U.S. patent application number 13/604718 was filed with the patent office on 2013-03-07 for conjugates comprising a gaba- or glycine compound, pharmaceutical compositions and combinations thereof as well as their use in treating cns disorders.
This patent application is currently assigned to Bar-Ilan University. The applicant listed for this patent is Irit Gil-Ad, Abraham NUDELMAN, Ada Rephaeli, Abraham Weizman. Invention is credited to Irit Gil-Ad, Abraham NUDELMAN, Ada Rephaeli, Abraham Weizman.
Application Number | 20130059910 13/604718 |
Document ID | / |
Family ID | 38621178 |
Filed Date | 2013-03-07 |
United States Patent
Application |
20130059910 |
Kind Code |
A1 |
NUDELMAN; Abraham ; et
al. |
March 7, 2013 |
CONJUGATES COMPRISING A GABA- OR GLYCINE COMPOUND, PHARMACEUTICAL
COMPOSITIONS AND COMBINATIONS THEREOF AS WELL AS THEIR USE IN
TREATING CNS DISORDERS
Abstract
A novel use of conjugates of psychotropic drugs (e.g.,
antidepressants or anti-epileptic drugs) and organic acids such as
GABA in the treatment of pain is disclosed. A novel GABA conjugate
and uses thereof is also disclosed.
Inventors: |
NUDELMAN; Abraham;
(Rechovot, IL) ; Rephaeli; Ada; (Herzlia, IL)
; Gil-Ad; Irit; (Herzlia, IL) ; Weizman;
Abraham; (Tel-Aviv, IL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
NUDELMAN; Abraham
Rephaeli; Ada
Gil-Ad; Irit
Weizman; Abraham |
Rechovot
Herzlia
Herzlia
Tel-Aviv |
|
IL
IL
IL
IL |
|
|
Assignee: |
Bar-Ilan University
Ramat-Gan
IL
Ramot at Tel-Aviv University Ltd.
Tel-Aviv
IL
|
Family ID: |
38621178 |
Appl. No.: |
13/604718 |
Filed: |
September 6, 2012 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
12309361 |
Feb 12, 2010 |
|
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PCT/IL2007/000903 |
Jul 17, 2007 |
|
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13604718 |
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60831195 |
Jul 17, 2006 |
|
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60831192 |
Jul 17, 2006 |
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Current U.S.
Class: |
514/547 ;
514/626 |
Current CPC
Class: |
A61P 25/00 20180101;
A61P 25/02 20180101; A61P 25/32 20180101; A61P 25/14 20180101; A61P
35/00 20180101; A61K 45/06 20130101; A61P 25/04 20180101; A61K
47/55 20170801; A61P 25/34 20180101; A61P 29/00 20180101; A61P
25/24 20180101; A61P 25/16 20180101; A61P 25/18 20180101; A61P
25/22 20180101; A61P 25/08 20180101; A61P 25/30 20180101; A61P
25/28 20180101 |
Class at
Publication: |
514/547 ;
514/626 |
International
Class: |
A61K 31/165 20060101
A61K031/165; A61P 25/34 20060101 A61P025/34; A61P 25/30 20060101
A61P025/30; A61P 25/32 20060101 A61P025/32; A61K 31/22 20060101
A61K031/22; A61P 29/00 20060101 A61P029/00 |
Claims
1. A method of treating pain, the method comprising administering
to a subject in need thereof a therapeutically effective amount of
a chemical conjugate which comprises a first chemical moiety
covalently linked to a second chemical moiety, wherein said first
chemical moiety is an antidepressant and wherein said second
chemical moiety is a GABA agonist.
2. The method of claim 1, wherein said second chemical moiety is
covalently linked to said first chemical moiety via a bond selected
from the group consisting of a carboxylic ester bond, an alkyloxy
carboxylic ester bond, an amide bond, an imine bond and a thioester
bond.
3. The method of claim 1, wherein said antidepressant is selected
from the group consisting of a tricyclic antidepressant, a
selective serotonin reuptake inhibitor, a serotonin and
noradrenaline reuptake inhibitor, a reversible monamine oxidase
inhibitor and a monoamine oxidase inhibitor.
4. The method of claim 1, wherein said antidepressant is
fluoxetine.
5. The method of claim 1, wherein said second chemical moiety is
GABA.
6. The method of claim 4, wherein said second chemical moiety is
GABA.
7. The method of claim 1, wherein said chemical conjugate is
flouxetine-GABA.
8. The method of claim 7, wherein said GABA is covalently linked to
said fluoxetine via an amide bond.
9. The method of claim 1, wherein said GABA agonist is selected
from the group consisting of (.+-.) baclofen, .gamma.-aminobutyric
acid (GABA), .gamma.-hydroxybutyric acid, aminooxyacetic acid,
.beta.-(4-chlorophenyl)-.gamma.-aminobutyric acid, isonipecotic
acid, piperidine-4-sulfonic acid, 3-aminopropylphosphonous acid,
3-aminopropylphosphinic acid, 3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic acid.
10. The method of claim 1, wherein said pain is a chronic pain.
11. The method of claim 10, wherein said chronic pain is a
neuropathic pain or a nociceptive pain.
12. The method of claim 1, wherein said pain is an acute pain.
13. An article-of-manufacture comprising a pharmaceutical
composition being packaged in a packaging material and identified
in print, in or on said packaging material for use in the treatment
of pain, said pharmaceutical composition including a
pharmaceutically acceptable carrier and, as an active ingredient, a
chemical conjugate which comprises a first chemical moiety
covalently linked to a second chemical moiety, wherein said first
chemical moiety is an antidepressant and wherein said second
chemical moiety is a GABA agonist.
14. The article-of-manufacture of claim 13, wherein said second
chemical moiety is covalently linked to said first chemical moiety
via a bond selected from the group consisting of a carboxylic ester
bond, an alkyloxy carboxylic ester bond, an amide bond, an imine
bond and a thioester bond.
15. The article-of-manufacture of claim 13, wherein said
antidepressant is selected from the group consisting of a tricyclic
antidepressant, a selective serotonin reuptake inhibitor, a
serotonin and noradrenaline reuptake inhibitor, a reversible
monamine oxidase inhibitor and a monoamine oxidase inhibitor.
16. The article-of-manufacture of claim 13, wherein said
antidepressant is fluoxetine.
17. The article-of-manufacture of claim 13, wherein said second
chemical moiety is GABA.
18. The article-of-manufacture of claim 16, wherein said second
chemical moiety is GABA.
19. The article-of-manufacture of claim 13, wherein said chemical
conjugate is flouxetine-GABA.
20. The article-of-manufacture of claim 19, wherein said GABA is
covalently linked to said fluoxetine via an amide bond.
21. The article-of-manufacture of claim 13, wherein said GABA
agonist is selected from the group consisting of (.+-.) baclofen,
.gamma.-aminobutyric acid (GABA), .gamma.-hydroxybutyric acid,
aminooxyacetic acid, .beta.-(4-chlorophenyl)-.gamma.-aminobutyric
acid, isonipecotic acid, piperidine-4-sulfonic acid,
3-aminopropylphosphonous acid, 3-aminopropylphosphinic acid,
3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic acid.
22. The article-of-manufacture of claim 13, wherein said pain is a
chronic pain.
23. The article-of-manufacture of claim 22, wherein said chronic
pain is a neuropathic pain or a nociceptive pain.
24. The article-of-manufacture of claim 13, wherein said pain is an
acute pain.
25. A method of treating an addictive disorder, the method
comprising administering to a subject in need thereof a
therapeutically effective amount of a chemical conjugate which
comprises a first chemical moiety covalently linked to a second
chemical moiety, wherein said first chemical moiety is an
antidepressant and wherein said second chemical moiety is a GABA
agonist.
26. The method of claim 25, wherein said second chemical moiety is
covalently linked to said first chemical moiety via a bond selected
from the group consisting of a carboxylic ester bond, an alkyloxy
carboxylic ester bond, an amide bond, an imine bond and a thioester
bond.
27. The method of claim 25, wherein said antidepressant is selected
from the group consisting of a tricyclic antidepressant, a
selective serotonin reuptake inhibitor, a serotonin and
noradrenaline reuptake inhibitor, a reversible monamine oxidase
inhibitor and a monoamine oxidase inhibitor.
28. The method of claim 25, wherein said antidepressant is
fluoxetine.
29. The method of claim 25, wherein said second chemical moiety is
GABA.
30. The method of claim 28, wherein said second chemical moiety is
GABA.
31. The method of claim 25, wherein said chemical conjugate is
flouxetine-GABA.
32. The method of claim 31, wherein said GABA is covalently linked
to said fluoxetine via an amide bond.
33. The method of claim 25, wherein said GABA agonist is selected
from the group consisting of (.+-.) baclofen, .gamma.-aminobutyric
acid (GABA), .gamma.-hydroxybutyric acid, aminooxyacetic acid,
.beta.-(4-chlorophenyl)-.gamma.-aminobutyric acid, isonipecotic
acid, piperidine-4-sulfonic acid, 3-aminopropylphosphonous acid,
3-aminopropylphosphinic acid, 3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic acid.
34. The method of claim 25, wherein said addictive disorder is
alcoholism or smoking.
35. An article-of-manufacture comprising a pharmaceutical
composition being packaged in a packaging material and identified
in print, in or on said packaging material for use in the treatment
of an addictive disorder, said pharmaceutical composition including
a pharmaceutically acceptable carrier and, as an active ingredient,
a chemical conjugate which comprises a first chemical moiety
covalently linked to a second chemical moiety, wherein said first
chemical moiety is an antidepressant and wherein said second
chemical moiety is a GABA agonist.
36. The article-of-manufacture of claim 35, wherein said second
chemical moiety is covalently linked to said first chemical moiety
via a bond selected from the group consisting of a carboxylic ester
bond, an alkyloxy carboxylic ester bond, an amide bond, an imine
bond and a thioester bond.
37. The article-of-manufacture of claim 35, wherein said
antidepressant is selected from the group consisting of a tricyclic
antidepressant, a selective serotonin reuptake inhibitor, a
serotonin and noradrenaline reuptake inhibitor, a reversible
monamine oxidase inhibitor and a monoamine oxidase inhibitor.
38. The article-of-manufacture of claim 35, wherein said
antidepressant is fluoxetine.
39. The article-of-manufacture of claim 35, wherein said second
chemical moiety is GABA.
40. The article-of-manufacture of claim 38, wherein said second
chemical moiety is GABA.
41. The article-of-manufacture of claim 35, wherein said chemical
conjugate is flouxetine-GABA.
42. The article-of-manufacture of claim 41, wherein said GABA is
covalently linked to said fluoxetine via an amide bond.
43. The article-of-manufacture of claim 35, wherein said GABA
agonist is selected from the group consisting of (.+-.) baclofen,
.gamma.-aminobutyric acid (GABA), .gamma.-hydroxybutyric acid,
aminooxyacetic acid, .beta.-(4-chlorophenyl)-.gamma.-aminobutyric
acid, isonipecotic acid, piperidine-4-sulfonic acid,
3-aminopropylphosphonous acid, 3-aminopropylphosphinic acid,
3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic acid.
44. The article-of-manufacturing of claim 35, wherein said
addictive disorder is alcoholism or smoking.
Description
RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 12/309,361 filed on Feb. 12, 2010, which is a
National Phase of PCT Patent Application No. PCT/IL2007/000903
having International filing date of Jul. 17, 2007, which claims the
benefit of priority of U.S. Provisional Patent Application Nos.
60/831,192 and 60/831,195, both filed on Jul. 17, 2006. The
contents of the above applications are all incorporated herein by
reference.
FIELD AND BACKGROUND OF THE INVENTION
[0002] The present invention relates to the field of pharmacology
and, more particularly, to the treatment of pain.
[0003] Inadequate pain management is widely prevalent and harmful
to patients. Numerous studies have demonstrated poor control of
post-operative and trauma pain, cancer pain and chronic non-cancer
pain. Thus, there is a continuing need for medications able to
provide high efficacy pain relief while reducing the possibility of
undesirable effects.
[0004] Pain can be classified into two main categories, acute and
chronic, differing in their etiology, pathophysiology, diagnosis
and treatment. Acute pain is nociceptive in nature (i.e. resulting
directly from local tissue injury). It is a normal, predictable
physiological response to an adverse chemical, thermal, or
mechanical stimulus associated with surgery, trauma, or acute
illness. It is normally self-limited such that when the condition
producing the pain resolves, the pain disappears. Chronic pain may
be defined as pain persisting longer than the expected time of
tissue healing. Injury or a disease process can trigger chronic
pain, but other factors besides the triggering event can perpetuate
the pain. Chronic pain may be either neuropathic (i.e. initiated or
caused by a primary lesion or dysfunction in the nervous system) or
nociceptive.
[0005] Nociceptive pain is typically treated with anti-inflammatory
or analgesic medications whereas neuropathic pain is usually
treated with medications that influence neurotransmitters. These
include, for example, antidepressants, and antiepileptic drugs.
Patients with refractory neuropathic pain are typically treated
with opioids.
[0006] One neurotransmitter which is known to be involved in the
complex circuitry underlying pain is the inhibitory
neurotransmitter, gamma-amino butyric acid (GABA). GABA is not
transported efficiently into the brain from the bloodstream because
of poor transport properties that prevent passage through the
blood-brain barrier. Consequently, brain cells synthesize virtually
all of the GABA found in the brain (by decarboxylation of glutamic
acid with pyridoxal phosphate).
[0007] GABA regulates neuronal excitability through binding to
specific membrane proteins (i.e., GABA receptors), which results in
opening of an ion channel. The entry of chloride ion through the
ion channel leads to hyperpolarization of the recipient cell, which
consequently prevents transmission of nerve impulses to other
cells.
[0008] Although most commonly used for the treatment of anxiety,
muscle spasms and epilepsy, GABA agonists have been shown to
alleviate the symptoms of pain through a number of mechanisms. For
example, pain may be reduced by potentiating GABA transmission.
