Note: Descriptions are shown in the official language in which they were submitted.
2 ~ 3
The invention relates to a liposomal dihydropyridine
formulation which can be administered parenterally and to
a process for its production.
Dihydropyridines are, in general, poorly water-soluble.
For parenteral administration, therefore, use with water-
miscible, tolerable organic solvents or use of particular
injection or infusion formulations is relied upon. These
include parenteral nanoemulsions, nanosuspensions
(hydrosols) and liposomes.
Organic solvent systems as carriers for dihydropyridines
have a number of disadvantages: on account of too low a
tolerability in the peripheral vessels they must either
be administered via a central catheter or diluted with a
simultaneously administered co-infusion. Complications
due to supersaturation can occur here owing to the
solubility ratios. Furthermore, depending on the type oi
dihydropyridine, considerable amounts of organic solvents
and possibly other auxiliaries had to be added. In
particular in th~ case of relatively long-term
intravenous therapies, this would lead to an amount of
undesired solvent which is no longer tolerable.
It is known that sparingly soluble active compounds can
be administered parenterally in a liposomal admini-
stration form (Liposome Technology, Vol I~III, ed.
Gregoriadis). It is also known that liposome dispersions
Le A 28 986 - l -
2 ~ 1 3
can be prepared by various techniques. ~xamples which may
be mentioned are: high-pressure homogenisation
(EP-A-0,101,007), ultrasonication, dialysis, extrusion,
microfluidisation and dilution processes~ Under the known
preparation techniques, processes which do not use
organic solvents are also described which, however, only
function under certain conditions and/or require a high
industrial outlay.
In particular for nimodipine, it is described, for
example, in DE 3,515,335 how this particularly sparingly
soluble dihydropyridine can be formulated as liposomes
using organic solvents.
It has surprisingly been found that with sparingly
soluble active compounds and even with the e~tremely low
solubility of nimodipine the use of organic solvents in
the preparation of liposomes can be avoided. The
liposomes according to the invention are prepared accord-
ing to the following process: cryoprotector, pH
stabiliser, antioxidant, phospholipid and active compound
are prehomogenised without the use of organic solvents in
a water receiver into which nitrogen is passed. The
predispersion is then subjected to high pressure
homogenisation at elevated temperature until the desired
liposome size is obtained.
It was not foreseeable that even with a dihydropyridine
which is as sparingly soluble as nimodipine, stable
liposomes can be formed without the active compound
Le A 28 936 - 2 -
recrystallising.
It is known that additions of polyalcohols ~O~fenlegungs-
schrift DE 3,515 r 335 A1) can prevent the recrystal-
lisation of active compounds such as dihydropyridines in
liposome dispersions.
It has surprisingly been found that active compounds, in
particular dihydropyridines, can be formulated keeping to
active compound:phospholipid ratios of 1:20 to 1:60
without the presence of organic solvents and without the
addition of polyalcohols to inhibit recrystallisation,
which is disadvantageous from the medical point of view.
At the said active compound:phospholipid ratios, the
active compound neither recrystallises during preparation
in the dispersion nor during storage in the lyophilisate.
Using the active compound:phospholipid ratio according to
the invention, a storage-stable liposome formulation is
thus `also achieved without the addition of polyalcohols.
In particular, it was not foreseeable that the active
compound does not recrystallise even if the liposome
lyophilisates are reconstituted after 2 years' storage at
6C.
The invention relates to stable pharmaceutical prepara-
tions which can be administered parenterally and are free
o~ organic solvents, based on liposomes with a phospho-
lipid membrane having an active compound:phospholipidratio between 1:20 and 1:100 and an average liposome
diameter between 40 and 200 nm which after reconstitution
Le A 28 936 - 3 -
have a pH between 4.5 and 6.5.
Pharmaceutical preparations are particularly preferred
whose liposome membrane contains between 20 and 60 parts
by weight of phospholipid and contains 1 part of active
compound, the preferred active compounds employed being
dihydropyridines of the formula (I)
~ X (I)
Y~Z
H3C 1I R3
in which
Ydenotes the group NO2, CN or -COORl, where
10R1 represents Cl-C4-alkyl, optionally substituted
by C1-C3~alkoxy/
Z represents the group CO-NH-cycloalkyl (3-7 C atoms)
or -COOR2, where
R2 denotes C1-Cl~-alkyl, optionally substituted by
15C1-C3-alkoxy, trifluoromethyl, N-methyl-N-
benzylamino or benzyl,
R3 represents Cl-C4-alkyl, cyano or hydroxyalkyl (1-4 C
atoms) and
X represents 1 or 2 identical or different sub-
20stituents from the group nitro, halogen, Cl-C4-alkyl,
Le A 28 936 - 4 -
2~
Cl-C4-alkoxy, benzoyl and a 2,3-ring member consist-
ing of =N-O-N=, 2-cycloalkylmethylthio, 2-(4-methyl-
benzyloxy) or trifluoromethyl.
