Note: Descriptions are shown in the official language in which they were submitted.
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TITLE OF THE INVENTION
LIQUID PHARMACEUTICAL FORMULATIONS FOR ORAL ADMINISTRATION OF A
CGRP ANTAGONIST
BACKGROUND OF THE INVENTION
The pharmaceutical industry is faced with the challenge of developing
formulations for an increasing number of active molecules that possess low
aqueous solubility
and/or intestinal epithelial permeability. In some cases, as in the case of
the CGRP receptor
antagonist 1-[1-({[(3R,6S)-6-(2,3-difluorophenyl)-2-oxo-1-(2,2,2-
trifluoroethyl)azepan-3-
yl]amino}carbonyl)piperidin-4-yl]-2-oxo-1,2-dihydroimidazo[4,5-b]pyridin-3-ide
(Compound I),
acceptable bioavailability can not be readily achieved by means of traditional
tablet or capsule
formulations. Therefore, there remains a need to develop stable oral
formulations of Compound
I that would maximize exposure and provide for a faster onset of exposure. A
formulation that
permits the presentation of large doses per capsule would also be a desirable
result. This
invention provides pharmaceutical compositions that are liquid solutions of
Compound I, said
solutions being orally administrable. The active solutions may be
administered, for example, as
fill in encapsulated dosage forms such as liquid filled and sealed hard
gelatin capsules or soft
gelatin capsules. Other technologies commonly used for delivering poorly water
soluble
compounds as oral agents, such as particle size reduction, amorphous
formulations (hot melt
extrusion or spray drying) are typically explored, as was the case with
Compound I. However,
due to physicochemichemical limitations these alternative formulation
approaches were found
not to be feasible.
SUMMARY OF THE INVENTION .
The compound 1-[1-({[(3R,6S)-6-(2,3-difluorophenyl)-2-oxo-1-(2,2,2-
trifluoroethyl)azepan-3-yl]amino} carbonyl)piperidin-4-yl]-2-oxo-l,2-
dihydroimidazo[4,5-
b]pyridin-3-ide (Compound I)
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CF3
N O
O
N
H O
F F N NH
d
is a CGRP receptor antagonist. I
Compound I, its free base form and various salt and solvate forms are
described in
Intemational Patent Application No. PCT/LJS07/008703, filed Apri16, 2007, and
published
International Patent Application W004/092166, published October 28, 2004.
This invention relates to liquid formulations of Compound I (and salts
thereof,
and solvate forms and free base forms of Compound I) for use in mammals,
especially humans,
especially in connection with encapsulated formulations including hard and
soft gelatin capsules.
One salt form of compound I is the potassium ethanolate salt.
These liquid formulations provide increased solubility of Compound I, and
salts
thereof, for absorption and hence higher bioavailability. Compositions of the
present invention
comprising Formulations of compound I, and salts thereof, are useful in the
treatment of diseases
mediated by the antagonism of the CGRP receptor, including migraine headache
and cluster
headache.
BRIEF DESCRIPTION OF THE DRAWINGS
FIGURE 1 is a graph of the mean plasma concentration of Compound I over time
generated in
the experiment described in Example 6, wherein healthy men ages 18 to 45 were
orally
administered a 400 mg dosage of Compound I in a liquid filled gelatin capsule
and in several
different carrier formulations.
DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to pharmaceutical compositions for the oral
administration of 1-[ 1-( { [(3R,6S)-6-(2,3-difluorophenyl)-2-oxo-1-(2,2,2-
trifluoroethyl)azepan-3-
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yl]amino} carbonyl)piperidin-4-yl]-2-oxo-l,2-dihydroimidazo[4,5-b]pyridin-3-
ide (Compound I),
or salts thereof, a compound with low aqueous solubility (<0.05 mg/mL). The
invention is a
formulation which increases the oral bioavailability, as determined by
elevated drug plasma
levels, while maintaining chemical integrity of Compound I in the formulation
upon long term
storage.
When dosed as a pin milled crystalline solid, in a dry blend with excipients
the
compound was found to be poorly bioavailable in dogs It has been found that
bioavailability is
increased dramatically by using a liquid dosage formulation (see Table, below)
in which the
compound is in solution in certain combinations of liquid excipients:
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Mean pharmacokinetic parameters after oral administration of
40 mg Compound I in dry filled capsules or Imwitor/Tween 80
solution filled capsules to male Beagle dogs (mean SD).
