U.S. patent application number 17/198479 was filed with the patent office on 2021-07-22 for delta opioid agonist, mu opioid antagonist compositions and methods for treating parkinsons disease.
The applicant listed for this patent is VERSI GROUP, LLC. Invention is credited to BRUCE REIDENBERG, EBRAHIM VERSI.
Application Number | 20210220349 17/198479 |
Document ID | / |
Family ID | 1000005497010 |
Filed Date | 2021-07-22 |
United States Patent
Application |
20210220349 |
Kind Code |
A1 |
VERSI; EBRAHIM ; et
al. |
July 22, 2021 |
DELTA OPIOID AGONIST, MU OPIOID ANTAGONIST COMPOSITIONS AND METHODS
FOR TREATING PARKINSONS DISEASE
Abstract
The present invention provides for compositions and methods for
the treatment of Parkinson's disease comprising a compound of
formula (i): ##STR00001## or a pharmaceutically effective salt or
ester thereof alone or in combination with L-DOPA to provide a
synergistic effect, thereby providing methods of (1) treating
patients with Parkinson's Disease for whom L-DOPA is no longer
effective, (2) treating patients with Parkinson's Disease who
developed dyskinesia due to L-DOPA, (3) treating patients with
Parkinson's Disease who have are receiving deep brain stimulation
and (4) treating patients with Parkinson's Disease whose symptoms
interfere with activities of daily living.
Inventors: |
VERSI; EBRAHIM; (GLADSTONE,
NJ) ; REIDENBERG; BRUCE; (RYE, NY) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
VERSI GROUP, LLC |
GLADSTONE |
NJ |
US |
|
|
Family ID: |
1000005497010 |
Appl. No.: |
17/198479 |
Filed: |
March 11, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
16077575 |
Aug 13, 2018 |
10973815 |
|
|
PCT/US2017/024989 |
Mar 30, 2017 |
|
|
|
17198479 |
|
|
|
|
62315717 |
Mar 31, 2016 |
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 25/16 20180101;
A61K 31/495 20130101; A61P 25/14 20180101 |
International
Class: |
A61K 31/495 20060101
A61K031/495; A61P 25/14 20060101 A61P025/14; A61P 25/16 20060101
A61P025/16 |
Claims
1. A method of treatment for a subject with Parkinson's disease
experiencing prolonged L-DOPA treatment and increasing episodes of
"off-time," the method comprises supplementing administered reduced
amount of L-DOPA dose with therapeutically effective amount of
formula ##STR00006##
4-((alpha-R)-alpha-(2S,5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)-
-3-hydroxybenzyl)-N,N-diethylbenzamide or a pharmaceutically
acceptable salt or ester thereof to maintain or achieve an
antiparkinsonian effect.
2. The method of claim 1 wherein the compound of formula (i) is
administered in a dose ranging from about 1 to about 100 mg/kg per
day.
3. The method of claim 1, wherein the reduced amount of L-DOPA
ranges from an amount from 1 to 10 mg/kg/day of L-DOPA.
4. The method of claim 2 wherein the compound of formula (i) is
administered in a dose ranging from about 5-20 mg/kg per day.
5. The method of claim 4 wherein the administered formula (i)
results in prolongation of the duration of effect of a single
sub-therapeutic dose of L-DOPA.
6. A method of treatment for a subject with Parkinson's disease
subjected to deep brain stimulation, the method comprising
postsurgical administration of formula (i): ##STR00007##
4-((alpha-R)-alpha-((2 S,
5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)-3-hydroxybenzyl)-N,N-d-
iethylbenzamide or a pharmaceutically acceptable salt or ester.
7. The method of claim 6, wherein the compound of formula (i) is
administered in a dose ranging from about 1 to about 100
mg/kg/day.
8. The method of claim 7, further comprising a dose of L-DOPA in an
amount from about 1 to 10 mg/kg/day.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is a divisional application and claims
priority to copending U.S. patent application Ser. No. 16/077,575
filed on Aug. 13, 2018, now U.S. patent Ser. No. ______, which in
turn claims priority to International Patent Application No.
PCT/US2017/0024989 filed on Mar. 30, 2017 which in turn claims
priority to U.S. Provisional Patent Application No. 62/315,717
filed on Mar. 31, 2016, the contents of which is hereby
incorporated by reference herein for all purposes.
BACKGROUND OF THE INVENTION
Field of the Invention
[0002] The present invention relates to compositions and methods of
treatment for Parkinson's disease, by administration to a subject
suffering or susceptible to same, a diarylmethylpiperazine compound
that exhibits both delta opioid receptor agonist and mu opioid
receptor antagonist activity and optionally in combination with
other active Parkinson treating agents.
Description of the Related Art
[0003] Parkinson's disease is a degenerative disorder identified by
the loss of motor control progressing to loss of motor functions as
well as Parkinsonian dementia in many cases. The pathophysiology of
the disease is dominated by a loss of dopamine containing neurons
in the brain, particularly in the basal ganglia areas.
[0004] Current medical therapy for Parkinson's disease is based
primarily around the replacement of the dopamine deficit through
the administration of L-DOPA with or without other dopamine
agonists. However, after 3-5 years of L-DOPA treatment alone
patients develop motor fluctuations due to "wearing-off" of the
therapeutic effect, "on-off" fluctuations in efficacy immediately
after dosing, or most commonly, an "overshoot" of effect manifested
as abnormal involuntary movements (AIMS; dyskinesias) (Hill et al.,
2000; Silverdale et al., 2003). The co-administration of dopamine
agonists (primarily ergot-derived) with L-DOPA slows the
development of these side-effects but does not prevent their
eventual onset (Hill et al., 2000; Silverdale et al., 2003).
Critically, once patients have experienced L-DOPA-induced
dyskinesias they are "primed" for dyskinesias in response to any
currently available dopamine-based therapeutic (Hill et al., 2000).
In light of these debilitating side-effects and the loss of
efficacy over extended L-DOPA use, non-dopaminergic therapy for
Parkinson's disease would be highly desirable, whether as a
monotherapy or as an adjunct therapy to low-dose L-DOPA
administration.
[0005] Some anti-parkinsonian effect has been demonstrated for
delta receptor activation by rodent and primate studies with the
delta receptor-selective agonist, SNC80 (Hille et al, 2001). SNC80
is a highly selective delta receptor agonist and is an analogue of
benzhydrylpiperazines such as BW373U86 (Chang et al., 1993). SNC80
at 10 mg/kg i.p. restored behavioral deficits observed in rats
treated with reserpine or dopamine receptor antagonists
(haloperidol or SCH23390) including ambulatory behavior, grooming,
rearing, social interaction and exploration, and static
investigation. When administered at similar dose levels to
MPTP-treated marmosets, SNC80 restored motor activity, bradykinesia
and disability scores to normal. The parkinsonian posture induced
by MPTP treatment in this model was not statistically reversed by
SNC80 although substantial improvements were observed in some
individual animals (Hille et al., 2001).
[0006] These reports, in both rodent and non-human primate models
of Parkinson's disease, indicate that the delta opioid receptor
agonist SNC80 has a powerful anti-parkinsonian effect.
Interestingly, even at relatively high doses behavior returned to
normal levels but did not show the hyperkinesias associated with
supra-optimal doses of L-DOPA (Hille et al, 2001). The mechanism of
action appears to be via the delta receptor (Hille et al, 2001).
However, SNC80 is neither orally active nor particularly safe for
use in a clinical setting. SNC80 has been shown to produce
seizure-like convulsions in mice and rats, and as such cannot be
pursued for clinical study (Broom, et al., 2002).
[0007] A highly selective mu antagonist cyprodime was shown to
reduce completely L-DOPA-induced dyskinesia in the MPTP-lesioned
primate (marmosets) model of Parkinson's disease without
attenuation of the anti-parkinsonian actions of L-DOPA (Henry et
al., 2001). It appears that the highly selective mu antagonist
cyprodime was more effective than less selective antagonists such
as naltrexone and naltrindole at the dose of 10 mg/kg i.p. This
data plus the demonstrated increase in the synthesis of
proenkephalin A and B in basal ganglia in the animal model of
L-DOPA-induced dyskinesia and in postmortem tissue from Parkinson's
disease patients treated with conventional therapy (Nisbet et al.,
1995; Maneuf et al., 1994; De Ceballos et al., 1993) provides
strong evidence that mu-receptor activation may contribute to the
development of dyskinesia in Parkinson's disease patients after
chronic L-DOPA based therapy.
[0008] Consequently, it follows that treatment a compound with dual
opioid action of delta receptor agonism and mu receptor antagonism
would not only have anti-Parkinsonian effects but also would reduce
or eliminate the potential of delta receptor agonist induced
dyskinesia. Further such mu antagonism may also reduce L-DOPA
induced dyskinesia.
[0009] Thus, there is a need for an effective opioid-based
treatment for Parkinson's disease that not only acts as a delta
opioid receptor agonist but also exhibits mu opioid receptor
antagonist activity and has the ability to overcome the negative
side effects of L-DOPA used in the treatment of Parkinson's
disease.
SUMMARY OF THE INVENTION
[0010] L-DOPA therapy is the gold standard for treating Parkinson's
disease. Unfortunately, the value of current L-DOPA based therapies
for Parkinson's disease is limited by significant complications of
long-term treatment, particularly dyskinesia. The present invention
provides for a novel therapy, with the ability, as monotherapy, to
alleviate or reduce parkinsonian symptoms to the same extent as
currently-available dopamine replacing agents, while producing
fewer side effects. Additionally, the present invention provides
for a compound that exhibits delta opioid receptor agonist and mu
opioid antagonist activity in combination with low-doses of an
existing dopaminergic such as L-DOPA that produces an equivalent
anti-parkinsonian response comparable to higher, optimal doses of
L-DOPA, but with fewer negative side-effects.
[0011] The present invention relates to a composition and a method
of treating Parkinson's disease comprising administering to said
subject an effective amount of a therapeutic composition comprising
a compound that exhibits delta opioid receptor agonist and mu
opioid antagonist activity, optionally in combination with other
active Parkinson treating agents. Preferably, the compound is a
diarylmethylpiperazine compound, wherein the compound has the
structure of formula (i) (DPI-289), as shown below:
##STR00002##
[0012]
4-((alpha-R)-alpha-((2S,5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-piper-
azinyl)-3-hydroxybenzyl)-N,N-diethylbenzamide or a pharmaceutically
acceptable salt or ester thereof.
[0013] In another aspect, the diarylmethylpiperazine compound
having the above structure of Formula (i) may be administered in
combination with other Parkinson's treating agents such as dopamine
agonists, including but not limited to apomorphine, bromocriptine,
cabergoline, ciladopa, dihydrexidine, dinapsoline, doxanthrine,
epicriptine, lisuride, piribedil, pramipexole propylnorapomorphine,
quinagolide, ropinirole, rotigotine, roxindole, sumanirole,
benztropine mesylate, entacapone, selegiline hydrochloride,
carbidopa, pergolide, amantadine hydrochloride and tolcapone.
Dopamine agonists rarely cause dyskinesia such as induced by L-DOPA
but they have poor efficacy and thus combining them with DPI-289
can provided increase efficacy and cause a synergistic
effectiveness.
