U.S. patent application number 17/191084 was filed with the patent office on 2021-06-24 for methods and compositions for treatment of asthma or parkinson's disease.
The applicant listed for this patent is InnoPharmaScreen Inc.. Invention is credited to Incheol KANG, Jeseong PARK.
Application Number | 20210186904 17/191084 |
Document ID | / |
Family ID | 1000005433116 |
Filed Date | 2021-06-24 |
United States Patent
Application |
20210186904 |
Kind Code |
A1 |
KANG; Incheol ; et
al. |
June 24, 2021 |
Methods and Compositions for Treatment of Asthma or Parkinson's
Disease
Abstract
The present invention relates to methods and compositions
comprising proguanil for treating asthma. The invention also
relates to methods and compositions comprising proguanil for
treating Parkinson's disease.
Inventors: |
KANG; Incheol; (Incheon,
KR) ; PARK; Jeseong; (Incheon, KR) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
InnoPharmaScreen Inc. |
Incheon |
|
KR |
|
|
Family ID: |
1000005433116 |
Appl. No.: |
17/191084 |
Filed: |
March 3, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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16562136 |
Sep 5, 2019 |
10966943 |
|
|
17191084 |
|
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62727858 |
Sep 6, 2018 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61K 31/155 20130101;
A61K 31/47 20130101; A61K 9/0053 20130101; A61P 11/06 20180101 |
International
Class: |
A61K 31/155 20060101
A61K031/155; A61P 11/06 20060101 A61P011/06; A61K 31/47 20060101
A61K031/47 |
Claims
1.-8. (canceled)
9. A pharmaceutical composition comprising a compound represented
by Formula 1 or a pharmaceutically acceptable salt thereof, a
compound represented by Formula 2 or a pharmaceutically acceptable
salt thereof, and a pharmaceutically acceptable excipient:
##STR00013##
10. The composition of claim 9, wherein the composition is suitable
for oral administration.
11. The composition of claim 9, wherein the composition comprises 1
mg to 1000 mg of each of the compound.
12. A method of treating Parkinson's disease in a subject in need
thereof, the method comprising administering to the subject a
pharmaceutical composition comprising a compound represented by
Formula 1 or a pharmaceutically acceptable salt thereof and a
pharmaceutically acceptable excipient: ##STR00014##
13. The method of claim 12, wherein the composition further
comprises a compound represented by Formula 2 or a pharmaceutically
acceptable salt thereof. ##STR00015##
14. The method of claim 12, wherein the composition is administered
orally to the subject.
15. The method of claim 13, wherein the composition is administered
orally to the subject.
16. The method of claim 12, wherein the composition comprises 0.001
mg to 100 mg of each of the compound per kg of the subject's body
weight.
17. The method of claim 13, wherein the composition comprises 0.001
mg to 100 mg of each of the compound per kg of the subject's body
weight.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims the benefit of the filing date of
U.S. Appl. No. 62/727,858, filed Sep. 6, 2018, which is herein
incorporated by reference intis entirety.
FIELD OF THE INVENTION
[0002] The present disclosure relates to methods and compositions
comprising proguanil for treating asthma. The present disclosure
also relates to methods and compositions comprising proguanil for
treating Parkinson's disease.
BACKGROUND OF THE INVENTION
[0003] Asthma is a chronic inflammatory disease of the lower
respiratory tract characterized by airway hyperresponsiveness and
mucous obstruction. Bronchial asthma is the most common chronic
disease affecting children and young adults. There is strong
evidence for a genetic component in asthma (Bleecker et al., Am. J.
Respir. Crit. Care. Med., 156: S113-6 (1997); Kauffmann et al.,
Chest, 121 (3 Supp.): 27S (2002)). Multiple environmental factors
are also known to modulate the clinical expression of asthma as
well as the asthma-associated phenotypes: bronchial
hyperresponsiveness, atrophy and elevated IgE (Koppelman et al.,
Eur. Resp. J., 13: 2-4 (1999); Cookson, Nature, 25: B5-11 (1999);
Holloway, Clin. Exp. Allergy, 29: 1023-1032 (1999)).
[0004] Pharmacologic analogues of cortisol (e.g., prednisone) have
been used clinically since 1948 and remain the standard of care for
the treatment of a variety of inflammatory diseases including
asthma. These glucocorticoids (GC) reduce pathological inflammation
that is central to asthma, and they are thought to control clinical
asthma symptoms through their anti-inflammatory effects (Expert
Panel Report 3 (EPR-3): Guidelines for the Diagnosis and Management
of Asthma-Summary Report 2007. J Allergy Clin. Immunol. 2007,
120:S94-138). There remains a need for improved methods for
treating asthma.
[0005] Parkinson's disease is a disturbance of voluntary movement
in which muscles become stiff and sluggish, movement becomes clumsy
and difficult and uncontrollable rhythmic twitching of groups of
muscles produces characteristic shaking or tremor. The condition is
believed to be caused by a degeneration of pre-synaptic
dopaminergic neurones in the brain. The absence of adequate release
of the chemical transmitter dopamine during neuronal activity
thereby leads to the Parkinsonian symptomatology.
[0006] The most widely used treatment for Parkinsonism is
administration of L-DOPA, a precursor of dopamine which acts
indirectly by replacing the missing dopamine. However,
disadvantages are associated with the use of L-DOPA, for example,
patients often suffer from side-effects such as dyskinesia and
on-off effects, and it is necessary to administer L-DOPA in
conjunction with a peripheral dopa-decarboxylase inhibitor such as
carbidopa or benzaseride. These inhibitors prevent the peripheral
degradation of levodopa to dopamine, thus enabling more drug to
enter the brain and limiting peripheral side-effects. Such
treatment improves quality of life for patients but does not halt
disease progression. Furthermore, such treatment is associated with
a number of adverse effects, including nausea, vomiting, abdominal
distension and psychiatric side-effects (for example, toxic
confusional state, paranoia, and hallucinations). There continues
to be a need for improved treatments for Parkinson's disease.
BRIEF SUMMARY OF THE INVENTION
[0007] Disclosed herein is a method of treating asthma in a subject
in need thereof, the method comprising administering to the subject
a pharmaceutical composition comprising a compound represented by
Formula 1 or a pharmaceutically acceptable salt thereof and a
pharmaceutically acceptable excipient:
##STR00001##
[0008] In some embodiments of the method, the composition further
comprises a compound represented by Formula 2 or a pharmaceutically
acceptable salt thereof:
##STR00002##
[0009] The asthma can be but is not limited to allergen-induced
asthma, viral-induced asthma, cold-induced asthma,
pollution-induced asthma and/or exercise-induced asthma.
