U.S. patent application number 15/426626 was filed with the patent office on 2017-08-24 for treatment of bdnf-related disorders using laquinimod.
This patent application is currently assigned to TEVA PHARMACEUTICAL INDUSTRIES, LTD.. The applicant listed for this patent is Liat Hayardeny. Invention is credited to Liat Hayardeny.
Application Number | 20170239234 15/426626 |
Document ID | / |
Family ID | 43535297 |
Filed Date | 2017-08-24 |
United States Patent
Application |
20170239234 |
Kind Code |
A1 |
Hayardeny; Liat |
August 24, 2017 |
TREATMENT OF BDNF-RELATED DISORDERS USING LAQUINIMOD
Abstract
This application provides for a method of increasing
brain-derived neurotrophic factor (BDNF) serum level in a human
subject comprising periodically administering to the subject an
amount of laquinimod or pharmaceutically acceptable salt thereof
effective to increase BDNF serum level in the human subject. The
method can further comprise periodically administering to the
subject an amount of a second BDNF-increasing agent. This
application also provides for a method for treating a human subject
suffering from a BDNF-related disease comprising periodically
administering laquinimod or a pharmaceutically acceptable salt
thereof in an amount effective to treat the human subject. This
application additionally provides for use of laquinimod in the
manufacture of a medicament for increasing BDNF serum level in a
human subject. This application further provides for a
pharmaceutical composition comprising an amount of laquinimod
effective for use in increasing BDNF serum level in a human
subject. This application also provides for a pharmaceutical
preparation comprising an amount of laquinimod and an amount of a
second BDNF-increasing agent effective for use in increasing BDNF
serum level in a human subject.
Inventors: |
Hayardeny; Liat; (Tel Aviv,
IL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Hayardeny; Liat |
Tel Aviv |
|
IL |
|
|
Assignee: |
TEVA PHARMACEUTICAL INDUSTRIES,
LTD.
Petach-Tikva
IL
|
Family ID: |
43535297 |
Appl. No.: |
15/426626 |
Filed: |
February 7, 2017 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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12806275 |
Aug 9, 2010 |
9585878 |
|
|
15426626 |
|
|
|
|
61273920 |
Aug 10, 2009 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61P 15/10 20180101;
A61K 9/0053 20130101; A61P 25/16 20180101; A61P 25/24 20180101;
A61P 25/14 20180101; A61K 31/138 20130101; A61P 25/18 20180101;
A61P 27/02 20180101; A61K 31/4704 20130101; A61K 31/428 20130101;
A61K 9/20 20130101; A61P 25/22 20180101; A61P 25/00 20180101; A61P
25/28 20180101; A61K 31/138 20130101; A61K 2300/00 20130101; A61K
31/428 20130101; A61K 2300/00 20130101; A61K 31/4704 20130101; A61K
2300/00 20130101 |
International
Class: |
A61K 31/4704 20060101
A61K031/4704; A61K 9/20 20060101 A61K009/20; A61K 9/00 20060101
A61K009/00 |
Claims
1. A method of increasing brain-derived neurotrophic factor (BDNF)
serum level in a human subject comprising periodically
administering to the subject an amount of laquinimod or
pharmaceutically acceptable salt thereof effective to increase BDNF
serum level in the human subject.
2. The method of claim 1, wherein the amount of laquinimod or
pharmaceutically acceptable salt thereof is administered to the
human subject once daily.
3. The method of claims 1 or 2, wherein the periodic administration
continues for at least 3 days.
4. The method of any one of claims 1-3, wherein the amount of
laquinimod administered is 0.1 mg/day-40.0 mg/day.
5. The method of claim 4, wherein the amount of laquinimod
administered is 0.6 mg/day.
6. The method of any one of claims 1-5, wherein the amount of
laquinimod is administered orally.
7. The method of any one of claims 1-6, wherein the subject is
suffering from a BDNF-related disease.
8. The method of claim 7, wherein the BDNF-related disease is
Parkinson's disease, Huntington's disease, amyotrophic lateral
sclerosis, a depressive disorder, an anxiety disorder, retinitis
pigmentosa, erectile dysfunction, a memory disorder, Rett syndrome,
Alzheimer's disease, bipolar disorder or acute mania.
9. The method of claim 8, wherein the depressive disorder is
depression, depression in cancer patients, depression in
Parkinson's disease patients, postmyocardia infarction depression,
depression in patients with human immunodeficiency virus (HIV),
subsyndromal symptomatic depression, depression in infertile women,
pediatric depression, major depression, single episode depression,
recurrent depression, child abused-induced depression, post-partum
depression, DSM-IV major depression, treatment-refractory major
depression, severe depression, psychotic depression, post-stroke
depression, neuropathic pain, manic depressive illness including
manic depressive illness with mixed episodes and manic depressive
illness with depressive episodes, seasonal affective disorder,
bipolar depression BP I, bipolar depression BP II, or major
depression with dysthymia.
10. The method of claim 8, wherein the anxiety disorder is
generalized anxiety, panic disorder, phobia, post traumatic stress
disorder, obsessive compulsive disorder, separation anxiety, or
childhood anxiety.
11. The method of any one of claims 1-10, the method further
comprising periodically administering to the subject an amount of a
second BDNF-increasing agent.
12. The method of claim 11, wherein the amount of the second
BDNF-increasing agent when taken alone is not effective to increase
BDNF serum level in the subject.
13. The method of claims 11 or 12, wherein the administration of
the laquinimod substantially precedes the administration of the
second BDNF-increasing agent.
14. The method of claims 11 or 12, wherein the administration of
the second BDNF-increasing agent substantially precedes the
administration of the laquinimod.
15. The method of any one of claims 11-14, wherein the effect on
the subject is greater than the effect on a subject treated with
the second BDNF-increasing agent alone.
16. The method of claim 15, wherein the increase in BDNF serum
level in the subject is greater than the increase in BDNF serum
level in a subject treated with the second BDNF-increasing agent
alone.
17. The method of any one of claims 1-16, wherein the
pharmaceutically acceptable salt of laquinimod is laquinimod
sodium.
18-21. (canceled)
Description
[0001] This application is a divisional of U.S. Ser. No.
12/806,275, filed Aug. 9, 2010, now U.S. Pat. No. 9,585,878, issued
Mar. 7, 2017, claiming the benefit of U.S. Provisional Application
No. 61/273,920, filed Aug. 10, 2009, the entire contents of each of
which are hereby incorporated by reference into the subject
application.
[0002] Throughout this application various publications are
referenced. The disclosures of these publications in their
entireties are hereby incorporated by reference into this
application in order to more fully describe the state of the art to
which this invention pertains.
BACKGROUND
[0003] Neurotrophic factors have a profound influence on
developmental events such as naturally occurring cell death,
differentiation and process outgrowth (Snider, 1989). The
brain-derived neurotrophic factor (BDNF) is a neurotrophic factor
which belongs to the neurotrophin family of growth factors. BDNF
acts on certain neurons of the central nervous system (CNS) and the
peripheral nervous system (PNS), helping to support the survival of
existing neurons and encourage the growth and differentiation of
new neurons and synapses (Acheson, 1995; Huang, 2001). Increasing
BDNF has been associated with the treatment of a number of
disorders including Parkinson's disease, Huntington's disease (HD),
amyotrophic lateral sclerosis (ALS), depressive disorders,
retinitis pigmentosa, erectile dysfunction, memory disorders, Rett
syndrome, Alzheimer's Disease, bipolar disorder and acute
mania.