[0014] In yet another aspect, the present invention relates to a
method of treating Parkinson's disease in subject that cannot take
L-DOPA because of the negative side effects, wherein administration
of the structure of formula (i) (DPI-289) as shown below:
##STR00003##
[0015] 4-((alpha-R)-alpha-((2
S,5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)-3-hydroxybenzyl)-N,N-
-diethylbenzamide or a pharmaceutically acceptable salt, in a
therapeutically effective amount having the effectiveness to delay
or eliminate the need for deep brain stimulation in a subject
suffering from Parkinson's disease.
[0016] In a still further aspect, the present invention provide for
a treatment for a subject who is L-DOPA experienced and having
dyskinesia adverse events including over-activity of movement,
dyskinesia, reduced "ON-time," postural instability and/or reduced
alertness, wherein the treatment comprises administering a
therapeutically effective amount of formula (i):
##STR00004##
[0017] 4-((alpha-R)-alpha-((2
S,5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)-3-hydroxybenzyl)-N,N-
-diethylbenzamide or a pharmaceutically acceptable salt or ester
thereof.
[0018] In another aspect, the present invention provides for
treatment of a subject who has developed L-DOPA induced dyskinesia
or unable to tolerate L-DOPA, the method comprising the
administration of formula (i) (DPI-289) in a single dose amount
ranging from about 1 mg/kg to about 100 mg/kg, preferably 5 to 20
mg/kg, administered orally several times a day to increase
improvement of movement and improvement in self-care including
feeding, dressing, personal hygiene without causing dyskinesia. The
daily dose of DPI-289 would be about 5 mg/kg to about 500 mg/kg,
preferably 25 mg/kg to 100 mg/kg.
[0019] In yet another aspect, the present invention provides for a
composition comprising a synergistic combination comprising a low
dose of L-DOPA, that being, from about 1 to 10 mg/kg of L-DOPA per
day, is combined with formula (i) (DPI-289) in an amount from about
1 mg/kg to 100 mg/kg, preferably 5 mg/kg to 20 mg/kg, where said
combination causes a synergistic effect that provides for extended
reduction in parkinsonian symptoms relative to either components
alone or a higher dose of L-DOPA alone (about 10 mg/kg to 100
mg/kg). The synergistic combination provides for [0020] Enhancement
of the efficacy of sub-therapeutic doses of L-DOPA [0021]
Enhancement of the efficacy of sub-therapeutic doses of L-DOPA
without causing dyskinesia [0022] Enhancement of the efficacy of
L-DOPA in patients who have become less responsive to previously
optimally response to L-DOPA [0023] Enhancement of the efficacy of
L-DOPA in patients who have become less responsive to previously
optimally response to L-DOPA without causing dyskinesia [0024]
Enhancement of "ON" time meaning L-DOPA is effective for a longer
period with limited reduction in symptomatic "on and off"
fluctuations [0025] Maintenance of efficacy of L-DOPA despite the
need to reduce L-DOPA doses to limit dyskinesia [0026] Prevention
or reduction of L-DOPA induced dyskinesia [0027] Prevention or
reduction of L-DOPA induced dyskinesia without reduction of e
efficacy of L-DOPA
[0028] In yet another aspect, the present invention provides for a
composition comprising a synergistic effect of formula (i) DPI-289
such that the duration of action of a single low dose of L-DOPA
(about 1 to 10 mg/kg) is prolonged. The dose of formula (i)
(DPI-289) in an amount from about 1 mg/kg to 25 mg/kg, preferably 2
mg/kg to 5 mg/kg, where said combination causes a synergistic
effect that provides for prolonged reduction in parkinsonian
symptoms. The synergistic combination provides for [0029]
Prolongation of the duration of effect of sub-therapeutic doses of
L-DOPA and the combination lasts longer than that of the high dose
(optimal) of L-DOPA [0030] Prolongation of the effect of DPI-289
with concomitant use of L-DOPA
[0031] As used herein, the term "a synergistic effect" is present
when the activity of the active compounds in a combination exceeds
the total of the action of the active compounds when applied
individually.
[0032] A therapeutic composition of the present invention
comprising a synergistic combination comprising formula (i)
(DPI-289) and a low dose of L-DOPA may be administered by any
suitable administrative mode, e.g., an administration modality
selected from the group consisting of oral, rectal, topical,
sub-lingual, mucosal, nasal, ophthalmic, subcutaneous,
intramuscular, intravenous, transdermal, spinal, intrathecal,
intracranial, intra-articular, intra-arterial, sub-arachnoid,
bronchial, lymphatic, vaginal and intra-uterine administration.
[0033] The synergistic composition may comprise formula (i)
(DPI-289) in a single dose amount from about 1 mg/kg to about 100
mg/kg, and more preferably, from about 5 mg/kg to about 20 mg/kg.
L-DOPA is included in an amount that does not cause negative side
effects such as dyskinesia, postural instability and/or reduced
alertness. Preferably a daily low dose ranges from about 100 mg to
1000 mg a day. Higher doses of L-DOPA can ranged from 2000 to 6000
mg per day. However, such higher doses can present negative
effects, and as such, reducing the amount to a lower dose in
combination with formula (i) provides for effective treatment of PD
without negative effects caused by higher doses of L-DOPA.
[0034] Another aspect of the present invention relates to use of a
synergistic combination comprising Formula (i) in combination with
a low dose of L-DOPA in the manufacture of a medicament or
pharmaceutical for the treatment of Parkinson's disease.
[0035] Various other aspects, features and embodiments of the
invention will be more fully apparent from the ensuing disclosure
and appended claims.
BRIEF DESCRIPTION OF THE FIGURES
[0036] FIG. 1 shows effect of Formula (i) (DPI-289) on movement
activity in L-DOPA naive MPTP-lesioned primates. This activity is
measured by an infra-red movement detector and captures all
movement, normal and abnormal. MPTP-lesioned cynomolgus monkeys
received acute oral administration of either vehicle or Formula (i)
(1, 10 or 20 mg/kg) according to a randomized incomplete Latin
Square-type. Levels of activity were assessed every minute over the
entire 6 h observation. FIG. 1A shows time course data as
mean.+-.s.e.m. (FIG. 1A; time-course) or cumulated shown in FIG. 1B
for 0-3 h; or FIG. 1C for 3-6 hr. N=8 for all treatment groups.
*/**/*** represents P<0.05, P<0.01 or P<0.001cf.
vehicle-treatment; 2-way RM ANOVA (FIG. 1A) or 1-way RM-ANOVA with
Holm-Sidak's Multiple Comparison test (FIGS. 1B and C) (ns-not
significant).
[0037] FIG. 2 shows the results on movement activity of
administration of DPI-289 in macaques with established motor
complications due to administered L-DOPA. This activity is measured
by an infra-red movement detector and captures all movement, normal
and abnormal. Data are mean.+-.s.e.m. (FIG. 2A shows time-course)
or cumulated shown in FIG. 2B for 0-3 h; or FIG. 2C for 3-6 hr. N=8
for all treatment groups. */**/*** represents P<0.05, P<0.01
or P<0.001cf. vehicle-treatment; 2-way RM ANOVA (FIG. 2A) or
1-way RM-ANOVA with Holm-Sidak's Multiple Comparison test (FIGS. 2B
and C). */**/*** represents P<0.05, P<0.01 or
P<0.001cf.
[0038] FIG. 3 shows effect of Formula (i) (DPI-289) on duration of
ON-time in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned
cynomolgus monkeys received acute oral administration of either
vehicle or Formula (i) (1, 10 or 20 mg/kg) according to a
randomized incomplete Latin Square-type. Levels of bradykinesia
were assessed every 10 min over the entire 6 h observation and
total duration of ON-time (minutes for which bradykinesia was
absent) was calculated. Data are mean.+-.s.e.m. N=8 for all
treatment groups. All P>0.05, vehicle-treatment; 1-way, RM-ANOVA
with Holm-Sidak's Multiple Comparison test.
[0039] FIG. 4 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA.
[0040] FIG. 5 shows effect of Formula (i) (DPI-289) on parkinsonian
disability in L-DOPA naive MPTP-lesioned primates. Parkinsonian
disability is scored by a trained neurologist who is blinded to
treatment allocation but is trained to differentiate normal from
abnormal movements and is able to distinguish diminution of
Parkinsonian disability from dyskinetic (abnormal) movements.
MPTP-lesioned cynomolgus monkeys received acute oral administration
of either vehicle or Formula (i) (1, 10 or 20 mg/kg) according to a
randomized incomplete Latin Square-type. Levels of parkinsonian
disability were assessed every 10 min over the entire 6 h
observation. Data are median values (FIG. 5A shows time-course) or
median bars with individual animal scores (cumulated 0-3 h shown in
FIG. 5B or 3-6 h shown in FIG. 3C). N=8 for all treatment groups.
*/P<0.05 cf. vehicle-treatment; 2-way RM ANOVA of ranked data
(FIG. 5A) or 1-way RM-ANOVA with Holm-Sidak's Multiple Comparison
test (FIGS. 5B and C).
[0041] FIG. 6 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA. Parkinsonian disability is scored by a trained neurologist
who is blinded to treatment allocation but is trained to
differentiate normal from abnormal movements and is able to
distinguish diminution of Parkinsonian disability from dyskinetic
(abnormal) movements. Data are mean.+-.s.e.m. (FIG. 6A shows
time-course) or cumulated for 0-3 h shown in FIG. 6B, or shown for
3-6 h in FIG. 6C. N=8 for all treatment groups. */**/*** represents
P<0.05, P<0.01 or P<0.001cf. vehicle-treatment; 2-way RM
ANOVA (FIG. 6A) or 1-way RM-ANOVA with Holm-Sidak's Multiple
Comparison test (FIGS. 6 B and C).
[0042] FIG. 7 shows effect of Formula (i) (DPI-289) on bradykinesia
in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned cynomolgus
monkeys received acute oral administration of either vehicle or
Formula (i) (1, 10 or 20 mg/kg) according to a randomized
incomplete Latin Square-type. Levels of bradykinesia were assessed
every 10 min over the entire 6 h observation. Data are median
values (FIG. 7A shows time-course) or median bars with individual
animal scores (cumulated for 0-3 h shown in FIG. 7B or 3-6 h shown
in FIG. 7C). N=8 for all treatment groups. */** represents
P<0.05 or P<0.01 cf. vehicle-treatment; 2-way RM ANOVA of
ranked data (FIG. 7A) or 1-way RM-ANOVA with Holm-Sidak's Multiple
Comparison test (FIGS. 7B and 7C).
[0043] FIG. 8 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA. Data are mean.+-.s.e.m. (FIG. 8A shows time-course) or
cumulated for 0-3 h shown in FIG. 8B or 3-6 h shown in FIG. 8C. N=8
for all treatment groups. *** represents P<0.001cf.
vehicle-treatment; 2-way RM ANOVA (FIG. 8A) or 1-way RM-ANOVA with
Holm-Sidak's Multiple Comparison test (FIGS. 8B and 8C).
[0044] FIG. 9 shows effect of Formula (i) (DPI-289) on range of
movement in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned
cynomolgus monkeys received acute oral administration of either
vehicle or Formula (i) (1, 10 or 20 mg/kg) according to a
randomized incomplete Latin Square-type. Levels of range of
movement were assessed every 10 min over the entire 6 h
observation. Data are median values (FIG. 9A shown for time-course)
or median bars with individual animal scores (cumulated for 0-3 h
shown in FIG. 9B or for 3-6 h shown in FIG. 9C). N=8 for all
treatment groups. All P>0.05 cf. vehicle-treatment; 2-way RM
ANOVA of ranked data (FIG. 9A) or 1-way RM-ANOVA with Holm-Sidak's
Multiple Comparison test (FIG. 9B and FIG. 9C).