[0010] In some embodiments of the method of treating asthma, the
compositions are administered orally to the subject. In some
embodiments of the method of treating asthma, the compositions
comprise 0.001 mg to 100 mg of each of the compound per kg of the
subject's body weight.
[0011] Also disclosed herein are pharmaceutical compositions
comprising a compound represented by Formula 1 or a
pharmaceutically acceptable salt thereof, a compound represented by
Formula 2 or a pharmaceutically acceptable salt thereof, and a
pharmaceutically acceptable excipient:
##STR00003##
[0012] In some embodiments, the compositions are suitable for oral
administration to subjects. In some embodiments, the compositions
comprise 1 mg to 1000 mg of each of the compound.
[0013] Also disclosed herein is a method of treating Parkinson's
disease in a subject in need thereof, the method comprising
administering to the subject a pharmaceutical composition
comprising a compound represented by Formula 1 or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable excipient:
##STR00004##
[0014] In some embodiments of the method, the composition further
comprises a compound represented by Formula 2 or a pharmaceutically
acceptable salt thereof:
##STR00005##
[0015] In some embodiments of the method of treating Parkinson's
disease, the compositions are administered orally to the subject.
In some embodiments of the method of treating Parkinson's disease,
the compositions comprise 0.001 mg to 100 mg of each of the
compound per kg of the subject's body weight.
BRIEF DESCRIPTION OF THE FIGURES
[0016] FIG. 1. Effect of IPS-07004 on the survival of A549 lung
carcinoma cells.
[0017] FIG. 2. Effect of IPS-07004 on the expression of IL-6 and
IL-8 in A549 lung carcinoma cells induced by LPS.
[0018] FIG. 3. Effect of IPS-07004 on the expression of IL-6 in
A549 lung carcinoma cells induced by IL-1 beta.
[0019] FIG. 4. Effect of IPS-07004 on the expression of IL-8 in
A549 lung carcinoma cells induced by IL-1 beta.
[0020] FIG. 5. Effect of the combined treatment of IPS-07004 and
Montelukast on the expression of IL-6 and IL-8 in A549 lung
carcinoma cells induced by LPS.
[0021] FIG. 6. Effect of the combined treatment of IPS-07004 and
Montelukast on the expression of IL-6 and IL-8 in A549 lung
carcinoma cells induced by IL-1beta.
[0022] FIG. 7. Inhibitory effect of IPS-07004 on TSLP expression in
mice and HMC-1 cells.
[0023] FIG. 8. Inhibitory effect of IPS-07004 on mast cell number
and serum IgE level.
[0024] FIG. 9. Effect of IPS-07005 on human Caspase-1 activity in
HMC-1 cells.
[0025] FIGS. 10A and 10B. Effect of IPS-07005 on the expression on
TL-6 and IL-8 in HMC-1 cells induced by PMA.
[0026] FIGS. 11A and 11B. Effect of the combined treatment of
IPS-07005 on the expression of IL-6 and IL-8 in A549 lung carcinoma
cells induced by IL-1.beta..
[0027] FIG. 12. Inhibitory effect of IPS-07005 on OVA-specific IgE
expression.
[0028] FIGS. 13A and 13B. Inhibitory effect of IPS-07005 on total
cell number and eosinophil number in bronchoalveolar lavage (BAL)
fluid. FIG. 13C. Histological images of eosinophils.
[0029] FIG. 14. Effect of IPS-07005 on OVA-induced lung
histological changes, as determined by H&E staining
(100.times.).
[0030] FIGS. 15A and 15B. Histological images of airways stained
with PAS for goblet cell visualization.
[0031] FIGS. 16A and 16B. Inhibitory effect of IPS-07005 on
expression of IL17A and CXCL1 in BAL fluid.
[0032] FIGS. 17A and 17B. Inhibitory effect of IPS-07005 on
expression of CCL3 and CCL2 in BAL fluid.
[0033] FIG. 18. Effect of IPS-07004 on behavior of Parkinson's
disease (PD) mice.
[0034] FIG. 19. Effect of IPS-07005 on the expression of tyrosine
hydroxylase in striatum and substantia nigra in PD brain.
[0035] FIG. 20. Effect of IPS-07005 on the expression of IL-1 in PD
brain.
DETAILED DESCRIPTION OF THE INVENTION
[0036] Proguanil, also known as chlorguanide, is a prodrug that is
converted by the liver to its active metabolite, cycloguanil, an
inhibitor of dihydrofolate reductase (DHFR). The present disclosure
provides compositions comprising a compound represented by Formula
1 or a pharmaceutically acceptable salt thereof:
##STR00006##
[0037] The compositions can further comprise montelukast (trade
name SINGULAIR), a compound represented by Formula 2 or a
pharmaceutically acceptable salt thereof:
##STR00007##
[0038] Montelukast is a leukotriene receptor antagonist (LTRA) used
for the maintenance treatment of asthma and to relieve symptoms of
seasonal allergies. Montelukast is a CysLT.sub.1 antagonist; it
blocks the action of leukotriene D4 (and secondary ligands LTC4 and
LTE4) on the cysteinyl leukotriene receptor CysLT.sub.1 in the
lungs and bronchial tubes by binding to it. This reduces the
bronchoconstriction otherwise caused by the leukotriene and results
in less inflammation.
[0039] Montelukast is used for a number of conditions including
asthma, exercise induced bronchospasm, allergic rhinitis, primary
dysmenorrhoea (i.e., dysmenorrhoea not associated with known
causes; see dysmenorrhea causes), and urticaria. It is mainly used
as a complementary therapy in adults in addition to inhaled
corticosteroids, if they alone do not bring the desired effect. It
is also used to prevent allergic reactions and asthma flare-ups
during the administration of intravenous immunoglobulin.
[0040] The compositions disclosed herein can comprise proguanil,
proguanil and montelukast, or proguanil and other anti-asthma
agents, such as zileuton, zafirlukast, terbutaline sulfate, and
albuterol. The compositions disclosed herein can also comprise
proguanil, montelukast and other agents for treating Parkinson's
disease, such as L-DOPA, carbidopa, and/or benzaseride.