[0004] Parkinson's disease is a chronic and progressive
degenerative disease of the brain that impairs motor control,
speech, and other functions. One of the most striking features of
Parkinson's disease is that it primarily affects a restricted
neuronal population in the brain. Although other neurons are also
affected, the dopaminergic neurons of the substantia nigra pars
compacta are the most vulnerable to the disease process (Chesselet,
2003) BDNF has potent effects on survival and morphology of
mesencephalic dopaminergic neurons, increasing their survival, and
thus its loss could contribute to death of these cells in
Parkinson's disease (PD) (Hyman, 1991; Howell, 2000).
[0005] Huntington's disease (HD) is a neurodegenerative disorder
characterized by motor, cognitive, and psychiatric symptoms and by
a progressive degeneration of neurons in basal ganglia in brain
cortex. Patients suffering from HD have significantly lower BDNF
levels in serum compared to healthy controls (Ciammola, 2007).
[0006] Amyotrophic lateral sclerosis (ALS) is a chronic and
debilitating neurodegenerative disease which involves degeneration
of cortical, bulbar and medullar motor neurons. Riluzole
(2-amino-6-[trifluoromethoxy]benzothiazole) is an antagonist of
glutamatergic neurotransmission that prolongs survival in ALS
(Riviere, 1998). Riluzole has also been shown to significantly
increase BDNF levels in the rat brain, thereby promoting precursor
proliferation (Katoh-Semba, 2002).
[0007] Depression is another indication in which BDNF has been
shown to have an effect. In a meta-analysis which encompasses many
studies, depressed patients were shown to have lower BDNF levels
than healthy control subjects, and anti-depressant therapy has been
shown to increase BDNF levels in depressed patients after treatment
(Sen, 2008).
[0008] Retinitis pigmentosa is a disease associated with retinal
photoreceptor cell loss. It has been shown that BDNF culturing of
retina explants from rd mice, who suffer from a mutation in the
same gene that has been found mutated in human autosomal recessive
retinitis pigmentosa, showed an increase in number of photoreceptor
nuclei in the outer nuclear layer (Caffe Romeo, 2001). This
suggests that increasing BDNF levels in humans suffering from
retinitis pigmentosa may slow the progression of the disease.
[0009] Erectile dysfunction is another disease which has been shown
to be associated with BDNF. In U.S. Pat. No. 7,485,311, example 3,
it was shown that treatment of rats in the bilateral cavernous
nerve freezing model using intracavernous injection of AAV-BDNF
improves maximum intracavernous pressure in response to bilateral
cavernous nerve electrostimulation.
[0010] BDNF has also been associated with learning and memory. In a
transgenic mouse model of BNF knockout mice, impairment of BDNF
production has been shown to cause impairments in learning and
memory in the adult stage and especially during early development
(Monteggia, 2007). In a rat model, it was shown that endogenous
BDNF in the hippocampus is involved in memory formation. Whereas
infusion of function-blocking anti-BDNF antibody was shown to
impair short term and long term memory in a model of fear-motivated
learning, infusion of recombinant human BDNF facilitated long-term
memory retention (Alonsa, 2005). Increasing BDNF levels in human
patients in need of learning or memory improvement may also improve
short and long term memory.
[0011] Rett Syndrome (RTT) is an X-linked neurodevelopmental
disorder and the leading cause of severe mental retardation in
females, affecting 1:10,000-15,000 births worldwide (Amaral, 2007).
The disease is associated with mutations in the gene MeCP2. One of
the targets of MeCP2 is the BDNF gene. This suggests that
deregulation of BDNF expression in Rett Syndrome may be the cause
of structural anomalies observed in patients, especially the
reduced dendritic branching and loss of dendritic spines (Amaral,
2007). This suggests that increasing BDNF levels in Rett syndrome
may be a viable therapy in treating the disease.
[0012] In bipolar disorder patients suffering from acute mania,
levels of BDNF were shown to vary depending upon whether the
patient was treated or untreated. Before treatment with known mania
treatments, BDNF levels were found to be lower than in healthy
controls, but upon treatment, the difference between BDNF levels in
serum of treated patients and controls was no longer significant
(Tramontina, 2009). It may be possible to treat acute mania by
increasing BDNF levels in patients in need thereof.
[0013] Several agents have been identified to increase BDNF levels
including riluzole and antidepressants such as fluoxetine
(Prozac.RTM.) (Katoh-Semba, 2002; Molteni, 2006).
SUMMARY OF THE INVENTION
[0014] Disclosed herein is another agent, i.e., Laquinimod, which
has been shown to increase BDNF in humans. Laquinimod is a novel
synthetic compound with high oral bioavailability, which has been
suggested as an oral formulation for Relapsing Remitting Multiple
Sclerosis (RRMS). The relationship between laquinimod and BDNF has
not been reported. Laquinimod is a compound whose chemical name is
N-ethyl-N-phenyl-1,2-dihydro-4-hydroxy-5-chloro-1-methyl-2-oxoquinoline-3-
-carboxamide, and its Chemical Registry number is 248281-82-7. The
process of synthesis of laquinimod and the preparation of its
sodium salt are disclosed in U.S. Pat. No. 6,077,851. Additional
process of synthesis of laquinimod is disclosed in U.S. Pat. No.
6,875,869 and in U.S. Patent Application Publication No.
2007-0088050. Pharmaceutical compositions comprising laquinimod
sodium are disclosed in PCT International Application Publication
No. WO 2005/074899 as well as in U.S. Patent Application
Publication Nos. 2007-0293537 and 2009-0162432.
[0015] This application provides for a method of increasing
brain-derived neurotrophic factor (BDNF) serum level in a human
subject comprising periodically administering to the subject an
amount of laquinimod or pharmaceutically acceptable salt thereof
effective to increase BDNF serum level in the human subject. The
method can further comprise periodically administering to the
subject an amount of a second BDNF-increasing agent.
[0016] This application also provides for a method for treating a
human subject suffering from a BDNF-related disease selected from
the group consisting of Parkinson's disease, Huntington's disease,
amyotrophic lateral sclerosis, depressive disorders, anxiety
disorders, retinitis pigmentosa, erectile dysfunction, memory
disorders, Rett syndrome, Alzheimer's disease, bipolar disorder and
acute mania comprising periodically administering laquinimod or a
pharmaceutically acceptable salt thereof in an amount effective to
treat the human subject.
[0017] This application also provides for use of laquinimod in the
manufacture of a medicament for increasing BDNF serum level in a
human subject.
[0018] This application also provides for a pharmaceutical
composition comprising an amount of laquinimod effective for use in
increasing BDNF serum level in a human subject.
[0019] This application also provides for a pharmaceutical
preparation comprising an amount of laquinimod and an amount of a
second BDNF-increasing agent effective for use in increasing BDNF
serum level in a human subject.
BRIEF DESCRIPTION OF THE FIGURES
[0020] FIG. 1: Shows one-way analysis of BDNF comparing the placebo
group and 0.6 mg laquinimod group at baseline.