Dry Filled Capsule Imwitor/Tween 80
Solution
AUCa-24
M-hr 0.229 0.293 2.073 1.427
Cmax
m 0.0495 0.560 0.637 0.377
Tmax
(hr) 2.2 1.8 2.0 0.0
However, the low solubility and/or chemical instability of Compound I in the
overwhelming
majority of vehicles typically used in the formulation of low aqueous
solubility compounds
necessitated extensive research and ingenuity to arrive at the surprising
invention disclosed
herein.
The formulations of the present invention possess the advantages of increased
bioavailability, excellent stability, increased potency per unit dose, good
safety and tolerability,
and ease of processing.
Thus, in one embodiment of the present invention, there is provided a liquid
formulation comprising Compound I or any pharmaceutically acceptable salt or
solvate thereof, a
surfactant component, a solvent component and other optionally present
components such as the
solvent water. In this embodiment the surfactant cornponerit may comprise one
or more
surfactants. Similarly, the solvent component may comprise one or more
solvents.
Suitable surfactants for use in the present invention include polyethoxylated
castor
oil (such as Cremophor EL (polyoxyl 35 castor oil), Cremophor RH60,
Cremophor RH40);
polysorbates (such as Tween-80, Tween-20); Polyethylene glycol ester
glycerides (such as
Labrasol , Labrifil 1944); sorbitan monooleate (such as Span -80, Span -20);
Vitamin E-
TPGS (vitamin E d-alpha-tocopherol polyethylene glycol succinate); saturated
polyglycolized
glycerides (such as Gelucire(D 44/14 and and Gelucire 50/13);
polypropoxylated stearyl
alcohols (such as Acconon MC-8, Acconon CC-6). Preferred surfactants for use
in the
present invention include Cremophor EL, Tween-80, Tween-20, and Labrasol .
Most
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preferred surfactants for use in the present invention include Tween 80
(polysorbate 80) and
Cremophor EL (polyoxyl 35 castor oil). The surfactants may be used alone or
in combination.
Most preferably, Tween 80 and Cremophor EL are used in combination with one
another.
Suitable solvents for use in the present invention include ethanol, propylene
glycol, glycerin, polyethylene glycols (such as PEG 200, PEG 400, PEG 600, PEG
900 and PEG
1000), DMSO, ethoxydiglycol (Transcutol ), diethyleneglycol, polyoxypropylene
block
copolymers (such as Polaxamer), and water. Preferred solvents for use in the
present invention
include ethanol and propylene glycols (PEG 400 and PEG 600). Most preferred
solvents for use
in the present invention include polyethylene glycol 400 (PEG-400) and
propylene glycol (PG).
The solvents may be used alone or in combination. Most preferably, PEG-400 and
PG are used
in combination with one another.
Within the first embodiment of the invention, there is a genus wherein the
liquid
forrnulation comprises Compound I, PEG-400, polysorbate 80, Cremophor EL, PG
and
optionally water, in the following amounts:
Component % (by wei ht
Compound I 5 to 40
PEG-400 0 to 95
Polysorbate 80 0 to 95
Cremo hor EL 0 to 95
Propylene GI col 0 to 95
Water 0 to 20
Within this genus there is a sub-genus wherein the liquid formulation
comprises
Compound I, PEG-400, polysorbate 80, Cremophorv EL, PG and optionally water,
in the
following amounts:
Component % (by wei ht
Compound I 20 to 35
PEG-400 0 to 50
Polysorbate 80 0 to 50
Cremo hor EL 0 to 50
Propylene Glycol 0 to 50
Water 0 to 10
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Within this sub-genus there is a further sub-genus wherein the liquid
formulation
comprises Compound I, PEG-400, polysorbate 80, Cremophor EL, PG and water, in
the
following amounts:
Component % (by wei ht
Compound I 28.5
PEG-400 23.3
Polysorbate 80 18.1
Cremo hor EL 18.1
Propylene Glycol 7
Water 5
The invention further relates to an oral pharmaceutical composition comprising
the liquid formulations described above.