[0041] The pharmaceutically acceptable salts of the compounds
disclosed herein include salts derived from inorganic bases such as
lithium (Li), sodium (Na), potassium (K), calcium (Ca), magnesium
(Mg), iron (Fe), copper (Cu), zinc (Zn), and manganese (Mn); salts
of organic bases such as N,N'-diacetylethylenediamine, glucamine,
triethylamine, chlorine, hydroxide, dicyclohexylamine, metformin,
benzylamine, trialkylamine, thiamine, and equivalents thereof;
chiral bases such as alkylphenylamine, glycinol, phenyl glycinol
and equivalents thereof; natural amino acid salts such as glycine,
alanine, valine, leucine, isoleucine, norleucine, tyrosine,
cystine, cysteine, methionine, proline, hydroxyl proline,
histidine, ornithine, lysine, arginine, serine, and equivalents
thereof; quaternary ammonium salts of compounds of the present
disclosure having an alkyl sulfate such as an alkyl halide, Me, and
(Me).sub.2SO.sub.4 and equivalents thereof; artificial amino acids
such as D-isomer, substituted amino acid, or the like; guanidine,
guanine substituted with one selected from nitro, amino, alkyl,
alkenyl, and alkynyl, ammonium or substituted ammonium salts, and
aluminum salts. Salts can include acid-added salts, and examples of
suitable salts include sulphates, nitrates, phosphates,
perchlorates, borates, hydrohalides, acetates, tartrates, maliates,
citrates, fumarates, succinates, palmoates, methanesulfonate,
benzoate, salicylate, benzenesulfonate, ascorbate,
glycerophosphate, ketoglutarate, and equivalents thereof.
Pharmaceutically acceptable solvent compounds include
crystallization solvents such as hydroxides or alcohols.
[0042] In addition to salt forms, the present disclosure provides
compounds that are in a prodrug form. Prodrugs of the compounds
described herein are those compounds that readily undergo chemical
changes under physiological conditions to provide the compounds of
the disclosure. Additionally, prodrugs can be converted to the
compounds of the present disclosure by chemical or biochemical
methods in an ex vivo environment. For example, prodrugs can be
slowly converted to the compounds of the disclosure when placed in
a transdermal patch reservoir with a suitable enzyme or chemical
reagent. Prodrugs are often useful because, in some situations,
they can be easier to administer than the parent drug. They can,
for example, be bioavailable by oral administration whereas the
parent drug is not. The prodrug can also have improved solubility
in pharmaceutical compositions over the parent drug. A wide variety
of prodrug derivatives are known in the art, such as those that
rely on hydrolytic cleavage or oxidative activation of the prodrug.
An example, without limitation, of a prodrug would be a compound of
the disclosure which is administered as an ester (the "prodrug"),
but then is metabolically hydrolyzed to the carboxylic acid, the
active entity. Additional examples include peptidyl derivatives of
a compound of the disclosure.
[0043] Certain compounds of the present disclosure can exist in
unsolvated forms as well as solvated forms, including hydrated
forms. In general, the solvated forms are equivalent to unsolvated
forms and are intended to be encompassed within the scope of the
disclosure. Certain compounds of the disclosure can exist in
multiple crystalline or amorphous forms. In general, all physical
forms are equivalent for the uses contemplated by the disclosure
and are intended to be within the scope of the disclosure.
[0044] Certain compounds of the present disclosure possess
asymmetric carbon atoms (optical centers) or double bonds; the
racemates, enantiomers, diastereomers, geometric isomers and
individual isomers are all intended to be encompassed within the
scope of the disclosure. These isomers can be resolved or
asymmetrically synthesized using conventional methods to render the
isomers "optically pure," i.e., substantially free of its other
isomers. For example, if a particular enantiomer of a compound of
the present disclosure is desired, it can be prepared by asymmetric
synthesis, or by derivation with a chiral auxiliary, where the
resulting diastereomeric mixture is separated and the auxiliary
group cleaved to provide the pure desired enantiomers.
Alternatively, where the molecule contains a basic functional
group, such as amino, or an acidic functional group, such as
carboxyl, diastereomeric salts are formed with an appropriate
optically-active acid or base, followed by resolution of the
diasteromers thus formed by fractional crystallization or
chromatographic means well known in the art, and subsequent
recovery of the pure enantiomers.
Compositions
[0045] Also disclosed herein are pharmaceutical compositions
comprising a compound represented by Formula 1 or a
pharmaceutically acceptable salt thereof, a compound represented by
Formula 2 or a pharmaceutically acceptable salt thereof, and a
pharmaceutically acceptable excipient:
##STR00008##
[0046] In some embodiments, the compositions are suitable for oral
administration to subjects. In some embodiments, the compositions
comprise 1 mg to 1000 mg of each of the compound.
[0047] In some aspects, the present disclosure provides
pharmaceutical compositions suitable for pharmaceutical use
comprising one or more compounds of the present disclosure and a
pharmaceutically acceptable excipient or carrier. The compositions
disclosed herein can comprise proguanil, proguanil and montelukast,
or proguanil and other anti-asthma agents, such as zileuton,
zafirlukast, terbutaline sulfate, and albuterol. The compositions
disclosed herein can also comprise proguanil, montelukast and other
agents for treating Parkinson's disease, such as L-DOPA, carbidopa,
and/or benzaseride.
[0048] As used herein, "pharmaceutically acceptable" or
"pharmacologically acceptable" mean molecular entities and
compositions that do not produce an adverse, allergic or other
untoward reaction when administered to an animal, or to a human, as
appropriate.
[0049] The term, "pharmaceutically acceptable excipient or carrier"
includes one or more inert excipients, which include starches,
polyols, granulating agents, microcrystalline cellulose, diluents,
lubricants, binders, disintegrating agents, and the like, and any
and all solvents, dispersion media, coatings, antibacterial and
antifungal agents, isotonic and absorption delaying agents and the
like. The use of such media and agents for pharmaceutical active
substances is well known in the art. Except insofar as any
conventional media or agent is incompatible with the active
ingredient, its use in the therapeutic compositions is
contemplated. Supplementary active ingredients can also be
incorporated into the compositions. "Pharmaceutically acceptable
excipient" also encompasses controlled release means.
[0050] Formulations can improve one or more pharmacokinetic
properties (e.g., oral bioavailability, membrane permeability) of a
compound or combinations of compounds of the disclosure (herein
referred to as the active ingredient(s)).
[0051] The composition, shape, and type of dosage form can
typically vary according to applications thereof. For example, a
dosage form suitable for mucosal administration can include a
smaller amount of the active ingredient than that in a dosage form
suitable for oral administration used in treating the same disease.
These aspects of the present disclosure will be fairly apparent to
those of ordinary skill in the art (reference: Remington's
Pharmaceutical Sciences (1990) 18th ed., Mack Publishing, Easton
Pa.).
[0052] Typical pharmaceutical compositions and dosage forms include
one or more excipients. Suitable excipients are apparent to those
of ordinary skill in the pharmaceutical art, and the present
disclosure is not limited to examples of suitable excipients
described herein.
[0053] Whether a particular excipient is suitable for a
pharmaceutical composition or a dosage form depends on various
factors well known in the art, including methods of formulating
preparations to be administered to a patient, but is not limited
thereto. For example, dosage forms for oral administration such as
tablets can include an excipient not suitable for use in
preparations for non-oral administration.
