U.S. patent application number 14/047679 was filed with the patent office on 2014-02-06 for methods of using ryanodine antagonists in treating neural injury.
This patent application is currently assigned to ALLERGAN, INC.. The applicant listed for this patent is ALLERGAN, INC.. Invention is credited to Cun-Jian Dong, William A. Hare.
Application Number | 20140039019 14/047679 |
Document ID | / |
Family ID | 29999695 |
Filed Date | 2014-02-06 |
United States Patent
Application |
20140039019 |
Kind Code |
A1 |
Dong; Cun-Jian ; et
al. |
February 6, 2014 |
Methods of Using Ryanodine Antagonists in Treating Neural
Injury
Abstract
The present invention provides a method of providing
neuroprotection to a mammal comprising administering to said mammal
suffering from or at risk of suffering a noxious action on its
nerve cells an effective amount of a ryanodine antagonist, e.g.
dantrolene, to inhibit or prevent nerve cell injury or death.
Inventors: |
Dong; Cun-Jian; (Irvine,
CA) ; Hare; William A.; (Tustin, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
ALLERGAN, INC. |
IRVINE |
CA |
US |
|
|
Assignee: |
ALLERGAN, INC.
IRVINE
CA
|
Family ID: |
29999695 |
Appl. No.: |
14/047679 |
Filed: |
October 7, 2013 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10189676 |
Jul 3, 2002 |
8557855 |
|
|
14047679 |
|
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Current U.S.
Class: |
514/390 |
Current CPC
Class: |
A61P 27/02 20180101;
A61K 31/4166 20130101; A61P 27/06 20180101; A61P 25/16 20180101;
A61P 25/28 20180101; A61K 31/4178 20130101; A61P 25/00
20180101 |
Class at
Publication: |
514/390 |
International
Class: |
A61K 31/4178 20060101
A61K031/4178 |
Claims
1-16. (canceled)
17. A method for treating retinal ganglion cell loss in a patient
comprising administering dantrolene to said patient.
18. The method according to claim 17, wherein said administering
comprises oral administration.
19. The method according to claim 17 wherein said administering
comprises intravitreal injection.
20. The method according to claim 17 wherein said administering
comprises topical administration of an ophthalmic solution of the
dantrolene to the eye.
21. The method according to claim 17, wherein the dantrolene is in
the form of an aqueous solution, a suspension, a gel, or a jelly.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This is a continuation of U.S. Ser. No. 10/189,676, filed
Jul. 3, 2002, which is incorporated by reference herein in its
entirety.
FIELD OF THE INVENTION
[0002] The present invention relates to neurology and
ophthalmology, and more specifically to protection of neural
tissues from injuries caused by abnormal elevation of intracellular
free calcium through calcium release from intracellular stores
under disease conditions, including stroke, acute brain trauma,
Alzheimer's disease, Parkinson's disease, glaucoma, diabetic
retinopathy, and age-related macular degeneration.
BACKGROUND OF THE INVENTION
[0003] There is compelling evidence that abnormally elevated
intracellular free calcium is one of the early events in the chain
of reactions leading to neuronal damage under pathological
conditions that range from acute neural injuries, such as stroke,
to more chronic indications, such as Alzheimer's disease. High
intracellular free calcium can cause mitochondrial injury and
activate various types of enzymes, such as proteases, nitric oxide
synthases and endonucleases. These calcium-induced/activated
cellular responses are believed to mediate cytotoxicity that
eventually leads to neuronal death.
[0004] There are two major mechanisms that can cause elevation of
intracellular free calcium: 1) calcium influx from extracellular
space through calcium and non-selective cation channels on the cell
membrane, and 2) calcium release from intracellular stores, such as
endoplasmic reticulum and mitochondria, through specialized
receptor-channel complex, such as ryanodine receptor channels.
These two mechanisms often interact. For example, calcium entered
the cell through ion channels on the cell membrane can trigger more
calcium release from intracellular stores. This calcium-induced
calcium release (CICR) has been demonstrated to contribute to
neuronal damage under pathological conditions.