In a second embodiment of the invention, there is provided a capsule
comprising
the liquid formulation of the first embodiment, wherein the capsule is filled
with 50 to 1200 mg,
and preferably 333 to 1050 mg of the liquid formulation of the first inventive
embodiment. The
fill amount corresponds to the presence of 100 to 300 mg of the active agent
Compound I or a
pharmaceutically acceptable salt or solvate thereof, 81.6 to 245 mg PEG-400,
47.5 to 190 mg
polysorbate 80, 47.5 to 190 mg Cremophor EL, 25 to 75 mg propylene glycol and
up to 50 mg
water; and preferably 300 mg the active agent Compound I, 245 mg PEG-400, 190
mg
polysorbate 80, 190 mg Cremophor EL, 75 mg propylene glycol and 50 mg water;
or 150 mg
the active agent Compound I, 122.5 mg PEG-400, 95 mg polysorbate 80, 95 mg
Cremophor
EL, 37.5 mg propylene glycol and 25 mg water.
Within the second embodiment, there is a genus wherein the capsule is selected
from a soft gelatin capsule and a hard gelatin capsule.
Within the second embodiment of the invention, there is a genus which is an
oral
pharmaceutical composition comprising a liquid filled capsule comprising
Compound I.
In a third embodiment of the invention, there is provided a method of making
the
liquid formulation of the first embodiment, and a method of making the liquid
filled capsule of
the second embodiment of the invention, wherein the method of making the
liquid formulation
comprises the steps of combining a pharmaceutically effective amount of
Compound I with one
or more solvents and one or more surfactants, and the method of making the
liquid filled capsule
further comprises filling a capsule with the liquid formulation and sealing
the capsule. In a
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preferred aspect of this embodiment, Compound I is combined with one or more
solvents
selected from ethanol, propylene glycol, glycerin, PEG 200, PEG 400, PEG 600,
PEG 900, PEG
1000, DMSO, Transcutol , diethyleneglycol, Polaxamer, water, and one or more
surfactants
selected from Cremophor EL, Tween-80, Tween-20, Labrasol , Labrifil 1944,
Cremophor
RH60, Cremophor RH40, Span -80, Vitamin E-TPGS, Geluicire 44/14, Acconon MC-
8,
Span&-20, Acconon CC-6, Gelucire 50/13, and the resulting liquid formulation
is filled into
a softgel and sealed. In a more preferred aspect, Compound I is combined with
the surfactants
Cremophor EL and polysorbate 80 and the solvents PEG 400 and propylene
glycol, and
optionally water, and the resulting liquid formulation is filled into a
softgel and sealed.
In a fourth embodiment of the invention, there is provided a method of
treating
one or more conditions selected from: migraine headache and cluster headache
with the capsule
of the second embodiment of the invention with the filled capsules of the
present invention
(containing Compound I). The method includes administering one or more
capsules to a patient
in need thereof, either to treat or prevent migraine headache or cluster
headache, or both.
Reference herein is made to a soft gelatin capsule, "softgel" being an
abbreviation
for soft gelatin capsules. It is understood that when reference is made to the
term "softgel" alone,
it shall be understood that the invention applies equally to all types of
gelatin and non-gelatin
capsules, regardless of hardness, softness, and so forth. In one embodiment of
the present
invention, the soft gelatin capsule contains plasticizers, such as glycerin
and sorbitol. Colorant
may be added to the gel mixture prior to encapsulation to produce soft gelatin
capsules of a
desired hue.
Referring to certain surfactants discussed herein, the terms Cremophor EL and
polyoxyl 35 castor oil are used interchangeably. Likewise, the terms Tween-80
and polysorbate
80 are interchangeable as used in this document. -
Referring to certain solvents discussed herein, the terms polyethylene glycol,
polyethylene glycol 400 and PEG 400 are used interchangeably. Similarly,
propylene glycol is
sometimes referred to as PG.
Reference to a specific weight or percentage of "active ingredient", or
Compound
I, is on the basis of the free base weight, absent the weight of any
counterion or solvate present,
unless otherwise indicated. For example, the phrase "1 mg 1-[1-({[(3R,6S)-6-
(2,3-
difluorophenyl)-2-oxo-1-(2,2,2-trifluoroethyl)azepan-3-yl]amino}
carbonyl)piperidin-4-yl]-2-
oxo-1,2-dihydroimidazo[4,5-b]pyridin-3-ide," or "1 mg Compound I" means that
the amount of
the compound selected is based on 1 mg of the free base form of Compound I
absent the weight
of the solvent present in the solvate.