The invention also relates to a process for the
production of the preparations according to the
invention, characterised in that 1 part by weight of the
active compound and 20-60 parts by weight of phospholipid
- are predispersed for an adequate period together with an
antioxidant, a cryoprotector and a pH stabiliser at
temperatures between about 10 and B0C, preferably 20 to
80C, and a pH between 5 and 7.5, preferably 6 and 7, in
water into which nitrogen has been passed using a stirrer
and the liposomes are then homogenised to an average size
between 40 and 200 nm in an abrasion-free high-pressure
jet homogeniser at temperatures between 20C and 80C and
a pressure between 400 and 1000 bar and the dispersion
thus obtained is optionally then poured into suitable
vials and rapidly frozen at temperatures between -50C
and -70C. It is particularly advantageous to employ
glucose as the cryoprotector and to select a ratio
between cryoprotector and phospholipid of between 1:1 and
4:1. If appropriate, the liposomes prepared in this way
can additionally be sterilised after homogenisation, for
example by filtration through a 0.2 ~m filter.
To achieve sho:rt drying times, it may be advantageous to
employ the shell-freeze or the spin-freeze technique.
Suitable reconstitution media which may be mentioned are
Le A 2~ 936 - 5 -
isotonic glucose solution (5% strengtn) or water for
injections. To improve the flocculation behaviour after
reconstitution it is recommended where appropriate that
the liposome membrane contains up to 10~ by weight of a
charge inducer. Those which may be mentioned as preferred
are DMPG-Na (dimyristoyl-phosphatidylglycerol sodium
salt), DPPG-Na (dipalmitoyl-phosphatidylglycerol sodium
salt) or DSPG-Na (distearoyl-phosphatidylglycerol sodium
salt).
The pharmaceutical preparations according to the inven-
tion are distinguished by excellent temperature and
storage stability. They can be stored, for example, at
temperatures below 30C for at least two years without
loss of quality occurring after reconstitution. The
reconstituted dispersions are stable for up to 24 h and
show no flocculation or aggregation phenomena of the
phospholipid.
The liposomes according to the invention are generally
suitable for lipophilic active compounds, in particular
for dihydropyridines such as nifedipine, nisoldipine,
nimodipine, nitrendipine, felodipine or amlodipine. The
technique is also particularly suitable for very
sparingly soluble dihydropyridines such as nimodipine.
The preparation of the liposomes according to the
invention comprises several sub-steps. First, after
initial weighing of the substances used, a prehomogenate
is prepared using conventional rotor-stator stirrers. The
Le A 28 936 - 6 -
predispersions here contain the phospholipid and
nimodipine in the said ratio between 20~1 and 40:1. The
predispersions furthermore contain known cryoprotectors
such as sucrose, glucose or fructose, antioxidants such
as ascorbic acid and pH stabilisers such as arginine.
Suitable phospholipids for the construction of the
liposomes for parenteral administration are all phospho-
lipids having the customary purity for this purpose. The
phospholipids can in this case be either unsaturated or,
to obtain other pharmacokinetic profiles, saturated. As
natural phospholipids, soya or egg phospnolipids are just
as suitable as synthetic phospholipids. To obtain par-
ticularly translucent liposome dispersions and to improve
the flocculation behaviour after reconstitution, the
addition of up to 10% of charge inducers such as Na-DMPG,
Na-DPPG or Na-DSPG is advantageous.
The prehomogenate is filtered through a 5 ~m filter
before high-pressure homogenisation and then homogenised
to the desired liposome size between 40 and 50 nm in a
customary high-pressure jet homogeniser at temperatures
between 20 and 80C, preferably between 40 and 80C,
under an inert gas, such as nitrogen protection.
For the purpose of microorganism reduction, the
homogenised liposome dispersion is filtered through a
customary 0.2 ~m filter and then lyophilised. For reasons
of optimum cryoprotector action, the phospholipid:cryo-
protector ratio should be between 1:1 and 1:4, preferably
Le A 28 936 - 7 -
between 1:1.8 and 1:4.
