Note: Descriptions are shown in the official language in which they were submitted.
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PHARMACEUTICAL COMPOSITIONS COMPRISING CEFQUINOME
The present invention is concerned with a pharmaceutical composition for
intrauterine administration of cefquinome and its use for the treatment of
metritis in mammalian animals.
Acute postpartum metritis ranks among the top health problems of fresh
cows in dairy farming. At least one-fourth of all fresh cows experience acute
metritis, an inflammation of the entire uterine wall caused by bacterial
infection. Acute puerperal metritis occurs within two weeks postpartum. It
results from contamination of the reproductive tract at parturition and often,
but not invariably, follows complicated parturition.
The causative organisms in cattle are most frequently Escherichia coli,
Arcanobacterium (Actinomyces) pyogenes in association with gram-negative
anaerobic bacteria such as Fusobacterium necrophorum and Prevotella spp.
Metritis can cause reduced fertility and can lead to higher culling rates of
female animals, especially cows. The severity of uterine inflammation varies
from an acute septic metritis involving the entire thickness of the uterine
wall
to a mild superficial endometritis. (Bretzlaff K, "Rationale for Treatment of
Endometritis in the Dairy Cow", Veterinary Clinics of North America: Food
Animal Practice - Vol 3, No. 3, 593-607, 1987).
For decades, metritis in cows has been treated with the intrauterine infusion
of antimicrobials. However, many preparations routinely administered into
the bovine uterus are detrimental to uterine tissue. Very often, they do not
restore fertility, they are not active against all major pathogens, and they
do
not persist for enough time to treat the infection. Therefore specific
formulations for intra-uterine administration are required.
Although antibiotic compositions for intrauterine application for the
treatment
of metritis have been described in prior art, such a composition for
cefquinome has not yet been allied in practice in the veterinary market.
Cefquinome is the first fourth-generation cephalosporin developed for use in
veterinary medicine. It is a semi-synthetic aminothiazolyl cephalosporin
resembling cefotaxime, but with a bicyclic pyridinium group at the C-3
position.
Cefquinome proved highly effective against the most commonly isolated
metritis pathogens (Sheldon, I et al "Minimum inhibitory concentrations of
some antimicrobial drugs against bacteria causing uterine infections in
cattle"
The Veterinary Record, 155, 383-387, 2004).
Injectable formulations of cefquinome and intramammary preparations of
cefquinome (Cobactan LC and Cobactan DC) for the use during the
lactation and dry period of cows are available on the veterinary market (sold
under the trademark Cobactan by Intervet International b.v., Boxmeer, The
Netherlands).
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International patent application WO 2004/037265 discloses a prolonged release
injectable formulation of cefquinome comprising an oil and aluminium
distearate.
International patent application WO 2003/063877 discloses a composition for
intramammary administration for the treatment and prevention of mammary
disorders during the dry period, comprising cefquinome in an oil/colloidal
silicon
dioxide base.
These formulations are however not optimal for intrauterine administration.
Thus, a need exists for a pharmaceutical composition for cefquinome that
overcomes one or more of the limitations of the prior art and is suited for
intrauterine administration. Such an advantageous pharmaceutical composition
needs to be active against the major pathogens present, non-irritant to the
uterine
tissue, and should have an activity persisting for enough time to treat the
infection
successfully.
The present invention provides a composition for cefquinome with such
advantageous properties that causes effective levels of the antibiotic in the
uterus
of the mammal that are suitable for the control of important pathogens over a
sufficient time.
The present invention provides a topical pharmaceutical composition for
intrauterine administration, characterised in that it comprises cefquinome in
a
base, said base comprising at least one medium chain triglyceride, at least
one
thickener and at least one macrogol cetostearyl ether.
According to one aspect of the present invention, there is provided an
intrauterine
liquid composition comprising cefquinome in a base, said base comprising at
least
one medium chain triglyceride, a thickener and at least one macrogol
cetostearyl
ether.