[0054] The pharmaceutical compositions include those suitable for
aerosol, pulmonary, inhalation, oral, parenteral (including
subcutaneous, intradermal, intramuscular, intravenous and
intraarticular), rectal and topical (including dermal, buccal,
sublingual and intraocular) administration. The most suitable route
can depend upon the condition and disorder of the recipient. The
compositions can conveniently be presented in unit dosage form and
can be prepared by any of the methods well known in the art of
pharmacy. In general, the compositions are prepared by uniformly
and intimately bringing into association the active ingredient with
liquid carriers or finely divided solid carriers or both and then,
if necessary, shaping the product into the desired composition.
[0055] Compositions of the present disclosure can also optionally
include other therapeutic ingredients, anti-caking agents,
preservatives, sweetening agents, colorants, flavors, desiccants,
plasticizers, dyes, and the like. Any such optional ingredient
must, of course, be compatible with the compound of the disclosure
to insure the stability of the composition.
[0056] The active agents, alone or in combination with other
suitable components, can be administered by pulmonary route
utilizing several techniques including but not limited to
intratracheal instillation (delivery of solution into the lungs by
syringe), intratracheal delivery of liposomes, insufflation
(administration of powder composition by syringe or any other
similar device into the lungs) and aerosol inhalation. Compositions
suitable for pulmonary route inhalation include sterile solutions
for nebulization comprising a therapeutically effective amount of
the compound dissolved in aqueous saline solution and optionally
containing a preservative such as benzalkonium chloride or
chlorobutanol, and aerosol compositions comprising a
therapeutically effective amount dissolved or suspended in an
appropriate propellant. Aerosols (e.g., jet or ultrasonic
nebulizers, metered-dose inhalers (MDIs), and dry-powder inhalers
(DPIs)) can also be used in intranasal applications. Aerosols can
be conveniently presented in unit dosage form and prepared by any
of the methods well-known in the art of pharmacy. Aerosol
compositions are stable dispersions or suspensions of solid
material and liquid droplets in a gaseous medium and can be placed
into pressurized acceptable propellants, such as hydrofluoroalkanes
(HFAs, i.e., HFA-134a and HFA-227, or a mixture thereof),
dichlorodifluoromethane (or other chlorofluocarbon propellants such
as a mixture of Propellants 11, 12, and/or 114), propane, nitrogen,
and the like. Pulmonary compositions can include permeation
enhancers such as fatty acids, and saccharides, chelating agents,
enzyme inhibitors (e.g., protease inhibitors), adjuvants (e.g.,
glycocholate, surfactin, span 85, and nafamostat), preservatives
(e.g., benzalkonium chloride or chlorobutanol), and ethanol
(normally up to 5% but possibly up to 20%, by weight). Ethanol is
commonly included in aerosol compositions as it can improve the
function of the metering valve and in some cases also improve the
stability of the dispersion. Pulmonary compositions can also
include surfactants, which include but are not limited to bile
salts and those described in U.S. Pat. No. 6,524,557 and references
therein. The surfactants described in U.S. Pat. No. 6,524,557,
e.g., a C8-C16 fatty acid salt, a bile salt, a phospholipid, or
alkyl saccharide are advantageous in that some of them also
reportedly enhance absorption of the compound in the
composition.
[0057] Also suitable are dry powder compositions comprising a
therapeutically effective amount of active compound blended with an
appropriate carrier and adapted for use in connection with a
dry-powder inhaler. Absorption enhancers which can be added to dry
powder compositions of the present disclosure include those
described in U.S. Pat. No. 6,632,456. WO02/080884 describes new
methods for the surface modification of powders. Aerosol
compositions can include U.S. Pat. Nos. 5,230,884, 5,292,499,
WO017/8694, WO01/78696, US2003019437, US20030165436, and WO96/40089
(which includes vegetable oil). Sustained release compositions
suitable for inhalation are described in US20010036481A1,
US20030232019A1, and US20040018243A1 as well as in WO01/13891,
WO02/067902, WO03/072080, and WO03/079885. Pulmonary compositions
containing microparticles are described in WO03/015750,
US20030008013, and WO00/00176. Pulmonary compositions containing
stable glassy state powder are described in US20020141945 and U.S.
Pat. No. 6,309,671. Other aerosol compositions are described in EP
1338272A1, WO90/09781, U.S. Pat. Nos. 5,348,730, 6,436,367,
WO91/04011, and U.S. Pat. Nos. 6,294,153 and 6,290,987 describes a
liposomal based composition that can be administered via aerosol or
other means. Powder compositions for inhalation are described in
US20030053960 and WO01/60341. The agents can be administered
intranasally as described in US20010038824.
[0058] While the pulmonary route is advantageous in most instances,
there can also be instances in which other routes of administration
can be advantageous. For example, oral administration can be
desirable. In that regard, one can contemplate administration using
a composition in which the compound is releasably encapsulated by
modified amino acids, as described in U.S. Pat. No. 5,811,127. One
can also contemplate administration as an implantable
sustained-release dosage form, such as described in
US20040115236.
[0059] The pharmaceutical compositions containing the active
ingredient can be in a form suitable for oral use, for example, as
tablets, troches, lozenges, aqueous or oily suspensions,
dispersible powders or granules, emulsions, hard or soft capsules,
or syrups or elixirs. Compositions intended for oral use can be
prepared according to any method known to the art for the
manufacture of pharmaceutical compositions. Such compositions can
contain one or more agents selected from sweetening agents,
flavoring agents, coloring agents and preserving agents in order to
provide pharmaceutically elegant and palatable preparations.
Tablets contain the active ingredient in admixture with other
non-toxic pharmaceutically acceptable excipients which are suitable
for the manufacture of tablets. These excipients can be, for
example, inert diluents, such as calcium carbonate, sodium
carbonate, lactose, calcium phosphate or sodium phosphate;
granulating and disintegrating agents, for example, corn starch, or
alginic acid; binding agents, for example starch, gelatin or
acacia, and lubricating agents, for example magnesium stearate,
stearic acid or talc. The tablets can be uncoated or they can be
coated by known techniques to delay disintegration and absorption
in the gastrointestinal tract and thereby provide a sustained
action over a longer period. For example, a time delay material
such as glyceryl monostearate or glyceryl distearate can be
employed. They can also be coated by the techniques described in
U.S. Pat. Nos. 4,256,108; 4,160,452; and U.S. Pat. No. 4,265,874 to
form osmotic therapeutic tablets for control release.