This may involve targeting GABA transporters as well as GABA
associated enzymes and receptors, such as the GABA-B receptors
[Frediani F. Neurol Sci. 2004, Suppl 3:S161-6]. In addition, GABA
agonists may alleviate pain by reducing glutamate-mediated
excitatory transmission and/or blocking voltage-activated ion
channels. The latter mechanism of action is exemplified by the
newer generation of antiepileptics such as lamotrigine and
gabapentin in the clinical treatment of neuropathic pain symptoms
[Blackburn-Munro G., et al., Curr Pharm Des. 2005;
11(23):2961-76].
[0009] The use of GABA agonists is limited since they typically
include hydrophilic functional groups (e.g., a free carboxylic acid
group and a free amino group) and therefore do not readily cross
the blood brain barrier (BBB). As such, invasive methods of
delivery are required for GABA agonists to have a therapeutic
effect. However, it was found that chemical conjugation of such
compounds with fatty amino acids or peptides could facilitate to
some extent their passage across the BBB [Toth I. J. Drug Target.
1994, 2, 217-39].
[0010] Despite considerable progress in developing new compounds,
the use of systemically acting GABA agonists (e.g., gabapentin,
pregabalin and various benzodiazepines) is limited by adverse
side-effects such as sedation and nausea.
[0011] The neurotransmitters norepinephrine and serotonin are
functionally inhibitory on pain transmission. Thus, as mentioned
hereinabove, monoamine up-regulators such as antidepressants and
antiepileptics are also used in the clinical management of
pain.
[0012] Tricyclic antidepressants are thought to affect pain
transmission in the spinal cord by inhibiting the reuptake of
norepinephrine and serotonin, both of which influence descending
pain pathways. In addition, histamine H1-receptor affinity
(associated with sedation) may be correlated with the analgesic
effect of antidepressants.
[0013] Tricyclic antidepressants may be categorized as secondary or
tertiary amines. Secondary amines such as nortriptyline (Pamelor)
and desipramine (Norpramin) show relatively selective inhibition of
norepinephrine reuptake. Tertiary amines such as amitriptyline and
imipramine (Tofranil) exhibit a more balanced inhibition of
norepinephrine and serotonin, however, they also have greater
anticholinergic side effects. The novel antidepressants venlafaxine
(Effexor) and duloxetine (Cymbalta) comprise a balanced inhibition
of serotonin and norepinephrine reuptake without blockade of other
neuroreceptors that are responsible for typical tricyclic side
effects. The mechanism of action of bupropion (Wellbutrin) is
uncertain but involves blockade of dopamine uptake.
[0014] The efficacy of antidepressant drugs varies dramatically for
neuropathic and non-neuropathic pain syndromes. In addition,
specific agents within each medication class can vary in
effectiveness.
[0015] For example, antidepressants with mixed-receptor or
noradrenergic activity appear to have the greatest therapeutic
effect in patients with neuropathic pain. Predominantly
serotoninergic drugs, such as selective serotonin reuptake
inhibitors (SSRIs), are ineffective in treating chronic pain
[McQuay H J, et al., Pain 1996; 68:217-27]. Amitriptyline and its
metabolite nortriptyline have the best documented efficacy in the
treatment of neuropathic and non-neuropathic pain syndromes [Bryson
H M, Wilde M I. Drugs Aging 1996; 8:459-76]. The novel
antidepressants bupropion, venlafaxine and duloxetine [Wernicke J,
et al., J Pain 2004; 5(3 suppl 1):S48] have been proven effective
in patients with neuropathic pain.
[0016] The efficacy of tricyclic antidepressants in the treatment
of neuropathic pain appears to be independent of their
antidepressant effect and patients with pain but no depression
respond to these agents [Max M B, et al., Neurology 1987;
37:589-96]. Although pain reduction occurs at dosages lower than
those typically required to treat depression, therapeutic doses are
still associated with a number of side effects including
drowsiness, dry mouth, blurred vision, constipation, weight gain,
low blood pressure after getting up, urinary problems, headaches,
impotence, loss of libido, tremor, dizziness, agitation and
insomnia.
[0017] U.S. Patent Application No. 20040242570 and International
PCT Patent Applications WO 03/026563 and WO 2005/092392, which are
incorporated by reference as if fully set forth herein, teach
conjugates of psychotropic drugs and GABA, for treating
psychotropic disorders, proliferative disorders and for enhancing
chemosensitization.
SUMMARY OF THE INVENTION
[0018] The present inventors have envisioned that conjugates of
psychotropic drugs such as anti-depressants and anti-epileptic
drugs and organic acids such as GABA agonists would exert an
improved therapeutic activity as compared with the psychotropic
drug alone and hence could be used in the treatment of pain.
[0019] While reducing the present invention to practice, it was
indeed found that GABA conjugated antidepressant and antiepileptic
drugs showed synergistic effects as compared to their parent
compounds both towards the central perception of pain and the
second, peripheral phase of pain. In addition, these conjugates
showed an accelerated onset of their protective effects and a
longer duration of their protective effects as compared to their
parent compounds.
[0020] Thus, according to one aspect of the present invention there
is provided a method of treating pain, the method comprising
administering to a subject in need thereof a therapeutically
effective amount of a chemical conjugate which comprises a first
chemical moiety covalently linked to a second chemical moiety,
wherein the first chemical moiety is selected from the group
consisting of a psychotropic drug and a GABA agonist and further
wherein the second chemical moiety is an organic acid, the organic
acid is selected so as to enhance the therapeutic effect induced by
the psychotropic drug when the psychotropic drug is administered
per se, thereby treating pain. According to one embodiment, the
first chemical moiety is an antidepressant or an antiepileptic drug
and the second chemical moiety is GABA or R--C(.dbd.O)--, where R
is an alkyl having 3-5 carbon atoms.
[0021] According to another aspect of the present invention there
is provided a use of a chemical conjugate comprising a first
chemical moiety covalently linked to a second chemical moiety,
wherein the first chemical moiety is selected from the group
consisting of a psychotropic drug and a GABA agonist and further
wherein the second chemical moiety is an organic acid, the organic
acid is selected so as to enhance the therapeutic effect induced by
the psychotropic drug when the psychotropic drug is administered
per se, for the manufacture of a medicament for treating pain.
According to one embodiment, the first chemical moiety is an
antidepressant or an antiepileptic drug and the second chemical
moiety is GABA or R--C(.dbd.O)--, whereas R is an alkyl having 3-5
carbon atoms.
[0022] According to yet another aspect of the present invention
there is provided an article-of-manufacture comprising a
pharmaceutical composition being packaged in a packaging material
and identified in print, in or on the packaging material for use in
the treatment of pain, the pharmaceutical composition including a
pharmaceutically acceptable carrier and a chemical conjugate which
comprises a first chemical moiety covalently linked to a second
chemical moiety, wherein the first chemical moiety is selected from
the group consisting of a psychotropic drug and a GABA agonist and
further wherein the second chemical moiety is an organic acid, the
organic acid is selected so as to enhance the therapeutic effect
induced by the psychotropic drug when the psychotropic drug is
administered per se. According to one embodiment, the first
chemical moiety is an antidepressant or an antiepileptic drug and
the second chemical moiety is GABA or R--C(.dbd.O)--, whereas R is
an alkyl having 3-5 carbon atoms.
[0023] According to an additional aspect of the present invention
there is provided a method of treating an addictive disorder, the
method comprising administering to a subject in need thereof a
therapeutically effective amount of a chemical conjugate which
comprises a first chemical moiety covalently linked to a second
chemical moiety, wherein the first chemical moiety is selected from
the group consisting of a psychotropic drug and a GABA agonist and
further wherein the second chemical moiety is an organic acid, the
organic acid is selected so as to enhance the therapeutic effect
induced by the psychotropic drug when the psychotropic drug is
administered per se, thereby treating pain. According to one
embodiment, the first chemical moiety is an antidepressant or an
antiepileptic drug and the second chemical moiety is GABA or
R--C(.dbd.O)--, whereas R is an alkyl having 3-5 carbon atoms.
[0024] According to still an additional aspect of the present
invention there is provided a use of a chemical conjugate
comprising a first chemical moiety covalently linked to a second
chemical moiety, wherein the first chemical moiety is selected from
the group consisting of a psychotropic drug and a GABA agonist and
further wherein the second chemical moiety is an organic acid, the
organic acid is selected so as to enhance the therapeutic effect
induced by the psychotropic drug when the psychotropic drug is
administered per se, for the manufacture of a medicament for
treating an addictive disorder. According to one embodiment, the
first chemical moiety is an antidepressant or an antiepileptic drug
and the second chemical moiety is GABA or R--C(.dbd.O)--, whereas R
is an alkyl having 3-5 carbon atoms.
[0025] According to yet an additional aspect of the present
invention there is provided an article-of-manufacture comprising a
pharmaceutical composition being packaged in a packaging material
and identified in print, in or on the packaging material for use in
the treatment of an addictive disorder, the pharmaceutical
composition including a pharmaceutically acceptable carrier and a
chemical conjugate which comprises a first chemical moiety
covalently linked to a second chemical moiety, wherein the first
chemical moiety is selected from the group consisting of a
psychotropic drug and a GABA agonist and further wherein the second
chemical moiety is an organic acid, the organic acid is selected so
as to enhance the therapeutic effect induced by the psychotropic
drug when the psychotropic drug is administered per se. According
to one embodiment, the first chemical moiety is an antidepressant
or an antiepileptic drug and the second chemical moiety is GABA or
R--C(.dbd.O)--, whereas R is an alkyl having 3-5 carbon atoms.
[0026] According to further features in embodiments of the
invention described below, the second moiety is further selected so
as to reduce adverse side effects induced by the psychotropic drug
when administered per se.
[0027] According to still further features in the described
preferred embodiments the second chemical moiety is a GABA
agonist.
[0028] According to still further features in the described
embodiments the second chemical moiety is covalently linked to the
first chemical moiety via a bond selected from the group consisting
of a carboxylic ester bond, an alkyloxy carboxylic ester bond, an
amide bond, an imine bond and a thioester bond.
[0029] According to still further features in the described
embodiments the psychotropic drug is an antidepressant or an
anti-epileptic drug.
[0030] According to still further features in the described
embodiments the antidepressant is selected from the group
consisting of a tricyclic antidepressant, a selective serotonin
reuptake inhibitor, a serotonin and noradrenaline reuptake
inhibitor, a reversible monamine oxidase inhibitor and a monoamine
oxidase inhibitor.
[0031] According to still further features in the described
embodiments the tricyclic antidepressant is a secondary amine
tricyclic antidepressant or a tertiary amine tricyclic
antidepressant.
[0032] According to still further features in the described
embodiments the secondary amine tricyclic antidepressant is
selected from the group consisting of nortriptyline and
desipramine.
[0033] According to still further features in the described
embodiments the tertiary amine tricyclic antidepressant is selected
from the group consisting of amitriptyline and imipramine.
[0034] According to still further features in the described
embodiments the selective serotonin reuptake inhibitor is selected
from the group consisting of fluoxetine, citalopram, paroxetine,
fluvoxamine, escitalopram (lexapro) and sertraline.
[0035] According to still further features in the described
embodiments the serotonin and noradrenaline reuptake inhibitor is
venlafaxine.
[0036] According to still further features in the described
embodiments the reversible monamine oxidase inhibitor is
moclobemide.
[0037] According to still further features in the described
embodiments the monoamine oxidase inhibitor is selected from the
group consisting of phenelzine and tranylcypromine.
[0038] According to still further features in the described
embodiments the antiepileptic drug is selected from the group
consisting of carbamazepine, valproate, ethosuximide and
phenyloin.
[0039] According to still further features in the described
embodiments the antidepressant drug is selected from the group
consisting of venlafaxine, duloxetine and bupropion.
[0040] According to still further features in the described
embodiments the psychotropic drug is selected from the group
consisting of nortryptiline, fluoxetine and valproic acid.
[0041] According to still further features in the described
embodiments the first chemical moiety is a GABA agonist.
[0042] According to still further features in the described
embodiments the second chemical moiety is a GABA agonist.
[0043] According to still further features in the described
embodiments the GABA agonist is selected from the group consisting
of (.+-.) baclofen, .gamma.-aminobutyric acid (GABA),
.gamma.-hydroxybutyric acid, aminooxyacetic acid,
.beta.-(4-chlorophenyl)-.gamma.-aminobutyric acid, isonipecotic
acid, piperidine-4-sulfonic acid, 3-aminopropylphosphonous acid,
3-aminopropylphosphinic acid, 3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic acid.
[0044] According to still further features in the described
embodiments the organic acid has a general formula:
--R--C(.dbd.O)--
wherein,
[0045] R is selected from the group consisting of a substituted or
non-substituted hydrocarbon having 1-20 carbon atoms, a substituted
or non-substituted hydrocarbon having 1-20 carbon atoms and at
least one heteroatom selected from the group consisting of oxygen,
nitrogen and sulfur and R.sub.1,
whereas,
[0046] R.sub.1 is a moiety of a general formula:
--Z--C(.dbd.O)O--CHR.sub.2--R.sub.3
wherein,
[0047] Z is selected from the group consisting of a single bond, a
substituted or non-substituted hydrocarbon having 1-20 carbon atoms
and a substituted or non-substituted hydrocarbon having 1-20 carbon
atoms and at least one heteroatom selected from the group
consisting of oxygen, nitrogen and sulfur;
[0048] R.sub.2 is selected from the group consisting of hydrogen
and an alkyl having 1-10 carbon atoms; and
[0049] R.sub.3 is selected form the group consisting of hydrogen, a
substituted or non-substituted hydrocarbon having 1-20 carbon atoms
and a substituted or non-substituted alkyl having 1-20 carbon atoms
and at least one heteroatom selected from the group consisting of
oxygen, nitrogen and sulfur.
[0050] According to still further features in the described
embodiments R is a substituted or non-substituted alkyl having 3-5
carbon atoms.