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As used herein, the term "treatment" or "treating" means any administration of
a
compound of the invention and includes (1) inhibiting the disease in an animal
that is
experiencing or displaying the pathology or symptomatology of the diseased
(i.e., arresting
further development of the pathology and/or symptomatology), or (2)
ameliorating the disease in
an animal that is experiencing or displaying the pathology or symptomatology
of the diseased
(i.e., reversing the pathology and/or symptomatology). The term "controlling"
includes
preventing treating, eradicating, ameliorating or otherwise reducing the
severity of the condition
being controlled.
The liquid formulations of the invention are solutions of Compound I, one or
more solvents, and optionally one or more surfactants. A "solution" refers to
a clear liquid state
where the drug is dispersed at the molecular level, dissolved with no apparent
solid particles, in a
liquid environment provided by the solvent.
The invention is not limited to a particular mechanism of action. However, it
is
believed that upon release into the gastrointestinal fluids that the
surfactants of the formulation
will form micelle assemblies and micro-emulsions which are able to solubilize
Compound 1
above the solubility limit without surfacants present. The solubilized drug is
available for
absorption through the intestinal membrane, and hence systemic exposure in
vivo. In the absence
of surfactants the drug precipitates from the gastric juices, and the poorly
soluble drug is not
bioavailable.
The composition can be formulated as a fill encapsulated in a gelatin capsule
of
appropriate gelatin composition, a hard gelatin capsule with an appropriate
seal, a non-gelatin
capsule such as a hydroxypropyl methylcellulose capsule, or an oral liquid or
emulsion by
methods commonly employed in the art. In one embodiment of the present
invention, the fill is
encapsulated in a sealed hard gelatin capsule or a soft gelatin capsule
containing plasticizers,
such as glycerin and sorbitol. In one class of this embodiment, the hard
gelatin capsule is sealed
by band sealing using a gelatin ribbon, or LEMS (i.e., spraying with a
hydroalcoholic solution to
locally melt and seal the gelatin capsule pieces). The fill is prepared by
mixing the excipients
and Compound I with heating if required.
In addition to the main softgel capsule ingredients previously noted, other
stabilizing additives, as conventionally known in the art of softgel
formulation, can be introduced
to the fill as needed, usually in relatively small quantities, such as
antioxidants (BHA (butylated
hydroxyanisole), BHT (t- butylhydroxytoluene), tocopherol, propyl gallate,
etc.) and other
preservatives such as benzyl alcohol or parabens. Preferably, the antioxidant
or preservative is
present in a weight percent range of 0.01 % to 0.1 %. The composition can be
formulated as a fill
encapsulated in a soft gelatin capsule, a hard gelatin capsule with an
appropriate seal, a non-
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gelatin capsule such as a hydroxypropyl methylcellulose capsule or an oral
liquid or emulsion by
methods commonly employed in the art.
Oral delivery of Compound I is particularly difficult because its aqueous
solubility
is extremely low, typically being less than 5 ug/mL. Achieving therapeutic
drug levels in the
blood by oral dosing of practical quantities of a drug generally requires a
large enhancement in
drug concentrations in the gastrointestinal fluid and a resulting large
enhancement in
bioavailability. The formulations of this invention will be administered in
such an amount that
an effective dose of Compound I is administered to the patient. The amount of
Compound I will
generally be known or determined by the attending physician. Thus, the amount
or volume of
administered dose will be determined by the amount of Compound I prescribed
and/or otherwise
desired as a dose and the solubility of the Compound I in the formulation. In
general, an
effective dose for Compound I is from 100 mg to about 1000 mg per day, in
single or divided
doses; preferably from about 200 mg to about 800 mg per day, more preferably
from about 100
mg to about 600 mg per day, even more preferably about 150 or 300 mg per day,
in single or
divided doses. For oral administration, the compositions are preferably
provided in the form of
one or more liquid-filled capsules containing (in total, if more than one
liquid-filled capsule is
administered) from 100 to 1000 mg, preferably 100, 200, 300, 400, 500, 600,
700, 800, 900 and
1000, most preferably 200, 300, 400, 500 and 600 milligrams of the active
ingredient, for the
symptomatic adjustment of the dosage to the patient to be treated.
The liquid formulations of the present invention are pre-concentrates which
are
generally administered orally, in soft or hard gelatin capsules, gelatin
encapsulation technology
being well known to the pharrnaceutical arts.