According to another aspect of the present invention, there is provided use of
the
intrauterine liquid composition described herein for treatment of metritis in
a
mammal.
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The term "cefquinome" when used herein includes pharmaceutically acceptable
salts and esters thereof.
A typical pharmaceutical composition according to the invention comprises 1
to 20% by weight of cefquinome. Preferably the pharmaceutical composition
comprises 2 to 10% by weight of cefquinome, especially 2.5 to 6.0%, most
preferred 3.0 to 4.2% by weight of cefquinome.
By "by weight" in this patent application it is meant a percentage by weight
of the
total composition.
In general all pharmaceutically acceptable cephalosporin salts can be
incorporated in the current pharmaceutical composition. Various crystalline
cephalosporin salts have been disclosed e.g. in European Patent No. EP 280157
and European Patent No. EP 711774.
In a preferred embodiment the current invention provides a pharmaceutical
composition characterised in that the cefquinome is cefquinome sulphate.
The base is selected so as to be non-irritant to the uterine tissue,
veterinary
acceptable, compatible with the antibacterial agent, and of a viscosity to
permit
administration, using a syringe at ambient temperature, including low winter
temperatures.
The base comprises a pharmaceutical acceptable low viscosity oily medium, such
as medium chain triglyceride or a mixture of medium chain triglycerides.
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Medium chain triglycerides (MCT oil) have fatty acid chains of 6 - 12 carbon
atoms and for the medically refined grades of MCT oil each chain has 8 - 10
carbon atoms. The MCT oil may comprise either triglycerides of the C8-C10
fatty acids, or propylene glycoldiesters of these fatty acids or a mixture of
both triglycerides and propylene glycol diesters. Preferably these C8 -C10
fatty acids are fully saturated, such as n-caprylic and n-capric acids. These
are conveniently prepared by the commercial fractionating of naturally
occurring vegetable (e.g. coconut) oil to give mainly C8-10 fatty acids
followed by esterification of these acids with a chosen alcohol.
Fractionated vegetable oil having the desired composition is commercially
available. Proprietary examples of such oils are Miglyol 812 as
capric/caprylic triglycerides and Miglyol 840 as propylene glycol
dicaprylate/caprate.
Equivalents of these oils are for example: Aldo MCT KFG, Aldo TO,
Calgene CC-33, Calgene CC-33-F, Calgene CC-33-L, Calgene CC-33-S,
Captex 300, Captex 355, Crodamol GTCC, Estasan GT 8-40 3578,
Estasan GT 8-60 3575, Estasan GT 8-60 3580, Estasan GT 8-65 3577,
Estasan GT 8-65 3581, Estasan GT 8-70 3579, Labrafac LIPO, Labrafac
lipophile WL 1349, Lexol GT-855, Lexol GT-865, Miglyol 810, Miglyol
812, Myritol 312, Myritol 318, Neobee 1053, Neobee M-5, Neobee
0, Pelemol CCT, Standamul 318, Standamul 7105 and Calgene CC-22,
Calgene CC-22-S, Captex 200, Lexol PG-865, Miglyol 840, Myritol
PC, Neobee 1054, Neobee M-20, Pelemol PDD, Standamul 302.
Most preferred is Miglyol grade 812.
The composition according to the invention comprises a thickener. A
thickener in a pharmaceutical formulation in general is useful to provide good
suspending properties and increases the viscosity of the composition without
negatively affecting the syringeability.
In one embodiment the thickener is hydrogenated castor oil. In another
embodiment the thickener is glycerol dibehenate. Alternatively combination
thereof or mixtures with other thickening agents known in the art are
employed.
It is preferred that the composition has a viscosity that is not very
temperature dependent to have a good syringeability also under cooler
temperature conditions as during winter time.
Typically the composition according to the present invention comprises 0.2 to
2.0% of the thickener.
The base further comprises one or more macrogol cetostearyl ether.