[0005] Glutamate is the major excitatory neurotransmitter in the
brain, including the retina. Its biological action is mediated by a
variety of glutamate receptors, including the NMDA receptor that is
an ionotropic receptor coupled with a non-selective cation channel
that has high calcium permeability. Under pathological conditions,
glutamate becomes a neurotoxin that causes neuronal damage in both
acute neural injuries, such as stroke, to more chronic indications,
such as Alzheimer's disease. This glutamate excitotoxicity is
mediated, to a large extent, by the NMDA receptor because of its
high calcium permeability. Over stimulation of the NMDA receptor
resulting from either excessive release or reduced reuptake of
glutamate causes intracellular calcium overload that can eventually
lead to neuronal death. Calcium entering the neuron through NMDA
channels can stimulate more calcium release from intracellular
stores via specialized ligand-activated channels, such as ryanodine
channels. This calcium-induced calcium release amplifies cellular
response triggered by NMDA receptor activation and has been shown
to contribute to excitotoxicity under pathological conditions.
[0006] Glaucoma is a neurodegenerative retinal disease
characterized by progressive death of retinal ganglion cells (RGCs,
the output neuron of the retina), which leads to progressive vision
loss and eventually to complete blindness. Glaucoma can be
classified into two major categories: hypertensive and
normotensive. The underlying causes for glaucoma are still not well
understood. The initial insults for the two types of glaucoma are
likely different. High intraocular pressure is believed to be a
major risk factor for the hypertensive glaucoma whereas the
vascular abnormality is though to play a significant role in
initiation and progression of the normotensive glaucoma. Despite
the difference in initial insults, progressive death of RGCs
appears to be a common feature shared by both types of
glaucoma.
[0007] There is increasing evidence that glutamate-induced
excitotoxicity plays a significant role in the pathology of
glaucoma. It has been demonstrated that glutamate concentration in
vitreous humor from the glaucoma patients is significantly higher
than that of normal subjects and the vitreal glutamate
concentration increases with the years with glaucoma. It has also
been shown that the NMDA receptor antagonist, memantine,
ameliorates RGC loss in glaucomatous monkeys, suggesting that the
NMDA receptor mediates, at least in part, glutamate-induced damage
to RGCs in glaucoma.
[0008] Diabetic retinopathy is another chronic degenerative retinal
disease that leads progressive vision loss. Recent studies provide
evidence that ischemia and glutamate excitotoxicity contribute to
neural injury in diabetic retinopathy. This suggests that calcium
release from intracellular stores is likely involved in the
pathology of diabetic retinopathy.
[0009] Thus, it is evident that there is an unmet need for agents
that have neuroprotective effects that can stop or retard the
progressive damage to CNS neurons resulting from abnormally
elevated intracellular free calcium caused by various noxious
provocations.
[0010] Dantrolene, a skeletal muscle relaxant, has been found to be
an antagonist of the ryanodine receptor-channel complex (See
Biochemistry 2001, 40, 531-542). Dantrolene blocks calcium release
from ryanodine channels when it binds to the receptor.
[0011] Dantrolene is
1-[[5-(p-Nitrophenyl)furfurylidene]amino]hydantoin.
##STR00001##
SUMMARY OF THE INVENTION
[0012] A new method of protecting the neurons in the retina and
other parts of the brain of a mammal from noxious provocations has
been discovered. The present method uses a ryanodine receptor
antagonist to prevent or ameliorate damage to CNS neurons caused by
noxious provocations that induce excessive calcium release from
intracellular stores via ryanodine receptor channels. These noxious
provocations, including excitotoxicity, ischemia, hypoxia,
mitochondrial dysfunction, and oxidative injury, are associated
with acute and chronic neural disorders, including glaucoma,
diabetic retinopathy, age-related macular degeneration (ARMD),
stroke, acute brain trauma, Alzheimer's disease, Parkinson's
disease, and Huntington's disease. The method comprises
administering to the mammal either systemically, topically,
epidurally or by intrabulbar injection an effective amount of one
or more ryanodine receptor antagonists, such as dantrolene (see
below for details).
[0013] For protection of retinal neurons in humans suffering from
glaucoma, diabetic retinopathy, and age-related macular
degeneration, the active compounds (or mixtures or salts thereof)
are administered in accordance with the present invention to the
eye admixed with an ophthalmically acceptable carrier. Any
suitable, e.g., conventional, ophthalmically acceptable carrier may
be employed. A carrier is ophthalmically acceptable if it has
substantially no long term or permanent detrimental effect on the
eye to which it is administered. Examples of ophthalmically
acceptable carriers include physiological saline and other aqueous
media. In accordance with the invention, the active compounds are
preferably soluble in the carrier which is employed for their
administration, so that the active compounds are administered to
the eye in the form of a solution. Alternatively, a suspension of
the active compound or compounds (or salts thereof) in a suitable
carrier may also be employed.