[0021] FIG. 2: Shows one-way analysis of BDNF comparing the placebo
group and 0.6 mg laquinimod group at week 12 (V3).
[0022] FIG. 3: Shows one-way analysis of BDNF comparing the placebo
group and 0.6 mg laquinimod group at week 36 (V9).
[0023] FIG. 4A-4D: Shows Forced Swim Test results conducted in
Example 2.1. (N=5/group) Laquinimod (1-25 mg/kg/dx3d po) showed
antidepressant activity with the 1 mg/kg p.o. dose being the most
active.
[0024] FIG. 4A: Effect of Laquinimod (1; 5; 10; 25 mg/kg/dx4,p.o.,
-90) and Fluoxetine (10 mg/kg/dx4, -90 min, p.o.) in the FST on
Distance moved [cm] on Balb C mice [4 min]
[0025] FIG. 4B: Effect of Laquinimod (1; 5; 10; 25 mg/kg/dx4, p.o.,
-90) and Fluoxetine (10 mg/kg/dx4, -90 min, p.o.) in the FST on
Velocity (cm/s) on Balb C mice [4 min]
[0026] FIG. 4C: Effect of Laquinimod (1; 5; 10; 25 mg/kg/dx4, p.o.,
-90) and Fluoxetine (10 mg/kg/dx4, -90 min, p.o.) in the FST on
Movement (Moving duration) on Balb C mice [4 min]
[0027] FIG. 4D: Effect of Laquinimod (1; 5; 10; 25 mg/kg/dx4, p.o.,
-90) and Fluoxetine (10 mg/kg/dx4, -90 min, p.o.) in the FST on
Immobility total duration [10%] on Balb C mice [4 min]
[0028] FIG. 5A-5D: Shows Forced Swim Test results conducted in
Example 2.2. (*p<0.01 vs. cont) Laquinimod (0.5-25 mg/kg/dx3d
po) showed antidepressant activity with doses at 5 and 25 mg/kg
inducing significant antidepressant activity.
[0029] FIG. 5A: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) in the FST on
Distance moved [cm] on Balb C mice [4 min]
[0030] FIG. 5B: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) in the FST on
velocity Balb C mice [4 min]
[0031] FIG. 5C: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) in the FST on
movement (total duration) on Balb C mices [4 min]
[0032] FIG. 5D: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) in the FST on
Immobility total duration [10%] on Balb C mice [4 min]
[0033] FIG. 6A-6D: Shows Open Field Test--motility parameter
results conducted in Example 3.1. (N=5/group) Laquinimod (1-25
mg/kg/dx3d po) did not modify general mobility parameters in open
field in Balb/c mice.
[0034] FIG. 6A: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Distance moved [cm] on BALB/c mice [20 min]
[0035] FIG. 6B: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Velocity mean [cm/s] on BALB/c mice [20 min]
[0036] FIG. 6C: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Immobility total duration on BALB/c mice [20
min]
[0037] FIG. 6D: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Strong mobility total duration [s] on BALB/c
mice [20 min]
[0038] FIG. 7A-7D: Shows Open Field Test--anxiety parameter results
(in zone 2) conducted in Example 3.1. (N=5/group) Laquinimod (1-25
mg/kg/dx3d po) showed tendency toward anxiolytic effect with
animals being more active in the center (zone 2).
[0039] FIG. 7A: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on In zone frequency (zone 2) on BALB/c mice [20
min]
[0040] FIG. 7B: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on In zone total duration (zone 2) on BALB/c mice
[20 min]
[0041] FIG. 7C: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Rearing Frequency (zone 2) on BALB/c mice [20
min]
[0042] FIG. 7D: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Distance moved in the center (zone 2) on BALB/c
mice [20 min]
[0043] FIG. 8A-8C: Shows Open Field Test--anxiety parameter results
(in zone 2+3) conducted in Example 3.1. (N=5/group) Laquinimod
(1-25/kg/dx3d po) showed tendency toward anxiolytic effect with
animals being more active in the center (zones 2+3). 1 mg/kg was
shown to be most potent dose.
[0044] FIG. 8A: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on In zone total duration (zone 3) on BALB/c mice
[20 min]
[0045] FIG. 8B: Effect of Laquinimod (1; 5; 10; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Distance Moved in the center (zone 3) on BALB/c
mice [20 min] FIG. 8C: Effect of Laquinimod (1; 5; 10; 25 mg/kg,
po; -90 min) and Fluoxetine (10 mg/kg, po; -90 min) and combination
in the Open field test on Rearing Frequency on BALB/c mice [20
min]
[0046] FIG. 9A-9D: Shows Open Field Test--motility parameter
results conducted in Example 3.2. (N=5/group; *p<0.05 vs. cont)
Laquinimod (0.5-25 mg/kg/dx3d po) induced a slight increase in
motility (0.5-5 mg/kg) in the open field in Balb/c mice.
[0047] FIG. 9A: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Distance moved [cm] on BALB/c mice [20 min]
[0048] FIG. 9B: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Immobility total duration on BALB/c mice [20
min]
[0049] FIG. 9C: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Rearing on BALB/c mice [20 min]
[0050] FIG. 9D: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Strong mobility total duration [s] on BALB/c
mice [20 min]
[0051] FIG. 10A-10C: Shows Open Field Test--anxiety parameter
results (in zones 2+3) conducted in Example 3.2. Laquinimod (0.5-25
mg/kg/dx3d po) shows significant anxiolytic effect with animals
showing more activity in the center (zones 2+3). Both fluoxetine
and laquinimod (0.5-5 mg/kg) showed significant anxiolytic effect
vs. the control. The most potent dose was laquinimod at 5
mg/kg.
[0052] FIG. 10A: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on In zone frequency (zone 2+3) on BALB/c mice [20
min]
[0053] FIG. 10B: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on In zone total duration (zone 2+3) on BALB/c mice
[20 min]
[0054] FIG. 10C: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, po; -90
min) and Fluoxetine (10 mg/kg, po; -90 min) and combination in the
Open field test on Rearing (zone 2+3) on BALB/c mice [20 min]
[0055] FIG. 11A-11D: Shows Elevated Plus Maze model results
conducted in Example 4.1. Laquinimod (5 and 25 mg/kg/dx3d po)
showed significant anxiolytic effect. Laquinimod showed a dose
dependent increase in activity on the open arm. The dose at 25
mg/kg p.o. was the most active.
[0056] FIG. 11A: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on In zone total duration[s] on Balb/C mice
[0057] FIG. 11B: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on Movement (Moving) Total duration(s) on Balb/C
mice
[0058] FIG. 11C: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on Strong mobility Frequency on Balb/C mice
[0059] FIG. 11D: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on Frequency to open arms on Balb/C mice
[0060] FIG. 12A-12D: Shows Elevated Plus Maze model results
conducted in Example 4.2. (Open arm parameters in Balb/c mice--5
min; N=5/group) Laquinimod (5-25 mg/kg/dx3d po) showed significant
anxiolytic effect.