[0060] A tablet can be made by compression or molding, optionally
with one or more accessory ingredients. Compressed tablets can be
prepared by compressing in a suitable machine the active ingredient
in a free-flowing form such as a powder or granules, optionally
mixed with a binder, lubricant, inert diluent, lubricating, surface
active or dispersing agent. Molded tablets can be made by molding
in a suitable machine a mixture of the powdered compound moistened
with an inert liquid diluent. The tablets can optionally be coated
or scored and can be formulated so as to provide sustained, delayed
or controlled release of the active ingredient therein.
[0061] Compositions for oral use can also be presented as hard
gelatin capsules wherein the active ingredient is mixed with an
inert solid diluent, for example, calcium carbonate, calcium
phosphate or kaolin, or as soft gelatin capsules wherein the active
ingredient is mixed with water or an oil medium, for example peanut
oil, liquid paraffin, or olive oil.
[0062] Aqueous suspensions contain the active materials in
admixture with excipients suitable for the manufacture of aqueous
suspensions. Such excipients are suspending agents, for example,
sodium carboxymethylcellulose, methylcellulose,
hydroxypropyl-methylcellulose, sodium alginate,
polyvinyl-pyrrolidone, gum tragacanth and gum acacia; dispersing or
wetting agents can be a naturally-occurring phosphatide, for
example lecithin, or condensation products of an alkylene oxide
with fatty acids, for example, polyoxyethylene stearate, or
condensation products of ethylene oxide with long chain aliphatic
alcohols, for example, heptadecaethyleneoxycetanol, or condensation
products of ethylene oxide with partial esters derived from fatty
acids and a hexitol such as polyoxyethylene sorbitol monooleate, or
condensation products of ethylene oxide with partial esters derived
from fatty acids and hexitol anhydrides, for example polyethylene
sorbitan monooleate. The aqueous suspensions can also contain one
or more preservatives, for example, ethyl, or n-propyl,
p-hydroxybenzoate, one or more coloring agents, one or more
flavoring agents, and one or more sweetening agents, such as
sucrose or saccharin.
[0063] Dispersible powders and granules suitable for preparation of
an aqueous suspension by the addition of water provide the active
ingredient in admixture with a dispersing or wetting agent,
suspending agent and one or more preservatives. Suitable dispersing
or wetting agents and suspending agents are exemplified by those
already mentioned above. Additional excipients, for example
sweetening, flavoring and coloring agents, can also be present.
[0064] The pharmaceutical compositions can be in the form of a
sterile injectable aqueous or oleagenous suspension. This
suspension can be formulated according to the known art using those
suitable dispersing or wetting agents and suspending agents which
have been mentioned above. The sterile injectable preparation can
also be a sterile injectable solution or suspension in a non-toxic
parenterally acceptable diluent or solvent, for example as a
solution in 1,3-butane diol. Among the acceptable vehicles and
solvents that can be employed are water, Ringer's solution and
isotonic sodium chloride solution. In addition, sterile, fixed oils
are conventionally employed as a solvent or suspending medium. For
this purpose any bland fixed oil can be employed including
synthetic mono- or diglycerides. In addition, fatty acids such as
oleic acid find use in the preparation of injectables.
[0065] The pharmaceutical compositions and methods of the
disclosure can further comprise other therapeutically active
compounds, as noted herein, useful in the treatment of asthma,
allergic diseases, and inflammatory conditions. In many instances,
compositions which include a compound disclosed herein and an
alternative agent have additive or synergistic effects when
administered.
[0066] The pharmaceutical compositions for the administration of
the compounds of this disclosure can conveniently be presented in
unit dosage form and can be prepared by any of the methods well
known in the art. All methods include the step of bringing the
active ingredient into association with the carrier which
constitutes one or more accessory ingredients. In general, the
pharmaceutical compositions are prepared by uniformly and
intimately bringing the active ingredient into association with a
liquid carrier or a finely divided solid carrier or both, and then,
if necessary, shaping the product into the desired composition. In
the pharmaceutical composition the active object compound is
included in an amount sufficient to produce the desired effect upon
the process or condition of diseases.
Methods of Use
[0067] The term "subject" or "patient" is defined herein to include
animals such as mammals, including, but not limited to, primates
(e.g., human or nonhuman), cows, sheep, goats, horses, dogs, cats,
rabbits, rats, mice and the like. In some embodiments, the subject
is a human.
[0068] The terms "prevent", "preventing" and "prevention", as used
herein, refer to a method of delaying or precluding the onset of a
disease and/or its attendant symptoms, barring a subject from
acquiring a disease or reducing a subject's risk of acquiring a
disease.
[0069] The terms "treat", "treating" and "treatment", as used
herein, are meant to include alleviating, reducing, or abrogating a
disease or condition and/or its attendant symptoms and alleviating,
reducing, or eradicating the cause of the disease itself.
[0070] The term "therapeutically effective amount" refers to the
amount of the subject compound that will elicit the biological or
medical response of a tissue, system, animal or human that is being
sought by the researcher, veterinarian, medical doctor or other
clinician. The term "therapeutically effective amount" includes
that amount of a compound that, when administered, is sufficient to
prevent development of, or alleviate to some extent, one or more of
the symptoms of the condition or disorder being treated. The
therapeutically effective amount will vary depending on the
compound, the disease and its severity and the age, weight, etc.,
of the mammal to be treated.
[0071] Disclosed herein are methods of treating asthma in a subject
in need thereof, the method comprising administering to the subject
a pharmaceutical composition comprising a compound represented by
Formula 1 or a pharmaceutically acceptable salt thereof and a
pharmaceutically acceptable excipient:
##STR00009##
[0072] Also disclosed herein are methods of treating asthma in a
subject in need thereof, the methods comprising administering to
the subject a pharmaceutical composition comprising a compound
represented by Formula 1 or a pharmaceutically acceptable salt
thereof, a compound represented by Formula 2 or a pharmaceutically
acceptable salt thereof, and a pharmaceutically acceptable
excipient:
##STR00010##
[0073] Also disclosed are methods of treating asthma in a subject
in need thereof, the methods comprising administering to the
subject a pharmaceutical composition comprising proguanil and other
anti-asthma agents, such as zileuton, zafirlukast, terbutaline
sulfate, and albuterol. Methods for preventing and/or treating
asthma includes allergen-induced asthma, viral-induced asthma,
cold-induced asthma, pollution-induced asthma and/or
exercise-induced asthma.
[0074] In some embodiments of the methods of treating asthma, the
compositions are administered orally to the subject. In some
embodiments of the methods of treating asthma, the compositions
comprise 0.001 mg to 100 mg of each of the compound per kg of the
subject's body weight.