[0051] Exemplary conjugates according to the present embodiments
include, without limitation, GABAoxymethylGABA (also referred to
herein as AN-214), GABA-oxymethylvalproate (also referred to herein
as AN-216), fluoxetine-GABA (also referred to herein as AN-227) and
nortriptyline-GABA (also referred to herein as AN-228).
[0052] According to still further features in the described
embodiments the pain is a chronic pain.
[0053] According to still further features in the described
embodiments the chronic pain is a neuropathic pain or a nociceptive
pain.
[0054] According to still further features in the described
embodiments the pain is an acute pain.
[0055] According to still further features in the described
embodiments the addictive disorder is alcoholism and/or
smoking.
[0056] According to a further aspect of the present invention there
is provided a chemical conjugate comprising a first chemical moiety
covalently linked to a second chemical moiety, wherein each of the
chemical moieties is independently a GABA agonist.
[0057] According to a further aspect of the present invention there
is provided a pharmaceutical composition comprising, as an active
ingredient, the chemical conjugate comprising a first chemical
moiety covalently linked to a second chemical moiety, wherein each
of the chemical moieties is independently a GABA agonist and a
pharmaceutically acceptable carrier.
[0058] According to yet a further aspect of the present invention
there is provided a method of treating a CNS disease or disorder,
the method comprising administering to a subject in need thereof a
therapeutically effective amount of the chemical conjugate
comprising a first chemical moiety covalently linked to a second
chemical moiety, wherein each of the chemical moieties is
independently a GABA agonist.
[0059] According to still a further aspect of the present invention
there is provided a use of the conjugate comprising a first
chemical moiety covalently linked to a second chemical moiety,
wherein each of the chemical moieties is independently a GABA
agonist, for the preparation of a medicament.
[0060] According to further features in embodiments of the
invention described below, the GABA agonists are covalently linked
therebetween via a bond selected from the group consisting of a
carboxylic ester bond, an alkyloxy carboxylic ester bond, an amide
bond, an imine bond and a thioester bond.
[0061] According to still further features in the described
embodiments the bond is an alkyloxy carboxylic ester bond.
[0062] According to still further features in the described
embodiments each of the GABA agonists is independently selected
from the group consisting of (.+-.) baclofen, .gamma.-aminobutyric
acid (GABA), .gamma.-hydroxybutyric acid, aminooxyacetic acid,
.beta.-(4-chlorophenyl)-.gamma.-aminobutyric acid, isonipecotic
acid, piperidine-4-sulfonic acid, 3-aminopropylphosphonous acid,
3-aminopropylphosphinic acid, 3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
y-vinyl-.gamma.-aminobutyric acid (4-aminohex-5-enoic acid, y-vinyl
GABA, vigabatrin), vigabatrin) and 3-(2-imidazolyl)-4-aminobutanoic
acid.
[0063] According to still further features in the described
embodiments each of the GABA agonists is a .gamma.-aminobutyric
acid (GABA).
[0064] According to still further features in the described
embodiments the pharmaceutical composition comprising, as an active
ingredient, the chemical conjugate comprising a first chemical
moiety covalently linked to a second chemical moiety, wherein each
of the chemical moieties is independently a GABA agonist and a
pharmaceutically acceptable carrier, is packaged in a packaging
material and identified in print, on or in the packaging material,
for use in the treatment of a CNS disease or disorder.
[0065] According to still further features in the described
embodiments the medicament is for treating a CNS disease or
disorder.
[0066] According to still further features in the described
embodiments the CNS disease or disorder is selected from the group
consisting of a pain disorder, a motion disorder, a dissociative
disorder, a mood disorder, an affective disorder, a
neurodegenerative disease or disorder and a convulsive
disorder.
[0067] According to still further features in the described
embodiments the CNS disease or disorder is selected from the group
consisting of Parkinson's, Multiple Sclerosis, Huntington's
disease, action tremors and tardive dyskinesia, panic, anxiety,
depression, alcoholism, insomnia and manic behavior, Alzheimer's
and epilepsy.
[0068] The present invention thus provides a novel method for
treating pain, and particularly neuropathic pain, which is
characterized by superior efficacy and reduced side effects as
compared to the presently known medications. The present invention
further provides novel conjugates which can be used in the
treatment of pain, as well as other CNS diseases and disorders.
[0069] Unless otherwise defined, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which this invention belongs. Although
methods and materials similar or equivalent to those described
herein can be used in the practice or testing of the present
invention, suitable methods and materials are described below. All
publications, patent applications, patents, and other references
mentioned herein are incorporated by reference in their entirety.
In case of conflict, the patent specification, including
definitions, will control. In addition, the materials, methods, and
examples are illustrative only and not intended to be limiting.
[0070] As used herein, the singular form "a," "an," and "the"
include plural references unless the context clearly dictates
otherwise. For example, the term "a compound" or "at least one
compound" may include a plurality of compounds, including mixtures
thereof.
[0071] Throughout this disclosure, various aspects of this
invention can be presented in a range format. It should be
understood that the description in range format is merely for
convenience and brevity and should not be construed as an
inflexible limitation on the scope of the invention. Accordingly,
the description of a range should be considered to have
specifically disclosed all the possible subranges as well as
individual numerical values within that range. For example,
description of a range such as from 1 to 6 should be considered to
have specifically disclosed subranges such as from 1 to 3, from 1
to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as
well as individual numbers within that range, for example, 1, 2, 3,
4, 5, and 6. This applies regardless of the breadth of the
range.
[0072] Whenever a numerical range is indicated herein, it is meant
to include any cited numeral (fractional or integral) within the
indicated range. The phrases "ranging/ranges between" a first
indicate number and a second indicate number and "ranging/ranges
from" a first indicate number "to" a second indicate number are
used herein interchangeably and are meant to include the first and
second indicated numbers and all the fractional and integral
numerals therebetween.
[0073] As used herein, the term "about" refers to .+-.10%.
[0074] The term "comprising" means that other steps and ingredients
that do not affect the final result can be added. This term
encompasses the terms "consisting of" and "consisting essentially
of".
[0075] The phrase "consisting essentially of" means that the
composition or method may include additional ingredients and/or
steps, but only if the additional ingredients and/or steps do not
materially alter the basic and novel characteristics of the claimed
composition or method.
[0076] The term "method" refers to manners, means, techniques and
procedures for accomplishing a given task including, but not
limited to, those manners, means, techniques and procedures either
known to, or readily developed from known manners, means,
techniques and procedures by practitioners of the chemical,
pharmacological, biological, biochemical and medical arts.
[0077] The term "active ingredient" refers to a pharmaceutical
agent including any natural or synthetic chemical substance that
subsequent to its application has, at the very least, at least one
desired pharmaceutical or therapeutic effect.
BRIEF DESCRIPTION OF THE DRAWINGS
[0078] The patent or application file contains at least one drawing
executed in color. Copies of this patent or patent application
publication with color drawing(s) will be provided by the Office
upon request and payment of the necessary fee.
[0079] The invention is herein described, by way of example only,
with reference to the accompanying drawings. With specific
reference now to the drawings in detail, it is stressed that the
particulars shown are by way of example and for purposes of
illustrative discussion of the preferred embodiments of the present
invention only, and are presented in the cause of providing what is
believed to be the most useful and readily understood description
of the principles and conceptual aspects of the invention. In this
regard, no attempt is made to show structural details of the
invention in more detail than is necessary for a fundamental
understanding of the invention, the description taken with the
drawings making apparent to those skilled in the art how the
several forms of the invention may be embodied in practice.
[0080] In the drawings:
[0081] FIG. 1 presents comparative dose response plots illustrating
the latency of nociceptive reaction to heat of mice treated p.o.
with nortriptyline and equimolar dose of nortriptyline-GABA
conjugate (AN-228). Male Balb-c mice (6/group) were treated p.o.
with the indicated doses of nortriptyline, respective equimolar
doses nortriptyline-GABA conjugate or vehicle. Two hours later the
animals were placed on the surface of the hot-plate
(52.+-.0.2.degree. C.). Response to the heat, manifested by shaking
or licking of the paws or jumping, was recorded as the index of
response latency.
[0082] FIGS. 2A-B present plots illustrating the time-course of
latency of the nociceptive reaction to heat of mice treated with
nortriptyline and the respective equimolar doses of the conjugate
nortriptyline-GABA (AN-228). In FIG. 2A, male Balb-c mice were
treated p.o. with 0.25 mg/kg nortriptyline, equimolar dose of its
GABA conjugate (0.32 mg/kg) or vehicle. In FIG. 2B, male Balb-c
mice were treated p.o. with 0.5 mg/kg nortriptyline, equimolar dose
of its GABA conjugate (0.64 mg/kg) or vehicle. Six mice were used
in all experimental groups. The animals latency response to heat
(sec), using a hot plate at 52.+-.0.2.degree. C., was measured
before and after treatment as indicated.
[0083] FIGS. 3A-E present graphs illustrating the effect of chronic
administration of nortriptyline (0.2 mg/kg) and an equimolar dose
of its GABA conjugate (AN-228) on latency of the nociceptive
reaction to heat. The latency of the nociceptive reaction was
measured prior to treatment (FIG. 3A); two hours following
treatment (FIG. 3B); three hours following treatment (FIG. 3C);
four hours following treatment (FIG. 3D); and five hours following
treatment (FIG. 3E).
[0084] FIG. 4 presents a graph illustrating the effect on latency
of the nociceptive reaction to heat of fluoxetine, equimolar GABA
and equimolar fluoxetine-GABA conjugate (AN-227). Male Balb-c mice
were placed on the surface of the hot-plate and the time between
the animal placement and its response was recorded as the index of
response latency. The reaction was recorded at: -60, 0, 60, 120,
180, 240, 300 min after p.o. administration of vehicle (n=18);
fluoxetine 10 mg/kg (n=11); equimolar fluoxetine-GABA conjugate
(n=11); and equimolar GABA (n=8).
[0085] FIGS. 5A-E present graphs illustrating the effect of chronic
administration (p.o.) of fluoxetine (10 mg/kg) and an equimolar
does of its GABA conjugate (AN-227) on latency of the nociceptive
reaction to heat. Male Balb-c mice (n=11 per treatment groups and
n=10 of control) were treated five times a week for 2 weeks. The
latency of the nociceptive reaction was measured prior to treatment
(FIG. 5A); two hours following treatment (FIG. 5B); three hours
following treatment (FIG. 5C); four hours following treatment (FIG.
5D); and five hours following treatment (FIG. 5E).
[0086] FIG. 6 presents a graph illustrating the effect of
GABA-oxymethylvalproate (AN-216) 0.24 mg/kg and an equimolar dose
of GABA (2.39 mg/kg) on heat sensation-time course. The latency
response to heat was recorded at: -60, 0, 60, 120, 180, 240 and 300
min, following treatment.
[0087] FIGS. 7A-E present graphs illustrating the effect of five
day treatment of GABA-oxymethylGABA (AN-214) 0.2 mg/kg (n=11) and
an equimolar dose of GABA-oxymethylvalproate (AN-216) (n=8) on
latency of the nociceptive reaction to heat. The latency of the
nociceptive reaction was measured prior to treatment (FIG. 7A); two
hours following treatment (FIG. 7B); three hours following
treatment (FIG. 7C); four hours following treatment (FIG. 7D); and
five hours following treatment (FIG. 7E) on days 1, 3 and 5.
[0088] FIGS. 8A-E present graphs illustrating the effect of five
day treatment of GABA on latency of the nociceptive reaction to
heat. Male Balb-c mice (n=8/group) were treated with the indicated
doses of GABA daily for five days. The latency of the nociceptive
reaction was measured prior to treatment (FIG. 8A); two hours
following treatment (FIG. 8B); three hours following treatment
(FIG. 8C); four hours following treatment (FIG. 8D); and five hours
following treatment (FIG. 8E).
[0089] FIGS. 9A-B present graphs illustrating the pain relieving
effect of fluoxetine and the fluoxetine-GABA conjugate (AN-227) on
both the early neurogenic phase--and the late inflammatory
peripheral phase to pain as measured by a formalin test. Balb/c
mice (5/group) were treated with: vehicle (DDW), 10 mg/kg
fluoxetine, 30 mg/kg fluoxetine and their respective equimolar
doses of fluoxetine-GABA conjugate. Three hours later 1% (20 ml)
formalin was injected to intraplantar of the right hind paw.
Immediately after injection, the mouse was placed in a Plexiglas
chamber for observation. The amount of time the animal spent
licking the injected paw was recorded as a quantitative indication
of nociception. (A) The early neurogenic phase (nociceptive
response) was measured between 0-5 min and (B) the late phase from
20-30 min after formalin injection (nociceptors and inflammatory
nociceptive responses).
[0090] FIGS. 10A-B present graphs illustrating the pain relieving
effect of nortriptyline and the nortriptyline-GABA conjugate
(AN-228) on both the early neurogenic phase--and the late
inflammatory peripheral phase to pain as measured by a formalin
test. Balb/c mice (5/group) were treated with: vehicle (DDW), 0.5
mg/kg nortriptyline, 5 mg/kg nortriptyline and their respective
equimolar doses of nortriptyline-GABA conjugate. Three hours later
1% (20 .mu.l) formalin was injected to intraplantar of the right
hind paw Immediately after injection, the mouse was placed in a
Plexiglas chamber for observation. The amount of time the animal
spent licking the injected paw was recorded as a quantitative
indication of nociception. (A) The early neurogenic phase
(nociceptive response) was measured between 0-5 min and (B) the
late phase from 10 min after formalin injection (nociceptors and
inflammatory nociceptive responses). The results are an average of
two experiments.
[0091] FIG. 11 presents a graph illustrating the anti-inflammatory
effect of fluoxetine and the fluoxetine-GABA conjugate (AN-227) as
measured by a carrageenan-induced paw edema test. Wistar rats
(250-350 g) were treated p.o. with fluoxetine 40 mg/kg (n=8),
equimolar of fluoxetine-GABA conjugate (n=6) and vehicle (n=8).
After 3 h they were injected to the plantar surface of the left
hind paw with 100 .mu.l 1% solution of .lamda.-carrageenan. The
edema developed in the injected area was measured 2 and 4 h later
with caliper.