Macrogol cetostearyl ethers (Polyoxyethylated cetylstearyl alcohols) are non-
ionic emulsifiers. In general they are used to stabilize oil in water
emulsions.
Macrogol cetostearyl ethers are manufactured by reacting higher saturated
fatty alcohols with ethylene oxide. In the current composition they promote
the dispersion of the oily suspension in the aqueous content of the uterus.
They are e.g. available under the trademark Cremophor A grades from
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BASF, Ludwigshafen; Eumulgin B1 PH and Eumulgin B2 PH from Cognis
Deutschland GmbH & Co KG, Dusseldorf; Simulsol 58 PHA and Simulsol 68
PHA from Selectchemie AG, Zurich.
In one embodiment a combination of Macrogol cetostearyl ether 12 (degree
of ethoxylation 12, e.g. Eumulgin B1 PH, US DMF No. 17079) and Macrogol
cetostearyl ether 20 (degree of ethoxylation 20, e.g. Eumulgin B2 PH, US
dMF 17198, or SIMULSOL 58 PHA, Fa. SEPPIC, Polyoxyl 20 cetostearyl
ether, 20 EO units) is employed. In the current composition they promote the
dispersion of the oily suspension in the aqueous content of the uterus. The
ratio of Macrogol cetostearyl ether 12 and Macrogol cetostearyl ether 20 is
from 1: 10 to 10:1, preferably 1:5 to 5:1, more preferably 1:2 to 2:1.
Alternatively mixtures with other emulsifiers known in the art are employed.
Typically the composition according to the present invention comprises 0.5 to
5.0% of the macrogol cetostearyl ether
The pharmaceutical composition according to the current invention may
further comprise additional pharmaceutical excipients known in the art. Such
pharmaceutical excipients are e.g. described in "Gennaro, Remington: The
Science and Practice of Pharmacy" (20. Edition, 2000) incorporated by
reference herein.
A more specific composition according to the present invention typically
contains 1 to 10 % of cefquinome sulphate, 0.5 to 3 % of macrogol
cetostearyl ether, 0.1 to 3 % hydrogenated castor oil and up to 100 % of
Miglyol grade 812.
The pharmaceutical composition contemplated herein can, if desired, include
more than one pharmacologically active ingredient.
The current invention furthermore provides a process for preparing a
veterinary composition as claimed in any of the preceding claims comprising
the steps of mixing the medium chain triglyceride, thickener and macrogol
cetostearyl ether to create the base and suspending the cefquinome in said
base.
More specifically the current invention provides a process according to the
invention characterised in that the hydrogenated castor oil and macrogol
cetostearyl ether are added to the medium chain triglyceride under gentle
stirring and heating to form the base and the mixture is allowed to cool
before adding the cefquinome.
A detailed description of the manufacturing process for a composition
according to the invention is provided in examples 1 to 5.
Furthermore the current invention provides the use of the pharmaceutical
composition according to the current invention for the manufacture of a
medicament for the treatment or prevention of metritis, especially acute
metritis in mammalian animals, especially in cows after parturition.
Example 6 shows a pharmacokinetic study after intrauterine administration
of the composition according to the invention.
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The composition according to the invention proved highly effective against
the most commonly isolated metritis pathogens, (E. coli, A. pyogenes,
Prevotella spp., Fusobacterium spp.).
To be efficacious in the control of a pathogen, an antibiotic should be
5 present at a minimum concentration level in tissues or biological fluids,
which
is characterized by its MIC (Minimum Inhibitory Concentration) against a
pathogen. The minimum concentration level, in the lochia and endometrium
of postpartum cows that is considered to be efficacious, is determined by the
MICs of cefquinome against the different pathogens.
As shown in example 6, after intrauterine administration of the
pharmaceutical composition according to the invention during the effective
period a therapeutic efficient level in the lochia and endometrium above the
MIC for the different pathogens was reached.
Example 7 shows another pharmacokinetic study after intrauterine
administration of the composition according to the invention compared with a
known cefquinome sulphate formulation for intramammary administration in
lactating cows. This Cobactan LC formulation contains 75 mg Cefquinome
sulphate in a paraffinum liquidum base.