[0061] FIG. 12A: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on In zone Frequency on Balb/C mice [5 min]
[0062] FIG. 12B: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on In zone total duration[s] on Balb/C mice [5
min]
[0063] FIG. 12C: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on Movement (Moving) Total duration(s) on Balb/C mice
[5 min]
[0064] FIG. 12D: Effect of Laquinimod (0.5; 1; 5; 25 mg/kg, p.o.,
-90) and Fluoxetine (10 mg/kg, -90 min, p.o.) combination in the
Open Arm test on Rearing total Frequency on Balb/C mice [5 min]
DETAILED DESCRIPTION OF THE INVENTION
[0065] This application provides for a method of increasing
brain-derived neurotrophic factor (BDNF) serum level in a human
subject comprising periodically administering to the subject an
amount of laquinimod or pharmaceutically acceptable salt thereof
effective to increase BDNF serum level in the human subject.
[0066] In one embodiment, the amount of laquinimod or
pharmaceutically acceptable salt thereof is administered to the
human subject once daily. In another embodiment, the periodic
administration continues for at least 3 days.
[0067] In one embodiment, the amount of laquinimod administered is
0.1 mg/day-40.0 mg/day. In another embodiment, the amount of
laquinimod administered is 0.6 mg/day. In another embodiment, the
amount of laquinimod is administered orally.
[0068] In one embodiment, the subject is suffering from a
BDNF-related disease. In another embodiment, the BDNF-related
disease is Parkinson's disease, Huntington's disease, amyotrophic
lateral sclerosis, a depressive disorder, an anxiety disorder,
retinitis pigmentosa, erectile dysfunction, a memory disorder, Rett
syndrome, Alzheimer's disease, bipolar disorder or acute mania.
[0069] In an embodiment, the depressive disorder is depression,
depression in cancer patients, depression in Parkinson's disease
patients, postmyocardia infarction depression, depression in
patients with human immunodeficiency virus (HIV), subsyndromal
symptomatic depression, depression in infertile women, pediatric
depression, major depression, single episode depression, recurrent
depression, child abused-induced depression, post-partum
depression, DSM-IV major depression, treatment-refractory major
depression, severe depression, psychotic depression, post-stroke
depression, neuropathic pain, manic depressive illness including
manic depressive illness with mixed episodes and manic depressive
illness with depressive episodes, seasonal affective disorder,
bipolar depression BP I, bipolar depression BP II, or major
depression with dysthymia.
[0070] In another embodiment, the anxiety disorder is generalized
anxiety, panic disorder, phobia, post traumatic stress disorder,
obsessive compulsive disorder, separation anxiety, or childhood
anxiety.
[0071] In one embodiment, the method further comprises periodically
administering to the subject an amount of a second BDNF-increasing
agent.
[0072] In one embodiment, the amount of the second BDNF-increasing
agent when taken alone is not effective to increase BDNF serum
level in the subject.
[0073] In one embodiment, the administration of the laquinimod
substantially precedes the administration of the second
BDNF-increasing agent. In another embodiment, the administration of
the second BDNF-increasing agent substantially precedes the
administration of the laquinimod.
[0074] In one embodiment, the effect on the subject is greater than
the effect on a subject treated with the second BDNF-increasing
agent alone. In another embodiment, the increase in BDNF serum
level in the subject is greater than the increase in BDNF serum
level in a subject treated with the second BDNF-increasing agent
alone.
[0075] In one embodiment, the pharmaceutically acceptable salt of
laquinimod is laquinimod sodium.
[0076] In one embodiment, the diagnosis of the subject prior to
administration excluded multiple sclerosis, insulin-dependent
diabetes mellitus, systemic lupus erythematosus, rheumatoid
arthritis, inflammatory bowel disease, psoriasis, asthma,
atherosclerosis, stroke and Alzheimer's disease.
[0077] This application also provides for a method for treating a
human subject suffering from a BDNF-related disease selected from
the group consisting of Parkinson's disease, Huntington's disease,
amyotrophic lateral sclerosis, depressive disorders, anxiety
disorders, retinitis pigmentosa, erectile dysfunction, memory
disorders, Rett syndrome, Alzheimer's disease, bipolar disorder and
acute mania comprising periodically administering laquinimod or a
pharmaceutically acceptable salt thereof in an amount effective to
treat the human subject.
[0078] This application also provides for use of laquinimod in the
manufacture of a medicament for increasing BDNF serum level in a
human subject.
[0079] This application also provides for a pharmaceutical
composition comprising an amount of laquinimod effective for use in
increasing BDNF serum level in a human subject.
[0080] This application also provides for a pharmaceutical
preparation comprising an amount of laquinimod and an amount of a
second BDNF-increasing agent effective for use in increasing BDNF
serum level in a human subject.
[0081] For the foregoing embodiments, each embodiment disclosed
herein is contemplated as being applicable to each of the other
disclosed embodiment.
Terms
[0082] As used herein, and unless stated otherwise, each of the
following terms shall have the definition set forth below.
[0083] As used herein, "administering to the subject" means the
giving of, dispensing of, or application of medicines, drugs, or
remedies to a subject to relieve or cure a pathological condition.
Oral administration is one way of administering the instant
compounds to the subject.
[0084] As used herein, "BDNF" means brain-derived neurotrophic
factor, a neurotrophic factor belonging to the neurotrophin family
of growth factors.
[0085] As used herein, a "BDNF-related disease" is a disease in
which a patient suffering from the disease has BDNF serum levels
which are lower than those of a corresponding healthy individual
and/or a disease in which the elevation of BDNF in a patient
suffering from the disease can be associated with amelioration of
the disease or of symptoms thereof.
[0086] As used herein, a "BDNF-increasing agent" is any agent which
directly or indirectly elevates BDNF level in a subject. For
example, a BDNF-increasing agent can be riluzole or an
antidepressant such as fluoxetine. As used herein, the term "agent"
includes any molecule, compound, protein, peptide, polypeptide,
nucleic acid, antibody, or drug or any combination thereof.
[0087] As used herein, an "amount" or "dose" of laquinimod as
measured in milligrams refers to the milligrams of laquinimod acid
present in a preparation, regardless of the form of the
preparation. For example, 0.6 mg of laquinimod means the amount of
laquinimod acid in a preparation is 0.6 mg, regardless of the form
of the preparation. Thus, when in the form of a salt, e.g. a
laquinimod sodium salt, the weight of the salt form necessary to
provide a dose of 0.6 mg laquinimod would be greater than 0.6 mg
due to the presence of the additional salt ion, but would be a
molar equivalent amount.
[0088] As used herein, "effective" as in an amount effective to
achieve an end means the quantity of a component that is sufficient
to yield an indicated therapeutic response without undue adverse
side effects (such as toxicity, irritation, or allergic response)
commensurate with a reasonable benefit/risk ratio when used in the
manner of this disclosure. For example, an amount effective to
treat a symptom of a disorder or disease without causing undue
adverse side effects. The specific effective amount will vary with
such factors as the particular condition being treated, the
physical condition of the patient, the type of mammal being
treated, the duration of the treatment, the nature of concurrent
therapy (if any), and the specific formulations employed and the
structure of the compounds or its derivatives.
[0089] A "salt" is salt of the instant compounds which have been
modified by making acid or base salts of the compounds. The term
"pharmaceutically acceptable salt" in this respect, refers to the
relatively non-toxic, inorganic and organic acid or base addition
salts of compounds of the present invention.