The liquid formulations of the present invention have utility in treating,
preventing, ameliorating, controlling or reducing the risk of one or more of
the following
conditions or diseases: headache; migraine; cluster headache; chronic tension
type headache;
pain; chronic pain; neurogenic inflammation and inflammatory pain; neuropathic
pain; eye pain;
tooth pain; diabetes; non-insulin dependent diabetes mellitus; vascular
disorders; inflammation;
arthritis; bronchial hyperreactivity, asthma; shock; sepsis; opiate withdrawal
syndrome; morphine
tolerance; hot flashes in men and women; allergic dermatitis; encephalitis;
brain trauma;
epilepsy; neurodegenerative diseases; skin diseases; neurogenic cutaneous
redness, skin
rosaceousness and erythema; tinnitus; inflammatory bowel disease, irritable
bowel syndrome,
cystitis; and other conditions that may be treated or prevented by antagonism
of CGRP receptors.
Of particular importance is the acute or prophylactic treatment of headache,
including migraine
and cluster headache.
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The subject compounds are fiarther useful in a method for the prevention,
treatment, control, amelioration, or reduction of risk of the diseases,
disorders and conditions
noted herein. The subject compounds are further useful in a method for the
prevention, treatment,
control, amelioration, or reduction of risk of the aforementioned diseases,
disorders and
conditions in combination with other agents.
The terms "administration of' and or "administering a" compound should be
understood to mean providing the composition of the invention to the
individual in need of
treatment.
The administration of the composition of the present invention to practice the
present methods of therapy is carried out by administering an effective amount
of the compound
of structural formula I to the patient in need of such treatment or
prophylaxis. The need for a
prophylactic administration according to the methods of the present invention
is determined via
the use of well known risk factors. The effective amount of an individual
compound is
detennined, in the final analysis, by the physician in charge of the case, but
depends on factors
such as the exact disease to be treated, the severity of the disease and other
diseases or conditions
from which the patient suffers, the chosen route of administration, other
drugs and treatments
which the patient may concomitantly require, and other factors in the
physician's judgment.
Representative experimental procedures are provided below. These are
exemplary only and should not be construed as being limitations on the novel
compositions and
processes of this invention.
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Example 1- Liquid Formulation
Component Amount
(grams)
Compound 297.7
Pol eth lene Gl co1400 245.0
Propylene Glycol 75.0
Cremo hor EL 190
Polysorbate 80 190
Total 997.7
A water bath was preheated 35 C (5 C variance). Liquids PEG 400, propylene
glycol,
Cremophor EL, and polysorbate 80 were dispensed directly into a stainless
steel vessel
(diameter is 5'/4 inches) and stirred at 600 RPM stirring using a 2 inch mixer
blade. Compound
1(powder) was added into the vortex. After addition of Compound I the contents
were mixed
for approximately 2 hours at 600 rpm while maintaining the temperature at 35 C
(5 C variance).
As the formulation was mixed it was periodically tested by visual inspection
for undissolved
particles. After 2 hours of mixing only air bubbles were present (which was
attributed to the high
level of surfactant), and Compound I was completely dissolved. The solution
was deaerated for
about 2.5 hours under vacuum, placed at 40 C (5 C variance) overnight, and
then again placed
under vacuum for an additional 1 hour under to complete deaeration. The
formulation was ready
for encapsulation.
Example 2 - Liquid Formulation
Component Amount kilo rams
Compound I 1.70
Pol eth lene Glycol 400 1.39
Propylene Glycol 0.42
Cremo hor EL 1.08
Polysorbate 80 1.08
Water 0.28
Total 5.95
Liquids PEG 400, Propylene Glycol, Cremophor EL, Polysorbate 80 and water were
placed into
a 5'/4 inch diameter container equipped and mixed with a 2 inch blade at 600
RPM. Compound I
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was added to the mixture and mixing continued for an additiona12.5 hours at 25
C :I: 5 C. The
mixture was sampled once per hour and examined under microscope to determine
visual end
point (the lack of solid particles). The solution was mixed for an additional
1 hr. The solution
was filtered through a-100 micron filter and de-aerated until a yellowish
clear solution free of
foam and bubbles was obtained.
Example 3- Soft Gelatin Capsule Fill
The fill solution obtained in Example 2 (1050 mg) and the gelatin component
was supplied by
Banner Pharmacaps and were fed into an encapsulation machine. To encapsulate
the fill
solution, the gelatin formulation was cast into sheets on two cooled rollers.