These experiments show that the intra-uterine formulation according to the
invention provides higher plasma concentrations and prolonged
concentrations in lochia that are helpful in achieving the therapeutic effect
of
the composition. Hence, example 7 shows that a topical formulation
developed for other body cavities (udder) is less suitable for intrauterine
administration.
The veterinary composition according to the invention can be applied in
general to all mammalian species that need treatment or prevention of
metritis and bacterial infections of the uterus such as e.g. pigs, cattle,
camel,
buffalo, horses, goats, sheep, and companion animals such as cats, dogs,
but especially to cows.
The chosen formulation may be filled into the tube or syringe packs of the
conventional type for intrauterine administration, i.e. connected to a
catheter
for insertion to allow extrusion directly into the uterus via the cervical
canal.
A single dose of the composition will normally contain 1 to 50 gram,
preferably 15 to 35 gram of the composition.
The particular amount of composition required for a particular treatment will
vary, depending upon the species, age and weight of the host animal being
treated, the particular disease to be guarded against, or treated, as well as
the specific antimicrobial agent selected for the treatment, the route and the
frequency of administration. For example the dose of cefquinome sulphate
for the treatment of acute metritis in post partum cows is 900 mg of
cefquinome for intrauterine administration.
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Examples
Example 1: Preparation of a cefquinome sulphate composition (100 kg)
3.60 kg Cefquinome as sulphate
1.11 kg Macrogol cetostearyl ether -12
0.47 kg Macrogol cetostearyl ether- 20
0.79 kg Hydrogenated castor oil
up to 100 kg Medium chain triglycerides
The manufacturing process encompasses the following steps:
1. The base of macrogol cetostearyl ether - 12 , macrogol cetostearyl ether
- 20, hydrogenated castor oil is weighted and mixed in medium chain
triglycerides, and homogenized under heating
2. the cefquinome sulphate is suspended under homogenisation
3. the product is filled in syringes.
Example 2: Preparation of cefquinome sulphate composition (100 kg)
3.60 kg Cefquinome as sulphate
2.22 kg Macrogol cetostearyl ether -12
0.94 kg Macrogol cetostearyl ether- 20
0.79 kg Hydrogenated castor oil
up to 100 kg Medium chain triglycerides
The composition is manufactured as disclosed in Example 1
Example 3: Preparation of cefquinome sulphate composition (100 kg)
3.60 kg Cefquinome as sulphate
1.66 kg Macrogol cetostearyl ether -12
0.70 kg Macrogol cetostearyl ether- 20
0.79 kg Hydrogenated castor oil
up to 100 kg Medium chain triglycerides
The composition is manufactured as disclosed in Example 1
Example 4: Preparation of a cefquinome sulphate composition (100 kg)
3.60 kg Cefquinome as sulphate
1.57 kg Macrogol cetostearyl ether -12
0.79 kg Hydrogenated castor oil
up to 100 kg Medium chain triglycerides
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The manufacturing process encompasses the following steps:
4. The base of macrogol cetostearyl ether and hydrogenated castor oil is
weighted and mixed in medium chain triglycerides, and homogenized
under heating
5. the cefquinome sulphate is suspended under homogenisation
6. the product is filled in syringes.
Example 5: Preparation of a cefquinome sulphate composition (100 kg)
3.60 kg Cefquinome as sulphate
1.57 kg Macrogol cetostearyl ether -20
0.79 kg Hydrogenated castor oil
up to 100 kg Medium chain triglycerides
The composition is manufactured as disclosed in Example 4
Example 6: In vivo pharmacokinetic study
Two pharmacokinetic studies were performed, during which plasma, lochia,
and endometrium cefquinome concentrations after administration of a
composition according to the invention (Examplel) were measured.
In parallel, bacteria isolated from field cases of acute metritis were tested
against cefquinome to determine MIC's.