[0090] A pharmaceutically acceptable salt of laquinimod can be
used. A pharmaceutically acceptable salt of laquinimod as used in
this application includes lithium, sodium, potassium, magnesium,
calcium, manganese, copper, zinc, aluminum and iron. Salt
formulations of laquinimod and the process for preparing the same
are described, e.g., in U.S. Patent Application Publication No.
2005-0192315 and PCT International Application Publication No. WO
2005/074899, which are hereby incorporated by reference into this
application.
[0091] As used herein, to "treat" or "treating" encompasses, e.g.,
inducing inhibition, regression, or stasis of the disorder and/or
disease. As used herein, "inhibition" of disease progression or
disease complication in a subject means preventing or reducing the
disease progression and/or disease complication in the subject.
[0092] As used herein, "pharmaceutically acceptable carrier" refers
to a carrier or excipient that is suitable for use with humans
and/or animals without undue adverse side effects (such as
toxicity, irritation, and allergic response) commensurate with a
reasonable benefit/risk ratio. It can be a pharmaceutically
acceptable solvent, suspending agent or vehicle, for delivering the
instant compounds to the subject.
[0093] A dosage unit as used herein may comprise a single compound
or mixtures of compounds thereof. A dosage unit can be prepared for
oral dosage forms, such as tablets, capsules, pills, powders, and
granules.
[0094] Laquinimod can be administered in admixture with suitable
pharmaceutical diluents, extenders, excipients, or carriers
(collectively referred to herein as a pharmaceutically acceptable
carrier) suitably selected with respect to the intended form of
administration and as consistent with conventional pharmaceutical
practices. The unit can be in a form suitable for oral
administration. Laquinimod can be administered alone but is
generally mixed with a pharmaceutically acceptable carrier, and
co-administered in the form of a tablet or capsule, liposome, or as
an agglomerated powder. Examples of suitable solid carriers include
lactose, sucrose, gelatin and agar. Capsule or tablets can be
easily formulated and can be made easy to swallow or chew; other
solid forms include granules, and bulk powders. Tablets may contain
suitable binders, lubricants, diluents, disintegrating agents,
coloring agents, flavoring agents flow-inducing agents, and melting
agents.
[0095] Specific examples of the techniques, pharmaceutically
acceptable carriers and excipients that may be used to formulate
oral dosage forms of the present invention are described, e.g., in
U.S. Patent Application Publication No. 2005/0192315, PCT
International Application Publication Nos. WO 2005/074899, WO
2007/047863, and WO/2007/146248, each of which is hereby
incorporated by reference into this application.
[0096] General techniques and compositions for making dosage forms
useful in the present invention are described-in the following
references: 7 Modern Pharmaceutics, Chapters 9 and 10 (Banker &
Rhodes, Editors, 1979); Pharmaceutical Dosage Forms: Tablets
(Lieberman et al., 1981); Ansel, Introduction to Pharmaceutical
Dosage Forms 2nd Edition (1976); Remington's Pharmaceutical
Sciences, 17th ed. (Mack Publishing Company, Easton, Pa., 1985);
Advances in Pharmaceutical Sciences (David Ganderton, Trevor Jones,
Eds., 1992); Advances in Pharmaceutical Sciences Vol 7. (David
Ganderton, Trevor Jones, James McGinity, Eds., 1995); Aqueous
Polymeric Coatings for Pharmaceutical Dosage Forms (Drugs and the
Pharmaceutical Sciences, Series 36 (James McGinity, Ed., 1989);
Pharmaceutical Particulate Carriers: Therapeutic Applications:
Drugs and the Pharmaceutical Sciences, Vol 61 (Alain Rolland, Ed.,
1993); Drug Delivery to the Gastrointestinal Tract (Ellis Horwood
Books in the Biological Sciences. Series in Pharmaceutical
Technology; J. G. Hardy, S. S. Davis, Clive G. Wilson, Eds.);
Modern Pharmaceutics Drugs and the Pharmaceutical Sciences, Vol. 40
(Gilbert S. Banker, Christopher T. Rhodes, Eds.). These references
in their entireties are hereby incorporated by reference into this
application.
[0097] Tablets may contain suitable binders, lubricants,
disintegrating agents, coloring agents, flavoring agents,
flow-inducing agents, and melting agents. For instance, for oral
administration in the dosage unit form of a tablet or capsule, the
active drug component can be combined with an oral, non-toxic,
pharmaceutically acceptable, inert carrier such as lactose,
gelatin, agar, starch, sucrose, glucose, methyl cellulose,
dicalcium phosphate, calcium sulfate, mannitol, sorbitol,
microcrystalline cellulose and the like. Suitable binders include
starch, gelatin, natural sugars such as glucose or beta-lactose,
corn starch, natural and synthetic gums such as acacia, tragacanth,
or sodium alginate, povidone, carboxymethylcellulose, polyethylene
glycol, waxes, and the like. Lubricants used in these dosage forms
include sodium oleate, sodium stearate, sodium benzoate, sodium
acetate, sodium chloride, stearic acid, sodium stearyl fumarate,
talc and the like. Disintegrators include, without limitation,
starch, methyl cellulose, agar, bentonite, xanthan gum,
croscarmellose sodium, sodium starch glycolate and the like.
[0098] It is understood that where a parameter range is provided,
all integers within that range, and tenths thereof, are also
provided by the invention. For example, "0.1 mg-40.0 mg" includes
0.1 mg, 0.2 mg, 0.3 mg, 0.4 mg, etc. up to 40.0 mg.
[0099] This invention will be better understood by reference to the
Experimental Details which follow, but those skilled in the art
will readily appreciate that the specific experiments detailed are
only illustrative of the invention as described more fully in the
claims which follow thereafter.
EXPERIMENTAL DETAILS
Example 1: Clinical Trial Showing Effect of Laquinimod on BDNF
Levels in Human Subjects Afflicted with Relapsing Remitting
Multiple Sclerosis
[0100] A study was initiated in relapsing remitting multiple
sclerosis (RRMS) patients using laquinimod, 0.6 mg daily, in the
form of the sodium salt, in an oral, once-daily tablet formulation.
The study was a multinational, multicenter, randomized,
double-blind, parallel-group, placebo controlled study assessing
the efficacy, tolerability and safety of two doses of laquinimod in
subjects with RRMS. Eligible subjects were randomized into the
following three groups: [0101] 1. 0.6 mg of laquinimod per os
(p.o.) once daily; [0102] 2. 0.3 mg of laquinimod per os (p.o.)
once daily; and [0103] 3. Matching placebo, per os (p.o.) once
daily.
[0104] Subjects must meet the following inclusion criteria to be
included in the study: [0105] 1. a confirmed MS diagnosis as
defined by the McDonald criteria; [0106] 2. a RRMS disease course;
[0107] 3. at least one documented relapse n the 12 months prior to
screening; [0108] 4. at least one Gadolinium-enhanced lesion on
their screening MRI scan; [0109] 5. ambulatory with a Kurtzke EDSS
score of 1.0-5.0 (Converted); [0110] 6. between 18-50 years of age
inclusive; [0111] 7. relapse-free and off corticosteroids or ACTH
for at least 30 days prior to the MRI scan at screening; [0112] 8.
relapse-free and off corticosteroids between screening and
randomization; [0113] 9. women of child-bearing potential practiced
a reliable method of birth control; [0114] 10. willing and able to
comply with the protocol requirements for the duration of the
study; and [0115] 11. able to give signed, written and informed
consent prior to entering the study.