[0092] FIG. 12 is a graph illustrating the effect of treatment with
Nortriptyline-GABA (AN-228) on latency of the nociceptive reaction
to heat as compared to Nortriptyline alone and Gabapentin
(1-(aminomethyl)cyclohexaneacetic acid) 2-48 hours following
injection.
[0093] FIGS. 13A-B are bar graphs illustrating the effect of
treatment with Nortriptyline-GABA (AN-228) on paw height as
compared to Nortriptyline alone and control (FIG. 13A) and as
compared to Nortriptyline alone, control and Gabapentin (FIG. 13B)
at particular time points following injection.
[0094] FIGS. 14A-B are bar graphs illustrating the effect of
treatment with Nortriptyline-GABA (AN-228) as compared to
Nortriptyline alone, control, Gabapentin and a mixture of
Nortriptyline and GABA on INF-.gamma. secretion (FIG. 14A) and
TNF-.alpha. secretion (FIG. 14B).
DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0095] The present invention is of a novel use of conjugates of
psychotropic drugs, particularly antidepressants and anti-epileptic
drugs, and organic acids such as GABA, for the treatment of pain.
The present invention is further of novel conjugates of GABA
agonists and their use in the treatment of CNS diseases and
disorders (e.g., pain).
[0096] The principles and operation of the chemical conjugates
according to the present invention may be better understood with
reference to the drawings and accompanying descriptions.
[0097] Before explaining at least one embodiment of the invention
in detail, it is to be understood that the invention is not limited
in its application to the details set forth in the following
description or exemplified by the Examples. The invention is
capable of other embodiments or of being practiced or carried out
in various ways. Also, it is to be understood that the phraseology
and terminology employed herein is for the purpose of description
and should not be regarded as limiting.
[0098] Psychotropic drugs are often encumbered by short and long
term adverse side effects. Conjugation thereof with GABA agonists
results in their enhanced therapeutic activity and a substantial
reduction of induced adverse side effects. Thus, U.S. Patent
Application No. 20040242570 and International PCT Patent
Applications WO 03/026563 and WO 2005/092392, by the present
inventors, teach GABA-conjugated psychotropic drugs for treating
psychotropic disorders, proliferative disorders and for enhancing
chemosensitization.
[0099] Whilst conceiving the present invention, it was hypothesized
that conjugates of organic acids such as GABA and antidepressants
and anti-epileptics may not only enhance the respective
antidepressant and antiepileptic therapeutic properties of these
drugs, but may also improve other characteristics thereof, such as
their pain relieving capabilities. Thus, as demonstrated in
Examples 2 and 3, GABA conjugated antidepressant and antiepileptic
drugs unexpectedly showed synergistic effects as compared to their
parent compounds both towards the central perception of pain and
the second, peripheral phase of pain. Not only did the GABA
conjugated drugs show enhanced pain relieving effects, but they
also showed an accelerated onset of their protective effects and a
longer duration of their protective effects as compared to their
non-GABA conjugated counterparts.
[0100] Without being bound to any particular theory, it is believed
that the enhanced palliative activities of the conjugates are due
to the simultaneous action of the psychotropic drug and the organic
acid at the same site in the brain resulting in a synergistic pain
relief. In addition, conjugation is believed to improve brain
permeability of both chemical moieties.
[0101] Thus, according to one aspect of the present invention,
there is provided a method of treating pain, which is effected by
administering to a subject in need thereof a therapeutically
effective amount of a chemical conjugate. The chemical conjugate
includes a first chemical moiety that is covalently linked to a
second chemical moiety. The first chemical moiety may be a
psychotropic drug or a GABA agonist, whereas the second chemical
moiety is an organic acid, selected so as to enhance the
therapeutic efficacy of the psychotropic drug and/or to reduce side
effects induced by the psychotropic drug when administered per
se.
[0102] By "administered per se" it is meant that a non-conjugated
psychotropic drug, corresponding to the same psychotropic drug that
constitutes the first moiety in the conjugate, is utilized. Thus,
the conjugates described herein are such that the therapeutic
efficacy thereof is enhanced as compared to the therapeutic
efficacy of a corresponding non-conjugated psychotropic drug and/or
the side effects induced thereby are reduced as compared to a
corresponding non-conjugated psychotropic drug.
[0103] As used herein, the term "pain" encompasses both acute and
chronic pain. As used herein, the term "acute pain" means
immediate, generally high threshold, pain brought about by injury
such as a cut, crush, burn, or by chemical stimulation such as that
experienced upon exposure to capsaicin, the active ingredient in
chili peppers. The term "chronic pain," as used herein, means pain
other than acute pain and includes, without limitation, neuropathic
pain, visceral pain, fibromyalgia pain, inflammatory pain, headache
pain, muscle pain and referred pain. It is understood that chronic
pain often is of relatively long duration, for example, months or
years and can be continuous or intermittent.
[0104] In one embodiment, the conjugates of the present invention
are used to treat "neuropathic pain," which, as used herein, means
pain resulting from injury to a nerve. Neuropathic pain can be
distinguished from nociceptive pain, which is pain caused by acute
tissue injury involving small cutaneous nerves or small nerves in
muscle or connective tissue. In contrast to neuropathic pain,
nociceptive pain usually is limited in duration to the period of
tissue repair and usually can be alleviated by available analgesic
agents or opioids [Myers, Regional Anesthesia 20:173-184
(1995)].
[0105] Neuropathic pain typically is long-lasting or chronic and
can develop days or months following an initial acute tissue
injury. Neuropathic pain can involve persistent, spontaneous pain,
as well as allodynia, which is a painful response to a stimulus
that normally is not painful, or hyperalgesia, an accentuated
response to a painful stimulus that usually is trivial, such as a
pin prick. Neuropathic pain generally is resistant to opioid
therapy [Myers, (1995) supra].
[0106] As used herein, the term "chemical moiety" refers to a
moiety derived from a chemical compound, which retains its
functionality.
[0107] Herein throughout, whenever a chemical moiety is described,
it is to be understood as that part of the chemical moiety that is
linked to the other chemical moiety in the conjugate, unless
otherwise indicated.
[0108] Thus, the phrase "psychotropic drug", whenever used with
respect to the first moiety in the conjugate, refers to a major
portion of a psychotropic drug that is covalently linked to another
chemical moiety, as this term is defined hereinabove.
[0109] As is described hereinabove, the phrase "psychotropic drug"
encompasses any agent or drug that exerts an activity in the
central nervous system and thereby can be used in the treatment of
various central nervous system diseases or disorders.
[0110] Hence, psychotropic drugs, according to the present
invention, include, for example, moieties derived from anxiolytic
drugs such as, but not limited to, benzodiazepines, phenothiazines
and butyrophenones, MAO inhibitors, anti-depressants,
anti-epileptic drugs, anti-convulsive drugs (also referred to as
anti-convulsants), anti-parkinsonian drugs, and acetylcholine
esterase inhibitors. The psychotropic drugs can be tricyclic,
bicyclic or monocyclic.
[0111] Preferred psychotropic drugs, according to the present
invention, are those having an amine group, a thiol group or a
hydroxyl group, as these terms are defined hereinbelow, which can
be reacted with the organic acid or a reactive derivative thereof.
Such groups can be present in the psychotropic drug, prior to its
incorporation in the conjugate, either as a free functional group
or as a part of another functional group, e.g., an amide group, a
carboxylic acid group and the like, as these terms are defined
hereinbelow.
[0112] According to a preferred embodiment of the present
invention, the psychotropic drug from which the first chemical
moiety is derived is known for its therapeutic effect on pain,
e.g., antidepressant or antiepileptic drugs. Both antidepressants
and antiepileptics have been shown to be efficacious in the
treatment of pain and more specifically neuropathic pain.
[0113] As used herein, the phrase "antidepressant drug" refers to
any drug which is known to alleviate the causes and/or symptoms of
depression regardless of its mechanism of action.
[0114] The phrase "anti-epileptic drug", which is also referred to
herein and in the art as "anti-epileptic", "anti-convulsant" or
"anti-convulsive drug", describes any drug which is known to
alleviate the causes and/or symptoms of convulsions and
particularly, convulsions caused by epilepsy.
[0115] As illustrated in the Examples section that follows,
conjugates of various psychotropic drugs have been tested and were
shown to exert enhanced therapeutic activity as compared to the
respective non-conjugates drugs, regardless of the pathway affected
thereby or of the receptor interacted therewith. Thus, for example,
it is demonstrated that conjugates of two known antidepressants,
fluoxetine and nortriptyline, acting on two entirely separate
pathways (selective serotonin reuptake inhibitor and selective
norepinephrine reuptake inhibitor, respectively) were shown to
induce pain relief in mouse in vivo studies.
[0116] In addition, a known antiepileptic drug (valproic acid)
acting on a GABA pathway was also shown to be effective in the
relief of pain. It is suggested that since valproic acid is a drug
that is capable of crossing the BBB, a valproate-GABA conjugate
(e.g., GABA-oxymethylvalproate) may carry the GABA into the brain
and the GABA in turn could potentiate the activity of valproic
acid.
[0117] Thus, according to this aspect of the present invention, the
antidepressant drug may be a tricyclic antidepressant (e.g.
secondary amine tricyclic antidepressant or a tertiary amine
tricyclic antidepressant), a selective serotonin reuptake
inhibitor, a serotonin and noradrenaline reuptake inhibitor, a
reversible monamine oxidase inhibitor and a monamine oxidase
inhibitor.
[0118] Non-limiting examples of antidepressants include
nortriptyline, desipramine, amitriptyline, imipramine, fluoxetine,
citalopram, paroxetine, fluvoxamine, escitalopram (lexapro),
sertraline, venlafaxine, moclobemide, phenelzine, duloxetine and
tranylcypromine
[0119] According to a preferred embodiment of this aspect of the
present invention, the antidepressant is nortriptyline or
fluoxetine.
[0120] Examples of anti-epileptic drugs include, but are not
limited to, carbamazepine, valproate (valproic acid), ethosuximide
and phenyloin.
[0121] Additional psychotropic drugs that are known to exert pain
relief are GABA agonists [Blackburn-Munro G., et al., Curr Pharm
Des. 2005; 11(23):2961-76]. Hence, the first chemical moiety may
alternatively be a GABA agonist. Examples of GABA agonists are
described hereinbelow. Conjugates of two GABA agonists that are
linked to one another have never been described hitherto.
[0122] As stated hereinabove, the psychotropic drug, according to
the present invention, is covalently coupled to a second chemical
moiety, which is an organic acid.
[0123] The phrase "organic acid" refers to a moiety, as defined
herein, that is derived from an organic acid that includes a free
carboxylic group.
[0124] The term "free carboxylic group" includes a "--C(.dbd.O)OH"
group either as is, in its protonated or in its ionized or salt
state.
[0125] The organic acid constituting the second moiety in the
conjugates described according to the present invention, can be,
for example, a moiety that has a general formula --R--C(.dbd.O)--,
where R can be, for example, a hydrocarbon that has 1-20 carbon
atoms.
[0126] The term "hydrocarbon" as used herein refers to an organic
compound that includes, as its basic skeleton, a chain of carbon
atoms and hydrogen atoms that are covalently linked
therebetween.
[0127] Thus, the hydrocarbon according to the present invention can
be, for example, alkyl or cycloalkyl.
[0128] As used herein, the term "alkyl" refers to a saturated
aliphatic hydrocarbon including straight chain and branched chain
groups. Preferably, the alkyl group has 1 to 20 carbon atoms.
[0129] Whenever a numerical range, e.g., "1-20", is stated herein,
it means that the group, in this case the alkyl group, may contain
1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and
including 20 carbon atoms. More preferably, the alkyl is a medium
size alkyl having 1 to 10 carbon atoms. Most preferably, the alkyl
has 3 to 5 carbon atoms.
[0130] As used herein, the term "cycloalkyl" includes an all-carbon
monocyclic or fused ring (i.e., rings which share an adjacent pair
of carbon atoms) group wherein one of more of the rings does not
have a completely conjugated pi-electron system. Examples, without
limitation, of cycloalkyl groups include cyclopropane, cyclobutane,
cyclopentane, cyclopentene, cyclohexane, cyclohexadiene,
cycloheptane, cycloheptatriene and adamantane.
[0131] The hydrocarbon, according to the present invention, can be
straight or branched. The hydrocarbon can further be saturated or
unsaturated. When unsaturated, the hydrocarbon can include a double
bond or a triple bond in its carbon chain. An unsaturated
hydrocarbon can further include an aryl.
[0132] As used herein, an "aryl" group refers to an all-carbon
monocyclic or fused-ring polycyclic (i.e., rings which share
adjacent pairs of carbon atoms) groups having a completely
conjugated pi-electron system. Examples, without limitation, of
aryl groups include phenyl, naphthalenyl and anthracenyl.
[0133] The hydrocarbon can further be substituted or
non-substituted. When substituted, the substituent can be, for
example, alkyl, cycloalkyl, aryl, heteroaryl, heteroalicyclic,
hydroxy, alkoxy, aryloxy, cyano, halo, oxo, amido and amino
[0134] A "heteroaryl" group refers to a monocyclic or fused ring
(i.e., rings which share an adjacent pair of atoms) group having in
the ring(s) one or more atoms, such as, for example, nitrogen,
oxygen and sulfur and, in addition, having a completely conjugated
pi-electron system. Examples, without limitation, of heteroaryl
groups, include pyrrole, furane, thiophene, imidazole, oxazole,
thiazole, pyrazole, pyridine, pyrimidine, quinoline, isoquinoline
and purine. The heteroaryl group may be substituted or
non-substituted. When substituted, the substituent group can be,
for example, alkyl, cycloalkyl, hydroxy, alkoxy, aryloxy, cyano,
halo, oxo, amido and amino
[0135] A "heteroalicyclic" group refers to a monocyclic or fused
ring group having in the ring(s) one or more atoms such as
nitrogen, oxygen and sulfur. The rings may also have one or more
double bonds. However, the rings do not have a completely
conjugated pi-electron system. The heteroalicyclic may be
substituted or non-substituted. When substituted, the substituted
group can be, for example, alkyl, cycloalkyl, aryl, heteroaryl,
halo, trihalomethyl, hydroxy, alkoxy, aryloxy, cyano, oxo, amido
and amino
[0136] A "hydroxy" group refers to an --OH group.