For the purpose of lyophilisation, the di~persion is
poured into vials or infusion bottles, the bottles are
placed in the freeze-dryer and the product is frozen
therein with the aid of the charging spaces pre-cooled
to -50C. To achieve particularly rapid drying times, it
is advantageous to freeze the bottles or vials while
rotating by the spin- or shell-free~ing principle.
The liposomes prepared according to the invention are of
between about 40 and 50 nm size even after homogenisation
and have a narrow distribution around this average value.
The liposome lyophilisates prepared according to the
invention can be reconstituted before administration
using the customary parenteral solutions. Preferably, the
reconstitution is carried out using 5% strength glucose
sol~tion or water for injections. The lyophilisates can
be reconstituted within one minute with repeated shaking.
The reconstituted liposome dispersions according to the
invention are preferably administered at the desired rate
using customary infusion pump systems such as Perfusor~,
Imed~ or Ivac~. For safety reasons, the reconstituted
liposome dispersion is filtered in-line through a com-
mercially available filter before administration.
Le A 2B 936 - 8 -
ExemPlarv embodiments
Example 1
Nitrogen is passed into aoo g of water for injection for
10 min. l g of ascorbic acid, 150 g of glucose and 1 g of
L-arginine are then dissolved in this amount of water.
1.667 g of nimodipine and 83.35 g o~ highly pure egg
phospholipid (for example Lipoid E PC) are dispersed in
this medium and it is made up to 1055.5 g with water into
which nitrogen has been passed (active compound: phospho-
lipid ratio = 1:50).
This dispersion is prehomogenised for 30 min at 75C and
under nitrogen protection using a high-speed stirrer (for
example Ultra-Turrax). The predispersion is filtered
through a 5 ~m filter.
The predispersion is then homogenised for one hour at
75C; 800 bar and under nitrogen protection in an
abrasion-free high-pressure jet homogeniser. Sterile
filtration (0.2 ~m) is carried out after cooling to room
temperature. The content is 100% of the theoretical
content.
The dispersion is poured to 15.3 ml into 250 or 50 ml
bottles and frozen to give a thin product cake with the
aid of spin- or shell-freezing at -65C (dry ice-ethanol
cooling mixture).
The frozen product is transferred to the charging spaces
Le A 28 936 - 9 -
~ ~ $ ~ 3
of a lyophiliser which is precooled to -50C and dried at
-30C and 0.05 mbar for 14 h. Afterdrying at 30C and
O.001 mbar lasts 7 h.
The lyophilisate is reconstituted with isotonic glucose
solution to 250 ml or with water for injection to 50 ml.
The content is 100% of the theoretical content.
Liposome properties:
before after recon-
lyophilisation stitution
_
Average s~ze 45 nm 50 nm
Dispersity index K2 0.350 0.500
Turbidity 170 units 40 units
Osmolarity 1.2 Osmol 360 mOsmol
pH 6.5 4.5
Particle load (HIAC) <10 particles
after in-line filtration (> 2 ~m/ml)
Example 2
Nitrogen is passed into 800 g of water for injection for
10 min. 1 g of ascorbic acid, 100 g of glucose and 1 g of
L-arginine are then dissolved in this amount of water.
1.667 g of nimodipine and 50.00 g of highly pure soya
phospholipid (for example Phospholipon 90) are dispersed
in this medium and it is made up to 1055.5 g with water
into which nitrogen has been passed (active compound:
Le A 28 936 - 10 -
phospholipid ratio = 1:30).
This dispersion is prehomogenised for 30 minutes at 75C
and under nitrogen protection using a high-speed stirrer
(for example Ultra-Turrax). The predispersion is filtered
through a 5 ~m filter.
The predispersion is then homogenised for one hour at
75C, 800 bar and under nitrogen protection in an
abrasion-free high-pressure jet homogeniser. Sterile
filtration (0.2 ~m) is carried out after cooling to room
temperature. ~he content is 100% of the theoretical
content.
In contrast to Example 1, after pouring into appropriate
bottles the dispersion is frozen at -65C without use of
spin- or shell-freezing.
The product is dried at +30C and 0.05 mbar for 7 h (main
drying) and afterdried at ~30C and 0.001 mbar. The
liposome properties from Example 1 are retained.