[0075] Also disclosed herein are methods of treating Parkinson's
disease in a subject in need thereof, the methods comprising
administering to the subject a pharmaceutical composition
comprising a compound represented by Formula 1 or a
pharmaceutically acceptable salt thereof and a pharmaceutically
acceptable excipient:
##STR00011##
[0076] Also disclosed herein are methods of treating Parkinson's
disease in a subject in need thereof, the methods comprising
administering to the subject a pharmaceutical composition
comprising a compound represented by Formula 1 or a
pharmaceutically acceptable salt thereof, a compound represented by
Formula 2 or a pharmaceutically acceptable salt thereof, and a
pharmaceutically acceptable excipient:
##STR00012##
[0077] In some embodiments of the methods of treating Parkin's
disease, the compositions are administered orally to the subject.
In some embodiments of the methods of treating Parkinson's disease,
the compositions comprise 0.001 mg to 100 mg of each of the
compound per kg of the subject's body weight.
[0078] Depending on the disease to be treated and the subject's
condition, the compounds of the disclosure can be administered by
aerosol, pulmonary, inhalation, oral, parenteral (e.g.,
intramuscular, intraperitoneal, intravenous, ICV, intracisternal
injection or infusion, subcutaneous injection or implant),
inhalation, nasal, vaginal, rectal, sublingual, or topical (e.g.,
transdermal, local) routes of administration and can be formulated,
alone or together, in suitable dosage unit compositions containing
conventional non-toxic pharmaceutically acceptable carriers,
adjuvants and vehicles appropriate for each route of
administration. The disclosure also contemplates administration of
the compounds of the disclosure in a depot composition, in which
the active ingredient is released over a defined time period.
[0079] The dose range of the active ingredient(s) or a
pharmaceutically acceptable salt thereof for adult humans is
generally from about 0.005 mg/day to about 10 g/day, from about 1
mg/day to about 1,000 mg/day, about 10 mg/day to about 750 m/day,
about 50 mg/day to about 500 mg/day, or about 75 mg/day to about
350 mg/day. Tablets or other forms of presentation provided in
discrete units can conveniently contain an amount of compound of
the disclosure which is effective at such dosage or as a multiple
of the same, for instance, units containing 5 mg to 500 mg, usually
around 10 mg to 200 mg. The precise amount of compound administered
to a patient will be the responsibility of the attendant physician.
However, the dose employed will depend on a number of factors,
including the age and sex of the patient, the precise disorder
being treated, and its severity.
[0080] In the treatment or prevention of asthma, an appropriate
dosage level will generally be about 0.001 to 100 mg per kg patient
body weight per day, which can be administered in single or
multiple doses. The dosage level can be about 0.01 mg/kg per day to
about 75 mg/kg per day or about 0.05 mg/kg per day to about 20
mg/kg per day. A suitable dosage level can be about 0.01 mg/kg per
day to about 50 mg/kg per day, about 0.05 mg/kg per day to about 20
mg/kg per day, or about 0.1 mg/kg per day to about 10 mg/kg per
day. Within this range the dosage can be 0.01 mg/kg per day to 0.1
mg/kg per day, 0.1 mg/kg per day to 1 mg/kg per day, 1 mg/kg per
day to 10 mg/kg per day. For oral administration, the compositions
can be provided in the form of tablets containing 1 mg to 1000 mg
of the active ingredient, such as but not limited to 1 mg, 5 mg, 10
mg, 15 mg, 20 mg, 25 mg, 50 mg, 75 mg, 100 mg, 150 mg, 200 mg, 250
mg, 300 mg, 400 mg, 500 mg, 600 mg, 750 mg, 800 mg, 900 mg, and
1000 mg of the active ingredient for the symptomatic adjustment of
the dosage to the patient to be treated. The compounds can be
administered on a regimen of 1 to 4 times per day, e.g., once,
twice, thrice, or four times per day.
[0081] Similarly, in the treatment or prevention of Parkinson's
disease, an appropriate dosage level will generally be about 0.001
to 100 mg per kg patient body weight per day, which can be
administered in single or multiple doses. The dosage level can be
about 0.01 mg/kg per day to about 75 mg/kg per day or about 0.05
mg/kg per day to about 20 mg/kg per day. A suitable dosage level
can be about 0.01 mg/kg per day to about 50 mg/kg per day, about
0.05 mg/kg per day to about 20 mg/kg per day, or about 0.1 mg/kg
per day to about 10 mg/kg per day. Within this range the dosage can
be 0.01 mg/kg per day to 0.1 mg/kg per day, 0.1 mg/kg per day to 1
mg/kg per day, 1 mg/kg per day to 10 mg/kg per day. For oral
administration, the compositions can be provided in the form of
tablets containing 1 mg to 1000 mg of the active ingredient, such
as but not limited to 1 mg, 5 mg, 10 mg, 15 mg, 20 mg, 25 mg, 50
mg, 75 mg, 100 mg, 150 mg, 200 mg, 250 mg, 300 mg, 400 mg, 500 mg,
600 mg, 750 mg, 800 mg, 900 mg, and 1000 mg of the active
ingredient for the symptomatic adjustment of the dosage to the
patient to be treated. The compounds can be administered on a
regimen of 1 to 4 times per day, e.g., once, twice, thrice, or four
times per day.
[0082] In some embodiments, the dosage of proguanil or montelukast
is provided at about 0.05 mg/day to about 1000 mg/day, including
any and all variations within this range, such as about 0.1 mg/day
to about 1000 mg/day, about 1 mg/day to about 1000 mg/day, about
2.5 mg/day to about 100 mg/day, about 5 mg/day to about 50 mg/day,
about 10 mg/day to about 750 mg/day, about 50 mg/day to about 500
mg/day, or about 75 mg/day to about 350 mg/day.
[0083] In some embodiments, montelukast is provided as a tablet, a
chewable tablet, and granules to be taken by mouth. Montelukast can
be taken once a day with or without food. Montelukast can be
administered as montelukast sodium, with 5.2 mg of montelukast
sodium being equivalent to 5 mg of montelukast.
[0084] It will be understood, however, that the specific dose level
and frequency of dosage for any particular patient can be varied
and will depend upon a variety of factors including the activity of
the specific compound employed, the metabolic stability and length
of action of that compound, the age, body weight, general health,
sex, diet, mode and time of administration, rate of excretion, drug
combination, the severity of the particular condition, and the host
undergoing therapy.
[0085] The compounds of the present disclosure can be combined or
used in combination with other agents useful in the treatment,
prevention, suppression or amelioration of the diseases or
conditions for which compounds of the disclosure are useful.
[0086] Such other agents, or drugs, can be administered, by a route
and in an amount commonly used therefor, simultaneously or
sequentially with a compound of the disclosure. When a compound of
the disclosure is used contemporaneously with one or more other
drugs, a pharmaceutical composition containing such other drugs in
addition to the compound of the disclosure is preferred.
Accordingly, the pharmaceutical compositions of the disclosure
include those that also contain one or more other active
ingredients or therapeutic agents, in addition to a compound of the
disclosure.
