NL8000139A - PHARMACEUTICAL PREPARATIONS COMPRISING LIPOSOMES AND THEIR PREPARATION. - Google Patents

Description

V, VO 8885

Title: Pharmaceutical preparations "consisting of liposcms and their" preparation.

The invention relates to pharmaceutical preparations (lyophilic liposcms), their preparation and purification. More particularly, the invention relates to a novel method for purifying liposcopic suspensions obtained according to known methods and for stabilizing them by lyophilization.

Liposcms are pharmaceutical preparations in which the drug is dispersed or otherwise present in aqueous concentric layers in addition to lipid layers (hydrophobic); the drug may be present in the water layer as well as in the lipid layer (inside or outside) or otherwise in the non-homogeneous system, commonly known as lino sane suspension.

The hydrophobic layer generally consists, / not exclusively, of phospholipids such as lecithin and sphynganyelin, steroids such as cholesterol, more or less ionic surfactants such as dicetyl phosphate, stearylamine or phosphatidic acid, and / or other hydrophobic iterials; the diameters of the liposcms are usually between 15 nm and 5 p ··

Preparations of this type are usually prepared in a manner known per se, mainly by two steps: The preparation of liposcms and purification of non-enclosed drug.

1) Preparation of liposcms; the lipid or lipophilic components are dissolved in an appropriate solvent, which is then usually evaporated to dryness under vacuum. The water layer with the drug is added to the vial with the residue therein as a thin layer, after which the whole is shaken mechanically or ultrasonically for 10 seconds to several hours. The non-homogeneous layer thus obtained is commonly referred to as a liposcopic suspension, and has yet to be purified from non-occluded drug.

2) Separation of the liposcms from the unclosed drug; this procedure is usually carried out by eluting over a 90% column graph using resins with exclusive molecular sieve function, such as Sepharose 2B, B or 6B or similar.

Liposams are first recovered while the free drug remains on the resin.

Another method followed is ultracentrifugation at 100,000 g and then washing, always under ultracentrifugation with a buffered solution. A further method consists of dialysis.

The present invention relates to a novel purification process of the non-shark layer known as liposcopic suspension containing both liquid and solid polymers, usually of a synthetic or organic nature with chemical functions, which can be used as ion exchangers, e.g. of styrene, divinylbenzene, acrylic acid or methacrylic acid, commonly known as ion exchange resins.

The exact amount of one or more of these resins is placed directly into the flask with the liposcme suspension and shaken for 10 to 60 minutes. After filtering through a sintered glass filter capable of holding back the ion exchanger on which the uncaptured drug has been adsorbed, a pure liposoan suspension is obtained, which is then lyophilized.

This purification procedure of the liposomic suspensions with ion-exchange resins has the great advantage that highly concentrated liposomic suspensions are retained (up to 5 mg / cm doxorubicin HCl) which cannot be retained by chromatography on a molecular sieve column (maximum 0.3 mg / cm ). The liposam suspension thus obtained is particularly stable and does not tend to settle, and is different from suspensions obtained by ultracentrifugation.

The same result is achieved by using instead of ion-exchange resins polymers and copolymers without specific chemical functions but which usually, but not exclusively, react according to Van der Waals forces; these are commonly known as adsorbent resins. These resins can be used in the purification of liposomal suspensions due to the difference in polarity between the hydrophobic substances that form the liposomes, the actual drugs having their more or less hydrophilic character.

Final chemical stabilization is achieved by lyophilizing the liposome suspension.

5 Example I:

The following amounts of lipids were dissolved in chloroform in a saponification flask and evaporated to dryness under vacuum: 1.5 g egg lecithin, Q, b g cholesterol and 0.2 g dicetyl phosphate.

A solution of doxorubicin hydrochloride (at a concentration of 10 mg / cm) was then poured into this flask in a phosphate buffer of 0.007 N and the suspension was shaken ultrasonically for 1 minute. After standing at room temperature under nitrogen for 30 min, 2 g of a resin, which had previously been brought into sodium form and obtained by a polymerization of methyl methacrylate, was cross-linked appropriately with a specific chemical such as divinylbenzene, with a carbaxyl functionality and a macroreticular structure allowing its use also in hydrophobic solutions, sold under the trade name IRC-50, and manufactured by Rohm and Haas,. then added (the weight 20 refers to the dry weight and is equivalent to 5 cm resin).

The flask was then shaken well for 30 min, after which the slurry was filtered through a glass filter G1. Liposams with a size ranging from 0.5 to 2 µ-h containing about 60% of the original amount of daxorubicin were thus obtained. and stabilized by lyophilization.

Example II;

Example I was repeated with the same amounts of lipids and doxorubicin, but the ultrasonic shaking time was increased to 10 min cm to obtain liposcms smaller than 1 / U.

Since the liposcms were not completely hcmogeneous in size, a non-granular resin was used which also functioned as a screen.