[0116] Subjects meeting any one of the following exclusion criteria
were excluded from the study: [0117] 1. suffered from any form of
progressive MS; [0118] 2. use of immunosuppressive or cytotoxic
treatments within 6 months prior to the screening visit (including
azathioprine, cyclophosphamide and methotrexate); [0119] 3. use of
experimental drugs, and/or participation in drug clinical studies
within the 6 months prior to screening; [0120] 4. previous
treatment with immunomodulators (including IFN-.beta. 1a and 1b,
glatiramer acetate, laquinimod and IVIG) within the 2 months prior
to screening; [0121] 5. use of potent inhibitors of CYP3A4, for
example, oral ketoconazole and erythromycin within 2 weeks prior to
baseline visit; [0122] 6. previous use of amiodarone; [0123] 7. use
of fluoxetine one month prior to baseline visit; [0124] 8. use of
substrates of CYP1A2 such as theophylline and warfarin within 2
weeks prior to screening; [0125] 9. previous treatment with
cladribine within the last 2 years prior to screening visit; [0126]
10. subjects for whom potential immunosuppression would be
contraindicated, for example, Hepatitis B/C or HIV. [0127] 11.
previous total body irradiation or total lymphoid irradiation;
[0128] 12. chronic corticosteroid treatment (30 or more consecutive
days) within the 2 months prior to screening; [0129] 13. pregnancy
or breastfeeding; [0130] 14. subjects with a clinically significant
or unstable medical or surgical condition that would preclude safe
and complete study participation as determined by medical history,
physical exams, ECG, abnormal laboratory tests and chest X-ray.
[0131] 15. inability to give informed consent, or to complete the
study, or if the subject is considered by the investigator to be,
for any reason, an unsuitable candidate for the study; [0132] 16. a
known history of sensitivity to Gd; or [0133] 17. inability to
successfully undergo MRI scanning.
[0134] The study duration was 40 weeks and consisted of 2 periods:
4 weeks of screening period (week -4 [screening] to week 0
[baseline]) and 36 weeks of double blind treatment (week 0
[baseline] to week 36 [termination]).
[0135] Subjects were evaluated at study sites at weeks -4, 0, 4, 8,
12, 16, 20, 24, 28, 32 and 36. Samples for inflammatory markers
were collected at all visits between 0 and 36 weeks.
[0136] BDNF concentration in serum was measured in patients in the
placebo group (102 patients were in the group) and in the 0.6 mg
laquinimod group (106 patients were in the group) at week 0 (V0),
week 12 (v3) and week 36 (V9). The analysis of BDNF content in the
plasma was performed using the sandwich ELISA method. The
concentration of BDNF in serum is expressed in terms of pg/mL.
Results
[0137] The results in the tables below represent mean BDNF levels
in both the laquinimod 0.6 mg/day group and the placebo group.
Baseline Values (at V0)
TABLE-US-00001 [0138] TABLE 1 Mean and Std Deviations Err Err Std
Lower Upper Level Number Mean Std Dev Mean 95% 95% Placebo 95
14110.7 6432.61 659.97 12800 15421 Laquinimod 96 13892.7 7267.89
741.78 12420 15365 0.6 mg
TABLE-US-00002 TABLE 2 t Test Laquinimod 0.6 mg-Placebo - Assuming
unequal variances Difference -218.0 t-Ratio -0.2196 Std Err Dif
922.9 DF 186.6984 Upper CL Dif 1740.7 Prob > |t| 0.8264 Lower CL
Dif -2176.7 Prob > t 0.5868 Confidence 0.95 Prob < t
0.4132
Week 12 Values (at V3)
TABLE-US-00003 [0139] TABLE 3 Mean and Std Deviations Err Err Std
Lower Upper Level Number Mean Std Dev Mean 95% 95% Placebo 96
13073.9 5712.02 582.98 11916 14231 Laquinimod 100 15519.6 7204.47
720.45 14090 16949 0.6 mg
TABLE-US-00004 TABLE 4 t Test Laquinimod 0.6 mg-Placebo - Assuming
unequal variances Difference 2445.70 t-Ratio 2.638935 Std Err Dif
926.77 DF 187.3752 Upper CL Dif 4273.95 Prob > |t| 0.0090* Lower
CL Dif 617.44 Prob > t 0.0045* Confidence 0.95 Prob < t
0.9955
Week 36 Values (at V9)
TABLE-US-00005 [0140] TABLE 5 Mean and Std Deviations Err Err Std
Lower Upper Level Number Mean Std Dev Mean 95% 95% Placebo 92
12783.6 6017.17 627.33 11538 14030 Laquinimod 92 15335.8 6699.89
698.51 13948 16723 0.6 mg
TABLE-US-00006 TABLE 6 t Test Laquinimod 0.6 mg-Placebo - Assuming
unequal variances Difference 2552.21 t-Ratio 2.718403 Std Err Dif
938.86 DF 179.9376 Upper CL Dif 4404.81 Prob > |t| 0.0072* Lower
CL Dif 699.61 Prob > t 0.0036* Confidence 0.95 Prob < t
0.9964
[0141] Starting from V3 there is a statistically significant
elevation of BDNF (at Confidence Level of 95%) between the placebo
group and the laquinimod 0.6 mg group. The difference was also
statistically significant at V9.
[0142] Laquinimod is effective in experimental autoimmune
encephalomyelitis (aEAE) model, and is currently being tested in MS
patients (on phase III clinical trial) thus suggesting the
possibility of its use in treatment of multiple sclerosis (MS).
Laquinimod is also effective in increasing BDNF levels in the
patients.
[0143] The ability of laquinimod to increase BDNF levels is
unexpected. A per se connection between treatment of MS and
increase in BDNF levels has not been established. Not all agents
which are effective in treating MS are effective in elevating BDNF
levels. A recent study showed that treatment with interferon-.beta.
1a and immunoglobulins, even after 12 months, and even in MS
patients benefiting from the treatment, did not show an increase in
plasma BDNF levels (Sarchielli, 2007). The finding that laquinimod
elevates BDNF levels in patients after only 12 weeks of treatment
is unexpected.
Example 2: Laquinimod Shows Antidepressant Activity in Mice
Models--the Forced Swim Test (FST)
[0144] Depression and anxiety disorders are burdensome conditions
with lifetime prevalence rates of approximately 7-20%. Animal
models are indispensable tools in the search to identify new
antidepressant drugs. Various paradigms have been developed and are
instrumental in detecting the antidepressant-like potential of
novel compounds in preclinical settings (Cryan, 2002; Ganbarana,
2001).
[0145] The Forced Swim Test (FST) is one of the most widely used
tools for screening antidepressant activity pre-clinically. The
acute test was first described by Porsolt et al (1977).