[0137] An "alkoxy" group refers to both an --O-alkyl and an
--O-cycloalkyl group, as defined herein.
[0138] An "aryloxy" group refers to both an --O-aryl and an
--O-heteroaryl group, as defined herein.
[0139] An "oxo" group refers to a --C(.dbd.O)--R' group, where R'
can be, for example, alkyl, cycloalkyl or aryl.
[0140] A "halo" group refers to fluorine, chlorine, bromine or
iodine.
[0141] A "trihalomethyl" group refers to a --CX.sub.3-- group
wherein X is a halo group as defined herein.
[0142] An "amino" or "amine" group refers to a --NH.sub.2
group.
[0143] An "amido" or "amide" group refers to a --C(.dbd.O)--NRaRb
group, where Ra and Rb can be, for example, hydrogen, alkyl,
cycloalkyl and aryl.
[0144] The hydrocarbon, according to the present invention, can
further include one or more heteroatoms interspersed within its
chain. The heteroatoms can be, for example, oxygen, nitrogen and/or
sulfur.
[0145] The organic acid can further have a general formula
--Z--C(.dbd.O)O--CHR.sub.2--R.sub.3, where Z can be, for example, a
single bond, or a substituted or non-substituted hydrocarbon as
described hereinabove; R.sub.2 can be, for example, hydrogen or an
alkyl having 1-10 carbon atoms; and R.sub.3 can be, for example,
hydrogen or a hydrocarbon as defined hereinabove.
[0146] Thus, representative examples of organic acids from which an
organic acid moiety according to the present invention can be
derived include oxalic acid, malonic acid, succinic acid, glutaric
acid, maleic acid, fumaric acid, phthalic acid, isophthalic acid,
tetraphthalic acid, butyric acid, 4-phenylbutyric acid,
4-aminobutyric acid (GABA), valeric acid, propionic acid, retinoic
acid, acetyl salicylic acid and ibuprofen.
[0147] According to another embodiment of the present invention,
the second chemical moiety of the chemical conjugates is a GABA
agonist. Since, as mentioned hereinabove, GABA agonists alone have
been shown to alleviate pain, conjugates containing such a moiety
may release the GABA agonist in the brain and thus exert a dual,
preferably synergistic, pain relief effect resulting from both the
GABA agonist and the psychotropic drug.
[0148] As used herein, the phrase "GABA agonist" describes
compounds that are capable of activating the GABA system in the
brain, either directly or indirectly, including compounds that
directly bind the GABA receptor or to any other receptor that
affects the GABA system, and are therefore pharmacologically
related to GABA. The term "GABA agonist" is hence understood to
include GABA itself.
[0149] Thus, GABA agonists, according to the present invention,
include, in addition to GABA (.gamma.-aminobutyric acid) itself,
other GABA agonist which can be covalently coupled to a
psychotropic drug.
[0150] Examples of such GABA agonists include (.+-.) baclofen,
isonipecotic acid, .gamma.-hydroxybutyric acid, aminooxyacetic
acid, .beta.-(4-chlorophenyl)-.gamma.-aminobutyric acid,
piperidine-4-sulfonic acid, 3-aminopropylphosphonous acid,
3-aminopropylphosphinic acid, 3-(aminopropyl)methylphosphinic acid,
1-(aminomethyl)cyclohexaneacetic acid (gabapentin),
4-amino-5-hexenoic acid (y-vinyl GABA, vigabatrin) and
3-(2-imidazolyl)-4-aminobutanoic acid.
[0151] The organic acid, according to the present invention, is
selected so as to enhance the therapeutic effect induced by the
psychotropic drug. In addition, the organic acid may be selected to
reduce the side effects that could be induced by the psychotropic
drug if administered alone.
[0152] The phrase "to enhance the therapeutic effect" as used
herein refers to the enhancement of a pain alleviating activity of
the conjugates of the present invention, which is higher than that
of the psychotropic agent and/or the organic acid that form the
conjugate, when administered per se. As is demonstrated in the
Examples section that follows, such an enhanced therapeutic
activity is typically characterized by reduced effective
concentrations of the drug that are required to achieve a certain
therapeutic activity, as compared to the effective concentrations
of the non-conjugated psychotropic agent and/or the organic
acid.
[0153] The phrase "side effects" as used herein refers to adverse
symptoms that may develop as a result of administering to a subject
a certain drug and particularly a psychotropic drug.
[0154] According to a preferred embodiment of the present
invention, the second chemical moiety in the chemical conjugates of
the present invention is covalently linked to the first chemical
moiety via a bond such as, for example, a carboxylic ester bond, an
oxyalkyl carboxylic ester bond, an amide bond or a thioester
bond.
[0155] As used herein, the phrase "carboxylic ester bond" includes
an "--O--C(.dbd.O)-" bond.
[0156] As used herein, the phrase "oxyalkyl carboxylic ester bond"
includes an "O--R--O--C(.dbd.O)-" bond, where R is an alkyl, as
defined hereinabove. Preferably R is methyl.
[0157] The phrase "amide bond" includes a "--NH--C(.dbd.O)-"
bond.
[0158] The phrase "thioester bond" includes a "--S--C(.dbd.O)-"
bond.
[0159] Such bonds are known to be hydrolizable by brain derived
enzymes, such as esterases and amidases, and it is therefore
assumed that the chemical conjugates of the present inventions act
as prodrugs that are metabolized in the brain and thereby
simultaneously release the psychotropic drug and the organic acid,
thus, providing for advantageous co-pharmacokinetics for the
psychotropic drug and the organic acid.
[0160] Alternatively, the second chemical moiety in the chemical
conjugates of the present invention is covalently linked to the
first chemical moiety via an imine bond.
[0161] As used herein, the term "imine bond" describes a
--C.dbd.NH-- bond. An imine bond is also known in the art as a
"Schiff base".
[0162] Herein, the term "treating" includes abrogating,
substantially inhibiting, slowing or reversing the progression of a
pain or substantially preventing the onset of a pain.
[0163] The term "administering" as used herein refers to a method
for bringing a chemical conjugate of the present invention into an
area or a site in the brain that has an effect on pain.
[0164] The chemical conjugate of the present invention can be
administered intraperitoneally. More preferably, it is administered
orally.
[0165] The term "subject" refers to animals, typically mammals
having a blood brain barrier, including human beings.
[0166] The term "therapeutically effective amount" refers to that
amount of the chemical conjugate being administered which will
relieve to some extent the sensation of pain.
[0167] Thus, for example a therapeutically effective amount of
nortriptyline-GABA ranges between 0.05 mg/kg body and 20 mg/kg
body, more preferably between 0.1 mg/kg body and 4 mg/kg body and
most preferably between 0.2 mg/kg body and 1.0 mg/kg body. A
therapeutically effective amount of fluoxetine-GABA ranges between
1 mg/kg body and 40 mg/kg body, more preferably between 5 mg/kg
body and 40 mg/kg body and most preferably between 10 mg/kg body
and 30 mg/kg body. Preferably, the therapeutically effective amount
of the conjugates of the present invention is lower than that used
by the parent compound to exert a similar effect.
[0168] According to an additional aspect of the present invention,
the conjugates of the present invention are used to treat an
addictive disorder. As used herein, the phrase "addictive disorder"
refers to a disorder characterized by a collection of symptoms
(i.e., a syndrome) that is caused by a pathological response to the
ingestion of mood altering substances. The symptoms include
euphoria, craving, loss of control, withdrawal and inability to
abstain. Examples of addictive disorders include but are not
limited to an eating disorder, an addiction to narcotics,
alcoholism and smoking.
[0169] Antidepressants (e.g. nortriptyline) were shown to be
effective in combination with transdermal nicotine in increasing
smoking cessation rate with little effect on withdrawal symptoms
[Prochazka et al., Arch Intern Med. 2004; 164:2229-2233].
Therefore, the GABA conjugates of the present invention can also
enhance this inherent therapeutic characteristic of an
antidepressant.
[0170] Likewise, both antidepressants and anticonvulsants have been
shown to be effective in the treatment of alcohol dependence
[Williams S. American Family Physician, Nov. 1, 2005, Volume 72,
Number 9]. Thus, the GABA conjugates of the present invention can
also enhance this inherent aspect of antidepressants and
anti-convulsants.
[0171] In addition, conjugates of the present invention may be used
to treat other diseases or disorders which are effectively treated
by antidepressants or anti-convulsants. For example, in a mouse
model of sudden unexpected death syndrome, antidepressants were
shown to decrease the incidence of respiratory arrest [Tupal, S.
Epilepsia, 47(1):21-26, 2006, Blackwell Publishing, Inc. C 2006
International League Against Epilepsy]. Thus, antidepressant-GABA
conjugates as described herein may be used to prevent this
side-effect of epilepsy.
[0172] According to another aspect of the present invention, there
is provided a chemical conjugate comprising a first chemical moiety
covalently linked to a second chemical moiety, wherein each of the
chemical moieties is an independent GABA agonist. According to one
embodiment of this aspect of the present invention, the chemical
conjugate comprises two independent .gamma.-aminobutyric acids
(GABA) linked by an alkyloxy carboxylic ester bond.
[0173] These chemical conjugates may be synthesized following the
general procedure described in U.S. Patent Application No.
20040242570 and WO 2005/092392. The synthesis of an exemplary
conjugate according to this aspect of the present invention is
described in Example 1 of the Examples section hereinbelow.
[0174] The conjugates according to this aspect of the present
invention can be beneficially utilized for treating various
CNS-associated diseases and disorders and are particularly
beneficial for treating CNS-associated diseases and disorders that
involve low levels of GABA in the brain. These include, for
example, a pain disorder, a motion disorder such as Parkinson's,
Multiple Sclerosis, action tremors and tardive dyskinesia, a
dissociative disorder, a mood disorder such as panic, anxiety,
depression, an addictive disorder, manic behavior, an affective
disorder, a neurodegenerative disease or disorder such as
Alzheimer's and a convulsive disorder such as epilepsy.
[0175] Hence, according to another aspect of the present invention,
there is provided a method of treating a CNS disease or disorder,
which is effected by administering to a subject in need thereof a
therapeutically effective amount of a chemical conjugate as
described in this context of the present invention.
[0176] Accordingly, these chemical conjugates can be used for
preparing a medicament, whereby such a medicament is preferably for
treating a CNS disease or disorder.
[0177] Representative examples of CNS diseases or disorders that
can be beneficially treated with the conjugates described herein
include, but are not limited to, a pain disorder, a motion
disorder, a dissociative disorder, a mood disorder, an affective
disorder, a neurodegenerative disease or disorder and a convulsive
disorder.
[0178] More specific examples of such conditions include, but are
not limited to, Parkinson's, Multiple Sclerosis, Huntington's
disease, action tremors and tardive dyskinesia, panic, anxiety,
depression, alcoholism, insomnia and manic behavior, Alzheimer's
and epilepsy.
[0179] In any of the methods described herein, the chemical
conjugate can be administered either as is, or, preferably as a
part of a pharmaceutical composition that further comprises a
pharmaceutically acceptable carrier.
[0180] As used herein a "pharmaceutical composition" refers to a
preparation of one or more of the chemical conjugates described
herein, with other chemical components such as pharmaceutically
suitable carriers and excipients. The purpose of a pharmaceutical
composition is to facilitate administration of a compound to a
subject.
[0181] Hereinafter, the term "pharmaceutically acceptable carrier"
refers to a carrier or a diluent that does not cause significant
irritation to a subject and does not abrogate the biological
activity and properties of the administered compound. Examples,
without limitations, of carriers are propylene glycol, saline,
emulsions and mixtures of organic solvents with water.
[0182] Herein the term "excipient" refers to an inert substance
added to a pharmaceutical composition to further facilitate
administration of a compound. Examples, without limitation, of
excipients include calcium carbonate, calcium phosphate, various
sugars and types of starch, cellulose derivatives, gelatin,
vegetable oils and polyethylene glycols.
[0183] According to a preferred embodiment of the present
invention, the pharmaceutical carrier is an aqueous solution of
lactic acid.
[0184] Techniques for formulation and administration of drugs may
be found in "Remington's Pharmaceutical Sciences," Mack Publishing
Co., Easton, Pa., latest edition, which is incorporated herein by
reference.
[0185] Suitable routes of administration may, for example, include
oral, rectal, transmucosal, transdermal, intestinal or parenteral
delivery, including intramuscular, subcutaneous and intramedullary
injections as well as intrathecal, direct intraventricular,
intravenous, intraperitoneal, intranasal, or intraocular
injections. Pharmaceutical compositions of the present invention
may be manufactured by processes well known in the art, e.g., by
means of conventional mixing, dissolving, granulating,
dragee-making, levigating, emulsifying, encapsulating, entrapping
or lyophilizing processes. Pharmaceutical compositions for use in
accordance with the present invention thus may be formulated in
conventional manner using one or more pharmaceutically acceptable
carriers comprising excipients and auxiliaries, which facilitate
processing of the active compounds into preparations which can be
used pharmaceutically. Proper formulation is dependent upon the
route of administration chosen.
[0186] For injection, the chemical conjugates of the invention may
be formulated in aqueous solutions, preferably in physiologically
compatible buffers such as Hank's solution, Ringer's solution, or
physiological saline buffer with or without organic solvents such
as propylene glycol, polyethylene glycol. For transmucosal
administration, penetrants are used in the formulation. Such
penetrants are generally known in the art.
[0187] For oral administration, the chemical conjugates can be
formulated readily by combining the active compounds with
pharmaceutically acceptable carriers well known in the art. Such
carriers enable the conjugates of the invention to be formulated as
tablets, pills, dragees, capsules, liquids, gels, syrups, slurries,
suspensions, and the like, for oral ingestion by a patient.