Example 3
Nitrogen is passed into 800 g of water for injection
purposes for 10 min. 1 g of ascorbic acid, 100 g of
glucose and 1 g of L-arginine are then dissolved in this
amount of water. 1.667 g of nimodipine, A5 g of highly
pure sakurated phospholipid (for example Epikuron 200 SH)
and 5 g of DMPG-Na are dispersed in this medium and it is
Le A 28 936
2~ g ~ ~
made up to lOS5.5 g with water into which nitrogen has
been passed (active compound: phospholipid ratio = 1:30).
~his dispersion i5 prehomogenised for 30 minutes at 75C
and under nitrogen protection using a high-speed stirrer
(for example Ultra-Turrax). The predispersion is filtered
through a 5 ~m filter.
The predispersion is then homogenised for one hour at
75C, 800 bar and under nitrogen protection in an
abrasion-free high-pressure jet homogeniser. Sterile
filtration (0.2 ~Lm) is carried out after cooling to room
temperature. The content is 100% of the theoretical
content.
The dispersion is poured to 15.3 ml into 250 ml or 50 ml
bottles and frozen to give a thin product cake with the
aid of spin- or shell-freezing at -65C (dry ice-ethanol
cooli~g mixture).
The frozen product is transferred to the charging spaces
of a lyophiliser which is precooled to -50~C and dried at
-30C and 0.05 mbar for 14 h. Afterdrying lasts for 7 h
at 30~C and 0.001 mbar.
The lyophilisate is reconstituted with isotonic glucose
solution to 250 ml or with water for injections to 50 ml.
The content is 100~ of the theoretical content.
Le A 28 936 - 12 -
Liposome properties:
before after lyo-
lyophilisation philisation
Average size 39 nm 58 nm
Dispersity index K2 0.530 0.630
Turbidity 130 units 40 units
Osmolarity 700 mOsmol 355 mOsmol
pH 6.5 6.1
Example 4
Nitrogen is passed into 800 g of water for injections for
10 min. 1 g of ascorbic acid, 50 g of glucose and 1 g of
L-arginine are then dissolved in this amount of water.
1.667 g of nimodipine and 25 g of egg phospholipid (for
example Lipoid E 80) are dispersed in this medium and it
is màde up to 1055.5 g with water into which nitrogen has
been passed (active compound: phospholipid ratio 1:15).
This dispersion is prehomogenised for 30 min at 75C and
under nitrogen protection using a high-speed stirrer (for
example Vltra-Turrax). The predispersion is filtered
through a 5 ~m filter.
The predispersion is then homogenised for one hour at
75C, 800 bar and under nitrogen protection in an
abrasion-free high-pressure jet homogeniser. Sterile
filtration (0.2 ~m~ is carried out after cooling to room
Le A 28 936 - 13 ~
2~3~3
temperature. The active compound content ~t this active
compound: phospholipid ratio (1:15) is 72~ of theory.
The dispersion is poured to 15.3 ml into 250 ml or 50 ml
bottles and frozen to give a thin product cake with the
aid of spin- or shell-freezing at -65C (dry ice-ethanol
cooling mixture)~
The froæen product is ~ransferred to the charging spaces
of a lyophiliser which is precooled to -50C and dried at
-30C and 0.05 mbar for 14 h. Afterdrying at 30~C and
0.001 mbar lasts 7 h.
The lyophilisate is reconstituted with isotonic glucose
solution to 250 ml or with water for injection to 50 ml.
The content of the reconstituted dispersion is 77% of the
theoretical content.
ExamPle 5
Nitrogen is passed into 800 g of water for injection for
10 min. 1 g of ascorbic acid, 23.34 g of glucose and l g
of L-arginine are then dissolved in this amount of water.
1.667 g of nimodipine and 11.68 g of egg phospholipid
(for example Lipoid E 80) are dispersed in this medium
and it is made up to 1055.5 g with water into which
nitrogen has been passed (active compound: phospholipid
ratio 1:7).
This dispersion i5 prehomogenised for 30 min at 75C and
Le A 28 936 - 14 -
under nitrogen protection using a high-speed stirrer (for
example Ultra-Turrax). The predispersion is filtered
through a 5 ~m filter. Active compound crystals can be
detected on the filter.
The predispersion is then homogenised for one hour at
75C, 800 bar and under nitrogen protection in an
abrasion-free high-pressure jet homogeniser. Sterile
filtration (0.2 ~m) is carried out after cooling to room
temperature. The content of the liposome dispersion is
18% of the theoretical content.
Le A 28 936 - 15 -