[0146] The test is based on the observation that rats and mice
develop an immobile posture when placed in an inescapable cylinder
of water. This behavior is considered to be a behavioral despair as
opposed to active form of coping with stressful conditions. An
antidepressant will reduce immobility and increase motivated
behavior of the rodent to escape from the despaired conditions.
This is evidenced by increase in time of swimming, distance moved,
velocity and attempts to climb the walls (strong mobility).
[0147] FST is considered a screening tool with high reliability and
predictive validity. The test can be performed in mice and
rats.
[0148] In example 2.1 and 2.2 described below, the FST was
conducted in male mice (Balb/c Harlan IL) after 3 days of drug
administration, 90 minutes after the last drug administration.
Round glass cylinders 18 cm diameter and 20 cm depth were used.
Water temperature was 24-28.degree. C. Motivated behavior was
defined by immobility, swimming and strong mobility.
[0149] Immobility in the animals was defined by activity lower than
10% movement of the center of gravity of the animal as determined
by the Noldus system. Swimming was defined by the distance and the
velocity of the animal, and climbing was related to strong mobility
(movement of center of gravity more than 30%). Animals were
released in the cylinder for 6 min and scoring was performed in the
last 4 min after 2 min of adaptation.
[0150] All results were analyzed by the Noldus (Holland) system
including a camera and software for animal behavior analyses.
Example 2.1
[0151] Balb/c mice were divided to 6 groups (5/group) and
administered daily for 3 days laquinimod (1, 5, 10 and 25 mg/kg per
os in the form of laquinimod sodium solution via gavage),
fluoxetine (positive control 10 mg/kg per os) or vehicle. On the
third day, 90 min following administration, mice were exposed to
the forced swim test. Behavior was video recorded and analyzed
using Ethovision software (Noldus Holland). The result is shown in
FIG. 4.
Conclusion:
[0152] The results show that laquinimod showed a significant
antidepressant activity as expressed by the increased mobility and
reduced immobility at 1 mg/kg, and a trend to significance at
higher doses (Laquinimod at 25 mg/kg). In this experiment, the
positive control fluoxetine did not show significant effect,
possibly due to non optimal conditions of time and dose.
Example 2.2
[0153] Balb/c mice were divided to 6 groups (5 mice/group) and
administered laquinimod daily for 3 days with (0.5, 1, 5 and 25
mg/kg p.o. in the form of laquinimod sodium solution via gavage),
fluoxetine (positive control 10 mg/kg po) or vehicle. On the third
day, 90 minutes following administration, mice were exposed to the
forced swim test. Behavior was video recorded and analyzed using
Ethovision software (Noldus Holland). The result is shown in FIG.
5.
Conclusion:
[0154] The results show that laquinimod showed a significant
antidepressant activity as expressed by the increased mobility and
reduced immobility at 5 and 25 mg/kg. The positive control
fluoxetine showed a trend to the same activity. This is possibly
due to non optimal conditions of time and dose.
Example 3: Laquinimod Shows Anxiolytic Activity in Mice Models--the
Open Field Test
[0155] Anxiety disorders are blanket terms covering several
different forms of abnormal and pathological fear and anxiety.
Current psychiatric diagnostic criteria recognize a wide variety of
anxiety disorders. Recent surveys have found that as many as 18% of
Americans may be affected by one or more of them (Kessler et al).
The disorders are divided to several classes including: Generalized
Anxiety, Panic disorders. Phobias, Post Traumatic Stress Disorders
(PTSD), Obsessive Compulsive Disorder (OCD), Separation Anxiety and
Childhood Anxiety.
[0156] Chronically administered antidepressant drugs, particularly
selective serotonin (5-HT) reuptake inhibitors (SSRIs), are
clinically effective in the treatment of all anxiety disorders,
including post traumatic stress disorder (PTSD) and obsessive
compulsive disorder (OCD). While the clinical effectiveness of
traditional anxiolytics, such as benzodiazepines (BDZs), is limited
to generalized anxiety disorder or acute panic attacks (Borsini et
al). Thus the potential anxiolytic effect of antidepressants is of
great relevance.
[0157] The Open Field Test--Exploratory locomotor activity used in
Examples 3.1 and 3.2 described below is one of the most popular in
evaluation of animals' behavior. It tests both motility parameters
and anxiety (Prut et al). An individual mouse is placed in a novel
plexiglass arena of 50.times.50 cm the floor of which is divided
into 3 digital zones: the outer peripheral zone 1, the medial zone
2 and the most central zone 3. The animal behavior in the open
field is recorded by videotaping for 20 min and is analyzed
subsequently digitally using Noldus software for animal behavior.
The measurements include general motility (distance moved, velocity
and strong mobility) and anxiety parameters (including frequency of
visits to the central area, time spent in the inner field, and
number of rearing events in the center). The more the animal stays
and performs in the center, the less anxious it is.
Example 3.1
[0158] Balb/c male mice were divided to 6 groups (5/group) and
administered laquinimod (1, 5, 10 and 25 mg/kg p.o. in the form of
laquinimod sodium solution via gavage), fluoxetine (positive
control 10 mg/kg po) or vehicle daily for 3 days. On the third day,
90 min following administration, mice were exposed to the open
field for 20 min. Behavior was video recorded and analyzed using
Ethovision software (Noldus Holland). The results with respect to
motility parameters are shown in FIG. 6. The results with respect
to anxiety parameters are shown in FIGS. 7 and 8.
Conclusion:
[0159] Motility Parameters--Laquinimod and fluoxetine did not
modify significantly the motility parameters in the field in this
model.
[0160] Anxiety Parameters (Zone 2)--Animals treated with laquinimod
and with fluoxetine tended to perform more in the center in terms
of frequency, time spent, distance moved and number of rearings.
The results show 1 mg/kg and 5 mg/kg were the most potent.
[0161] Anxiety Parameters (Zones 2 and 3)--Animals treated with
laquinimod tended to perform more in the center as evidenced by
time spent, distance moved and number of rearings. The results show
1 mg/kg was the most potent dosage of laquinimod in this
experiment.
[0162] This model shows that laquinimod shows anxiolytic
effects.
Example 3.2
[0163] Balb/c male mice were divided to 6 groups (5/group) and
administered laquinimod (0.5, 1, 5 and 25 mg/kg p.o. in the form of
laquinimod sodium solution via gavage), fluoxetine (positive
control 10 mg/kg po) or vehicle daily for 3 days. On the third day,
90 min following administration, mice were exposed to the open
field test for 20 minutes. Behavior was video recorded and analyzed
using Ethovision software (Noldus Holland). The motility parameter
results are shown in FIG. 9. The anxiety parameter results are
shown in FIG. 10.
Conclusion:
[0164] Motility Parameters--Laquinimod (0.5-25 mg/kg/dx3d po)
induced a slight increase in motility (0.5-5 mg/kg) in the open
field in Balb/c mice.
[0165] Anxiety Parameters--Animals treated with laquinimod (0.5-5
mg/kg po) and with Fluoxetine (10 mg/kg po) performed significantly
more in the center as judged by frequency, time spent and number of
rearings. 5 mg/kg was the most potent.
[0166] This model shows that laquinimod shows anxiolytic
effects.