Pharmacological preparations for oral use can be made using a solid
excipient, optionally grinding the resulting mixture, and
processing the mixture of granules, after adding suitable
auxiliaries if desired, to obtain tablets or dragee cores. Suitable
excipients are, in particular, fillers such as sugars, including
lactose, sucrose, mannitol, or sorbitol; cellulose preparations
such as, for example, maize starch, wheat starch, rice starch,
potato starch, gelatin, gum tragacanth, methyl cellulose,
hydroxypropylmethyl-cellulose, sodium carbomethylcellulose and/or
physiologically acceptable polymers such as polyvinylpyrrolidone
(PVP). If desired, disintegrating agents may be added, such as
cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt
thereof such as sodium alginate.
[0188] Dragee cores are provided with suitable coatings. For this
purpose, concentrated sugar solutions may be used which may
optionally contain gum arabic, talc, polyvinyl pyrrolidone,
carbopol gel, polyethylene glycol, titanium dioxide, lacquer
solutions and suitable organic solvents or solvent mixtures.
Dyestuffs or pigments may be added to the tablets or dragee
coatings for identification or to characterize different
combinations of active compound doses.
[0189] Pharmaceutical compositions, which can be used orally,
include push-fit capsules made of gelatin as well as soft, sealed
capsules made of gelatin and a plasticizer, such as glycerol or
sorbitol. The push-fit capsules may contain the active ingredients
in admixture with filler such as lactose, binders such as starches,
lubricants such as talc or magnesium stearate and, optionally,
stabilizers. In soft capsules, the active compounds may be
dissolved or suspended in suitable liquids, such as fatty oils,
liquid paraffin, or liquid polyethylene glycols. In addition,
stabilizers may be added. All formulations for oral administration
should be in dosages suitable for the chosen route of
administration.
[0190] For buccal administration, the compositions may take the
form of tablets or lozenges formulated in conventional manner.
[0191] For administration by inhalation, the chemical conjugates
for use according to the present invention are conveniently
delivered in the form of an aerosol spray presentation from a
pressurized pack or a nebulizer with the use of a suitable
propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane,
dichloro-tetrafluoroethane or carbon dioxide. In the case of a
pressurized aerosol, the dosage unit may be determined by providing
a valve to deliver a metered amount. Capsules and cartridges of,
e.g., gelatin for use in an inhaler or insufflator may be
formulated containing a powder mix of the compound and a suitable
powder base such as lactose or starch.
[0192] The chemical conjugates described herein may be formulated
for parenteral administration, e.g., by bolus injection or
continuous infusion. Formulations for injection may be presented in
unit dosage form, e.g., in ampoules or in multidose containers with
optionally, an added preservative. The compositions may be
suspensions, solutions or emulsions in oily or aqueous vehicles,
and may contain formulatory agents such as suspending, stabilizing
and/or dispersing agents.
[0193] Pharmaceutical compositions for parenteral administration
include aqueous solutions of the active compound in water-soluble
form. Additionally, suspensions of the active compounds may be
prepared as appropriate oily injection suspensions. Suitable
lipophilic solvents or vehicles include fatty oils such as sesame
oil, or synthetic fatty acids esters such as ethyl oleate,
triglycerides or liposomes. Aqueous injection suspensions may
contain substances, which increase the viscosity of the suspension,
such as sodium carboxymethyl cellulose, sorbitol or dextran.
Optionally, the suspension may also contain suitable stabilizers or
agents which increase the solubility of the conjugates to allow for
the preparation of highly concentrated solutions. Alternatively,
the active ingredient may be in powder form for constitution with a
suitable vehicle, e.g., sterile, pyrogen-free water, before
use.
[0194] The chemical conjugates of the present invention may also be
formulated in rectal compositions such as suppositories or
retention enemas, using, e.g., conventional suppository bases such
as cocoa butter or other glycerides.
[0195] The pharmaceutical compositions herein described may also
comprise suitable solid of gel phase carriers or excipients.
Examples of such carriers or excipients include, but are not
limited to, calcium carbonate, calcium phosphate, various sugars,
starches, cellulose derivatives, gelatin and polymers such as
polyethylene glycols.
[0196] Pharmaceutical compositions suitable for use in context of
the present invention include compositions wherein the active
ingredients are contained in an amount effective to achieve the
intended purpose. More specifically, a therapeutically effective
amount means an amount of chemical conjugate effective to prevent,
alleviate or ameliorate pain.
[0197] Determination of a therapeutically effective amount is well
within the capability of those skilled in the art, especially in
light of the detailed disclosure provided herein.
[0198] For any chemical conjugate used in the methods of the
invention, the therapeutically effective amount or dose can be
estimated initially from activity assays in cell cultures and/or
animals. For example, the effectiveness of the conjugates may be
assayed on rats which have undergone an L5/L6 spinal nerve ligation
surgical procedure. Specifically, left paraspinal muscles are
dissected from the spinous processes at the levels of L4 through S2
of anaesthetized rats and their L5 and L6 spinal nerves are
isolated. Each spinal nerve is tightly ligated with e.g. a 4-0 silk
suture distal to the dorsal root ganglion. Following spinal nerve
ligation, the wound is sutured, and the skin is closed. Ten to 14
days following spinal nerve ligation, the rats are typically placed
in individual Plexiglas chambers on an elevated wire mesh where
they are allowed to acclimate. Following the acclimation period,
rats are tested for tactile allodynia by applying a series of
calibrated von Frey filaments to the plantar aspect of the left
hind paw ipsilateral to the site of nerve injury. The mean 50%
withdrawal threshold (g) is determined [Li Y, Dorsi M J, Meyer R A,
Belzberg A J. Pain. 2000; 85(3):493-502]. Rats that display a
predrug withdrawal threshold >4 g are generally not considered
allodynic and are excluded from the study. Following determination
of predrug withdrawal thresholds, rats may be treated with either
conjugates of the present invention or vehicle, and effects on
tactile allodynia are determined over time by measuring hind paw
withdrawal thresholds for time intervals such as 30, 60, 90, and
120 min post-injection.
[0199] The conjugates of the present invention were also tested on
rats or mice using the hot plate test or formalin test as described
below in Examples 2, 3 and 5 respectively.
[0200] The conjugates of the present invention were also tested by
analyzing their ability to prevent secretion of inflammatory
cytokines as described in Example 6 hereinbelow.
[0201] For example, a dose can be formulated in animal models to
achieve a circulating concentration range that includes the 1050 as
determined by activity assays (e.g., the concentration of the test
compound, which achieves a half-maximal inhibition of the
proliferation activity). Such information can be used to more
accurately determine useful doses in humans.
[0202] Toxicity and therapeutic efficacy of the chemical conjugates
described herein can be determined by standard pharmaceutical
procedures in experimental animals, e.g., by determining the 1050
and the LD50 (lethal dose causing death in 50% of the tested
animals) for a subject compound. The data obtained from these
activity assays and animal studies can be used in formulating a
range of dosage for use in human.
[0203] Side effects of the conjugates of the present invention at
doses which are effective at alleviating pain may be assayed using
standard assays known in the art. For example, in order to
investigate the possible non-specific muscle relaxant or sedative
effects of the conjugates of the present invention mice or rats may
be treated with the conjugates of the present invention, and
subsequently tested on a rota-rod for motility. This apparatus
consists of a bar with a diameter of 3.5 cm mice and 7 cm for rats,
subdivided into four compartments. The bar rotates at a constant
speed of 0.5-22 revolutions per minute and the animals are
evaluated for the time taken to fall from the bar. The animals may
be selected 24 hours previously by eliminating those which do not
remain on the bar for 60 seconds. Results are typically expressed
as the length of time animals remain on the rota-rod bar.
[0204] The dosage may vary depending upon the dosage form employed
and the route of administration utilized. The exact formulation,
route of administration and dosage can be chosen by the individual
physician in view of the patient's condition. (See e.g., Fingl, et
al., 1975, in "The Pharmacological Basis of Therapeutics", Ch. 1 p.
1).
[0205] Dosage amount and interval may be adjusted individually to
provide plasma levels of the active moiety which are sufficient to
maintain the pain alleviating effects, termed the minimal effective
concentration (MEC). The MEC will vary for each preparation, but
can be estimated from in vitro and/or in vivo data, as described
hereinabove. Dosages necessary to achieve the MEC will depend on
individual characteristics and route of administration. HPLC assays
or bioassays can be used to determine plasma concentrations.
[0206] Dosage intervals can also be determined using the MEC value.
Preparations should be administered using a regimen, which
maintains plasma levels above the MEC for 10-90% of the time,
preferable between 30-90% and most preferably 50-90%.
[0207] Depending on the severity and responsiveness of the pain to
be treated, dosing can also be a single administration of a slow
release composition described hereinabove, with course of treatment
lasting from several days to several weeks or until cure is
effected or diminution of the disease state is achieved.
[0208] The amount of a composition to be administered will, of
course, be dependent on the subject being treated, the severity of
the pain, the manner of administration, the judgment of the
prescribing physician, etc.
[0209] Compositions of the present invention may, if desired, be
presented in a pack or dispenser device, such as a FDA approved
kit, which may contain one or more unit dosage forms containing the
active ingredient. The pack may, for example, comprise metal or
plastic foil, such as a blister pack. The pack or dispenser device
may be accompanied by instructions for administration. The pack or
dispenser may also be accompanied by a notice associated with the
container in a form prescribed by a governmental agency regulating
the manufacture, use or sale of pharmaceuticals, which notice is
reflective of approval by the agency of the form of the
compositions or human or veterinary administration. Such notice,
for example, may be of labeling approved by the U.S. Food and Drug
Administration for prescription drugs or of an approved product
insert. Compositions comprising the chemical conjugates of the
invention formulated in a compatible pharmaceutical carrier may
also be prepared, placed in an appropriate container, and labeled
for the treatment of pain. Compositions comprising conjugates of
two GABA agonists may also be labeled for the treatment of other
disorders e.g. motion disorders such as Parkinson's, Huntingdon's
disease, Multiple Sclerosis, action tremors and tardive dyskinesia,
dissociative disorders, mood disorders such as panic, anxiety,
depression, alcoholism, insomnia and manic behavior, affective
disorders, neurodegenerative diseases or disorders such as
Alzheimer's and convulsive disorders such as epilepsy.
[0210] Hence, according to preferred embodiments of the present
invention, the pharmaceutical compositions of the present invention
are packaged in a packaging material and are identified in print,
on or in the packaging material, for alleviating pain.
[0211] According to a further aspect of the present invention,
there is provided an article-of-manufacture, which comprises a
pharmaceutical composition as described hereinabove, being packaged
in a packaging material and identified in print, in or on the
packaging material for use in the treatment of pain, as described
herein. The pharmaceutical composition includes a pharmaceutically
acceptable carrier, as described herein and any of the chemical
conjugates described herein.
[0212] According to yet a further aspect of the present invention,
there is provided an article-of-manufacture, which comprises a
pharmaceutical composition as described hereinabove, being packaged
in a packaging material and identified in print, in or on the
packaging material for use in the treatment of an addictive
disorder, as described herein.
[0213] Additional objects, advantages, and novel features of the
present invention will become apparent to one ordinarily skilled in
the art upon examination of the following examples, which are not
intended to be limiting. Additionally, each of the various
embodiments and aspects of the present invention as delineated
hereinabove and as claimed in the claims section below finds
experimental support in the following examples.
EXAMPLES
[0214] Reference is now made to the following examples, which
together with the above descriptions, illustrate the invention in a
non limiting fashion.
Example 1
Preparation of GABA Conjugates of Antidepressant and Antiepileptic
Drugs
Synthesis of 2-Propyl-pentanoic acid (valproic acid)
4-aminobutyryloxymethyl ester hydrochloride (AN-216)
[0215] To a solution of 2-propyl-pentanoic acid
N-t-boc-4-amino-butyryloxymethyl ester (AN-217, prepared as
described in WO 2005/092392, 1.7 grams, 4.7 mmol) in ethyl acetate,
a solution of 4N HCl in ethyl acetate was added. The obtained
mixture was stirred for 4 hours at room temperature, the solvent
was thereafter evaporated and the residue was further dried under
high vacuum. The residue was dissolved in ether, and addition of
hexane lead to precipitation of the desired product AN-216 (0.75
gram, 62%) as an amorphous solid having a melting point of
35-37.degree. C.
[0216] .sup.1H-NMR (CD.sub.3OD): .delta.=0.9 (t, J=7.1 Hz, 6H, two
CH.sub.3), 1.2-1.64 (m, 8H, two CH.sub.2CH.sub.2Me), 1.95 (q, J=7.5
Hz, 2H, CH.sub.2CH.sub.2CH.sub.2), 2.4-2.5 (m, 1H, CHCO), 2.53 (t,
J=7.2 Hz, 2H, CH.sub.2CO), 2.99 (t, J=7.2 Hz, 2H, CH.sub.2N), 5.77
(s, 2H, OCH.sub.2O).
[0217] .sup.13C-NMR (CD.sub.3OD): .delta.=14.2 (two CH.sub.3), 21.5
(two MeCH.sub.2), 23.5 (CH.sub.2CH.sub.2CH.sub.2), 31.3
(COCH.sub.2), 35.5 (two CH.sub.2CH), 39.9 (NCH.sub.2), 46.2 (CH),
80.6 (OCH.sub.2O), 172.5 (CH.sub.2CO), 176.4 (CHCO.sub.2).
[0218] MS (CI/NH.sub.3): m/z (%)=260 (MH.sup.+, 100).
Synthesis of
N-(3-(4-(trifluoromethyl)phenoxy)-3-phenylpropyl)-4-amino-N-methylbutanam-
ide Hydrochloride (AN-227)
[0219] tert-Butyl
3-(N-(3-(4-(Trifluoromethyl)phenoxy)-3-phenylpropyl)-N-methyl
carbamoyl)propylcarbamate (AN-229) was prepared as described in WO
2005/092392. AN-229 (0.5 mmol) was added to a solution of HCl in
EtOAc (35 ml) and the resulting mixture was stirred for 3 hours.