[0045] FIG. 10 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA. Data are mean.+-.s.e.m. (FIG. 10A shows time-course) or
cumulated 0-3 h shown in FIG. 10B or for 3-6 h shown in FIG. 10C.
N=8 for all treatment groups. */**represents P<0.05 or P<0.01
cf. vehicle-treatment; 2-way RM ANOVA
[0046] (FIG. 10A) or 1-way RM-ANOVA with Holm-Sidak's Multiple
Comparison test (FIGS. 10B and C).
[0047] FIG. 11 shows effect of Formula (i) (DPI-289) on postural
impairment in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned
cynomolgus monkeys received acute oral administration of either
vehicle or Formula (i) (1, 10 or 20 mg/kg) according to a
randomized incomplete Latin Square-type. Postural impairment was
assessed every 10 min over the entire 6 h observation. Data are
median values (FIG. 11A shows time-course) or median bars with
individual animal scores (cumulated for 0-3 h shown in FIG. 11B, or
for 3-6 h shown in FIG. 11C). N=8 for all treatment groups.
**/P<0.01 cf. vehicle-treatment; 2-way RM ANOVA of ranked data
(FIG. 11A) or 1-way RM-ANOVA with Holm-Sidak's Multiple Comparison
test (FIGS. 11B and C).
[0048] FIG. 12 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA. Data are mean.+-.s.e.m. (FIG. 12A shows time-course) or
cumulated for 0-3 h shown in FIG. 12B or for 3-6 h shown in FIG.
12C. N=8 for all treatment groups. */**/*** represents P<0.05,
P<0.01 or P<0.001cf. vehicle-treatment; 2-way RM ANOVA (FIG.
12A) or 1-way RM-ANOVA with Holm-Sidak's Multiple Comparison test
(FIGS. 12B and C).
[0049] FIG. 13 shows effect of Formula (i) (DPI-289) on alertness
in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned cynomolgus
monkeys received acute oral administration of either vehicle or
Formula (i) (1, 10 or 20 mg/kg) according to a randomized
incomplete Latin Square-type. Alertness was assessed every 10 min
over the entire 6 h observation. Data are median values (FIG. 13A
shows time-course) or median bars with individual animal scores
(cumulated for 0-3 h shown in FIG. 13B or for 3-6 h shown in FIG.
13C). N=8 for all treatment groups. All P>0.05 cf.
vehicle-treatment; 2-way RM ANOVA of ranked data (FIG. 13A) or
1-way RM-ANOVA with Holm-Sidak's Multiple Comparison test (FIGS.
13B and C).
[0050] FIG. 14 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA. Data are mean.+-.s.e.m. (FIG. 14A shows time-course) or
cumulated for 0-3 h shown in FIG. 14B or for 3-6 h shown in FIG.
14C. N=8 for all treatment groups. */**/*** represents P<0.05,
P<0.01 or P<0.001cf. vehicle-treatment; 2-way RM ANOVA (FIG.
14A) or 1-way RM-ANOVA with Holm-Sidak's Multiple Comparison test
(FIGS. 14B and C).
[0051] FIG. 15 shows effect of Formula (i) (DPI-289) on dyskinesia
in L-DOPA naive MPTP-lesioned primates. MPTP-lesioned cynomolgus
monkeys received acute oral administration of either vehicle or
Formula (i) (1, 10 or 20 mg/kg) according to a randomized
incomplete Latin Square-type. Dyskinesia was assessed every 10 min
over the entire 6 h observation. Data are median values (FIG. 15A
shows time-course) or median bars with individual animal scores
(cumulated for 0-3 h shown in FIG. 15B or for 3-6 h shown in FIG.
15C). N=8 for all treatment groups. All zero values.
[0052] FIG. 16 shows the results of administration of DPI-289 in
MPTP treated macaques with established motor complications due to
administered L-DOPA. Data are mean.+-.s.e.m. (FIG. 16A shows
time-course) or cumulated for 0-3 h shown in FIG. 16B or for 3-6 h
shown in FIG. 16C. N=8 for all treatment groups. 2-way RM ANOVA
(FIG. 16A) or 1-way RM-ANOVA with Holm-Sidak's Multiple Comparison
test (FIG. 16B and FIG. 16C).
[0053] FIG. 17 shows the motor activity effect of acute DPI-289
administered in combination with low and high dose of L-DOPA in
macaques with established motor complications due to administered
L-DOPA. The dose of L-DOPA was individualized to each animal, the
low dose being sub-therapeutic but which caused minimal dyskinesia
and the high dose being a therapeutic but which also caused
disabling dyskinesia. This activity is measured by an infra-red
movement detector and captures all movement, normal and abnormal.
Data are mean.+-.s.e.m. (FIG. 17A shows time-course) or cumulated
for 0-6 h shown in FIG. 17B. N=8 for all treatment groups. 2-way RM
ANOVA (FIG. 17A) or 1-way RM-ANOVA with Holm-Sidak's Multiple
Comparison test (FIG. 17B).
[0054] FIG. 18 shows the parkinsonian disability effect of acute
DPI-289 administered in combination with low and high dose of
L-DOPA. Parkinsonian disability is scored by a trained neurologist
who is blinded to treatment allocation but is trained to
differentiate normal from abnormal movements and is able to
distinguish diminution of Parkinsonian disability from dyskinetic
(abnormal) movements. Data are mean.+-.s.e.m. (FIG. 18A shows
time-course) or cumulated for 0-6 h shown in FIG. 18B. N=8 for all
treatment groups. 2-way RM ANOVA (FIG. 18A) or Friedman's test with
Dunn's post-hoc (FIG. 18B).
[0055] FIG. 19 shows the level of dyskinesia effect of acute
DPI-289 administered in combination with low and high dose of
L-DOPA in the same animals as displayed in FIGS. 17 & 18. Data
are mean.+-.s.e.m. (FIG. 19A shows time-course) or cumulated for
0-6 h shown in FIG. 19B. N=8 for all treatment groups. 2-way RM
ANOVA (FIG. 19A) or Friedman's test with Dunn's post-hoc (FIG.
19B).
[0056] FIG. 20 shows the postural instability effect of acute
DPI-289 administered in combination with low and high dose of
L-DOPA in the same animals as displayed in FIGS. 17 & 18. Data
are mean.+-.s.e.m. (FIG. 20A shows time-course) or cumulated for
0-6 h shown in FIG. 20B. N=8 for all treatment groups. 2-way RM
ANOVA (FIG. 20A) or Friedman's test with Dunn's post-hoc (FIG.
20B).
[0057] FIG. 21 shows the alertness effect of acute DPI-289
administered in combination with low and high dose of L-DOPA in the
same animals as displayed in FIGS. 17 & 18. Data are
mean.+-.s.e.m. (FIG. 21A shows time-course) or cumulated for 0-6 h
shown in FIG. 21B. N=8 for all treatment groups. 2-way RM ANOVA
(FIG. 21A) or Friedman's test with Dunn's post-hoc (FIG. 21B).
[0058] FIG. 22 shows the range of movement impairment effect of
acute DPI-289 administered in combination with low and high dose of
L-DOPA in the same animals as displayed in FIGS. 17 & 18. Data
are mean.+-.s.e.m. (FIG. 22A shows time-course) or cumulated for
0-6 h shown in FIG. 22B. N=8 for all treatment groups. 2-way RM
ANOVA (FIG. 22A) or Friedman's test with Dunn's post-hoc (FIG.
22B).
[0059] FIG. 23 shows the bradykinesia effect of acute DPI-289
administered in combination with low and high dose of L-DOPA in the
same animals as displayed in FIGS. 17 & 18. Data are
mean.+-.s.e.m. (FIG. 23A shows time-course) or cumulated for 0-6 h
shown in FIG. 23B. N=8 for all treatment groups. 2-way RM ANOVA
(FIG. 23A) or Friedman's test with Dunn's post-hoc (FIG. 23B).
[0060] FIG. 24 shows the total ON-time effect of acute DPI-289
administered in combination with low and high dose of L-DOPA. FIG.
24B shows the total ON-time, FIG. 24 shows the Good ON-time and
FIG. 24C shows the BAD ON-time.
[0061] FIG. 25 shows the increase in contralateral and ipsilateral
rotations by DPI-289 in 6-OHDA-lesioned rats. N=6 rats per group,
Newman-Keuls post hoc test was used for statistical
significance.
[0062] FIG. 26 shows augmentation of L-DOPA-induced contralateral
rotations by coadministration of DPI-289 in 6-OHDA-lesioned Rats.
N=8 per group, Newman-Keuls post hoc test for statistical
significance.
[0063] FIG. 27A shows reduction in postural asymmetry of unilateral
6-OHDA-lesioned Rats by DPI-289 or L-DOPA. FIG. 27B shows the
absence of dyskinesia following repeated administration of DPI-289
to unilateral 6-OHDA-lesioned Rats. N=8 rats per group. Statistical
significance was assessed by a two-way ANOVA and Holm-Sidak's
multiple comparisons test.
DETAILED DESCRIPTION OF THE INVENTION
[0064] In one broad method aspect of the present invention, a
diarylmethylpiperazine compound that exhibits delta opioid receptor
agonist activity and also mu opioid receptor antagonist activity as
hereinafter more fully described is administered to a subject in
need of treatment for Parkinson's disease.
[0065] The invention broadly contemplates the treatment of
Parkinson's disease by using a mono-therapy treatment, involving
compounds of the invention as singular therapeutic agents in
administered therapeutic compositions, or co-therapy treatment,
wherein a compound in accordance with the present invention is
administered contemporaneously, e.g., simultaneously, or
sequentially, with another therapeutic Parkinson agent.
[0066] In a particularly preferred method aspect of the invention,
Parkinson's disease is treated by administering to a subject in
need of such treatment an effective amount of a compound of Formula
(i) (DPI-289) or a pharmaceutically acceptable ester or salt
thereof.
[0067] Examples of pharmaceutically acceptable esters of compounds
of formula (i) include carboxylic acid esters of the hydroxyl group
in the compounds of Formula (i) (DPI-289) where OH on any of the
rings in which the non-carbonyl moiety of the carboxylic acid
portion of the ester grouping is selected from straight or branched
chain alkyl (e.g. n-propyl, t-butyl, n-butyl), alkoxyalkyl (e.g.
methoxymethyl), arylalkyl (e.g. benzyl), aryloxyalky (e.g.
phenoxymethyl), and aryl (e.g. phenyl); alkyl-, aryl-, or
arylalkylsulfonyl (e.g. methanesulfonyl); amino acid esters (e.g.
L-valyl or L-isoleucyl); dicarboxylic acid esters (e.g.
hemisuccinate); carbonate esters (e.g. ethoxycarbonyl); carbamate
esters (e.g. dimethylaminocarbonyl, (2-aminoethyl)aminocarbonyl);
and inorganic esters (e.g. mono-, di- or triphosphate). However,
esters that are not pharmaceutically acceptable may also find use,
for example, in the preparation or purification of a
pharmaceutically acceptable compound. All salts, whether or not
derived from a pharmaceutically acceptable acidic moiety, are
within the scope of the present invention.
[0068] Examples of pharmaceutically acceptable salts of the
compounds of Formula (i) include salts derived from an appropriate
base, such as an alkali metal (for example, sodium, potassium), an
alkaline earth metal (for example, calcium, magnesium), ammonium
and NR'.sub.4.sup.+ (wherein R' is C.sub.1-C.sub.4 alkyl).