Cefquinome concentrations from the pharmacokinetic studies were
compared to MIC's of cefquinome against acute metritis pathogens.
Material and methods:
One to six days after calving, fourteen healthy cows received an intra-uterine
administration of 900mg cefquinome in a composition of example 1.
Lochia samples and endometrium biopsies were collected at 7, 24, 48, and
72 hours after treatment. Blood samples were collected just before
treatment, and then for up to 48 hours after treatment.
Cefquinome concentrations in lochia and endometrium were determined with
validated microbiological assays. Cefquinome concentrations in plasma were
determined with a validated HPLC assay.
The strains used to determine the MIC's were isolated between 2000 to
2005, from cows with acute metritis from different European countries
(France, Germany, Hungary and Netherlands). Strains were collected by
intra-uterine swabbing, before any treatment.
The pathogens identified were used to determine cefquinome MIC's, using
standard broth micro-dilution or agar dilution techniques.
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Results:
One hour after treatment the median maximum plasmatic concentration was
1.28pg/mL. Plasmatic concentrations then steadily decreased to values
below 0.03pg/mL (limit of quantification) over 48 hours (Figure 1).
Cefquinome was detected in lochia and endometrium samples (Figure 1) for
up to 72 hours after treatment.
From the 194 field cases of acute metritis sampled for bacteriology, 504
microbial strains were identified. MIC's of cefquinome against the most
frequent pathogens (E. coli, A. pyogenes, Prevotella spp., Fusobacterium
spp.) are presented in Table 1.
Table 1: MIC50 and MIC9o (pg/mL of cefquinome) against key acute metritis
pathogens.
Bacteria species Number of MIC50 MIC90
strains
E. coli 131 0.032 0.125
A.pyogenes 120 0.25 0.5
Prevotella spp. 103 1 8
Fusobacterium 75 0.062 0.125
sPp.
Discussion: After intra-uterine administration there is a moderate diffusion
of cefquinome to the plasma. Antibacterial concentrations of cefquinome in
lochia and endometrium are reached after a single intra-uterine
administration.
In lochia the median cefquinome concentration was above or equal to the
MIC50 of 3 of the main acute metritis pathogens for up to 72 hours after
treatment, and above the MIC90 for at least 48 hours after treatment.
In endometrium, the median concentration of cefquinome was above the
MIC90 of the main acute metritis pathogens for more than 24 hours after
treatment.
Prolonged presence of cefquinome at antibacterial concentrations in
endometrium and lochia can prevent dissemination of the bacteria beyond
the uterus cavity and can decrease the intra-uterine bacterial load, allowing
the cow to recover from the infection.
Conclusion:lntra-uterine administration of 900 mg of cefquinome in the
composition provides prolonged local antibacterial concentrations of
cefquinome.
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Example 7: In vivo pharmacokinetic (PK) study
Two pharmacokinetic studies were performed, during which plasma and
lochia cefquinome concentrations after administration of a composition
according to the invention (Examplel) in comparison with a commercially
available intramammary formulation of Cefquinome sulphate for lactating
cows (Cobactan LC) were measured.
The results are illustrated in Figure 2 and Figure 3.
.After intra uterine administration the cefquinome plasma concentrations
measured can be considered as a marker for the endometrial tissue
cefquinome concentration. The availability of cequinome within endometrial
tissue is important for the successful treatment of metritis.
The PK studies show that Cobactan IU formulation according to the
invention (with 600 mg cefquinome) gives more plasmatic diffusion than the
Cobactan LC formulation (also 600 mg), and therefore, leads to higher
endometrial tissue concentrations.
The same formulations were compared by analysing samples of lochia (intra
uterine fluid).
Compared to Cobactan LC (with 600 mg cefquinome), Cobactan IU,
provides a lower concentration (sufficient for antimicrobial activity), and
with
a prolonged release, which makes the Cobactan IU formulation more
suitable for the control of metritis than the Cobactan LC formulation.