These sheets were
passed through a series of rolls where a food grade lubricant was applied. The
sheets were then
fed through the rotary die rolls where the softgel was formed. As the lower
edge of the softgel
was formed, a reciprocating pump injected 1050 mg of the fill solution into
the center of the
softgel, after which the upper edge of the die came together to seal the
softgel. The newly
formed softgels were dislodged from the sheet and pneumatically conveyed to a
tumble dryer
where they stayed for 45-60 minutes. Upon exiting the dryer, the softgels were
spread on trays
and placed in a drying tunnel (low humidity chamber) and dried. Upon
completion of the drying
process, the softgels were visually inspected for defects. Subsequently, the
capsules were sized
to remove oversized and undersized capsules and polished. The resulting filled
softgel capsules
each contained 300 mg Compound I.
Example 4- Hard Gelatin Capsule Fill
Using a peristaltic pump the solution of Example 1 was pumped into a hopper
for encapsulation.
The liquid formulation was dispensed into the size 1, white, opaque hard
gelatin capsules
(CAPSUGEL, containing gelatin and titanium dioxide) to a target fill weight of
667 mg. The
filled capsules were transferred to a LEMS 30 capsule sealer and they were
sealed by spraying
with a mixture of 1:1 (weight:weight) water:ethanol (dehydrated, 190 proof)
solution. After
spraying the capsules were dried by gentle heating to approximately 45 C. The
sealed capsules
were placed onto trays lined with tray paper and were placed into a depression
chamber
(ZANASI 40E vacuum trap). After the completion of the vacuum cycle the
capsules were
visually inspected for leaking. The acceptable capsules were passed through a
ZANASI capsule
sorter to remove empty capsules. The finished capsules were then packaged into
appropriate
containers. The resulting filled capsules each contain 200 mg Compound I.
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Example 5 - Stability
Weeks Conditions Mg per Composite
Capsule Assay %
Present
0 - 208.0m 104.0%
14 5C/Amb 207.3mg 103.6%
14 25C/60%R 205.1 mg 102.5%
H
39 25C/60%R 201.2 mg 100.6
H
200 mg hard gelatin capsules containing the liquid formulation of Compound I
(capsules and
liquid fill prepared according to procedures similar to those described in
Examples 1 and 4) were
manufactured and stored under the conditions: 5 C at ambient humidity, and at
25 C. at 60%
relative humidity. Composite assay results at 14 and 39 weeks show excellent
stability.
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Example 6 - Bioavailability
Mean (S.D.) Pharmacokinetic Parameter Values Following
400-mg Single Oral Doses of Compound I(n=20)
Apparent
AUCõ AUCp,qw Cmarz T,naxt t'/2
Formulation (uM-hr) (!iM-hr) (!.-M) (hr) (hr)
17.64 (7.57) 8.08 (3.43) 3.15 (1.29) 1.5 (0.7-4) 6.2
LFC (PEG) 43 !o CV 41 % CV
15.92 (7.77) 7.05 (3.37) 2.97 (1.44) 1.75 (0.7-4) 6.4
LFC (PEG/PG) 49% CV 49% CV
26.35 (9.72) 14.63 (4.39) 6.68 (2.26) 1.5 (0.7-2) 5.7
LFC (PEG/PG/Crem/Tween) 37% CV 34% CV
1.45 (0.56) 0.51 (0.26) 0.21 (0.10) 2.0 (0.7-3) 7.8
DFC (Dynomilled Neutral) 38% CV 50% CV
14.38 (8.03) 6.48 (2.83) 2.65 (1.04) 1.75 (0.7-4) 6.4
DFC (Eudragit) 56% CV 39% CV
Adjusted for assayed potencies of the formulations
t Median (range)