Pharmaceutically acceptable salts of an amino group include salts
of: organic carboxylic acids such as acetic, lactic, tartaric,
malic, lactobionic, fumaric, and succinic acids; organic sulfonic
acids such as methanesulfonic, ethanesulfonic, isethionic,
benzenesulfonic and p-toluenesulfonic acids; and inorganic acids
such as hydrochloric, hydrobromic, sulfuric, phosphoric and
sulfamic acids. Pharmaceutically acceptable salts of a compound
having a hydroxyl group consist of the anion of said compound in
combination with a suitable cation such as Na.sup.+,
NH.sub.4.sup.+, or NR'.sub.4.sup.+ (wherein R' is for example a
C.sub.1-4 alkyl group).
[0069] For therapeutic use, salts of the compound of Formula (i)
will be pharmaceutically acceptable, i.e., they will be salts
derived from a pharmaceutically acceptable acid or base. However,
salts of acids or bases that are not pharmaceutically acceptable
may also find use, for example, in the preparation or purification
of a pharmaceutically acceptable compound. All esters, whether or
not derived from a pharmaceutically acceptable acid or base, are
within the scope of the present invention.
[0070] The present invention also contemplates pharmaceutical
formulations, both for veterinary and for human medical use, which
comprise as the active agent one or more compound(s) of the
invention.
[0071] In such pharmaceutical formulations, the
diarylmethylpiperazine compound preferably is utilized together
with one or more pharmaceutically acceptable carrier(s) therefore
and optionally any other therapeutic ingredients. The carrier(s)
must be pharmaceutically acceptable in the sense of being
compatible with the other ingredients of the formulation and not
unduly deleterious to the recipient thereof. The active agent is
provided in an amount effective to achieve the desired
pharmacological effect, as described above, and in a quantity
appropriate to achieve the desired daily dose.
[0072] The formulations include those suitable for parenteral as
well as non-parenteral administration, and specific administration
modalities include oral, rectal, topical, sub-lingual, mucosal,
nasal, optic, ophthalmic, subcutaneous, intramuscular, intravenous,
transdermal, spinal, intrathecal, intracranial, intra-articular,
intra-arterial, sub-arachnoid, bronchial, lymphatic, vaginal and
intra-uterine administration.
[0073] When the active agent of the present invention is utilized
in a formulation comprising a liquid solution, the formulation
advantageously may be administered parenterally. When the active
Parkinson agent of the present invention is employed in a liquid
suspension formulation or as a powder in a biocompatible carrier
formulation, the formulation may be advantageously administered
orally, rectally, or bronchially.
[0074] When the active Parkinson agent of the present invention is
utilized directly in the form of a powdered solid, the active agent
may advantageously be administered orally. Alternatively, it may be
administered bronchially, via nebulization of the powder in a
carrier gas, to form a gaseous dispersion of the powder that is
inspired by the patient from a breathing circuit comprising a
suitable nebulizer device.
[0075] In some applications, it may be advantageous to utilize the
active Parkinson agent of the present invention in a "vectorized"
form, such as by encapsulation of the active agent in a liposome or
other encapsulant medium, or by fixation of the active Parkinson
agent of the present invention, e.g., by covalent bonding,
chelation, or associative coordination, on a suitable biomolecule,
such as those selected from proteins, lipoproteins, glycoproteins,
and polysaccharides.
[0076] The formulations comprising the active Parkinson agent of
the present invention may conveniently be presented in unit dosage
forms and may be prepared by any of the methods well known in the
art of pharmacy. Such methods generally include the step of
bringing the active Parkinson compound(s) of the present invention
into association with a carrier that constitutes one or more
accessory ingredients. Typically, the formulations are prepared by
uniformly and intimately bringing the active compound(s) into
association with a liquid carrier, a finely divided solid carrier,
or both, and then, if necessary, shaping the product into dosage
forms of the desired formulation.
[0077] Formulations of the present invention suitable for oral
administration may be presented as discrete units such as capsules,
cachets, tablets, or lozenges, each containing a predetermined
amount of the active ingredient as a powder or granules; or a
suspension in an aqueous liquor or a non-aqueous liquid, such as a
syrup, an elixir, an emulsion, or a draught.
[0078] A tablet may be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets may be
prepared by compressing in a suitable machine, with the active
compound being in a free-flowing form such as a powder or granules
which optionally is mixed with a binder, disintegrant, lubricant,
inert diluent, surface active agent, or discharging agent. Molded
tablets comprised of a mixture of the powdered active compound with
a suitable carrier may be made by molding in a suitable
machine.
[0079] A syrup may be made by adding the active Parkinson agent of
the present invention to a concentrated aqueous solution of a
sugar, for example sucrose, to which may also be added any
accessory ingredient(s). Such accessory ingredient(s) may include
flavorings, suitable preservative, agents to retard crystallization
of the sugar, and agents to increase the solubility of any other
ingredient, such as a polyhydroxy alcohol, for example glycerol or
sorbitol.
[0080] Formulations suitable for parenteral administration
conveniently comprise a sterile aqueous preparation of the active
compound, which preferably is isotonic with the blood of the
recipient (e.g., physiological saline solution). Such formulations
may include suspending agents and thickening agents and liposomes
or other microparticulate systems which are designed to target the
compound to blood components or one or more organs. The
formulations may be presented in unit-dose or multi-dose form.
[0081] Nasal spray formulations comprise purified aqueous solutions
of the active Parkinson agent of the present invention with
preservative agents and isotonic agents. Such formulations are
preferably adjusted to a pH and isotonic state compatible with the
nasal mucous membranes.
[0082] Formulations for rectal or vaginal administration may be
presented as a suppository or pessary with a suitable carrier such
as cocoa butter, hydrogenated fats, or hydrogenated fatty
carboxylic acids.
[0083] Ophthalmic formulations are prepared by a similar method to
the nasal spray, except that the pH and isotonic factors are
preferably adjusted to match that of the eye.
[0084] Topical formulations comprise the active Parkinson agent of
the present invention dissolved or suspended in one or more media,
such as mineral oil, petroleum, polyhydroxy alcohols, or other
bases used for topical pharmaceutical formulations.
[0085] The compounds of the invention may also be delivered through
the skin or muscosal tissue using conventional transdermal drug
delivery systems, i.e., transdermal "patches" wherein a composition
of the present invention is typically contained within a laminated
structure that serves as a drug delivery device to be affixed to
the body surface. In such a structure, the pharmaceutical
composition is typically contained in a layer, or "reservoir,"
underlying an upper backing layer. The laminated device may contain
a single reservoir, or it may contain multiple reservoirs. In one
embodiment, the reservoir comprises a polymeric matrix of a
pharmaceutically acceptable contact adhesive material that serves
to affix the system to the skin during drug delivery. Examples of
suitable skin contact adhesive materials include, but are not
limited to, polyethylenes, polysiloxanes, polyisobutylenes,
polyacrylates, polyurethanes, and the like. Alternatively, the
active agent-containing reservoir and skin contact adhesive are
present as separate and distinct layers, with the adhesive
underlying the reservoir which, in this case, may be either a
polymeric matrix as described above, or it may be a liquid or gel
reservoir, or may take some other form. The backing layer in these
laminates, which serves as the upper surface of the device,
functions as the primary structural element of the laminated
structure and provides the device with much of its flexibility. The
material selected for the backing layer should be substantially
impermeable to the active agent and any other materials that are
present.
[0086] Transdermal formulations may be prepared by incorporating
the active Parkinson agent of the present invention in a
thixotropic or gelatinous carrier such as a cellulosic medium,
e.g., methyl cellulose or hydroxyethyl cellulose, with the
resulting formulation then being packed in a transdermal device
adapted to be secured in dermal contact with the skin of a wearer.
The transdermal formulation would optimally be comprised of a
backing, a reservoir layer containing the composition, and an
adhesive. The transdermal system would be optimized to allow
prolonged wear (up to 7 days) and consistent composition delivery
through the skin. The excipients necessary in the transdermal
system would support composition stability. The adhesive would be
permeable to the compound and to water to avoid occlusion damage to
the skin. The backing layer would also be permeable to water vapor,
but not wettable, to both avoid occlusion damage to the skin and to
support prolonged wear.
[0087] In addition to the aforementioned ingredients, formulations
of this invention may further include one or more accessory
ingredient(s) selected from diluents, buffers, flavoring agents,
binders, disintegrants, surface active agents, thickeners,
lubricants, preservatives (including antioxidants), and the
like.
[0088] Depending on the specific condition to be treated, animal
subjects may be administered compounds of the present invention at
any suitable therapeutically effective and safe dosage, as may
readily be determined within the skill of the art, and without
undue experimentation.
[0089] In general, while the effective dosage of Formula (i)
(DPI-289) of the invention for therapeutic use may be widely varied
in the broad practice of the invention, depending on the specific
condition involved, the delivery modes that being either oral,
transdermal or other modes, the stage of the individual's disease
and the duration and dosage of the individual's prior exposure to
L-DOPA as readily determinable within the skill of the art,
suitable therapeutic doses of the compounds of the invention, for
each of the appertaining compositions described herein, and for
achievement of therapeutic benefit in treatment of each of the
conditions described herein, will be in the range of or
approximately 50 micrograms (.mu.g) to 500 milligrams (mg) per
kilogram body weight of the recipient per day, preferably in sub
doses. The desired oral dose may be presented as one, two, three,
four, five, six, or more sub-doses administered at appropriate
intervals throughout the day.
[0090] The mode of administration and dosage forms will of course
affect the therapeutic amounts of the Parkinson agent of the
present invention which are desirable and efficacious for the given
treatment application.
[0091] In oral administration, dosage levels for compounds of the
present invention may be on the order of 1-100 mg/kg body
weight/day preferably 5-25 mg/kg body weight/day. In tablet dosage
forms, typical active agent dose levels are on the order of
100-1000 mg per tablet.
[0092] The following examples are illustrative of synthetic
procedures that may be advantageously utilized to synthesize the
compound of the present invention.
Example 1 (Formula (i) (DPI-289)
##STR00005##
[0093]
4-((alpha-R)-alpha-02S,5R)-2,5-Dimethyl-4-(3-fluorobenzyl)-1-pipera-
zinyl)-3-hydroxybenzyl)-N,N-diethylbenzamide
[0094] Method A--Alkylation with Phenol Protection
[0095] A solution of 3-bromophenol (400 g, 2.31 mol),
tert-butylchlorodimethylsilane (391 g, 2.54 mol), and imidazole
(346 g, 5.08 mol) in 5000 mL of dichloromethane was stirred
overnight at room temperature. The reaction solution was poured
into water (2000 mL) and the layers separated. The organic layer
was washed with 1N aqueous sodium hydroxide solution (3.times.1500
mL) and water (2.times.1500 mL) before passing through a pad of
silica gel (400 g, silica 60, 230-400 mesh). The silica gel was
washed with dichloromethane (2.times.500 mL), the filtrates
combined and the solvent removed under reduced pressure to give
3-(bromophenoxy)-tert-butyldimethylsilane (669 g, 98.4%) as a clear
pale yellow liquid. NMR (300 MHz, CDCl.sub.3): .delta. 0.2 (s, 6H);
1.0 (s, 9H); 6.75 (m, 1H); 7.0 (br s, 1H); 7.1 (m, 2H).