[0014] In accordance with the invention the active compounds (or
mixtures or salts thereof) are administered in an ophthalmically
acceptable carrier in sufficient concentration so as to deliver an
effective amount of the active compound or compounds to the eye.
Preferably, the ophthalmic, therapeutic solutions contain one or
more of the active compounds in a concentration range of
approximately 0.0001% to approximately 10% (weight by volume) and
more preferably approximately 0.005% to approximately 0.5% (weight
by volume).
[0015] Any method of administering drugs directly to a mammalian
eye may be employed to administer, in accordance with the present
invention, the active compound or compounds to the eye to be
treated. By the term "administering directly" is meant to exclude
those general systemic drug administration modes, e.g., injection
directly into the patient's blood vessels, oral administration and
the like, which result in the compound or compounds being
systemically available. The primary effect on the mammal resulting
from the direct administering of the active compound or compounds
to the mammal's eye is preferably a reduction in intraocular
pressure. More preferably, the active useful compound or compounds
are applied topically to the eye or are injected directly into the
eye. Particularly useful results are obtained when the compound or
compounds are applied topically to the eye in an ophthalmic
solution (ocular drops).
[0016] Topical ophthalmic preparations, for example ocular drops,
gels or creams, are preferred because of ease of application, ease
of dose delivery, and fewer systemic side effects, such as
cardiovascular hypotension. An exemplary topical ophthalmic
formulation is shown below in Table 1. The abbreviation q.s. means
a quantity sufficient to effect the result or to make volume.
TABLE-US-00001 TABLE I Ingredient Amount (% W/V) Active Compound in
accordance about 0.0001 to about 1 with the invention, Preservative
0-0.10 Vehicle 0-40 Tonicity Adjustor 1-10 Buffer 0.01-10 PH
Adjustor q.s pH 4.5-7.5 Antioxidant as needed Purified Water as
needed to make 100%
[0017] Various preservatives may be used in the ophthalmic
preparation described in Table I above. Preferred preservatives
include, but are not limited to, benzalkonium chloride,
chlorobutanol, thimerosal, phenylmercuric acetate, and
phenylmercuric nitrate. Likewise, various preferred vehicles may be
used in such ophthalmic preparation. These vehicles include, but
are not limited to, polyvinyl alcohol, povidone, hydroxypropyl
methyl cellulose, poloxamers, carboxymethyl cellulose, hydroxyethyl
cellulose, and purified water.
[0018] Tonicity adjustors may be added as needed or convenient.
They include, but are not limited to, salts, particularly sodium
chloride, potassium chloride, mannitol, and glycerin, or any other
suitable ophthalmically acceptable tonicity adjustor.
[0019] Various buffers and means for adjusting pH may be used so
long as the resulting preparation is ophthalmically acceptable.
Accordingly, buffers include but are not limited to, acetate
buffers, citrate buffers, phosphate buffers, and borate buffers.
Acids or bases may be used to adjust the pH in these formulations
as needed.
[0020] In a similar vein, ophthalmically acceptable antioxidants
include, but are not limited to, sodium metabisulfite, sodium
thiosulfate, acetylcysteine, butylated hydroxyanisole, and
butylated hydroxytoluene.
[0021] The ophthalmic solution (ocular drops) may be administered
to the mammalian eye as often as necessary to obtain the desired
concentration intravitreally that affords neuroprotection. For
acute neuroprotective effect such as photoprotection in laser
treatment for ARMD, the protective agent would be administered in
advance of the treatment to provide optimal protection during the
laser procedure. For chronic treatments such as in protection of
the retinal ganglion cells against damage from the neuropathic
effects of, for example, glaucoma or dry ARMD, the drug would be
administered as frequently as necessary to maintain desired
intravitreal concentration or range of concentrations at all times.
In other words, the ophthalmic solution (or other formulation)
which contains the ryanodine antagonist as the active ingredient,
is administered to the mammalian eye as often as necessary to
maintain the beneficial neuroprotective effect of the active
ingredient in the eye. Those skilled in the art will recognize that
the frequency of administration depends on the precise nature of
the active ingredient and its concentration in the ophthalmic
formulation. Within these guidelines it is contemplated that the
ophthalmic formulation of the present invention will be
administered to the mammalian eye approximately once or twice
daily.