Harmonic mean
Five different capsule formulations (refer to above table) of Compound I were
administered
orally as a single 400 mg dose in twenty healthy male subjects -between the=
ages of 18 and 45 in
an open label, randomized, 5-period brossover study. In each period subjects,
after an overnight
fast, were administered with a single oral dose of 1 of the 5 formulations
with a cup of 240 mL
water and then followed with a minimum washout of 72 hours. Blood was drawn in
the presence
of sodium heparin as an anticoagulant at predose and at 20, 40 minutes, 1,
1.5, 2, 3, 4, 6, 8, 12,
16, 24, 48 hours after dosing. The plasma was separated by centrifugation and
kept frozen at -
70 C until analysis by LC-MS/MS.
An online-extraction assay using Cohesive high-turbulence liquid
chromatography (HTLC) under
quick elution mode coupled with liquid chromatography/tandem mass spectrometry
for the
determination of Compound I concentration in human plasma was developed and
validated. A
stable isotope labeled D5-Compound I was used as the internal standard. Plasma
samples were
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prepared on a Packard Mu1tiPROBE II liquid-handling system, and directly
injected on a
Cohesive TX1 or TX2 Flux 2300 system (the TX2 system allowed parallel
operation of two sets
of extraction and analysis to increase sample throughput). The analyte and its
internal standard
were on-line extracted with a Cohesive C18 column (0.5x50 nun, 60u) using
10:90
acetonitrile:0.1% formic acid, back-eluted with 50:50 acetonitrile:0.1% formic
acid, and
analyzed on a Thermo-Keystone FluophaseRP column (100x2.1mm, 5u) using ramping
gradient
from 50:50 to 70:30 acenitonitrile:0.1 % formic acid. Detection was
accomplished on a Sciex
API4000 mass spectrometer in multiple-reaction monitoring (MRM) mode using
positive
ionization with a turbo ion-spray interface. Urider these conditions, the
diastereomer was well
separated from Compound I and no interference was observed for either Compound
I or the
internal standard from the endogenous components of human plasma. The assay
had a lower
limit of quantification (LOQ) 5 nM based on 0.05-mL aliquots of human plasma.
The standard
curve range was from 5 to 5000 nM. The analysis time was 10.0 minutes per
sample on the TX1
system.
Example 7 - Solubility
The solubility of the potassium salt ethanolate of Compound I in various
liquid components and
mixtures commonly used in making capsule formulations is shown below. The
units are
expressed in free acid equivalents (unionized drug) and the values were
ascertained by visual
inspection by optical microscopy_ The combinations were studied since the
highest solubility was
found in these individual excipeints.
Solvent I Surfactant m !
45% PEG 400/55% PEG 400 200-250
Tween 80 250-300
PEG 400 300-350
5% PG:95% PEG 400 350-400
5% PG:40% PEG 400:55% 250-300
Tween 80
100% PG >450
10% PG :90% PEG 400 >400
10% PG:35% PEG 400:55% <300
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WO 2008/030524 PCT/US2007/019461
Tween 80
9.2% PG: 35 % PEG 400: >400
27.9%
Tween 80: 27.9% Cremophor@)
EL
Example 8 - Phase Stability
Because mixtures of PEG 400 and surfactants such as polysorbate 80 are not
misicble in all
proportions, considerable ingenuity was required to develop a workable
surfactant / solvent
system. Phase separation is the common term for the point when the two liquid
solvents are
immiscible and the mixture becomes cloudy. In order to have a completely
miscible PEG 400 /
polysorbate 80 mixture the surfactant must be above 40% by weight. However,
this level of
surfactant is above the allowable limit to dose to human patients. Thus,
another surfactant was
required in the formulation to allow for a proper balance of PEG and
surfactant. On the basis of
keeping a single liquid phase a number of potential ingredients were tested as
a replacement for
about '/z of the polysorbate 80. The preferred ingredients in terms of phase
separation include
ethanol, polysorbate 20, Labrasol@, Cremophor EL, and Carpryol 90.
Possible Function FDA Phase Stability with Redisperibility
Ingredients Allowed PEG/Polysorbate
>400 mg 80/PG
PEG 200 Solvent Poor
PEG 400 Solvent Poor
PEG 600 Solvent Poor
PEG 1000 Solvent Poor
Propylene Solvent Poor
Glycol
Ethanol Solvent No Good
Gylcerin Solvent Poor
Transcutol Solvent Poor
Polysorbate Surfactant No Good
Polysorbate Surfactant Yes (but not Good
80 >800 mg)
Labrasol Surfactant Excellent
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WO 2008/030524 PCT/US2007/019461
Cremophor8 Surfactant Yes Excellent Excellent
EL
Cremophor Surfactant Poor
RH
Labrifil Triglyceride Incompatible
Oil
lmwitor 742 Triglyceride Poor
Oil
Capryol 90 Triglyceride Good
Oil
Gelucire Semisolid Poor
44/14 Oil
While the invention has been described and illustrated with reference to
certain particular embodiments thereof, those skilled in the art will
appreciate that various
changes, modifications and substitutions can be made therein without departing
from the
spirit and scope of the invention. It is intended, therefore, that the
invention be defined
by the scope of the claims which follow and that such claims be interpreted as
broadly as
is reasonable.
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