Example 4: Laquinimod Shows Anxiolytic Activity in Mice
Models--Elevated Plus Maze (EPM)
[0167] The EPM model used in Examples 4.1 and 4.2 described below
utilizes the natural fear of rodents to avoid open and elevated
places. The apparatus consists of a plus-maze with two enclosed and
two opposite open arms, elevated above the floor. Naive animals
spend only about 30% of the test time on open arms, while treatment
with benzodiazepines significantly increases open-arm exploration
(Pellow et al). This is one of the most widely used models to study
effects of anxiety-like behavior.
[0168] The maze consisted of two opposing open arms (40.times.10
cm) and two opposing closed arms (40.times.10 cm, with 40 cm walls)
on a platform 50 cm above the ground. Mice were placed in the
center square (10.times.10 cm) facing an open arm and videotaped
during a 5 min exploration. Arm entries and duration were scored
when all four paws enter the arm.
Example 4.1
[0169] Balb/c male mice were divided to 6 groups (5/group) and
administered laquinimod (0.5, 1, 5 and 25 mg/kg p.o. in the form of
laquinimod sodium solution via gavage), fluoxetine (positive
control 10 mg/kg po) or vehicle daily for 3 days. On the third day,
90 min following administration, mice were exposed to the EPM for 5
minutes. Mice were placed in the center square (10.times.10 cm)
facing an open arm and videotaped during a 5 minute exploration.
Arm entries and duration was scored when all four paws enter the
arm. Behavior was video recorded and analyzed using Ethovision
software (Noldus Holland). The results are shown in FIG. 11.
Conclusion: Mice treated with laquinimod show anxiolytic activity
in a dose dependent manner. Maximal effect was obtained with the
dose of 25 mg/kg po.
Example 4.2
[0170] Balb/c male mice were divided to 6 groups (5/group) and
administered laquinimod (0.5, 1, 5 and 25 mg/kg p.o. in the form of
laquinimod sodium solution via gavage), fluoxetine (positive
control 10 mg/kg po) or vehicle daily for 3 days. On the third day,
90 min following administration, mice were exposed to the EPM for 5
min. Behavior was video recorded and open arms parameters were
analyzed using Ethovision software (Noldus Holland). The results
are shown in FIG. 12.
Conclusion:
[0171] Mice treated with laquinimod show anxiolytic activity in a
dose dependent manner. Maximal effect was obtained with 5-25 mg/kg
po.
REFERENCES
[0172] 1. PCT International Application Publication No. WO
2007/047863, published Apr. 26, 2007, international filing date
Oct. 18, 2006. [0173] 2. PCT International Application Publication
No. WO 2007/146248, published Dec. 21, 2007, international filing
date Jun. 12, 2007. [0174] 3. Acheson, A, et al. (1995). "A BDNF
autocrine loop in adult sensory neurons prevents cell death".
Nature, 374(6521):450-3. [0175] 4. Alonsa, M, et al. (2005)
"Endogenous BDNF is required for long-term memory formation in the
rat parietal cortex". Learning & Memory, 12:504-510. [0176] 5.
Amaral, M D, et al. (2007) "TRPC channels as novel effectors of
BDNF signaling: Potential implications for Rett syndrome".
Pharmacol Ther, 113(2):394-409. [0177] 18. Borsini F, et al. (2002)
"Do animal models of anxiety predict anxiolytic-like effects of
antidepressants?" Psychopharmacology. 163:121-41. [0178] 6. Caffe
Romeo, A, et al. (2001) "A combination of CNTF and BDNF rescues rd
photoreceptors but changes rod differentiation in the presence of
RPE in retinal explants". Investigative Ophthalmology & Visual
Science, 42:275-82. [0179] 7. Chesselet, M F (2003) "Dopamine and
Parkinson's disease: is the killer in the house?" Molecular
Psychiatry, 8:369-370. [0180] 8. Ciammola, A, et al. (2007) "Low
brain-derived neurotrophic factor (BDNF) levels in serum of
Huntington's disease patients". Am J Med Gent Part B, 144b:574-577.
[0181] 9. Cryan J F, et al. (2002) "Assessing antidepressant
activity in rodents: recent developments and future needs". Trend
in Pharmacological Science. 23:238-45. [0182] 10. Gambarana, C., et
al. (2001) "Animal models for the study of antidepressant
activity". Brain Res. Protocol. 7:11-20. [0183] 11. Howells, D W,
et al. (2000) "Reduced BDNF mRNA expression in the Parkinson's
disease substantia nigra". Experimental Neurology, 166(1):127-135.
[0184] 12. Huang, E J and Reichardt, L F (2001) "Neurotrophins:
roles in neuronal development and function". Annu. Rev. Neurosci,
24:677-736. [0185] 13. Hyman, C. et al., (1991) "BDNR is a
neurotrophic factor for dopaminergic neurons of the substantia
nigra". Nature, 350(6315):230-2. [0186] 14. Katoh-Semba, R, et al.
(2002) "Riluzole enhances expression of brain-derived neurotrophic
factor with consequent proliferation of granule precursor cells in
the rat hippocampus". FASEB J, 16:1328-30. [0187] 19. Kessler R C,
et al. (June 2005) "Prevalence, severity, and comorbidity of
12-month DSM-IV disorders in the National Comorbidity Survey
Replication". Arch. Gen. Psychiatry. 62(6):617-27. [0188] 15.
Molteni, R, et al. (2006) "Abstract: Chronic treatment with
fluoxetine [Prozac.RTM.] up-regulates cellular BDNF mRNA expression
in rat dopaminergic regions". Int J Neuropsychopharmacol.
9(3):307-17. [0189] 16. Monteggia, L M (2007) "Elucidating the role
of brain-derived neurotrophic factor in the brain". Am J
Psychiatry, 164:1790. [0190] 20. Pellow S, et al. (1985)
"Validation of open:closed arm entries in an elevated plus-maze as
a measure of anxiety in the rat". J Neurosci Methods. 14:149-167.
[0191] 21. Porsolt R. D., et al. (1977) "Behavioral despair in
mice: a primary screening test for antidepressants". Arch. Int.
Pharmacodyn. Ther. 229: 327-336. [0192] 22. Prut L, and Belzung C.
(2003) "The open field as a paradigm to measure the effects of
drugs on anxiety-like behaviors: a review". Eur J Pharmacol.
463:3-33. [0193] 17. Riviere, M (1998) "An analysis of extended
survival in patients with amyotrophic lateral sclerosis treated
with riluzole". Arch Neurol, 55:526-8. [0194] 18. Sarchielli, P, et
al. (2007) "Production of Brain-derived neurotrophic factor by
mononuclear cells of patients with multiple sclerosis treated with
glatiramer acetate, interferon-.beta. 1a, and high doses of
immunoglobulins". Multiple Sclerosis, 13:313-331. [0195] 19. Sen,
S, et al. (2008) "Serum brain-derived neurotrophic factor,
depression, and antidepressant medications: meta-analyses and
implications". Biol Psychiatry, 64:527-532. [0196] 20. Snider, et
al., (1989) "Neurotrophic molecules". Ann Neurol, 26(4):489-506.
[0197] 21. Tramontina, J F, et al. (2009) "brain-derived
neurotrophic factor serum levels before and after treatment for
acute mania". Neuroscience Letters, 452:111-3.
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