[0096] 3-tert-Butyldimethylsilyloxyphenylmagnesium bromide was
formed by the slow addition of a mixture of
3-bromophenoxy-tert-butyldimethylsilane (27.3 g, 92.6 mmol) and
dibromoethane (3.45 g, 18.4 mmol) in inhibitor-free anhydrous
tetrahydrofuran (100 mL) to a solution of magnesium turnings (3.57
g, 147 mmol) in inhibitor-free anhydrous tetrahydrofuran (200 mL)
at reflux. After stirring for one hour at reflux the light brown
clear mixture was cooled to room temperature.
[0097] 4-Carboxybenzaldehyde (100.3 g, 0.67 mol) was
dissolved/suspended in toluene (1200 mL), dimethylformamide (0.15
mL) was added and the suspension was stirred during the dropwise
addition of thionyl chloride (53.5 mL, 87.2 g, 0.73 mol). The
reaction mixture was heated to reflux under nitrogen and stirred
for 2 h, during which time much, but not all of the aldehydo-acid
passed into solution. A further quantity of thionyl chloride (20
mL, 32.6 g, 0.27 mol) was added and reflux continued overnight. The
clear reaction mixture was evaporated, and the residue dissolved in
anhydrous tetrahydrofuran (1500 mL). The solution was cooled in an
ice/water bath and diethylamine (173 mL, 122 g, 1.67 mol (2.5
equivalents)) was added dropwise to the stirred solution. The
ice-bath was removed and stirring continued for 2.5 h. The reaction
mixture was filtered to remove the white crystalline diethylamine
hydrochloride by-product. The crystals were washed with ethyl
acetate (2.times.600 mL), and the washings set aside. The
tetrahydrofuran filtrate was evaporated, and the residue dissolved
in the ethyl acetate washings. The solution was washed sequentially
with 1 M-hydrochloric acid (2.times.600 mL), water (2.times.300
mL), dilute sodium carbonate solution (saturated
Na.sub.2CO.sub.3:H.sub.2O, 1:1, 2.times.600 mL), water (2.times.300
mL) and saturated sodium chloride solution (300 mL). The organic
layer was separated, dried over anhydrous sodium sulfate and
evaporated to yield 4-formyl-N,N-diethylbenzamide as a pale brown
oil (115.7 g, 84%) which was used without further purification.
[0098] In a 1000 mL round bottom flask fitted with a condenser and
Dean-Stark trap were combined 4-formyl-N,N-diethylbenzamide (9.50
g, 46.3 mmol), benzotriazole (5.51 g, 46.3 mmol), and
(2R,5S)-1-allyl-2,5-dimethylpiperazine (7.15 g, 46.3 mmol,
Chirotech Division of Dowpharma, The Dow Chemical Company,
Cambridge, England) with toluene (400 mL). The reaction mixture was
heated to reflux under nitrogen until no additional water was
observed in the trap (ca. 2 hours). The reaction mixture was cooled
to room temperature and concentrated under vacuum to leave a volume
of approximately 50 mL. Anhydrous tetrahydrofuran (100 mL) was
added to the flask under nitrogen with stirring to dissolve all
residue. The solution of benzotriazole adduct was added to the
solution of 3-tert-butyldimethylsilyloxyphenylmagnesium bromide
(above) at room temperature via double-ended needle. After stirring
for 2 hours, the reaction was quenched by addition of saturated
aqueous ammonium chloride (20 mL). Anhydrous magnesium sulfate (20
g) was added and the reaction was filtered. Solvent was removed
under vacuum and the residue was redissolved in ethyl acetate (800
mL). The ethyl acetate solution was washed with 1 M sodium
hydroxide (4.times.200 mL), water (200 mL), and saturated aqueous
sodium chloride (200 mL). The organic layer was dried over
anhydrous magnesium sulfate and the solvent removed to give a dark
oil. The oil was dissolved in tetrahydrofuran (250 mL) and 3 M
hydrochloric acid (350 mL) and stirred for 2 hours at room
temperature. The reaction solution was extracted with a mixture of
diethyl ether/ethyl acetate (2:1, 3.times.250 mL). Ethyl acetate
(300 mL) was added to the aqueous layer and pH was adjusted to 8
with aqueous sodium hydroxide. Layers were separated and the
aqueous portion was extracted with another ethyl acetate
(3.times.300 mL). The combined organic extracts were washed with
saturated aqueous sodium chloride, dried over anhydrous sodium
sulfate, and the solvent removed under vacuum to give a brown
residue (12.4 g). The residue was purified by chromatography on
silica gel (300 g), eluting with a gradient of ethanol in
dichloromethane (1-15%), to give
4-((alpha-R)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-hy-
droxybenzyl)-N,N-diethylbenzamide as a colorless gum (5.54 g, 27%
from 4-formyl-N,N-diethylbenzamide).
[0099] 4-((alpha-R)-alpha-((2
S,5R)-4-Allyl-2,5-dimethyl-1-piperazinyl)-3-hydroxybenzyl)-N,N-diethylben-
zamide (4.87 g, 11.2 mmol) was dissolved in methylene chloride (60
mL) and triethylamine (5.15 mL, 3.73 g, 37 mmol) added. N-Phenyl
bis(trifluoromethanesulfonimide) (4.40 g, 12.3 mmol) was added and
the reaction mixture sealed under nitrogen and stirred at room
temperature overnight. The reaction mixture was evaporated to
dryness, the residue dissolved in ethyl acetate (100 mL), and the
solution extracted with aqueous sodium carbonate solution (5%,
2.times.75 mL). The organic layer was separated, dried over
anhydrous sodium sulfate and evaporated to yield a viscous amber
oil. The residue was dissolved in methylene chloride (30 mL),
applied to a column of silica gel (1000 g), and eluted with
ethanol/methylene chloride (2:98 v/v). Pure fractions containing
desired product, as evidenced by t.l.c. (silica gel, EM60F.sub.254,
2% NH.sub.4OH in ethyl acetate, R.sub.f=0.78) were evaporated to
dryness to yield 4-((alpha-R)-alpha-((2
S,5R)-4-Allyl-2,5-dimethyl-1-piperazinyl)-3-trifluoromethyl
sulfonyloxybenzyl)-N,N-diethylbenzamide (4.03 g) as a yellow/amber
oil. .sup.1H NMR (CDCl.sub.3, 500 MHz); .delta. 1.00 (d, J=6.2 Hz,
3H); 1.12 (br m, 3H); 1.21 (d, J=6.1 Hz, 3H); 1.25 (br m, 3H); 1.83
(t, J=10.6 Hz, 1H); 2.60 (m, 3H); 2.91 (dd J=11.4, 2.7, 1H); 3.02
(m, 1H); 3.18 (br s, 2H); 3.28 (br m, 2H); 3.46 (dd, J=13.7, 5.5
Hz, 1H); 3.55 (br m, 2H); 5.25 (m, 2H); 5.31 (s, 1H); 5.88 (m, 1H);
7.02 (d, J=7.7 Hz, 1H); 7.05 (s, 1H); 7.23 (m, 2H); 7.32 (d, J=8.1
Hz, 2H); 7.40 (d, J=8.1 Hz, 2H); 7.46 (t, J=7.9 Hz, 1H).
[0100] A solution of
4-((alpha-R)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-trifluo-
romethylsulfonyloxybenzyl)-N,N-diethylbenzamide (4.03 g, 7.20 mmol)
and thiosalicylic acid (1.32 g, 8.52 mmol) in anhydrous
tetrahydrofuran (25 mL) was stirred under nitrogen for 3 h at room
temperature with a catalyst solution prepared by dissolution of
bis(dibenzylidineacetone)palladium (204 mg, 0.355 mmol) and
1,4-bis(diphenylphosphino)butane (151 mg, 0.355 mmol) in
tetrahydrofuran (3 mL). The reaction mixture was evaporated to
dryness, the residue dissolved in a mixture of ethyl acetate/ether
(1:3, 125 mL) and extracted with aqueous sodium carbonate solution
(5%, 2.times.75 mL). The organic layer was diluted with two volumes
of pentane and extracted with 3M-hydrochloric acid (5.times.25 mL).
The aqueous solution was adjusted to pH 9-10 with concentrated
ammonia solution and extracted with methylene chloride (3.times.50
mL). The combined organic extracts were dried over anhydrous sodium
sulfate and evaporated to yield
4-((alpha-R)-alpha-((2S,5R)-2,5-dimethyl-1-piperazinyl)-3-trifluoromethyl-
-sulfonyloxybenzyl) --N,N-diethylbenzamide as a brittle pale yellow
foam (3.53 g). The product showed a single spot on thin layer
chromatography (silica gel, EM60F.sub.264, 2% NH.sub.4OH in ethyl
acetate, R.sub.f=0.33). .sup.1H NMR (CDCl.sub.3, 500 MHz); .delta.
0.95 (d, J=6 Hz, 3H); 1.13 (br m, 3H); 1.20 (d, J=6.1 Hz, 3H); 1.26
(br m, 3H); 1.50 (t, J=9.7 Hz, 1H); 2.31 (m, 1H); 2.64 (dd J=11.3,
2.5, 1H); 2.71 (m, 1H); 2.95 (m, 1H); 3.29 (br m, 2H); 3.56 (br m,
2H); 5.43 (s, 1H); 7.04 (m, 1H); 7.21 (d, J=7.7, 1H); 7.24 (dd,
J=8.2, 2.2 Hz, 1H); 7.34 (d, J=8.2 Hz, 2H); 7.42 (d, J=8.1 Hz, 2H);
7.48 (t, J=8 Hz, 1H).
[0101] A solution of
4-((alpha-R)-alpha-((2S,5R)-2,5-dimethyl-1-piperazinyl)-3-trifluoromethyl-
-sulfonyloxybenzyl)-N,N-diethylbenzamide (3.522 g, 6.0 mmol) and
sodium iodide (90 mg, 0.6 mmol) in acetonitrile (30 mL) was stirred
during the addition of triethylamine (3.0 mL, 2.186 g, 21.6 mmol)
followed by 3-fluorobenzyl bromide (1.472 mL, 2.268 g, 12.0 mmol).
An immediate turbidity was observed, thickening to a white
crystalline precipitate as the reaction progressed. The reaction
mixture was sealed under nitrogen and stirred at room temperature.