The solvent was thereafter evaporated and the product, AN-227, was
obtained in quantitative yield.
[0220] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta.=1.9 (quint,
J=7.25 Hz, 2H), 2.52 (t, J=6.9 Hz, 2H), 2.87 (m, t J=7.38 Hz, 2H,
minor), 2.93 (s, 3H, NMe minor), 2.97 (t, J=7.13 Hz, 3H, NMe,
major), 3.03 (s, 3H, major), 3.59-3.67 (m, 2H), 5.37 (dd, J=8.6,
4.18 Hz, 1H, major), 5.42 (dd, J=8.57, 4.12 Hz, 1H, minor),
6.99-7.05 (m, 2H), 7.24-7.49 (m, 7H) ppm.
[0221] .sup.13C NMR (300 MHz, CDCl.sub.3): .delta.=23.7 (major),
23.9 (minor), 30.9 (minor), 31.5 (major), 33.8 (minor), 36.1
(major), 37.1 (major), 37.9 (minor), 40.42, 46.3 (major), 47.5
(minor), 78.5 (minor), 79.4 (major), 117.2, 124.1, 127.3, 127.8,
129.0 (major), 129.2 (minor), 129.7, 129.9 ppm.
Synthesis of
10,11-Dihydro-5-(3-methylaminopropylidene)-5H-dibenzo[a,d][1,4]cyclohepte-
ne 4-Amino-N-butanamide Hydrochloride (AN-228)
[0222] tert-Butyl
3-(10,11-Dihydro-5H-dibenzo[a,d]cycloheptane-5-ylidene-N-methyl-1-propana-
mine-3-(methylcarbamoyl)propyl carbamate (AN-230) was prepared as
described in WO 2005/092392. AN-230 (0.5 mmol) was added to a
solution of HCl in EtOAc (35 ml) and the resulting mixture was
stirred for 3 hours. The solvent was thereafter evaporated and the
product was obtained in quantitative yield in the form of two
rotamers.
[0223] .sup.1H NMR (300 MHz, CDCl.sub.3): .delta.=1.67 (t, J=6.9
Hz, 1H), 1.8 (t, J=7.07 Hz, 1H), 2.1-2.35 (m, 2H), 2.4 (q, J=6.7
Hz, 2H), 2.6-2.9 (m, 5H), 3.3-3.4 (m, 2H), 5.76 (dt, J=7.86, 7.76
Hz, 1H), 6.7-7.3 (m, 8H) ppm.
Synthesis of GABA oxymethylGABA (AN-214)
Preparation of 4-tert-Butoxycarbonylamino-butyric acid
##STR00001##
[0225] To a solution of .gamma.-aminobutyric acid (1 equivalent) in
t-BuOH (4 ml/gram) and H.sub.2O (3 ml/gram) was added NaOH (1
equivalent), BOC.sub.2O (1 equivalent), and the mixture was stirred
at room temperature for 24 hours. The solvent was thereafter
evaporated and the residue was partitioned between hexane and
water. The aqueous layer was acidified with KHSO.sub.4 1N to reach
pH=2, and was thereafter extracted with CH.sub.2Cl.sub.2. The
organic layer was dried over MgSO.sub.4 and the solvent was
evaporated to give the desired product as a white solid (85%
yield).
[0226] .sup.1H-NMR (200 MHz) CDCl.sub.3: .delta.=1.44 (s, 9H,
t-Bu), 1.81 (quint, J=7.5 Hz, 2H, CH.sub.2CH.sub.2CH.sub.2), 2.37
(t, J=7.5 Hz, 2H, CH.sub.2CO.sub.2), 3.17 (m, 2H, CH.sub.2NH), 4.71
(bs, 1H, NH) ppm.
[0227] .sup.13C-NMR (200 MHz) CDCl.sub.3: .delta.=25.3
(CH.sub.2CH.sub.2CH.sub.2), 28.4 (Me.sub.3C), 31.1
(CH.sub.2CO.sub.2H), 39.7 (CH.sub.2NH), 79.65 (CMe.sub.3), 156.2
(NCO.sub.2), 176.9 (CO.sub.2H) ppm.
[0228] MS (CI/NH.sub.3): m/z (%)=204 (MH.sup.+, 24.3), 148
(MH.sup.+-C.sub.4H.sub.8, 100), 130
(MH.sup.+-C.sub.4H.sub.10O).
Preparation of 4-tert-Butoxycarbonylamino-butyric acid chloromethyl
ester
##STR00002##
[0230] To a mixture of chloromethyl chlorosulfate (1.1 equivalent)
and 4-tert-Butoxycarbonylamino-butyric acid (1 equivalent) was
added NaHCO.sub.3 (3.6 equivalents) and Bu.sub.4NHSO.sub.4
(catalytic amount) in water/CH.sub.2Cl.sub.2 (1:1). The mixture was
stirred at room temperature overnight. The organic phase was
thereafter separated and the aqueous phase was extracted with
CH.sub.2Cl.sub.2 (three times). The combined organic layer was
washed with NaHCO.sub.3 (.times.3), brine (.times.3), dried with
MgSO.sub.4, filtered and evaporated.
[0231] The product was purified by flash chromatography, using a
5:1 hexane:EtOAc as an eluent, and was isolated as a yellow oil
(64% yield).
[0232] .sup.1H-NMR (200 MHz) CDCl.sub.3: .delta.=1.44 (s, 9H,
t-Bu), 1.86 (quint, J=6.3 Hz, 2H, CH.sub.2CH.sub.2CH.sub.2), 2.44
(t, J=6.93 Hz, 2H, CH.sub.2CO.sub.2), 3.18 (m, 2H, CH.sub.2NH),
4.75 (bs, 1H, NH), 5.69 (s, 2H, CH.sub.2Cl) ppm.
[0233] .sup.13C-NMR (300 MHz) CDCl.sub.3: .delta.=24.9
(CH.sub.2CH.sub.2CH.sub.2), 28.3 (Me.sub.3C), 31.1
(CH.sub.2CO.sub.2), 39.4 (CH.sub.2NH), 68.6 (CH.sub.2Cl), 79.2
(CMe.sub.3), 155.9 (NCO.sub.2), 171.3 (CO.sub.2H) ppm.
[0234] MS (ES+): m/z (%)=274 (MNa.sup.+, 40.6), 252 (MH.sup.+,
40.2), 196 (MH.sup.+-C.sub.4H.sub.8, 25.9).
Preparation of 4-tert-Butoxycarbonylamino-butyric acid
4-tert-butoxycarbonylamino-butyroloxymethyl ester
##STR00003##
[0236] A mixture of 4-tert-Butoxycarbonylamino-butyric acid (1.2
equivalents) and the chloromethyl ester described above (1
equivalent) in dry ethylmethylketone (EMK) was stirred under
nitrogen atmosphere while triethylamine (1.2 equivalents) was added
dropwise, and the reaction mixture was refluxed overnight. The
formed white precipitate was thereafter filtered, washed with EtOAc
and the filtrate was evaporated. The residue was dissolved in EtOAc
and was washed with NaHCO.sub.3 (.times.3), and brine (x 3), dried
with MgSO.sub.4, filtered and evaporated. The crude product (brown
oil) was purified by flash chromatography, using a 8:1 hexane:EtOAc
mixture as eluent, to give the desired product as a yellow oil (38%
yield).
[0237] .sup.1H-NMR (200 MHz, CDCl.sub.3): .delta.=1.44 (s, 18H, two
t-Bu), 1.83 (quint, J=7.1 Hz, 4H, two CH.sub.2CH.sub.2CH.sub.2),
2.41 (t, J=7.3 Hz, 4H, two CH.sub.2CO.sub.2), 3.16 (t, J=6.8 Hz,
4H, two CH.sub.2NH), 4.67 (bs, 2H, two NH), 5.75 (s, 2H,
OCH.sub.2O) ppm.
[0238] .sup.13C-NMR (300 MHz, CDCl.sub.3): .delta.=25.0
(CH.sub.2CH.sub.2CH.sub.2), 28.4 (Me.sub.3C), 31.1
(CH.sub.2CO.sub.2), 39.7 (CH.sub.2NH), 79.3 (OCH.sub.2O), 155.9
(NCO.sub.2), 172.9 (CO.sub.2CH.sub.2) ppm.
[0239] MS (ES+): m/z (%)=441 (MNa.sup.+, 17.3), 319 (MH.sup.+-BOC,
19), 196 (MH.sup.+-BOC-C.sub.4H.sub.8, 25.9).
Preparation of 4-Amino-butyric acid 4-amino-butyryloxymethyl ester
dihydrochloride (AN-214)
##STR00004##
[0241] To a solution of the N-tert-Boc protected compound in EtOAc,
a solution of 4N HCl in EtOAc was added. The reaction mixture was
stirred for 4 hours at room temperature and the solvent was
thereafter evaporated to give the crude product. The crude product
was recrystallized from a MeOH-ether mixture, filtered and dried
over P.sub.2O.sub.5 under vacuum to give the pure product as a
white solid (90% yield).
[0242] mp: 155-158.degree. C.
[0243] .sup.1H-NMR (200 MHz, CD.sub.3OD): .delta.=1.96 (quint,
J=7.37 Hz, 4H, two CH.sub.2CH.sub.2CH.sub.2), 2.55 (t, J=7.15 Hz,
4H, two CH.sub.2CO.sub.2), 2.99 (t, J=7.6 Hz, 4H, two CH.sub.2NH),
5.76 (s, 2H, OCH.sub.2O) ppm.
[0244] .sup.13C-NMR (300 MHz, CD.sub.3OD): .delta.=23.4
(CH.sub.2CH.sub.2CH.sub.2), 31.3 (CH.sub.2CO.sub.2), 39.9
(CH.sub.2NH.sub.2), 80.8 (OCH.sub.2O), 172.7
(CO.sub.2CH.sub.2).
[0245] MS (ES+): m/z (%)=219 (MH.sup.+, 100), 242 (MNa.sup.+, 90),
104 (MH.sup.+-C.sub.5H.sub.10O.sub.2N, 85) ppm.
[0246] Table 1 below presents the chemical conjugates synthesized
by the methods described hereinabove.
TABLE-US-00001 TABLE 1 AN-214 GABA oxymethylGABA
C.sub.9H.sub.20Cl.sub.2N.sub.2O.sub.4 291.17 ##STR00005## AN-216
GABA-oxymethyl- valproate C.sub.13H.sub.26ClNO.sub.4 295.80
##STR00006## AN-227 Flouxetine-GABA
C.sub.21H.sub.26ClF.sub.3N.sub.2O.sub.2 430.16 C, 58.54; H, 6.08;
Cl, 8.23; F, 13.23; N, 6.50; O, 7.43 ##STR00007## AN-228
Nortriptyline- GABA C.sub.23H.sub.29ClN.sub.2O 384.2 C, 71.76; H,
7.59; Cl, 9.21; N, 7.28; O, 4.16 ##STR00008##
Example 2
Effects of GAB-Antidepressant Drug and GABA-Anti-Epileptic Drug
Conjugates as Determined by the Hot Plate Test
[0247] Central pain perception was assessed by the analgesic
response on a hot plate. The hot-plate test used to measure latency
in response to heat was essentially carried out according to the
method described by Eddy and Leimbach (1953), with the
Materials and Methods
[0248] Animals:
[0249] Male Balb-c mice (8-14 weeks old) were obtained from Harlan,
Israel. The mice were housed under conditions of controlled
temperature (23.+-.3.degree. C.) and humidity (55.+-.15%) with a 12
hour light/12 hour dark cycle. All experiments were carried out in
accordance with the ethical guidelines of the International
Association for the Study of Pain, and for the Committee on the
Care and Use of Laboratory Animals of Tel Aviv University.
[0250] Experimental Procedure:
[0251] Nortriptyline (Sigma, Aldrich), fluoxetine, GABA (Sigma,
Israel), the Nortriptyline-GABA conjugate (AN-228), the
fluoxetine-GABA conjugate (AN-227) and valproyloxymethyl-GABA
conjugate (AN-216) were solubilized in saline and administered to
the mice by gastric gavage. After the indicated time periods, the
animals were placed on a hot plate, (MRC, model-MH-4, 230 V/50 Hz,
750 W) which was maintained at 52.+-.1.degree. C. The time of
response to the heat sensation was detected by one or more of the
following reactions: raising paw, licking of the paw, jumping or
running. Antinociception or analgesia reaction was measured as the
latency to withdrawal evoked by exposing the mice paws to the
thermal stimulus. Data was collected at the following time points:
-60, 0, 120, 180, 240, and 300 minutes following administration of
the drugs.
[0252] Statistical Analysis:
[0253] Data is expressed as mean.+-.SD. The data was analyzed by
the Student's t-test. Values of P<0.05 were considered
statistically significant.
Experimental Results
[0254] Male Balb-c mice (6/group) were treated p.o. with the
indicated doses of nortriptyline, respective equimolar doses
nortriptyline-GABA conjugate or vehicle. Two hours later the
animals were placed on the surface of the hot-plate
(52.+-.1.degree. C.). Response to the heat, manifested by shaking
or licking of the paws or jumping, was recorded as the index of
response latency.
[0255] As illustrated in FIG. 1, 0.5 mg/kg of nortriptyline was
significantly less effective two hours following administration
than an equimolar dose of its GABA conjugate. At this dose,
nortriptyline did not differ significantly (p>0.05) from control
mice treated with vehicle only. At the dose of 4.5 mg/kg
nortriptyline and equimolar dose of its GABA conjugate displayed a
significant delay in the response compared to control mice treated
with vehicle (p<0.05). These results demonstrate that the
GABA-nortriptyline conjugate, imparted antinociceptive activity at
a lower concentration than nortriptyline, demonstrating its
advantage. Saturation in response by nortriptyline GABA was seen
since equimolar dose of 0.5 mg/kg did not differ significantly from
equimolar dose of 4.5 mg/kg.