[0087] The weight ratio of the compound of the disclosure to the
second active ingredient can be varied and will depend upon the
effective dose of each ingredient. Generally, an effective dose of
each will be used. Thus, for example, when a compound of the
disclosure is combined with an NSAID, the weight ratio of the
compound of the disclosure to the NSAID will generally range from
about 1000:1 to about 1:1000, or about 200:1 to about 1:200.
Combinations of a compound of the disclosure and other active
ingredients will generally also be within the aforementioned range,
but in each case, an effective dose of each active ingredient
should be used.
[0088] Hereinafter, the present disclosure will be described in
further detail with reference to the following non-limiting
examples. However, these examples are provided for illustrative
purposes only and are not intended to limit the scope of the
present disclosure.
EXAMPLES
Example 1. In Vitro Cell Viability Assay (MTT)
[0089] IPS-07004 (proguanil) was purchased from Key Organics
(London, UK).
[0090] A549 lung carcinoma cells (1.times.10.sup.3/well) were
seeded on a 96-well plate for 16 hrs. After starvation for 24 hrs.,
the cells were pretreated with LPS or IL-1b for 4 hrs. The cells
were then treated with IPS-07004 for 24 hrs. in a dose-dependent
manner. After 48 hrs. of incubation, MTT
[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide]
solution (5 mg/ml) was added to each well in an amount of 1/10 of
total volume, and the cells were incubated for 2 hrs. at 37.degree.
C.
[0091] To examine the survival rate for A549 cells treated with
IPS-07004, an in vitro cell viability assay was performed using
MTT. There were little or no effect of IPS-07004 on the viability
when the cells were induced by LPS (lipopolysaccharide) and
IL-1beta (interleukin-1beta) (FIG. 1).
Example 2. Enzyme-Linked Immunosorbent Assay (ELISA) of IL-6 and
IL-8
[0092] A549 cells (1.times.10.sup.5/well) were seeded on a 96-well
plate for 16 hrs. After starvation for 24 hrs., the cells were
pretreated with LPS or IL-1beta for 4 hrs. The cells were then
treated with IPS-07004 for 24 hrs. in a dose-dependent manner.
Cytokines levels (IL-6 and IL-8) in media were analyzed in
accordance with the manufacturer's specification (BD Biosciences
Pharmingen, San Diego, Calif., USA).
[0093] To assess whether IPS-07004 inhibits the expression of IL-6
and IL-8, which play a critical role in asthma pathology, ELISA of
the cytokines in A549 cells was employed.
[0094] IPS-07004 reduced the expression levels of IL-6 and IL-8 in
A549 cells induced by LPS (FIG. 2) and IL-1beta (FIGS. 3 and 4) in
a dose-dependent manner. These data indicate that IPS-07004 is
applicable for treating asthma.
Example 3. Effect of the Combined Treatment of IPS-07004 and
Montelukast on the Expression of IL-6 and IL-8 in A549 Lung
Carcinoma Cells Induced by LPS or IL-1Beta
[0095] To examine the synergistic effect of IPS-07004 combined with
Motelukast (an anti-asthma agent) on the anti-asthmatic function,
cytokine expression assay in A549 cells was carried out.
[0096] The combination of IPS-07004 with Montelukast appeared to be
more effective in suppressing the expression of IL-6 and IL-8 in
A549 cells induced by LPS (FIG. 5) and IL-1beta (FIG. 6) compared
with treatment with Motelukast alone. These data indicate that
IPS-07004 can be used for combined therapy with Montelukast for
treating asthma.
Example 4. Compound Profile of IPS-07005
[0097] Indication: Asthma
[0098] Mode of action target: NLRP3 inflammasome-derived Caspase-1
inhibitor
[0099] Lead material: IPS-07004
[0100] Composition: IPS-07004 10 mg/kg (mpk)+Montelukast 1.0
mpk
[0101] Development step: Phase I
[0102] FIG. 7 shows the inhibitory effect of IPS-07004 on Caspase-1
activity and TSLP (Thymic Stromal Lymphopoietin) expression in
HMC-1 cells and DNCB (2,4-dinitrochlorobenzene) mice.
[0103] FIG. 8 shows the inhibitory effect of IPS-07004 on mast cell
number and serum IgE level.
Example 5. Inhibitory Effect of IPS-07005 on Asthma--In Vitro
Cell-Based Assay
[0104] Results
[0105] Using HMC-1 (human mast cell line) cell-based assay,
IPS-07005 ((IPS-07004+Montelukast) inhibited the activity of
Caspase-1, a critical component of inflammasome, to suppress the
conversion of Pro-TSLP, IL-1beta, and -IL-18 to active forms as
shown in FIG. 9.
[0106] IPS-07004 alone inhibited caspase-1 activity in HMC-1 cells
in a dose-dependent manner and IPS-07005 provided a synergistic
effect.
[0107] FIG. 10 shows the effect of IPS-07005 on the expression of
IL-6 and IL-8 in HMC-1 cells induced by PMA (Phorbol 12-myristate
13-acetate) and A23187, a calcium ionophore (C1). IPS-07005
inhibited the expressions of IL-6 and IL-8 compared with IPS-07004,
montelukast alone or dexamethasone in HMC-1 cells.
[0108] To assess whether IPS-07005 inhibits the expression of IL-6
and IL-8 which play a critical role in asthma pathology, ELISA of
the cytokines in A549 lung carcinoma cells was employed. IPS-07005
reduced the expression levels of IL-6 and IL-8 in A549 cells
induced by IL-10 (FIG. 11) in a dose-dependent manner. IPS-07005
inhibited the expression of IL-6 and IL-8 compared to IPS-07004,
montelukast alone, or dexamethasone in A549 cells (human lung
cancer cells).
[0109] These data indicate that IPS-07005 is applicable for
treating asthma.
Example 6. Inhibitory Effect of IPS-07005 on Asthma--In Vivo Animal
Data
[0110] FIG. 12 shows the inhibitory effect of IPS-07005 on
OVA-specific IgE expression. IPS-07005 (IPS-07004 10
mpk+montelukast 1 mpk) showed significant reduction of OVA-induced
IgE level in bronchoalveolar lavage fluid (BALF).
[0111] FIGS. 13A and 13B show the inhibitory effect of IPS-07005 on
total cell number and eosinophil number in bronchoalveolar lavage
(BAL) fluid. FIG. 13C shows decreased number of eosinophils in
OVA-challenged cells when treated with IPS-07004 or IPS-07005.
Analysis of the inflammatory cells in the BALF samples revealed
that total cell numbers were significantly increased by OVA
sensitization. However, total cell numbers were reduced by
treatment with IPS-07005. Specifically, administration of IPS-07005
also significantly reduced the total and eosinophil cell counts in
BALF in the corresponding group compared with the OVA-challenged
group (FIGS. 13A and 13B).