After 18 h the solvent was removed by evaporation under reduced
pressure and the residue partitioned between ethyl acetate (30 mL)
and saturated sodium bicarbonate solution (10 mL). The organic
layer was separated and the aqueous portion further extracted with
ethyl acetate (3.times.15 mL). The combined organic extract and
washings were dried over sodium sulfate, the solution evaporated to
dryness and re-dissolved in ethyl acetate (.about.5 mL). The
solution was applied to an intermediate (4.times.15 cm) Biotage
column and eluted with ethyl acetate, collecting fractions of 20
mL. Fractions containing pure material as evidenced by thin layer
chromatography (silica, EM60F.sub.254, developed with ethyl
acetate, R.sub.f 0.9) were pooled and evaporated to yield a
yellow/orange oil (3.01 g). The oil was dissolved in ethanol (30
mL) and aqueous sodium hydroxide solution (10.0 mL, 2.5-M, 25 mmol)
was added. The initially cloudy suspension clarified to a yellow
solution that was set aside at room temperature for 3 h. The
mixture was evaporated under reduced pressure to remove ethanol,
and evaporation continued until condensation of water indicated
complete removal of ethanol. The cloudy suspension of the oily
sodium salt of the phenol was diluted to 20 mL with water to yield
a clear yellow solution. The pH of the strongly basic solution was
adjusted to 8.5-9 by passage of carbon dioxide gas (from dry ice)
to yield a dense white flocculent precipitate. The solid was
removed by filtration and washed thoroughly with cold water,
including twice re-slurrying of the precipitate on the sinter with
fresh water. The solid was air-dried on the sinter overnight, then
dried under vacuum at 1 mm Hg at room temperature to yield
4-((alpha-R)-alpha-((2S,5R)-2,5-dimethyl-4-(3-fluorobenzyl)-1-piperazinyl-
)-3-hydroxybenzyl)-N,N-diethylbenzamide as a white solid (2.062 g,
67%) Calc. for C.sub.31H.sub.38FN.sub.3O.sub.2 0.5 H.sub.2O: C,
72.63; H, 7.67; N, 8.20; F, 3.71. Found C, 72.77; H, 7.52; N, 8.18;
F, 3.61%. .sup.1H NMR (CDCl3, 300 MHz); .delta. 1.05 (d, J=5.9 Hz,
6H); 1.11 (br m, 3H); 1.23 (br m, 3H); 2.00 (m, 2H); 2.59 (br m,
2H); 2.62 (d, J=11.4 Hz, 1H); 2.68 (d, J=11.0 Hz, 1H); 3.19 (d,
J=13.6 Hz, 1H); 3.28 (br m, 2H); 3.54 (br m, 2H); 3.89 (d, J=13.9
Hz, 1H); 5.01 (s, 1H); 6.15 (v br s, 1H); 6.63 (s, 1H); 6.70 (m,
2H); 6.91 (t, J=8.8 Hz, 1H); 7.07 (m, 2H); 7.14 (t, J=7.8 Hz, 1H);
7.22 (m, 1H); 7.28 (d, J=8.2 Hz, 2H); 7.44 (d, J=8.1 Hz, 2H).
[0102] Method B--Reductive Alkylation
[0103] A solution of
4-[(R)-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)-met-
hyl]-N,N-diethylbenzamide (10.89 g, 25 mmol, from Method A) and
thiosalicylic acid (4.63 g, 30 mmol) in anhydrous tetrahydrofuran
(50 mL) was stirred with a catalyst solution prepared by
dissolution of bis(dibenzylidineacetone)palladium (0.718 g, 1.25
mmol) and 1,4-bis(diphenylphosphino)butane (0.533 g, 1.25 mmol) in
tetrahydrofuran (10 mL) at room temperature under nitrogen for 1.5
hours (J. P. Genet, S. Lemaire-Audoire, M. Savignac, Tetrahedron
Letters, 36, 1267-1270 (1995)). The reaction mixture was
concentrated under reduced pressure and the residue was partitioned
between ethyl acetate (150 mL) and aqueous sodium carbonate
solution. The layers were separated and diethyl ether (250 mL) was
added to the organic layer. This was extracted with 5% sodium
carbonate solution (2.times.150 mL). The organic layer was diluted
with pentane (500 mL) and extracted with 3 M hydrochloric acid
(6.times.30 mL). The aqueous solution was adjusted to pH 9-10 with
saturated aqueous sodium carbonate solution and extracted with
methylene chloride (3.times.100 mL). The combined organic extracts
were dried over anhydrous sodium sulfate and the solvent was
removed under reduced pressure to yield 4-[(R)-((2
S,5R)-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)methyl]-N,N-diethylben-
zamide as a brittle pale yellow foam (10.24 g). The product showed
a single peak on HPLC (Zorbax C-8, isocratic 40% 0.01 M NH.sub.4OAc
in MeOH, 3 min; linear gradient to 100% MeOH, 45 min; isocratic
MeOH, 5 min; 1.0 mL/min; .delta..sub.obs=210 nm, Rt=19.24 min).
Calc. for C.sub.24H.sub.33N.sub.3O.sub.2.0.1 EtOAc.0.4
CH.sub.2Cl.sub.2: % C, 67.96; H, 7.96; N, 9.59. Found: % C, 67.90;
H, 8.03; N, 9.54. .sup.1H NMR (CDCl.sub.3, 300 MHz): .delta.7.42
(d, J=8.1 Hz, 2H); 7.26 (d, J=8.3 Hz, 2H); 7.11 (t, J=7.8 Hz, 1H);
6.72 (d, J=8.1 Hz, 1H); 6.65 (s, 1H); 6.59 (d, J=7.6 Hz, 1H); 5.16
(s, 1H); 4.93 (v br s, 2H); 3.51 (br m, 2H); 3.27 (br m, 2H);
3.02-2.97 (m, 1H); 2.92 (d, J=10.5 Hz, 1H); 2.66 (br d, J=8.5 Hz,
2H); 2.60-2.45 (m, 1H); 1.84 (dd, J=11.3, 8.3 Hz, 1H); 1.27-1.15
(m, 3H); 1.10 (d, J=6.1 Hz, 3H overlapping with m, 3H); 1.02 (d,
J=6.1 Hz, 3H).
[0104] Glacial acetic acid (0.635 mL, 11.1 mmol) was added to a
solution of
4-[(R)-((2S,5R)-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)methyl]-N-
,N-diethylbenzamide (1.98 g, 5 mmol) and 3-fluorobenzaldehyde (1.24
g, 10 mmol) in anhydrous tetrahydrofuran (35 mL). While stirring
briskly, sodium triacetoxyborohydride (2.12 g, 10 mmol) was added
in 50-100 mg portions, allowing effervescence to subside after each
addition. The reaction was monitored for absence of starting
material by HPLC. After stirring at room temperature for 16 hours,
additional 3-fluorobenzaldehyde (0.62 g, 5 mmol), acetic acid
(0.318 mL, 5 mmol), and sodium triacetoxyborohydride (1.06 g, 5
mmol) were added. After stirring an additional 4 hours, the
reaction mixture was concentrated under reduced pressure and the
residue was partitioned between ethyl acetate (50 mL) and 3 M
hydrochloric acid (25 mL). The layers were separated and the
organic layer was extracted again with 3 M hydrochloric acid (25
mL). The aqueous solution was adjusted to pH 9-10 with 5 M sodium
hydroxide solution, resulting in the formation of chunky white
precipitate. Brief sonication followed by continuous stirring for
60 hours allowed hydrolysis of any residual ethyl acetate, as well
as complete precipitation of product. The white solid was filtered,
washed with cold water, and further dried under reduced pressure to
yield
4-[(R)-((2S,5R)-2,5-dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)(3-hydroxyp-
henyl)-methyl]-N,N-diethylbenzamide (1.78 g, 82.3% yield from
4-[(R)-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)-met-
hyl]-N,N-diethyl-benzamide). The product showed a single peak on
HPLC (Zorbax C-8, isocratic 40% 0.01 M NH.sub.4OAc in MeOH, 3 min;
linear gradient to 100% MeOH, 45 min; isocratic MeOH, 25 min; 1.0
mL/min; .delta..sub.obs=210 nm, Rt=43.51 min). Calc. for
C.sub.31H.sub.38FN.sub.3O.sub.2.0.5 H.sub.2O: % C, 72.63; H, 7.67;
N, 8.20; F, 3.71. Found: % C, 72.77; H, 7.52; N, 8.18; F, 3.61.
[0105] Method C--Direct Alkylation
[0106] To
4-[(R)-((2S,5R)-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)
methyl]-N,N-diethyl-benzamide (0.79 g, 2 mmol) and sodium iodide
(30 mg, 0.2 mmol) in acetonitrile (10 mL) was added triethylamine
(1.0 mL, 7.2 mmol), followed by 3-fluorobenzylbromide (0.76 g, 4
mmol). The reaction mixture was sealed under nitrogen and stirred
at ambient temperature for 20 hours. The reaction mixture was
concentrated under reduced pressure and the residue was partitioned
between ethyl acetate (150 mL) and saturated sodium carbonate
solution (15 mL) diluted with water (50 mL). The organic layer was
separated and the aqueous layer was extracted with ethyl acetate
(3.times.75 mL). The combined organic extracts were dried over
anhydrous sodium sulfate and magnesium sulfate. The solvent was
removed under reduced pressure to yield a light yellow solid (0.90
g). This solid was dissolved in isopropanol (30 mL) upon heating to
boiling, and water (20 mL) was added slowly while swirling and
keeping the solution hot. Crystallization occurs as solution begins
to cool. Crystallization was allowed to proceed overnight. Crystals
were collected and washed sparingly with isopropanol:water/1:1.
Drying under 5 mm Hg at 40.degree. C. for 48 hours yielded
4-[(R)-((2S,5R)-2,5-dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)(3-hydroxyp-
henyl)methyl]-N,N-diethyl-benzamide as old ivory colored crystals
(0.77 g, 76.4% from
4-[(R)-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-(3-hydroxyphenyl)-met-
hyl]-N,N-diethylbenzamide). Calc. for
C.sub.31H.sub.38FN.sub.3O.sub.2.0.25 H.sub.2O. 0.1 .sup.iPrOH: % C,
73.12; H, 7.70; N, 8.17; F, 3.69. Found: % C, 73.13; H, 7.73; N,
8.24; F, 3.53.
[0107] Method D--Assembly from
(2R,5S)-1-(3-fluorobenzyl)-2,5-dimethylpiperazine
[0108] In a 3 L round bottom flask fitted with a condenser and
Dean-Stark trap are combined 4-formyl-N, N-diethylbenzamide (20.53
g, 100 mmol), benzotriazole (11.91 g, 100 mmol), and
(2R,5S)-1-(3-flourobenzyl)-2,5-dimethylpiperazine (22.23 g, 100
mmol, Chirotech Division of Dowpharma, The Dow Chemical Company,
Cambridge, England) with 1000 mL of toluene. The reaction is heated
to reflux under nitrogen until no additional water is observed in
the trap (ca. 3 hours). The reaction is cooled to room temperature
and concentrated under vacuum to leave a volume of approximately
300 mL. Anhydrous tetrahydrofuran (500 mL) is added to the
benzotriazole adduct under nitrogen with stirring until dissolved.
To this solution is added to the solution of
3-tert-butyldimethylsilyloxyphenylmagnesium bromide (from Method A)
at room temperature via double-ended needle. After stirring for
approximately 2 hours, the reaction is quenched by the addition of
saturated aqueous ammonium chloride solution (50 mL) and stirred
for 15 minutes. Anhydrous magnesium sulfate (50 g) is added,
stirred for approximately 1 hour, and the reaction is filtered. The
solvent is removed under reduced pressure and the residue is
dissolved in ethyl acetate (1000 mL). The ethyl acetate solution is
washed with 1 M sodium hydroxide (5.times.400 mL), water
(4.times.400 mL), and saturated aqueous sodium chloride solution
(400 mL). The organic layer is dried over anhydrous magnesium
sulfate and the solvent is removed to give a viscous oil. The oil
is dissolved in 500 mL of tetrahydrofuran and 300 mL of 3 M
hydrochloric acid and stirred for approximately 1.5 hours at room
temperature. Upon completion of the desilylation, the reaction is
diluted with water (300 mL) and concentrated under vacuum to about
half the original volume. This solution is extracted with pentane
(2.times.500 mL). The aqueous layer is adjusted to pH 8-9 with 5 M
sodium hydroxide and extracted with ethyl acetate (250 mL). The
layers are separated and the aqueous portion is extracted with more
ethyl acetate (250 mL). The combined organic extracts are washed
with saturated aqueous sodium chloride solution, dried over
anhydrous sodium sulfate, and the solvent is removed under reduced
pressure to give a viscous oil. This residue is dissolved in ethyl
acetate (25 mL), seeded with crystals of the authentic compound,
and allowed to crystallize overnight (seed crystals can be obtained
from hot 2-propanol with addition of water). The crystals are
filtered and washed sparingly with cold ethyl acetate. Drying under
5 mm Hg at room temperature yields the desired product 4-[(R)-((2
S,5R)-4-(3-fluorobenzyl)-2,5-dimethyl-1-piperazinyl)-(3-hydroxy-phenyl)-m-
ethyl]-N,N-diethylbenzamide as tan to off-white crystals, free from
the undesired epimer. This solid is dissolved in isopropanol (300
mL) upon heating to boiling, and water (200 mL) is added slowly
while swirling and keeping the solution hot. Crystallization occurs
as solution begins to cool and is allowed to proceed overnight.
