermal administration of the liposomal compositions. Figure 7: Serum concentrations calculated from radioactivity (unchanged substances and metabolites) after transdermal administration of different doses of the CHL-2 composition.

Figure 8: Concentration of a radioactive substance (selegiline + metabolites) 168 hours after the transdermal administration of the CHL-2 composition. Figure 9: The radioactivity of the organs different from the guinea pigs and from that of the application site 168 hours after the transdermal administration of the composition CHL-2.

Claims (11)

1. A liposomal composition, characterized in that it contains as the active ingredient (-) - N-a-dimethyl-N- (2-propynylphenylethylamine) (selegiline) and / or a salt thereof.

2. The liposomal composition, characterized in that it contains 0.1-40% by weight of selegiline (-) - Na-dimethyl-N- (2-propynylphenylethylamine)] and / or a salt thereof, from 2 to 40% by weight of lipids, preferably phospholipids, from 0 to 10% by weight of cholesterol, from 0 to 20% by weight of an alcohol, from 0 to 25% by weight of a glycol, from 0 to 3% by weight of an antioxidant, from 0 to 3% by weight of a preservative agent, from 0 to 2% by weight of an agent influencing viscosity, from 0 to 50% by weight of cyclodextrin or a derivative of cyclodextrin and from 30 to 90% by weight of water .

3. The composition according to claim 1 or 2, characterized in that it contains 0. 1 to 20% by weight preferably 0.1 to 10% by weight of selegiline and / or a salt thereof.

4. The composition according to claim 1 or 2, characterized in that it contains at least 10% by weight of the selegiline and / or a salt thereof in a single or multilane elad and an additional amount of selegiline necessary for 100% by weight in solution and in solid form.

5. The composition according to claim 2, characterized in that it contains as a lipid phospholipid, preferably phosphatidylcholine and / or lysophosphatidyl choline and / or phosphatidylserine and / or phosphatidyl ethanolamine and / or phosphatidylinositol.

6. The composition according to claim 2, characterized in that it contains as an alcohol preferably ethanol or isopropanol; as a glycol, preferably a propylene glycol or polyethylene glycol; as an antioxidant preferably tocopherol or butylated hydroxyanisole; as a preservative agent, preferably germaben; and as the agent influencing the viscosity, preferably carbapol.

7. The composition according to claim 2, characterized in that it contains as a cyclodextrin and / or a cyclodextrin derivative preferably a, ß or gamma-cyclodextrin, a water-soluble cyclodextrin polymer, a hydroxypropylated methylated cyclodextrin derivative, or succinylmethylated or any mixture thereof.

8. The pharmaceutical composition characterized in that they contain the liposomal composition according to claims 1 or 2 and, if desired, customary fillers and diluents and other auxiliaries, preferably orally, parenterally or transdermally.

9. The process for the preparation of liposomal compositions containing selegiline and / or a salt thereof, characterized in that by evaporating the organic solvent from an organic solvent mixture containing liposoluble components, having at least one lipid between them and selegiline, then they combine them with an aqueous solution of the water-soluble components according to claim 2, under stirring.

10. The process according to claim 9, characterized in that a mixture of chloroform and methanol is used as the organic solvent.

11. The composition according to any of claims 1, 2 and 8, characterized in that it is used for the treatment of Alzheimer's disease, Parkinson's disease, depression, attack, movement sickness or myelitis.