O

10 cm of a D0WEX 50-X U 100-200 mesh resin, previously activated to the sodium form, was added thereto.

After filtration, a. suspension obtained with liposcms with a.--'800 0 1 39 -u size of 0.2-0.8 µ containing approximately 75% of all starting doxorubicin:. contain. These liposomes were stabilized by lyophilization. Example III:

A solution of 5-fluorouracil at a concentration of 105 mg / cm in a phosphate buffer of 0.007 IT with pH 8 was poured into an saponification flask containing a lipid phase prepared as described above. The suspension was then treated as shown in Example I using 10 cm of the filter resin Amberlite IRA-1200 (Cl) previously activated as hydrochloride.

The obtained liposcms were then stabilized by lyophilization.

Example IV:

The liposomes of 5-fluorouracil were obtained as described above using the following amounts of lipids: 1.5 g egg lecithin, 0.1 g cholesterol, 0.2 g stearylamine.

. . 3

As a purification system, 10 cm of the resin DOWEX 1 (50-100 mesh) pre-activated was used. The liposcms thus obtained were stabilized by lyophilization.

Example V: Example II was repeated with the difference that the DOWEX 50 W-X resin was replaced with 15 g of the adsorbent resin of Rohm and Haas XAD 7 and the shaking time was extended to 50 minutes.

After filtering through a sintered glass G1, a suspension of liposomes containing about 50% of the starting amount of daxoru-bicin was obtained. These liposomes were stabilized by lyophilization.

Example VI:

1.5 g of soy lecithin, 0.1 g of cholesterol and 0.3 g of dicetyl phosphate were dissolved in CHgClg and to this solution a further solution of aminosidine sulfate in 0.02 M phosphate buffer with pH

O

6.5 at a concentration of 3 mg / cnr.

The two phases were emulsified and nitrogen was injected into the shaken emulsion at room temperature until all methylene chloride had evaporated.

The suspension was kept at room temperature for U hours for 80 0 0 1 39 -5-, then poured into the flask an amount of resin known as IRC-50 from Rohm and Haas, equivalent to 5 g dry resin.

After a further 1 hour of shaking, the liposoaceous suspension was filtered through a sintered glass filter to remove the resin, the latter retaining unclosed drug. This liposoam suspension was then stabilized.

8000139

Claims (10)

Method for the purification of non-homogeneous systems, known as liposomal suspensions, of inclusions, characterized in that these suspensions contain liquid or solid polymers, usually of a synthetic or organic nature, usually but not necessarily cross-linked with an appropriate chemical agent , and are treated with chemical functionality used as ion exchangers, such as of styrene, divinylbenzene, acrylic acid, methacrylic acid and others commonly known as ion-exchanging resins. Method according to claim 1, characterized in that the ge. said cation-type ion exchange resins are of low, medium or high ionic strength, usually determined, but not necessarily limited to carboxyl, phosphon and sulfone functions of various matrices, e.g. of phenol formaldehyde, styrene-divinylbenzene, acrylates, methacrylates, various hydrocarbons and various condensation resins in a liquid or solid form. 3. Process according to claim 1, characterized in that the * ion exchangers are of the strong anion type, first and second type, of weak or medium strength, usually described as, but not necessarily limited to various salt forms of quaternary ammonium, primary, secondary or tertiary amines, phosphine functions or other functions with different matrices, e.g. of the type phenol oldehyde, styrene-divinylbenzene, acrylates, methacrylates, various hydrocarbons and various condensation resins in a liquid or solid form. k. Process according to claim 1, characterized in that the resins are used in a salt form or an activated form. 5. Method for the total or partial extraction of the dissolved drugs, after destruction of the lipid structure, from liposomes according to the procedure described in the above claims. 800 0 1 39 «k, - '' ........... * s \ -7- 6. A method for purifying non-hogenous systems known as liposome suspensions of unclosed drug, characterized in that these suspensions are treated with polymers and / or copolymers without specific chemical function, which usually but not exclusively react according to Van der Waals forces, and commonly known as adsorbents. Method according to claim 6, characterized in that these adsorbents are of aliphatic, aromatic or inorganic origin. Method for the total or partial extraction of the drug, whether or not enclosed by liposcms, according to the procedure set out in claims 6 and 7 9. Process for the preparation of liposomic suspensions, characterized in that these suspensions are obtained by blowing inert gas (eg nitrogen or others) into a two-phase emulsion type system, or by a simple co-presence of immiscible solvents. 10. A method of stabilizing liposomal suspensions, characterized in that the purified liposomic suspensions are subsequently lyophilized. 11. Pharmaceutical preparation consisting of lyophilic liposcms of doxorubicin hydrochloride. 12. Pharmaceutical preparation consisting of lyophide liposomes of aminosidine sulfate. 25 13 · Pharmaceutical preparation consisting of lyophilic liposomes 5-fluorouracil. 8000? "" • · ->