Crystals are collected and washed sparingly with cold
isopropanol:water/1:1. Drying under 5 mm Hg at 40.degree. C. for 48
hours yields
4-[(R)-((2S,5R)-2,5-dimethyl-4-(3-fluorobenzyl)-1-piperazinyl)(3-hydroxyp-
henyl)methyl]-N,N-diethyl-benzamide as ivory colored crystals
(expected yield .about.30% from 4-formyl-N,N-diethylbenzamide).
Example 2
[0109] Monotherapy with Formula (i) in L-DOPA Naive MPTP-Lesioned
Macaques and MPTP-Lesioned Macaques with Established Motor
Complications.
[0110] Eight cynomolgus monkeys (Macaca fascicularis, 6.4-6.8 years
of age, 2.8-4.2 kg were used in this study. Animals were housed two
or three per cage in the same housing room. The housing rooms were
subject to a 12 hour light-dark cycle (lights on 7 a.m.),
temperature 20-25.degree. C. in a room containing only animals of
the same sex. Fresh fruit, primate pellets and water were available
ad libitum other than at times of overnight fasting prior to PK or
behavioral assessment days.
[0111] The animals were rendered parkinsonian via once-daily
subcutaneous injection of MPTP (0.2 mg/kg), administered for
between 9 and 12 days, until the appearance of parkinsonism
symptoms. After this time, a parkinsonian syndrome reached a
moderate to marked level, over approximately 30 days, and
stabilized. The monkeys were allowed to recover for a minimum of
further 40 days until their parkinsonism was demonstrated as being
stable. L-DOPA naive animals have received no L-DOPA and those
showing established motor complication have received L-DOPA in an
amount 25 mg/kg orally twice daily for at least 2 months. L-DOPA is
given with the decarboxylase inhibitor benserazide (as
Madopar.TM.).
[0112] Activity
[0113] In L-DOPA naive MPTP-lesioned macaques Formula (i) resulted
in a significant increase in activity counts during the second hour
(10 and 20 mg/kg, by 62% and 63% respectively) and third hour (20
mg/kg only, by 82%) of observation (third and fourth hours after
administration) compared to that seen following vehicle treatment.
In L-DOPA naive MPTP-lesioned macaques, levels of activity observed
following treatment with vehicle averaged 2249.+-.658 counts over
the 0-6 h period of observation. Examining the whole 6 h
time-course period of observation revealed no significant effect of
Formula (i) treatment (F (3, 21)=0.6734, P=0.5779) or the
interaction of treatment and time (F (15, 105)=1.295, P=0.2184) but
did show an effect of time alone (F (5, 35)=7.928, P<0.0001) on
levels of activity (2-way, RM-ANOVA, FIG. 1A). Post-hoc
Holm-Sidak's analysis revealed a significant increase in activity
in the 1-2 h period following start of observations (2-3 h period
following treatment) with 10 and 20 mg/kg doses of Formula (i) (by
62% and 63%, P<0.05 and P<0.01 respectively cf. vehicle)
while in the 2-3 h period there was a significant increase in
activity levels (by 82%) in response to high-dose Formula (i) (20
mg/kg; P<0.05 cf. vehicle). Examining levels of average activity
cumulated over each of the three-hour periods 0-3 and 3-6 h after
start of observations revealed no significant effect of Formula (i)
treatment during either period (0-3 h; F (1.170, 8.189)=0.9758,
P=0.3672, 3-6 h; F (1.262, 8.836)=0.4690, P=0.5550, FIGS. 1B and
1C).
[0114] FIG. 2 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA and there is an increase in activity relative to the vehicle
in the first few hours which decreased in the 3 to 6 hours after
administration of the DPI-289. Clearly the oral administration of
single dose of 10 or 20 mg/kg of DPI 289 significantly alleviated
motor impairment for 1 to 4 hours. The differences in results of
FIGS. 1A and 2A are primarily due to differences in response to the
vehicle.
[0115] Total ON-Time
[0116] In L-DOPA naive MPTP-lesioned macaques Formula (i)
administration provided no significant increase in total duration
of ON-time at any dose assessed. (On-time is defined as the number
of minutes wherein no bradykinesia (slowness of movement) is
observed) L-DOPA naive MPTP-lesioned macaques treated with vehicle,
displayed a mean total duration of ON-time of 5.+-.4 min. There was
no significant effect of DPI-289 treatment on total ON-time between
the 1 and 10 mg/kg; 1 mg/kg; 46.+-.48 min, 10 mg/kg; 49.+-.48 min,
20 mg/kg; 1.+-.1 min (F (1.002, 7.016)=1.106, P=0.3281, one-way,
RM-ANOVA, FIG. 3). FIG. 4 shows the results of administration of
DPI-289 in macaques with established motor complications due to
administered L-DOPA and with only limited on-time at a dose of 20
mg/kg of DPI-289. ON-time represents lack of bradykinesia so mere
improvement in bradykinesia would not be reflected in changes in
ON-time. The oral bioavailability of the drug in these animals was
only 1-2% so the lack of effect on ON-time is likely due to
inadequate systemic exposure levels. Higher systemic levels of the
drug may have a more profound effect on bradykinesia and which
could translate into changes in ON-time.
[0117] Parkinsonian Disability
[0118] In L-DOPA naive MPTP-lesioned macaques Formula (i) (10
mg/kg) provided a modest but significant reduction in parkinsonian
disability (to absent-mild levels) during the second hour of
observation (third hour after administration) compared to vehicle
treatment (mild-moderate levels). In L-DOPA naive MPTP-lesioned
macaques, levels of parkinsonian disability observed following
treatment with vehicle were of mild to moderate levels during the
0-6 h period of observation. Examining the whole 6 h time-course
period of observation revealed no significant effect of DPI-289
treatment (F (3, 28)=1.004, P=0.4057) or time (F (5,
140)=0.0,P>0.9999) but did show a significant effect of the
interaction of the two (F (15, 140)=2.062, P=0.0151) on levels of
disability (2-way, RM-ANOVA, FIG. 5A). Post-hoc Holm-Sidak's
analysis revealed a significant decrease in disability to
absent-mild levels in the 1-2 h period of observation (2-3 h
following treatment with DPI-289, 10 mg/kg only, P<0.05 cf.
vehicle; mild-moderate levels). Examining levels of average
parkinsonism cumulated over each of the three-hour periods 0-3 and
3-6 h after start of observations revealed no significant effect of
DPI-289 treatment during either period (0-3 h; Friedman Statistic
(FS)=5.582, P=0.1338, 3-6 h; FS=1.71, P=0.635, Friedman test, FIGS.
5B and 5C).
[0119] FIG. 6 A shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
of L-DOPA with a significant reduction in parkinsonian disability
at a dose of 20 mg of/kg of DPI-289 in two to three hours after
dosing with DPI-289 (FIG. 6B). Overall, there was a dose effect of
DPI 289 on parkinsonian disability and a single dose of 20 mg/kg
had an effect for about 4 hours.
[0120] Bradykinesia
[0121] In L-DOPA naive MPTP-lesioned macaques Formula (i) (10
mg/kg) provided a significant reduction in levels of bradykinesia
(to mild-moderate levels) during the first four hours of
observation (1-5 hours after administration) compared to vehicle
treatment (moderate-marked levels). In L-DOPA naive MPTP-lesioned
macaques, levels of bradykinesia observed following treatment with
vehicle were of moderate to marked levels during the 0-6 h period
of observation. Examining the whole 6 h time-course period of
observation revealed a significant effect of Formula (i) treatment
(F (3, 28)=3.029, P=0.0459) and the interaction of time and
treatment (F (15, 140)=2.236, P=0.0077) but not time alone (F (5,
140)=0.0,P>0.9999) on levels of bradykinesia (2-way, RM-ANOVA,
FIG. 7A). Post-hoc Holm-Sidak's analysis revealed a significant
decrease in bradykinesia to mild-moderate levels in the 0-1, 1-2,
2-3 and 3-4 h periods of observation (1-2, 2-3, 3-4 and 4-5 h
following treatment with DPI-289; 10 mg/kg only, all P<0.05 cf.
vehicle). Examining levels of average bradykinesia cumulated over
the first three-hour period (0-3 h) after start of observations
revealed a significant effect of DPI-289 treatment (0-3 h; Friedman
Statistic (FS)=8.681, P=0.034, Friedman test, FIG. 7B). However,
post-hoc Dunn's analysis was unable to distinguish any difference
between the four treatment groups (all P>0.05). During the 3-6 h
period after start of observations there were no significant
effects of DPI-289 treatment (3-6 h; Friedman Statistic (FS)=4.385,
P=0.2228, Friedman test, FIG. 7C).
[0122] FIG. 8 shows the results of administration of DPI-289 in
macaques with established motor complications (bradykinesia) due to
administered L-DOPA with a significant change only in the first
hour at a dose of 20 mg of/kg of DPI-289 as shown in FIG. 8A.
[0123] Range of Movement
[0124] In L-DOPA naive MPTP-lesioned macaques Formula (i)
administration resulted in no reduction in levels of impaired range
of movement at any dose assessed. Equally, at no time during the
study did treatment with Formula (i) decrease range of movement. In
L-DOPA naive MPTP-lesioned macaques, levels of range of movement
observed following treatment with vehicle were typically of mild to
moderate levels during the 0-6 h period of observation. Examining
the whole 6 h time-course period of observation revealed no
significant effect of Formula (i) treatment (F (3, 28)=0.6160,
P=0.6104), time (F (5, 140)=0.0, P>0.9999) or the interaction of
the two (F (15, 140)=1.215, P=0.2670) on levels of range of
movement (2-way, RM-ANOVA, FIG. 9A). Examining average levels of
range of movement cumulated over each of the three-hour periods 0-3
and 3-6 h after start of observations revealed no significant
effect of DPI-289 treatment during either period (0-3 h; Friedman
Statistic (FS)=4.13, P=0.248, 3-6 h; FS=1.18, P=0.759, Friedman
test, FIGS. 9B and 9C).
[0125] FIG. 10 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA with an increase in the range of movement three to four
hours after a dose of 1, 10 or 20 mg of/kg of DPI-289 shown in FIG.
10A.