[0022] This new method is particularly effective when administered
as a prophylactic treatment, i.e. before damage to the nerve has
taken place, or before long-term progression of the disease state,
such as glaucoma, diabetic retinopathy, or ARMD, has taken place.
Without wishing to be held to a particular theory regarding the
role that the compounds of the present invention play in
neuroprotection, applicants hypothesize that the compounds and
methods described inhibit the intracellular Ca+2 release. (See for
example U.S. Pat. No. 5,891,911.)
[0023] Thus it is further contemplated that the compounds of the
present invention can advantageously be used in combination with
compounds that inhibit cell death. Such cell death inhibiting
compounds include NMDA antagonists especially memantine.
DETAILED DESCRIPTION OF THE INVENTION
[0024] The present invention is related to methods of using
ryanodine receptor antagonists to protect CNS neurons, particularly
the retinal neurons, from injuries caused by acute and chronic
noxious provocations. The drawing will first be briefly
described.
DRAWINGS
[0025] FIG. 1 is a bar graph showing the neuroprotective effect of
dantrolene on NMDA-induced injury of retinal ganglion cells in vivo
in rabbits. Intravitreal injection of NMDA caused retinal ganglion
cell loss in control animals. Application of dantrolene ameliorated
NMDA-induced damage to ganglion cells.
[0026] As mentioned above, excessive release of calcium from
intracellular stores under disease conditions is cytotoxic to
neurons. NMDA receptor mediated excitotoxicity is believed to be a
common cause that can trigger excessive calcium release from
intracellular stores in acute and chronic disorders mentioned
above. For example, NMDA receptor antagonist memantine protects
RGCs in glaucomatous monkeys, suggesting that the NMDA receptor
mediates, at least in part, glutamate excitotoxicity in
glaucoma.
[0027] There is strong evidence that damage to CNS neurons often
has two stages: Primary and secondary degeneration. Initially,
direct neuronal insults, such as local ischemia, trauma etc., lead
to degeneration of the affected neurons. However, the associated
pathophysiological and biochemical events occurring in the injured
neurons are probably responsible for the subsequent progressive
(secondary) degeneration of the neighboring neurons that are not
directly affected by the primary insults. These secondary effects
largely determine the long-term functional outcome.
[0028] The immediate injury-induced response strongly influences
the subsequent degenerative response. Treatment that reduces or
attenuates the injury to the primary insults is therefore likely to
generate optimal results by preventing or delaying the secondary
degenerative processes.
[0029] It has now been discovered that neuroprotection is conferred
upon retinal neurons by administration of a ryanodine antagonist,
e.g. dantrolene, to the retina of a mammal within a period prior
to, or following an primary insult to the retinal neurons but prior
to cell death.
[0030] The terms noxious actions or noxious provocations are
defined as an occurrence which is harmful or destructive to a nerve
cell. It is not limited to events extrinsic to the mammal being
treated but includes disease states and pathological occurrences or
events, such as, for example, stroke or heart attack, that are
harmful or destructive to the nerve cell via a chain of events.
Non-limiting examples of noxious actions include: compressive or
mechanical effects or trauma or stress factors, such as glutamate
neurotoxicity, impaired blood flow to the nerves (ischemia) and
with respect to the retina, glaucoma, diabetic retinopathy,
retinitis pigmentosa and age-related macular degeneration.
[0031] Human Dosage and Administration
[0032] The methods of this invention are useful in treating any
mammal, including humans.
[0033] According to this invention, mammals are treated with
pharmaceutically effective amount of a neuroprotective agent for a
period of time and at a time such that noxious provocations do not
kill or permanently damage the nerve cells. Protective agents may
be administered orally, topically to the eye or by any other
appropriate means of delivery described below or known in the
art.
[0034] In accordance with this invention, pharmaceutically
effective amounts of a protective agent can be administered alone
to treat neural injury or to prevent nerve cell death.
Alternatively a protective agent may be administered sequentially
or concurrently with another drug. For example, it may be used with
an antiglaucoma drug, such as a beta-blocker, an alpha.sub.2
agonist, a muscarinic agent such as pilocarpine, a carbonic
anhydrase inhibitor (CAI), or other intraocular pressure (TOP)
lowering drugs. It may also be used with an anti-angiogenesis drug
for the treatment of ARMD and diabetic retinopathy. The most
effective mode of administration and dosage regimen of protective
agent will depend on the type of disease to be treated, the
severity and course of that disease, previous therapy, the
patient's health status, and response to the drug and the judgment
of the treating physician. Generally, the neuroprotective agent
should be administered in a dose to achieve a serum or intravitreal
concentration of 0.01 nM to 5 M. Preferably the neuroprotective
agent is administered prior to injury to the nerve, but can be
administered after injury has occurred with lessened effect.