[0112] FIG. 14 shows the effects of IPS-07005 on OVA-induced lung
histological changes, as determined by H&E staining
(100.times.). FIG. 14 shows the histological images of airways
stained with PAS for goblet cell visualization. The lung sections
were stained with PAS (Periodic acid-Schiff) to evaluate their
levels of goblet cell hyperplasia. We observed noticeable
differences in the sizes of the purple areas, i.e., the areas of
lung tissue stained with PAS, among the three groups. We noted
goblet cell hyperplasia and mucus overproduction in the bronchial
passages of OVA-challenged mice. However, we noted a significantly
lower number of goblet cells in the IPS-07005 and high dose of
Montelukast-treated groups than in the OVA-challenged group (FIGS.
15A and 15B).
[0113] FIG. 16 shows the inhibitory effect of IPS-07005 on
expression of IL17A and CXCL1 in BAL fluid. FIG. 17 shows the
inhibitory effect of IPS-07005 on expression of CCL3 and CCL2 in
BAL fluid.
[0114] To determine the anti-asthmatic effect of IPS-07005 on
cytokine levels in mice with asthma, IL-17A, CXCL1, CCL2, and CCL3
production in BAL fluid was measured. Exposure to OVA caused a
marked increase of IL-17A, CXCL1, CCL2, and CCL3 in BAL fluid
compared with the mice in the normal group (FIGS. 16 and 17).
However, the levels of IL-17A, CXCL1, which play an important role
in neutrophil infiltration, in BAL fluid following OVA exposure
were decreased in the presence of IPS-07005 in OVA-exposed mice
(FIG. 16).
[0115] This result strongly suggested that IPS-07005 can suppress
neutrophilic asthma, which is a medical unmet need for treating
asthma. The levels of CCL2 and CCL3, which play a key role in
macrophage activity in BAL fluid were decreased by IPS-07005 (FIG.
17). It appeared to be more effective in reducing the expression of
these cytokines compared to IPS-07004 or Montelukast. The
inhibitory effect of IPS-07005 on the expression of these cytokines
is due to the synergistic function of IPS-07004 and Montelukast.
Taken together, these findings demonstrate that the
anti-inflammatory effect of IPS-07005 is mediated by the regulation
of multiple inflammatory factors.
[0116] Methods
[0117] 25 mg Ovalbumin (OVA) and 1 mg aluminum hydroxide hydrate
(AlOH3) were dissolved in 300 mL of sterile saline solution and
sensitized by intraperitoneal administration at 0 and 21 days. At
the end of the observation period, all living animals were
anesthetized using Isoflurane, and blood was collected from the
vena cava/abdominal aorta. Lungs are removed and washed with
sterile saline solution. 250 mL of sterile saline was filled in a 1
mL syringe, and the injection and recovery were repeated three
times. The washed solution was lightly centrifuged (300 rpm, 10
minutes) and the supernatant was frozen and used for analysis of
CXCL1 (ab216951, Abcam PLC) IL-17A (ab199081, Abcam PLC) and CCL3
(ab100726, Abcam PLC). The precipitate was resuspended in the same
amount of sterile saline solution and used for eosinophil count and
total cell count. The collected blood was left at room temperature
for 30 minutes and centrifuged at 3,000 rpm for 20 mins. to
separate serum and used for total IgE and Ova-specific IgE
analysis. Histopathological examination was performed using lung
tissue fixed in formalin. Lung tissues extracted at autopsy were
placed in a cassette and submerged in 10% Neutral buffered formalin
(10% NBF) at least 20 times the tissue volume. After fixation, the
tissue was cut to about 4 mm in thickness and fabricated into
paraffin blocks after general tissue treatment and embedding. After
that, the slice was cut to 4 .mu.m thickness using a rotary
microtome, and Hematoxylin & Eosin (H&E) staining and PAS
staining were performed.
Example 7. Inhibition of Parkinson's Disease
[0118] Methods
[0119] For 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)
intoxication, the mice received four intraperitoneal (i.p.)
injections of MPTP-HCl (20 mg/kg, free base in saline;
Sigma-Aldrich, St. Louis, Mo.) dissolved in PBS at 2 h intervals.
Twelve hours after the last MPTP injection, the MPTP-injected mice
were received IPS-07004, Montelukast, or IPS-07005 once daily
through p.o. administrations.
[0120] To examine the effect of IPS-07004 on abnormal behavior of
PD mice, Pole test (T-turn and T-LA time) was carried out. PD was
induced in mice treated with MPTP (20 mg/kg, IP) and the mice were
administered with IPS-07004 at a dose of 10 mg/kg. And then pole
test was performed in the mice. which can measure bradykinesia, a
hallmark of Parkinson's disease. The T-turn measures the time the
black mouse turns down from the top of the pole. IPS-07004 reduced
the T-turn and T-LA time compared with MPTP-treated control group
and L-Dopa-treated group, as shown in FIG. 18.
[0121] To determine the anti-PD function of IPS-07004 and IPS-07005
in PD mice, we analyzed the expression of tyrosine hydroxylase (TH)
in striatum and Substantia nigra in PD brain. Immunohistochemical
analysis in the brain showed that TH level was decreased in
IPS-07004 and 07005-treated mice compared with MPTP-treated control
group and L-Dopa-treated group, as shown in FIG. 19.
[0122] To assess whether the anti-PD function of IPS-07004 and
07005 is due to inhibit inflammation in PD brain, the analysis of
cytokine expression was performed. IPS-07004 and IPS-07005 reduced
IL-1b expression in PD mice, as shown in FIG. 20.
[0123] The foregoing description of the specific embodiments will
so fully reveal the general nature of the invention that others
can, by applying knowledge within the skill of the art, readily
modify and/or adapt for various applications, without departing
from the general concept of the invention. Therefore, such
adaptations and modifications are intended to be within the meaning
and range of equivalents of the disclosed embodiments, based on the
teaching and guidance presented herein. It is to be understood that
the phraseology or terminology herein is for the purpose of
description and not of limitation, such that the terminology or
phraseology of the present specification is to be interpreted by
the skilled artisan in light of the teachings and guidance.
[0124] The breadth and scope of the present invention should not be
limited by any of the above-described exemplary embodiments but
should be defined only in accordance with the following claims and
their equivalents.
[0125] All of the various aspects, embodiments, and options
described herein can be combined in any and all variations.
[0126] All publications, patents, and patent applications mentioned
in this specification are herein incorporated by reference to the
same extent as if each individual publication, patent, or patent
application was specifically and individually indicated to be
incorporated by reference.
* * * * *