[0126] Postural Impairment
[0127] In L-DOPA naive MPTP-lesioned macaques Formula (i) (10
mg/kg) provided a significant reduction in postural impairment (to
absent-mild levels) during the second hour of observation (third
hour following administration) compared to vehicle treatment
(mild-moderate levels). In L-DOPA naive MPTP-lesioned macaques,
levels of postural impairment observed following treatment with
vehicle were of moderate to marked levels during the 0-6 h period
of observation. Examining the whole 6 h time-course period of
observation revealed a significant effect of Formula (i) treatment
(F (3, 28)=3.733, P=0.0225) and the interaction of time and
treatment (F (15, 140)=1.776, P=0.0437) but not time alone (F (5,
140)=0.0,P>0.9999) on levels of postural impairment (2-way,
RM-ANOVA, FIG. 11A). Post-hoc Holm-Sidak's analysis revealed a
significant decrease in postural impairment to absent-mild levels
in the 1-2 h period of observation (2-3 h following treatment with
DPI-289; 10 mg/kg only, P<0.01 cf. vehicle).
[0128] Examining levels of average postural impairment cumulated
over the first three-hour period (0-3 h) after start of
observations revealed a significant effect of Formula (i) treatment
(0-3 h; Friedman Statistic (FS)=8.87, P=0.031, Friedman test, FIG.
11B). However, post-hoc Dunn's analysis was unable to distinguish
any difference between the four treatment groups all P>0.05).
During the 3-6 h after start of observations there were no
significant effects of Formula (i) treatment (3-6 h; Friedman
Statistic (FS)=1.350, P=0.717, Friedman test, FIG. 11C.
[0129] FIG. 12 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA with a significant reduction in postural impairment at a
dose of 1, 10 and 20 mg of/kg of DPI-289 in two to four hours after
dosing with DPI-289 shown in FIG. 12A.
[0130] Alertness
[0131] In L-DOPA naive MPTP-lesioned macaques Formula (i)
administration resulted in no increase in levels of alertness at
any dose assessed. Equally, at no time during the study did
treatment with Formula (i) exacerbate any deficits in alertness. In
L-DOPA naive MPTP-lesioned macaques, average levels of alertness
observed following treatment with vehicle were of absent to mild
levels during the 0-6 h period of observation. Examining the whole
6 h time-course period of observation revealed no significant
effect of Formula (i) treatment (F (3, 28)=2.493, P=0.0806) or time
(F (5, 140)=0.0,P>0.9999) but did show a significant effect of
the interaction of the two (F (15, 140)=1.786, P=0.0422) on levels
of alertness (2-way, RM-ANOVA, FIG. 13A). Examining average levels
of alertness cumulated over each of the three-hour periods 0-3 FIG.
13B and 3-6 h FIG. 13C after start of observations revealed no
significant effect of Formula (i) treatment during either period
(0-3 h; Friedman Statistic (FS)=5.5, P=0.1386, 3-6 h; FS=5.73,
P=0.1257, Friedman test, FIGS. 13B and 13C).
[0132] FIG. 14 shows the results of administration of DPI-289 in
macaques with established motor complications due to administered
L-DOPA with a significant reduction in deficits in alertness at a
dose of 10 or 20 mg of/kg of DPI-289 in two to four hours after
dosing with DPI-289 as shown in FIG. 14A.
[0133] Dyskinesia
[0134] In L-DOPA naive MPTP-lesioned macaques Formula (i)
administration did not evoke dyskinesia at any of the doses
assessed as shown in FIG. 15A. In L-DOPA naive MPTP-lesioned
macaques there was no dyskinesia evident following treatment with
vehicle. Examining the whole 6 h time-course period of observation
revealed no induction of dyskinesia in response to Formula (i)
treatment at any time-point. As all values were zero no statistical
analyses were undertaken (FIGS. 15 A-C). This is also true for the
results shown in FIG. 16.
Example 3
Behavioral Effect of Acute DPI-289 Administered in Combination with
Low and High Dose of L-DOPA
[0135] This study utilized female MPTP-lesioned cynomolgus macaques
that have received chronic repeat-treatment with L-DOPA and
manifested stable and reproducible dyskinesia, of choreic and
dystonic nature, in response to subsequent L-DOPA-treatments. Eight
cynomolgus monkeys (Macaca fascicularis, 7.6-13.1 years of age,
3.0-4.5 kg) were used in this study. In a prior experiment, for
each individual animal a low and high dose of L-DOPA was
established such that the low dose (LDl) provided modest
anti-parkinsonian action (1-2 h) and elicited predominantly
non-disabling dyskinesia, the high dose (LDh) provided the best
anti-parkinsonian effect achievable but also elicited disabling
dyskinesia and an anti-parkinsonian benefit of 2-3 h duration
[0136] The behavioral effects of acute DPI-289 administered in
combination with low-dose L-DOPA (LDl) were assessed. Also higher
dose of L-DOPA was also assessed without the addition of DPI-289.
The low dose (LDl) was about 10 to 11 mg/kg of L-DOPA which is
considered to be sub-optimal in terms of anti-parkinsonian benefit.
The dose of DPI-289 was 20 mg/kg and the high dose of L-DOPA was
about 30 to 31 mg/kg and considered to an optimal does in terms of
anti-parkinsonian benefit but with disabling dyskinesia. The
DPI-289 was administered one (1) hour prior to the L-DOPA and start
of observation. Activity counts were collected and parkinsonian
disability and dyskinesia was assessed via post-hoc analysis of
video-recordings by a movement disorder neurologist blinded to
treatment.
[0137] On the day before behavioral observations, food was removed
overnight, from 5 p.m. On days of behavioral assessment, treatment
was administered to the animals in their home cage at approximately
9 a.m. A 6 h period of observation then commenced 1 h after oral
dosing.
[0138] The animals were rendered parkinsonian by once daily
subcutaneous injection of 0.2 mg/kg MPTP, administered for 8-12
days, until the first appearance of parkinsonism symptoms. After
this time, a parkinsonian syndrome reached a moderate to marked
level, over approximately 30 days, and stabilized. Additional
administrations of MPTP were given to some animals to titrate to
similar degrees of parkinsonism in individuals across the group.
The monkeys were allowed to recover for a minimum of further 30
days until their parkinsonism was demonstrated as being stable.
Sixty days after commencing MPTP administration, L-DOPA (25 mg/kg)
was administered orally twice daily for at least two months. L-DOPA
was given with the decarboxylase inhibitor benserazide (as
Madopar.TM.). This treatment leads to the development of motor
fluctuations, including dyskinesia.
[0139] FIG. 17 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for
extended activity into four and five hours (arrow) and beyond the
use of the high dose of L-DOPA.
[0140] FIG. 18 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for an
even and consistent level of fairly mild parkinsonian disability
and as viewing the results in FIG. 18A, it is evident that with
higher doses of L-DOPA the parkinsonian disability increased in the
last four to six hours. The synergistic combination provides the
most consistent results as shown by the arrow, wherein the
combination of sub-therapeutic L-DOPA in combination with DPI-289
lasted at least 6 hours.
[0141] FIG. 19 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided no
additional dyskinesia to that exhibited by low-dose L-DPOA alone
while the high dose of L-DOPA exhibited severe dyskinesia in the
first three hours after dosage. The figure shows that augmentation
effect of DPI was not associated with any increase in Dyskinesia.
It is evident that the therapeutic dose of L-DOPA causes
significant dyskinesia which last the duration of the therapeutic
effect. In sharp contrast addition of Dpi 289 which augmented the
effect of sub-therapeutic doses of L-DOPA did not result in any
increased dyskinesia. FIGS. 15 & 16 demonstrated that DPI-289
does not cause dyskinesia in neither L-DOPA naive nor in L-DOPA
primed animals. This data shows that DPI-289 does not increase
L-DOPA induced dyskinesia and may even lower it. It is theorized
that DPI-289 having both delta opioid agonist activity and mu
opioid antagonist activity eliminates or greatly reduces dyskinesia
activity.
[0142] FIG. 20A shows that the synergistic combination of DPI-289
at 20 mg/kg in combination with low-dose of L-DOPA provided for
reduced postural instability from about 2 to 6 hours after the
dosing and clearly the synergistic combination showed that such a
combination was more effective than the high dose of L-DOPA or
DPI-289 alone as shown in FIG. 12.
[0143] FIG. 21 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for an
improvement in alertness value that mirrored that of the high dose
of L-DOPA and importantly the beneficial effects actually lasted
longer than for high dose L-DOPA alone.
[0144] FIG. 22 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for
reduced range of movement impairment than low dose L-DOPA alone but
this difference did not reach statistical significance. It would
appear that the combination therapy resulted in more consistent
reduction in range of movement impairment than low or high dose
L-DOPA alone.
[0145] FIG. 23 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for the
same level of bradykinesia as that of the high dose of L-DOPA for
the hours of three to six hours after dosing. Further for the first
three hours showed less bradykinesia from that shown in FIG. 8.
Again, as seen in FIG. 22, it would appear that the combination
therapy resulted in more consistent levels of bradykinesia than low
or high dose L-DOPA alone (arrow).
[0146] FIG. 24 shows that the synergistic combination of DPI-289 at
20 mg/kg in combination with low-dose of L-DOPA provided for
comparable `good` time as that of the high dose of L-DOPA and
considerably less of `bad` time (ON-time with disabling dyskinesia)
relative to the high dose of L-DOPA and certainly an increase in
good on-time relative to the results shown in FIG. 4.
Example 4
[0147] Rodent Model of PD Showed the Use of Dpi-289 Increased
Locomotor Activity without Causing Dyskinesia.
[0148] In a rat model of PD, focal injection of the neurotoxin
6-hydroxydopamine (6-OHDA) was used to create a unilateral lesion
of the nigrostriatal dopamine pathway. When challenged with
dopamine agonists, rats rotate in the direction contralateral to
the lesion and the number of rotations may be used as a measure of
anti-parkinsonian efficacy. Administration of DPI-289 (1-10 mg/kg
i.p.) caused a dose-related increase in both contralateral and
ipsilateral rotations, a profile that suggests the effect is not
mediated through dopaminergic mechanism; a higher dose of 30 mg/kg
also increased rotations but the effect on contralateral rotations
appeared to be diminished, possibly due to sedation at this dose as
shone in FIG. 25. When co-administered with a sub-optimal dose of
L-DOPA (25 mg/kg i.p.), DPI-289 caused a marked potentiation of
contralateral rotations, suggesting that it has utility as an
adjunct to augment the anti-parkinsonian activity of L-DOPA, as
shown in FIG. 26.
[0149] Rats with unilateral 6-OHDA lesions also have motor
incoordination when they are placed on a slowly rotating drum
(rotarod). Daily administration of DPI-289 (3 mg/kg p.o.) or L-DOPA
(6 mg/kg i.p.) for 15 days reduced or reversed the postural
asymmetry and this was maintained throughout treatment duration as
shown in FIG. 27A. Animals receiving L-DOPA developed abnormal
involuntary movements (AIMS: dyskinesia) on the side of the body
opposite the lesion, characterized by twisting of the neck and
trunk, jerky movements of the forelimb and paw, and orofacial
movements (chewing, tongue protrusion). Unlike L-DOPA, daily
administration of DPI-289 for 15 days did not elicit dyskinesia in
6-OHDA-lesioned rats, shown in FIG. 27B.
[0150] Notably, the erratic responses to L-DOPA, shown in FIG. 27A,
were likely due to the dyskinetic movements interfering with the
scoring for asymmetry.
[0151] While the invention has been described herein in reference
to specific aspects, features and illustrative embodiments of the
invention, it will be appreciated that the utility of the invention
is not thus limited, but rather extends to and encompasses numerous
other aspects, features and embodiments. Accordingly, the claims
hereafter set forth are intended to be correspondingly broadly
construed, as including all such aspects, features and embodiments,
within their spirit and scope.
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