[0035] Conventional modes of administration and standard dosage
regimens of neuroprotective agents can be used. Optimal dosages for
coadministration of a drug, e.g. an IOP-lowering drug, with a
neuroprotective agent can be determined using methods known in the
art. Dosages of neuroprotective agents may be adjusted to the
individual patient based on the dosage of the drug with which the
agent is coadministered and the response of the patient to the
treatment regimen. The neuroprotective agent may be administered to
the patient at one time or over a series of treatments.
[0036] The agent may be administered locally, e.g. intravitreally
by intrabulbar injection for ocular neuroprotection, or by
intrathecal or epidural administration for spinal protection. Many
of the agents of the invention can be administered systemically,
e.g., orally, or intravenously, or by intramuscular injection.
Additionally, agents for protection of the retina and optic nerve
that are capable of passing through the cornea, and achieving
sufficient concentration in the vitreous humor, may also be
administered topically to the eye.
[0037] The composition used in these therapies may also be in a
variety of forms. These include, for example, solid, semi-solid,
and liquid dosage forms, such as tablets, pills, powders, preserved
or non-preserved liquid solution or suspension, liposomes,
suppositories, injectable and infusible solutions. The compositions
also preferably include conventional pharmaceutically acceptable
carriers which are known to those of skill in the art.
[0038] The following non-limiting examples describe assays and
measurements used in 1) evaluating efficacy of neuroprotecting
agents and 2) selecting ryanodine antagonists other than
dantrolene.
Example 1:
[0039] Experimental Procedure for Measuring Neural Protection in
Rabbit Model.
[0040] To evaluate in vivo neuroprotective effects of dantrolene
(DTL) on NMDA-induced injury of RGCs an imaging method to count
cell numbers at the RGC layer in the isolated retinas was
developed. Briefly, two weeks following intravitreal injection of
vehicle or various test agents, a rabbit was euthanized and the
treated eye was enucleated. A piece of retina (8 mm in diameter)
was cut immediately below the optic nerve head, flat-mounted in a
plastic chamber filled with HEPES-buffered Ames medium, and imaged
at 25 fields in a 5.times.5 array with a 40.times. water immersion
objective using an Olympus microscope (BX50WI) equipped with an
epi-fluorescence unit. The images were taken with a Hamamatsu
C4742-95 digital camera and Image-Pro Plus software (V4.5). The
total number of neurons at the ganglion cell layer in these 25
fields was counted. The same measurements were conducted in one
control group (rabbits treated with vehicle) and 4 test groups
treated with 1) NMDA, 2) dantrolene+NMDA, 3) veratridine (Verat) a
neurotoxin that damages retinal ganglion cells with intracellular
sodium overload, and 4) dantrolene+veratridine. The results from
the 4 test groups are normalized with respect to that of
control.
[0041] The results are reported in FIG. 1. NMDA caused a loss of
53% of cells at the RGC layer. Pretreatment with dantrolene
significantly reduced NMDA-induced cell death to 35%. Dantrolene
also reduced cell loss caused by veratridine from 56% to 50%.
Example 2:
[0042] Assay for Selecting Ryanodine Antagonists other than
Dantrolene.
[0043] Assays for determining ryanodine antagonist may be conducted
following procedures modified from that described by Laver et al.,
(J. Physiol. 537:763-778, 2001). Briefly, purified ryanodine
receptor-channel complexes are incorporated into planar
phospholipid bilayers with resting calcium gradient similar to that
in a normal neuron at rest (100 nM cytoplasmic and 1 mM luminal).
The level of channel activation can be determined in the presence
of various ligands that activate ryanodine receptors. Effective
antagonistic action of the compounds to be selected can be
determined by a reduction of agonist-induced activation of the
channel. The specificity of the antagonists can be determined by
commercially available standard screens, such as NovaScreens.
[0044] While this invention has been described with respect to
various specific examples and embodiments, it is to be understood
that the invention is not limited thereby and should only be
construed by interpretation of the scope of the appended
claims.
* * * * *