[0256] A time course of the response to heat by mice treated with
nortriptyline (0.25 mg/kg and 0.5 mg/kg) and an equimolar dose of
nortriptyline-GABA (0.32 mg/kg and 0.64 mg/kg, respectively) is
shown in FIGS. 2A-B. At a dose of 0.25 mg/kg, 4 hours following
treatment, the analgesic effect of nortriptyline and its GABA
conjugate increased significantly the latency of response compared
to untreated mice (FIG. 2A). The conjugate exhibited significantly
greater delay in response compared to the untreated mice at 4 and 6
hours post treatment and at 4 hours its antinociceptive effect was
significantly greater than that of nortriptyline. This indicates
that nortriptyline-GABA, compared to nortriptyline, is a more
potent analgesic drug and its effect lasts longer. Two hours
following a 0.5 mg/kg treatment, the conjugate caused a significant
antinociceptive effect (compared to nortriptyline and vehicle
treated mice) as illustrated in FIG. 2B. Un-conjugated
Nortriptyline on the other hand, imparted a significant latency,
(compared to vehicle treated animals), after 4 hours only. This
indicates that not only does the additional GABA moiety increase
the antinociceptive activity, but it also accelerates the onset of
the protective effect.
[0257] The effect of repeated oral treatment with the drugs on
response to heat was tested in mice (6/group) for nortriptyline
(0.2 mg/kg) and its GABA conjugate (AN-228) at equimolar doses. The
drugs were administered daily for 15 days and the treatment effect
on heat sensation was measured trice a week. As illustrated in FIG.
3A, during the first week, the animals response to heat sensation
prior to treatment (time 0), did not differ and was unaffected by
the treatment. From the eighth day, a sustained latency response
was noted prior to treatment and mostly in the nortriptyline-GABA
conjugate and to a lesser extent in the nortriptyline treated mice.
From the third day of treatment and during the five hours of
testing (FIGS. 3B-E), typically, the conjugate imparted a
significantly better antinociceptive effect compared to untreated
animals or those treated with nortriptyline. A significantly
improved antinociceptive effect compared to untreated animals and
those treated with nortriptyline was observed 4 and 5 h after
treatment with the daily dose from the first day of treatment. The
results indicate that the nortriptyline-GABA conjugate produced a
sustained and more effective analgesic response than nortriptyline.
Repeated administration shortened the onset of the antinociceptive
effect as well as increased the latency of the response. The
antinociceptive effects of the SSRI fluoxetine and its GABA
conjugate (AN-227) were tested and compared in the hot-plate test.
A time course of the response to heat by mice treated with
fluoxetine (10 mg/kg) and equimolar dose of fluoxetine-GABA is
shown in FIG. 4. At 2, 3 and 4 hours following treatment, the
analgesic effect of fluoxetine-GABA conjugate on mice was
significantly greater than that of untreated mice or mice treated
with equimolar dose of GABA or fluoxetine. The observation
indicates that fluoxetine-GABA, compared to fluoxetine, is a more
potent analgesic and its effect lasts longer.
[0258] The effect of repeated treatment with fluoxetine and its
GABA conjugate on response to heat was tested. Results are
indicated in FIGS. 5A-E. Fluoxetine (10 mg/kg) and its conjugate at
an equimolar dose were given orally to mice daily for 11 days and
the treatment effect on heat sensation was tested at days 3, 7, 9
and 11. Delayed response to heat sensation in mice receiving
fluoxetine-GABA was noted on days 3, 9 and 11 prior to treatment
(FIG. 5A). A delay was noted only on day 11 with fluoxetine treated
mice compared to vehicle treated animals. Typically during the 11
days of treatment, the GABA-fluoxetine conjugate exerted a
significantly improved antinociceptive effect compared to
fluoxetine or untreated animals. Fluoxetine treated mice also
showed a significantly better antinociceptive effect compared to
untreated animals. The results indicate that the fluoxetine-GABA
conjugate produced a lasting and more effective analgesic response
than fluoxetine.
[0259] Antiepileptic drugs, encompassing valproate and GABA and
their analogs, have been shown to be effective analgesics. Since
valproic acid crosses the BBB, a valproate-GABA conjugate could
carry the GABA to the brain and the GABA in turn could potentiate
the activity of valproic acid. 3, 4 and 5 hours following oral
administration of 0.2 mg of valproyloxymethyl GABA, a significant
latency in response to heat sensation compared to the untreated
control mice, was observed as illustrated in FIG. 6. In parallel,
GABA administered at >10-folds higher dose (2.39 mg/kg), had no
activity.
[0260] A conjugate comprising two independent GABAs (AN-214) was
also assayed for its pain relieving activities. Male Balb/c mice
were treated orally for 5 consecutive days with either
GABAoxymethylGABA (n=11), an equivalent molar dose of
valproyloxyethyl GABA (n=11) or vehicle (n=8). As illustrated in
FIG. 7, no antinociceptive activity was detected during the entire
period.
[0261] Chronic administration of GABA neither had a cumulative
effect on analgesia nor contributed to the antinociceptive effect
as illustrated in FIG. 8. The mice were treated orally for 5
consecutive days with three different doses of GABA. No
antinociceptive activity was detected during the entire period.
Example 3
Effects of GABA-Antidepressant Drug and GABA-Anti-Epileptic Drug
Conjugates as Determined by the Formalin Test
[0262] Intraplantar injection of formalin was used to assess the
second, peripheral phase of pain.
Materials and Methods
[0263] Animals:
[0264] Balb-c mice at 10 and 12 weeks of age were obtained from
Harlan, Israel. Mice were housed under conditions of controlled
temperature (23.+-.3.degree. C.) and humidity (55.+-.15%) with a 12
hour light/12 hour dark cycle. All experiments were carried out in
accordance with the ethical guidelines of the International
Association for the Study of Pain, and for the Committee on the
Care and Use of Laboratory Animals of Tel Aviv University.
[0265] Experimental Procedure
[0266] (A) Fluoxetine:
[0267] Five mice were used in each group and three hours following
oral administration of either fluoxetine (10 or 30 mg/kg) or
equimolar doses of the fluoxetine conjugate (AN-227), a 1% formalin
solution was injected subcutaneously into the dorsal surface of the
right hind paw.
[0268] (B) Nortriptyline:
[0269] Eight mice were used in each group and three hours following
oral administration of either nortriptyline (0.5 or 5 mg/kg) or
equimolar doses of the nortriptyline conjugate (AN-228), a 1%
formalin solution was injected subcutaneously into the dorsal
surface of the right hind paw.
[0270] The formalin induced typical biphasic flinching behavior of
the injected paw. The animals were returned to a glass chamber and
the total time spent by the animal licking or biting the injected
paw was counted. The frequency of pain-related behaviors was
recorded during the early phase (0-5 min after injection) and the
late phase (25-35 min after injection).
Experimental Results
[0271] As illustrated in FIGS. 9A-B, both doses of the fluoxetine
conjugates of the present invention decreased significantly
(p<0.05) the early neurogenic response to pain as compared to
fluoxetine alone and the higher dose of the fluoxetine conjugate
(equivalent to 30 mg/kg) reduced significantly (p<0.05) the late
inflammatory peripheral response. As further illustrated in FIGS.
10A-B, both doses of the nortriptyline conjugates (equivalent of
0.5 and 5 mg/kg) of the present invention decreased significantly
(p<0.05) the early neurogenic response and the late inflammatory
peripheral response to pain as compared to nortriptyline alone.
Example 4
Effects of GABA Fluoxetine Conjugate as Determined by the
Carrageenan-Induced Paw Edema Test
[0272] Intraplantar injection of carrageenan was used to assess the
anti-inflammatory effects of GABA-fluoxetine conjugate
(AN-227).
Materials and Methods
[0273] Experimental Procedure:
[0274] Wistar rats (340-400 g) were marked with a permanent marker
at the ankle of their left hind paws to define the area of the paw
to be monitored. The rats were treated p.o. with fluoxetine 40
mg/kg (n=8) and an equimolar concentration of fluoxetine GABA (n=6)
and control vehicle (n=8). After three hours, paw edema was induced
by injecting 100 .mu.l of a 1% solution of .lamda.-carrageenan
(Sigma, USA) in normal saline into the plantar surface of the left
hind paw of the rats. The area of the induced edema was measured 2
and 4 hours later using caliper. The anti-inflammatory activity was
expressed as area of edema calculated by the measured of its length
and width (area=LXW).
Experimental Results
[0275] The fluoxetine-GABA conjugate of the present invention
reduced edema both at two hours and four hours as compared to
control and fluoxetine alone (FIG. 11). While 40 mg/kg of
fluoxetine had no effect on reducing the inflammatory area,
fluoxetine GABA conjugate significantly reduced the inflammatory
area. Although in this animal model high doses of fluoxetine were
previously shown to reduce oedema response in rats (Omar et al.
Pharmacol. Res. 49 (2004) 119-131), the present example teaches
that in side to side comparison, fluoxetine GABA conjugate acts at
a lower dose.
Example 5
Effects of GABA-Nortriptyline Drug Conjugate Compared with
Gabapentin and Nortriptyline as Determined by the
Carrageenan-Induced Paw Edema Test and Hot Plate Test
[0276] Intraplantar injection of carrageenan and the hot plate test
was used to assess the anti-inflammatory effects of
GABA-nortriptyline as compared to nortriptyline alone and to
Gabapentin (1-(aminomethyl)cyclohexane acetic acid).
Materials and Methods
[0277] Experimental Procedure:
[0278] 12 week old Wistar rats, (250-300 g; Harlan, Israel) were
divided in four groups (n=8-10) and treated as follows:
[0279] 1. Control--vehicle
[0280] 2. Nortriptyline 5 mg/kg, po. (14 mg+2.8 ml DDW)
[0281] 3. Nortriptyline-GABA (AN-228), molarequivalent to 5 mg/kg,
po, ((14.4 mg*1.28) 18.432 mg+2.8 ml DDW).
[0282] 4. Gabapentin 100 mg/kg, po.
[0283] Wistar rats (250-300 g) were marked with a permanent marker
at the ankle of their left hind paws to define the area of the paw
to be monitored. The rats were treated as described herein above.
After three hours, paw edema was induced by injecting 100 .mu.l of
a 1% solution of .lamda.-carrageenan (Sigma, USA) in normal saline
into the plantar surface of the left hind paw of the rats. The area
of the induced edema was measured using caliper at the specified
time points. The anti-inflammatory activity was measured by
analyzing the increase in height.
[0284] The animals were also tested on hot-plate at -60, 0, 120,
240 min, 24 h and 48 hours.
Experimental Results
[0285] As illustrated in FIG. 12, a significantly delayed response
to heat sensation was noted in rats receiving Nortriptyline-GABA as
compared to Nortriptyline alone and Gabapentin 4 hours following
injection.
[0286] As illustrated in FIG. 13A, paw height was significantly
lower in rats having received Nortriptyline-GABA as compared to
Nortriptyline alone and control at 4 hours, 24 hours and 48 hours
following edema induction. As illustrated in FIG. 13B, paw height
was significantly lower in rats having received Nortriptyline-GABA
as compared to Nortriptyline alone, Gabapentin and control at 24
hours and 48 hours following edema induction.
Example 6
Effects of GABA-Nortriptyline Conjugate Compared with Gabapentin
and Nortriptyline as Determined by Inflammatory Cytokine
Secretion
[0287] Cytokine secretion was used to assess the anti-inflammatory
effects of GABA-nortriptyline (AN-228) as compared with
nortriptyline alone or Gabapentin (1-(aminomethyl)cyclohexane
acetic acid).
Materials and Methods
Measurement of TNF-.alpha. and INF-.gamma. in Paw Skin of
Mice:--
[0288] Balb-c mice were divided to four groups (n=8). Two hours
prior to formalin injection they were treated orally with 0.5 mg/kg
nortriptyline, equimolar dose of AN-228 nortriptyline-GABA, a
mixture of 0.5 mg/kg nortriptyline and equimolar dose of GABA, and
50 mg/kg gabapentin. After 4 hours the mice were sacrificed and
tissues from the injection site were collected and analyzed for
TNF-.alpha. or INF-.gamma.. The collected tissues were homogenized
in 300 .mu.l of ice-cold PBS containing 0.4 mM NaCl, 0.05% Tween
20, 0.1 mM phenylmethylsulfonyl fluoride (PMSF) and 0.1 mM protease
inhibitor cocktail (Calbiochem, Darmstadt, Germany). The homogenate
was centrifuged 10,000 g for 30 min at 4.degree. C. The supernatant
was removed and assayed by the mouse TNF-.alpha. ELISA kit (BD
OptEIA, CA, USA) and mouse INF-.gamma. immunoassay (R&D
Systems, Minneapolis, Minn., USA) according to the manufacturer's
instruction. The results are expressed as ng TNF-.alpha. or
INF-.gamma. per mg protein).
Experimental Results
[0289] The amount of both INF-.gamma. (FIG. 14A) and TNF-.alpha.
(FIG. 14B) was significantly lower in mice treated with
AN-228Nortriptyline-GABA than in mice treated with an equimolar
dose of Nortriptyline or an equimolar dose of
Nortriptyline+GABA.
[0290] It is appreciated that certain features of the invention,
which are, for clarity, described in the context of separate
embodiments, may also be provided in combination in a single
embodiment. Conversely, various features of the invention, which
are, for brevity, described in the context of a single embodiment,
may also be provided separately or in any suitable
subcombination.
[0291] Although the invention has been described in conjunction
with specific embodiments thereof, it is evident that many
alternatives, modifications and variations will be apparent to
those skilled in the art. Accordingly, it is intended to embrace
all such alternatives, modifications and variations that fall
within the spirit and broad scope of the appended claims. All
publications, patents and patent applications mentioned in this
specification are herein incorporated in their entirety by
reference into the specification, to the same extent as if each
individual publication, patent or patent application was
specifically and individually indicated to be incorporated herein
by reference. In addition, citation or identification of any
reference in this application shall not be construed as an
admission that such reference is available as prior art to the
present invention.
* * * * *